key: cord- -f hm v g authors: micklitz, hans-w. title: the covid- threat: an opportunity to rethink the european economic constitution and european private law date: - - journal: nan doi: . /err. . sha: doc_id: cord_uid: f hm v g nan the "political" through the member states. this is exactly what critiques of european neoliberalism were asking for. the nation states are taking action within the boundaries of their national democratic constitutions and the eu legal order. they act through legislative packages to save the lives of their citizens and eu citizens in border regions through the establishment of a safety net for the economy, for employees and for companies. the covid- threat dictates the order of assistancehealth first, then money; society first, the economy second. this order demonstrates and explains the weakness of the eu as a quasi-state with a quasi-constitutional legal order built around enumerated powers and the still-remaining foundational powers of the member states as the guardians of the treaty in terms of the management of societal crises. the eu has no competence-competence. the covid- threat offers legal scholars a unique opportunity to seriously think about the legal order that should govern the society we want to live in and an economy that serves the expectations of people in the post-covid- world. the covid- threat has opened a window of opportunity for transgressing boundaries, for thinking the unthinkable: a fundamental revision of the european economic constitution and therewith european private law. the window will not remain open for long. the longer the shutdown lasts, the stronger the voices will be to return to normal, to bring people back to work, to get the economy going again; and the more the political debate around crisis management turns to its economic implications, the easier it will become for the eu to gain ground again and to integrate national crisis management into the european legal order and into the set of competences the eu has and that allows it to manage the economic effects of the covid- threat. depending on the depth and length of the expected recession, one might even think of a much more radical scenario, one where the member states use the eu as a catalyst to promote a hard-core version of economic neoliberalism where the social and environmental achievements of recent decades are blown away as barriers in order to relaunch the economy. the revival of economics will come. what is therefore needed here and now is a rethinking of the european economic constitution and european private law. in the potential euphoria surrounding new perspectives, it should not be forgotten that the current crisis management by the eu member states relies on the full functioning of key economic sectorsfood production and supply, banking, healthcare, transport, the internet, energymore often than not provided by multinational, transnational corporations, national incumbents and supermarket chains. the internal market has not been suspended, at least not with regards to the cross-border trade in goods and services. what is mainly suspended is personal freedomin eu language, the free movement of persons and non-essential business activities in areas where small and medium-sized companies dominate. there will be lessons to learn as to who will benefit from the crisis -member state politics, national governments, multinationals, online business, transport of goodsand who will suffer: the eu as an institution, the european legal order based on the four freedoms and competition, national parliaments, small and medium-sized companies and non-essential economic sectors. a stocktaking will be needed on what remains of the european legal order after covid- and whether and to what extent the state of emergency the member states have proclaimed could be brought into line with eu law. this is the kind of clean-up that lawyers are accustomed to doing as a repair service for society and the economy. what, then, does rethinkingthinking the unthinkableimply? thinking the unthinkable does not relate to krastev's "after europe" the dissolution of the eu overnight. the majority of european legal scholars might in one way or the other sympathise with baquero cruz, with the deeper historical origins of the "law after auschwitz" (the heading of cruz's chapter ): its peace through trade mission, a rejection of the return to the nation state and the political and theoretical opportunity for a serious and substantial reform of the eu. in rethinking, there is no need to start from scratch. from the s onwards, growing critical legal scholarship has accompanied the eastern enlargement of the eu and the feasibility of a european constitution and of a european civil code. by and large, the critique has fallen on deaf ears: the eastern enlargement was pushed through against warnings about the impact of the "one-size-fits-all" approach on the former socialist economies, let alone the lack of democratic institutional structures and a society that subscribes to liberal democratic values. the failed european constitution was saved in the lisbon treaty, and the failed european civil code was broken down into a bunch of regulations and directives that breathe the same spirit. in the covid- threat, the criticism appears in a different light, not as a laborious attempt to change the direction of european politics, but as a possible scenario for a new beginning. in what follows, i would like to tie three strands of discussion together: the covid- threat, the un sustainable development goals and the circular economy, as well as the digital economy and society. only such a holistic perspective opens up pathways for a new european economic constitution and a new european private law post-covid- . the covid- threat has laid bare the missing dimension of all of the european economic constitution and european private law rhetoric of the last years: the inclusion of the societal dimension of the european integration process into the european legal order. the exclusion of the social impact of the internal market on national societies and on an infant european society builds the first strand of critique that unites philosophers and legal theorists. in his letter to p. van parijs, rawls asks whether "meaningless consumerism" could be a legitimate aim for the eu and for a european society. davies and taguri highlight the societal and cultural implications of ever-increasing consumer choice in the internal market. the one-sided promotion of online business, which empties our cities and transforms physical communication into online orders, is just one visible expression of what is excluded from the toolbox of european lawnot only to investigate potential gains in economic growth through increased efficiency and increased effectiveness of political objectives through fully harmonising european private law rules, but also to seriously look into their impacts on society. what can law contribute to the building of a national society? the eastern enlargement seems to teach the lesson that there are limits to the role of law in post-communist states, where society had been suppressed for over years. this does not mean that european law cannot play a much larger role in the making of a european society. the nonexistence of a fully-fledged european state opens up morenot feweropportunities for the law to enable society-building. this implies understanding the law of the internal market as the law of the internal market society. the addressee of such a legal order is the consumer citizen, the worker citizen, the employer citizen. such an understanding breaks down the fragmentation of the legal person in the eu and establishes a common umbrella for status-related european laws (non-discrimination, labour and consumer law). whether it is feasible to combine citizenship with economic rights is subject to controversy. in an admittedly optimistic reading, i have tried to formulate out of the bits and pieces of european private law de lege lata a tripartite distinction between universal services, the law of the labour market society and consumer market society and societal private law. the second potential for rewriting the european economic constitution is the everlouder call for sustainability, which reached the eu through adoption of the circular economy in . the covid- threat provides the earth with breathing space. the juncker commission has brought to a "successful" end a whole range of regulations and directives that cover broad fields of economic and private law. these newly adopted legislative measures follow the internal market philosophy, the rationality of market efficiency piggy-backing on social regulations on consumer and environmental protection. they are united in that sustainability and the circular economy are downgraded to a few window-dressing references in the recitals without any conceptual rethinking, let alone re-conceptualising. sustainability goals translated into the circular economy are kept strictly separated from economic (internal market) and private law. there is no attempt to initiate a debate that the un sustainable development goals and the circular economy, if taken seriously, might require a different economic constitution and a different private law. what is urgently needed is a serious stocktaking of the state of affairs in terms of european private law in the light of the sustainable development goals that highlight the dark side of one-sided rules that protect employees and consumers without taking their counterproductive effects on the environment into account. in his "many constitutions", k. tuori distinguishes between the framing of juridical and political constitutions and the sectoral constitutions: micro-and macro-economic, social and security constitutions. neither he nor von bogdandy and bast in their compendium mention the possible existence of an environmental/sustainable constitution. the bits and pieces enshrined in article of the treaty on the functioning of the european union (tfeu) and article tfeu render it difficult if not impossible to deduce concrete consequences of the positioning of the environment in relation to the micro-and macro-economic constitution. here is the place for a political commitment by the member states to give shape to a new european economic constitution. the covid- threat is supposed to strengthen the transformation of the nation state into a steering state -"l'État providence" in the format of the precautionary state. the eu member states will have to make sure that the healthcare sector will never again suffer from shortages in life-saving medical devices and pharmaceuticals and that essential industries that are indispensable for fighting the next pandemic threat are located within the eu if not on national territory. the transformation will affect the european economic constitution and the interplay between private entrepreneurship and de facto nationalised industrial sectors. the huge safety net that the member states are currently building with taxpayers' money is the means through which the eu member states intervene in the economy. the very same means could and should be understood as an opportunity to build a sustainable economy that deserves its name. financial aid could be bound to sustainability. the overall assumption is that the covid- threat will boost the digitisation of society and the economy in personal and business communication, in education at schools and universities and in online and offline business. the private law of contracts and private regulation through contracts, codes and standards beyond the nation state enables and promotes digitisation. the eu is at the forefront in setting legal standards through the general data protection regulation, the copyright directive, the platform-to-business regulation and, last but not least, through the directive on digital content. the rationale is again to promote economic growth in the internal (digital) market through digitisation. the bits and pieces are pillars of an emerging european digital constitution in which the proclaimed human-centred artificial intelligence risks falling by the wayside. the post-covid- wonderland should not be entered without a deeper understanding of the political economy behind digitisation. digitisation will be used ever more strongly to consider if, how and to what extent algorithms, machine learning techniques and neuronal nets can reduce the social costs of labour, which will have a huge impact on our societies. the covid- threat will accelerate and legitimise the digital transformation of society and the economy. however, the very same crisis must be seized as an opportunity to make sure that technology serves, in the words of brownsword, "the anterior conditions for humans to exist and for them to function as a community of agents". in the scholarly debate on digitisation and the impact of economic and private law, there seems to be agreement that red lines are needed that cannot be crossed by anyone and that should not be submitted to a balancing test. the digital constitution is supposed to set benchmarks for all sorts of european, national and private regulation that governs private relations in the digital society and economy. who is the addressee? can the eu take over the proclaimed stewardship responsibility? zuboff stressed a move towards surveillance capitalism; the covid- threat presents the risk of a surveillance state. it is certainly not enough to expect red lines to be established by states or by the eu. much will depend on pressure from civil society and from critical legal scholarship to develop models that could be translated into legislative programmes on whether the lex digitalis can be turned into a joint exercise of a transnational community. this window of opportunity can only be fully exploited if the three strands of discussion are brought together in the design of a post-covid- european economic constitution and european private law. the working programme of the european commission, adopted in january , aims at bringing sustainability and digitisation together, without stressing the need, however, to involve national societies and the infant european society. the covid- threat provides critical legal scholarship with a unique opportunity, despite all of the current and future uncertainties and speculations, to move far beyond the mainstream understanding of the economic constitution and private law. squaring the circle? reconciling consumer law and the circular economy the many constitutions of europe principles of european constitutional law there is an abundant literature on the deficits in the lisbon treaty; see a kiss environment in the future european constitution der staat wird zum vorsorgestaat, frankfurter allgemeine zeitung the role of technology in political economy the global governance of knowledge: patent offices and their clients legal tech book series: tomorrow's lawyers by responsible regulation: prudence, precaution and stewardship the age of surveillance capitalism: the fight for a human future at the new frontier of power political disruption, technological disruption, and the future of eu private law big other: surveillance capitalism and the prospects of an information civilization key: cord- -z sxr dx authors: coveri, andrea; cozza, claudio; nascia, leopoldo; zanfei, antonello title: supply chain contagion and the role of industrial policy date: - - journal: j doi: . /s - - - sha: doc_id: cord_uid: z sxr dx the covid- pandemic triggered a major disruption in global value chains (gvcs) that pushed the global economy into a recession that promises to be worse than the crisis. this article illustrates the mechanisms through which the covid- pandemic affected gvcs in the context of a changing configuration of the global economy. in particular, it is argued that gvcs became the main transmission channels of “economic contagion”. finally, we posit that the pandemic provides an opportunity to revive the role of industrial policy as to govern the landslides of a world economy constantly pressured by globalization and deglobalization forces. the diffusion of covid- represents the most serious pandemic since the spanish flu over the - period. after the outbreak in china in the second half of february , the pandemic spread worldwide in a few weeks. the large lockdown of production that followed has immediately shown its disruptive impact and promises to trigger the most serious economic crisis since the great recession of . among the most affected countries are in fact the largest industrialized economies in the world, primarily the united states, spain, italy, germany, the united kingdom and france, as well as china itself and japan, responsible overall for over % of global gdp (baldwin and tomiura ) . this short contribution illustrates the impact of the covid- pandemic on the current configuration of global economy (sect. ), stressing the role of global value chains (gvcs) as the main transmission channels of "economic contagion" worldwide (sect. ). in this context, we provide a focus on italy, which has been the first economy to be affected in europe, and most substantially. section highlights the challenges imposed by the pandemic and discusses the key role that industrial policy should play to set the global economy on a more resilient and sustainable path. finally, the last section draws some conclusions. supply-and demand-side contagion channels pandemic occurs in the midst of a transformation that has been characterizing the world economy over the last four decades. since the s, a series of technological, political and institutional factors gave rise to what baldwin ( ) called the " nd unbundling", meaning a powerful push towards the geographical dispersion of value-added tasks required to get the realization of final products. this vertical fragmentation of production on a global scale has given rise to global value chains (gvcs), along which east asia and china in particular represent a crucial node (baldwin ; gereffi and fernandez-stark ; zanfei et al. ) . in other words, the sophisticated interconnection of the productive structures of the different countries at the international level led the interdependence of the economies to increase remarkably. as a consequence, a production bottleneck in a hub of the dislocated supply chain can trigger a synchronized economic slowdown and, therefore, a global downturn (baldwin and weder di mauro ; boehm et al. ; inoue and todo ; sforza and steininger ; unctad ) . therefore, the first channel through which the pandemic has affected the global economy concerns the interruption of the supply chains, hitting what has been identified as the productive heart of the world, that is east asia, and china in particular (buckley and strange ) . in this regard, consider that chinese industrial production has fallen by . % on an annual basis between january and february , i.e. the biggest drop in production in china since the post-deng xiaoping era, especially in the transport equipment (− . %), general equipment (− . %), textiles (− . %) and machinery (− . %) (data from national bureau of statistics of china). the second, and related, channel concerns the amplification effect of the contagion induced by the chains of global subcontracting of intermediate goods, since even the manufacturing sectors of the least affected countries have had difficulty in acquiring (importing) the intermediate inputs necessary for domestic production (baldwin and tomiura ; bonadio et al. ) . in this context, the loosening of the production lockdown in china (in march), while it was being strengthened in europe and the united states (in april), adds further complexity to this scenario. on the one hand, the attempts of western importing companies to reduce their dependence on intermediate goods from china, the so-called decoupling, might turn out a bad strategy, as china is now recovering faster than the rest of world. on the other hand, the global spread of covid- is ultimately harmful also for china, as it imports a large part of the intermediate goods necessary for production for its domestic market, as well as for foreign markets (world bank ). in addition, the demand-side of the gvcs-pandemic nexus should not be underestimated. firstly, governments' measures aimed at containing the virus, such as the drastic reduction of people mobility and the shutdown of almost all commercial and leisure activities, entail an immediate reduction of consumptions. some of them will be postponed but many others will be probably never recovered. the heaviest effects will concern the services sector, in particular transport, tourism, accommodation and catering sector, thus affecting some countries (e.g. italy) more than others. secondly, the sharp slowdown in production is expected to generate an increase in the unemployment rate, which will result in a reduction in households' disposable income, starting with those who are hired on a temporary basis. if this reduction mainly affects the poorer segments of the population-those with a higher marginal propensity to consume-the consequences on aggregate consumption will be even more significant. in addition, due to the uncertainty caused by the speed and spread of the infection, it is reasonable to expect a reduction in investment rates and an increase in precautionary saving as a form of protection for a dark future. thirdly, a lower utilization rate of production capacity by firms could make it more difficult for them to amortize fixed costs (a trivial example is the rental cost of buildings). this in turn could lead to an increase in unit costs, a reduction in the profit rates and therefore a further contraction in investment expenditure. finally, the global reduction in consumption and investments amplifies the value added contraction simultaneously, further restricting foreign outlet markets and therefore slowing down the dynamics of net exports of both final and intermediate goods. although our focus is on economic consequences of the pandemic, it might be worth mentioning an important feedback effect. in fact, covid- may not only affect gvcs, but its diffusion might have been favored by the increasing resort to international production networks. one often emphasized mechanism is through increase in pollution. some scientists suggest that industrial production is altering ecosystems and, above all, reducing biodiversity (wallace ) . this reduction would result in a greater probability of the cross-species transmission of viruses. however, the probability of virus transmission is not only likely to be connected to industrial production in general, but more specifically to the modern hyper-globalized world economy. this link from gvcs to covid- holds for at least two reasons. first, consider the extent to which the gvcs have increased the concentration of manufacturing production in certain geographical areas, particularly in east asia but also in latin america. this concentration is accompanied by an accentuated specialization of such regions in the most energy-intensive manufacturing stages of production, taking advantage of cost reductions with little consideration of environmental issues. this is often due to the very low cost of local labor, which in turn depends on the large pool of workers in emerging countries and the less restrictive legislation compared to western economies. environmental regulation is also less restrictive than in other parts of the world, attracting polluting manufacturing activities which could contribute to increasing the likelihood of new viruses. second, the logic of gvcs meant a transnational dislocation of production which has at least partially required greater workers' mobility. given that gvcs are dominated by global actors, the international mobility of their workers, especially managers of multinational corporations (mncs), has likely played a role on the spread of the virus. although anecdotal, the first detected cases of "patient one" in germany, a manager infected by a colleague from shanghai who allegedly brought the virus to germany on january , (rothe et al. ) ; or the circumstance that the first case in the italian village of codogno was a manager of unilever, are emblematic. the shock caused by covid- pandemic occurred at a time when the global economy was already on the verge of a recession. figure reports the progressive reduction in the gdp growth rates of major economic regions-especially european ones-from to along with the imf growth projection to (imf ). according to the latter, the outbreak of the pandemic will lead to a global gdp contraction of % for . notably, the world gdp contraction during the great recession in was equal to . %. ( ) with regard to the european union, imf ( ) estimates a . % reduction in gdp, with italy called to suffer the biggest backlash: a . % decrease with respect to the previous year. to fully acknowledge the magnitude, consider that in , i.e. the worst year of the past global financial crisis, italy's gdp fell by just under % and at that time the unemployment rate in italy soared to almost % at the end of (data from oecd). moreover, preliminary data provided by the italian national institute of statistics (istat) show a decrease of . % in industrial production in march as compared to february for italy-much worse than in other industrialized countries, as reported in fig. -with a drop of . % in durable consumption goods and of . % in capital goods on a yearly basis. in addition, preliminary estimations provided by prometeia report that the pandemic would lead to an overall reduction of the italian industrial production for march and april equal to % . as stressed in the previous section, the strong interdependence of the economies has been crucial in spreading the economic contagion worldwide, following a domino scheme. in this context, the fact that china was the first economy affected by the covid- represented a factor of paramount importance because of its central role in modern global production networks. an empirical illustration of this phenomenon is shown by fig. , which reports the backward participation of prominent industrialized countries with respect to china for , and . this index is often used to approximate the level of international fragmentation of production, as it measures the percentage share of added value produced abroad (in china in this case) which is contained in the gross exports of each individual country over the total gross exports of the latter. in other words, this indicator measures to what extent the exports of countries under observation depend on the import of intermediate goods from china. as fig. focusing on italy, it is worth mentioning that the industries showing the highest level of backward participation with respect to china report much higher and increasing values over time, which in spanned from . % for machinery and equipment to . % for textiles and wearing apparel sector, up to a level equal to . % for computer and electronic and . % for electrical equipment industry (data from oecd-tiva). unsurprisingly, with the outbreak of the pandemic, this complex global interconnection of production-and the crucial role that the "factory of the world" plays in this context-has prompted part of the economic literature to warn about the need for many countries to promote supplier diversification with the aim of reducing their dependence on imports of intermediate goods from china (javorcik ; monga ). it is worth noting that, on the one hand, this diversification had been to a small extent already implemented by some mncs, which had begun to diversify their supply sources in response to trade restrictions due to the trade war between the united states and china. in particular, some mncs have moved production to other countries mainly in south east asia and/or have undertaken reshoring policies-i.e. return of production back to the country of origin from overseas-a phenomenon we will briefly discuss below. on the other hand, it should not be forgotten that the breadth of the chinese market makes the complete abandonment of the asian giant very risky and unprofitable for many companies. however, the italian backward linkages are not limited to asian-centered production networks. although the latter are gaining ever more prominence over time, the italian industrial fabric shows a higher level of integration in european and north american gvcs (wto ). indeed, italy reported an overall backward participation equal to . % in , of which the chinese share accounted for less than % (data from oecd-tiva). it follows that the success in fighting the contagion by china since march has only alleviated the gvcs-disruption burden on several italian production activities. in fact, after being imposed in china, the lockdown hit even harder european economies and the us, with respect to which italy shows an even stronger vertical integration. in this regard, fig. shows the italian industries with the highest level of backward participation, i.e. those relying the most on foreign production, with the exclusion of the manufacturing industry of coke and refined petroleum products (whose backward index reaches almost % in ). the figure reports clear-cut evidence about the crucial role of imports from foreign suppliers for the production activity of key italian industries such as basic metals, automotive, electrical equipment as well as chemicals and pharmaceutical products; for these sectors the amount of foreign value added the italian firms process to export products to third countries is more than % of their gross exports. in other words, and beyond what happens in china, if the world stops because of covid- pandemic, italy can only suffer highly negative consequences, at least in the short period. notably, the evolution of the backward participation index of the italian automotive industry (sector ) is the one showing the most marked increase from to , which goes from to %. this growing trend mirrors the offshoring of manufacturing stages of production increasingly carried out by a share of italian firms towards central and eastern european countries (ceecs) since the nineties (celi et al. ) . furthermore, the growing vertical fragmentation of the automotive sector and the changing position of italy in the global production network related to motor vehicles represents a good example to illustrate the potential implications of the pandemic on italy's economy. especially after the great recession of , the italian automotive industry has clearly made a shift in gvcs, playing a greater role as a global supplier of components for car industry, while losing positions in the final markets of cars (fig. ) . although the car production is still important in the national economy, employing around , workers in firms in , it exhibits a lower export propensity than the car electrical components industry. in particular, after , the latter largely increased its share of exports, shifting the functional role of the italian automotive industry into gvcs and increasing the country's specialization in the production of some parts of the automotive production chain, namely electrical components and bodies for motor vehicles. the traditional car industry, which used to be highly concentrated and dependent upon the italian domestic market, has thus moved towards a specialization pattern more and more focused on the exports of motor vehicles components, while the national production of cars is stable if not declining (being its volume in equal to the one in ). hence, as the world stops because of pandemic, the italian car industry suffers the interruption of purchases from foreign markets of its components while the home market has diminished its capacity to absorb them, due to the relatively low growth of its assembly activities. quite symmetrically, the lockdown firstly imposed by italian government has represented inevitably a supply chain disruption also for the other countries relying on import of intermediate goods from italy for production, exacerbating the economic crisis. alongside the vulnerabilities of gvcs highlighted by the pandemic, in italy the latter also showed how the austerity policies pursued in the last decade have made the healthcare system unable to successfully deal with emergencies (prante et al. ) . although an ageing population requires larger health expenditures, the public health system shrank. several healthcare profiles, like doctors and nurses, had to deal with a labour market with few professional opportunities and highly relying on temporary positions due to the outsourcing of health services to the private sector, especially in lombardy-i.e. the italian region most affected by covid- . the pandemic has thus revealed the shortage of health personnel, of medical infrastructures like intensive care units (icu) and of individual protection devices (ipds), like face masks for the medical staff (ranney et al. ). in this context, the pandemic caused a further reduction of the available medical staff due to a large amount of contagions because of the individual protection devices shortage, pushing the italian government to set up a call to scale up the medical staff. in addition, the restriction on international trade and the export limits imposed by several governments, aimed at ensuring national self-sufficiency, partially disrupted the supply chain of medical devices. although the medical industry was not included into the lockdown policy due to its relevance for the emergency, offshoring of relevant segments of the manufacturing chain by italian firms might have reduced the sourcing capacity of key medical equipment. the pandemic also highlights how the extension of gvcs impacts on the "knowledge chain". in fact, the covid- emergency has shown that de-specializing in some key segments of healthcare equipment manufacturing is detrimental to the overall efficiency of the system and to its resilience in particular. for example, only a few italian firms still have the knowledge to rapidly set up fully equipped icus if necessary, despite the long lasting italian industrial tradition in the biomedical sector. another example is provided by the united kingdom; the country's long-term deindustrialisation process made it very difficult to cope with the pandemic. the prime minister boris johnson had to struggle to find national companies able to manufacture icus, leading him to negotiate with the automotive industry. at the same time, he faced a shortage of personal protective equipment and of health workers, who are essential for managing any icu. the pandemic has thus highlighted the fragilities of the vertical disintegration of production across national borders, questioning at least partially the merits of fragmenting supply chains occurred in the last three decades. from this perspective, the global diffusion of covid- is likely to contribute to a reduction of trade in gvc, a process that has already been recorded in the aftermath of the great recession of , giving rise to a number of studies on production reshoring and back-shoring phenomena (rodrik ) . there are several motivations underlying such a re-thinking of gvcs. first, a sluggish global demand and market saturation phenomena reduced the (domestic and cross-border) investment opportunities for firms. second, china's involvement in the global production networks appears to have decreased over the past decade-as documented by the sharp fall in backward participation of the country from . % in to . % in (data from oecd-tiva). third, reshoring and back-shoring strategies have become more common for mncs, as a reaction to: fewer arbitrage opportunities on labour cost; higher coordination costs associated to global networking; and, last but not least, the diffusion of digital production technologies, making it convenient to re-internalize production without incurring in higher costs (dachs and zanker ; dachs et al. ; ilo ; seric and winkler ) . alongside these factors, policy-driven anti-globalization forces as brexit, trade wars and protectionist measure by the us have likely played a role (dachs and seric ) . in this scenario, the challenge for a new industrial policy concerns the capability to govern the "landslides" of a world economy constantly pressured by globalization and deglobalization forces. in other terms, leaving aside calls for hyper-globalization on one side and national self-sufficiency on the other, the mission of a modern industrial policy should be the consolidation of resilient gvcs on a reduced, largely regional, scale to pursue common development objectives. the strategy we are suggesting is not so different from what china is promoting. the asian country in the last decade addressed huge public and private resources to diversify and upgrade its production matrix, moving from low-to high-value added segments of production while building an ever more complete value chain within the south-east asian borders (european union chamber of commerce in china ). this allowed china to partially shorten its gvcs and its dependence upon imports-especially for those inputs it strongly needed in the first times of its industrialisation-without giving up the productivity gains from specialization and trade (de backer et al. ; duan et al. ; stollinger ) . in doing this, china has been building brand new production capacity with the aim of expanding and diversifying its productive matrix and enter in global markets from which chinese firms were absent until a couple of decades ago. in front of this competitive challenge, the european economy needs to simultaneously combine greater resilience of the productive system to major disruptive events as pandemics, with a strengthened international competition of its firms on global markets. we suggest that this will inevitably require a vast investment plan at the continental level. more precisely, we argue that a well-targeted european industrial policy aimed at consolidating its continental supply chains could take four birds with one stone. first, aiming to fill in the gaps-even expanded by the pandemic-along its continental value chains, the european economy would enjoy the efficiency gains arising from the division of labour while fundamentally augmenting the resilience of its supply chains. this would be ever more important with regard to strategic productions, reason why europe should act cooperatively sharing resources, knowledge and managing capabilities in industries like health, food supply, infrastructure, environmental and renewable energy and social security (see also pianta et al. ) . moreover, the pandemic-induced reorganization of the global industrial structure could be an opportunity for the selection and evolution of small-and medium-size enterprises (juergensen et al. ) , also through their repositioning into shorter supply chains marked by lower power asymmetries. second, such european industrial plan could favour economic convergence among country members, finally closing the long-term structural asymmetries which triggered the european debt crisis of - and doomed europe to a sluggish recover in the following years (celi et al. ) . it follows that investments needed to pursue such industrial strategy should be firstly addressed towards those european countries which in the last decades have suffered the most the progressive impoverishment of their production matrix (celi et al. ; landesmann and stollinger ; lucchese and pianta a) . from this perspective, the consolidation of a european block of converging and well-integrated economies, effectively participating in regional level gvcs, could constitute a stronger basis to selectively develop intercontinental production networks. the latter could be aimed at gaining access to valuable resources and knowledge assets that might not be available and cost effectively developed within one region, and that can be conveniently sought elsewhere. european economies would then benefit from specialization and trade at a wider scale, while exerting a greater bargaining power in their effort to access such resources. third, while the hyper-specialization of national economies in a few phases of the value chain increases the mutual dependence of economies-up to exposing single countries to whatever shock occurring wherever in the global economy-it also risks to make it more difficult to convert domestic production in case of shortage of critical supplies. the industrial policy design we are advancing should aim to reduce such interdependence while increasing the responsiveness and adaptation of the productive system to both internal and external conditions. finally, an industrial policy aimed at strengthening the position of european national industries in the value chains of strategic productions could help reduce the social and economic costs and risks of hyper-specialization. in fact, a draw-back of the extreme fragmentation of production is that it has reduced the incentives of a part of firms to make investments for the development of those value chain activities-other than core competencies-which appeared too risky, not enough profitable, and largely offshorable (antràs ; dottling et al. ; gutiérrez and philippon ; hummels and klenow ) . this has reduced their knowledge base while decreasing the resilience of european production networks. in fact, during the booming period of the international fragmentation of production and of greater expansion of gvcs, i.e. at the beginning of the xxi century with the entry of china in the global capitalistic market, the european economy witnessed a stagnant dynamics of investment in fixed assets, both tangible and intangible (arrighetti ; pisano and shih ). the hyper-specialization of firms thus produced an incentive to outsourcing (and geographical dispersion of) the other value chain business activities (baldwin and gu ; bernard et al. ) . the consequence has been that also fixed capital investments have been concentrated on a increasingly narrow production matrix, causing a reduction of their productive capacity and of the pool of knowledge on which their ability to compete was built on. such a dynamics points out a structural shortcoming in the process of cross-border vertical disintegration of production: not only key gaps in value chains involving strategic industries emerge, but the whole evolution of the technological matrix of european companies comes out depressed. furthermore, all that goes hand in hand with stagnant productivity, weak employment dynamics and an impoverishment of the technological base of european companies vis-à-vis emerging companies overseas. this further clarifies the necessity and feasibility of a continental industrial policy plan aimed at recovering and expanding the technological matrix of companies and at the same time filling the gaps in the value chains involving strategic productions. these considerations suggest that current tendencies of globalisation and gvcs should not be addressed with a "tina" (there is no alternative) slogan; the same slogan which contributed to the acceleration of globalisation itself over the s. conversely, alternatives to overcome the weaknesses of modern global economy exist, as also mentioned in a world investment report of a few years ago: "are active promotion of gvcs and gvc-led development strategies the only available options or are there alternatives? [the alternative] is an industrial development strategy aimed at building domestic productive capacity, including for exports, in all stages of production […] to develop a vertically integrated industry that remains relatively independent from the key actors of gvcs for its learning and upgrading processes. […] it thus appears that countries can make a strategic choice whether to promote or not to promote gvc participation. to do so, they need to carefully weigh the pros and cons of gvc participation" (unctad , p. ). in this contribution we illustrated the supply-and demand-side mechanisms through which the covid- pandemic has triggered the economic contagion which led to a sharp reduction of production worldwide. we showed that between the pandemic and globalization a close and multi-faceted relationship exists. on the one hand, the outbreak of the pandemic has an impact on global production with a domino effect. on the other hand, global production has played a role in the amplification of the pandemic, especially accelerating its spread up to a rate never experienced before. further, we provided empirical evidence about the crucial role of gvcs in bringing about the leap forward in the level of interdependence of the economies on global scale. most of this empirical evidence concerns the impact of the pandemic in italy, one of the most affected countries. in this context, we underlined the weaknesses of italian participation in gvcs as a pivotal factor in causing the collapse of production. the debate about the future trend of international production and gvcs is open (eiu ; lucchese and pianta b; seric and winkler ; strange ). many economists support the idea of an even increasing expansion of gvcs, as their widespread diversification might help reducing the global impact of national shocks (baldwin and evenett ; baldwin and weder di mauro ; monga ). however, the idea that "firms are eager, willing and able to meet any demand" (baldwin ) proved to be at least over-optimistic. conversely, one can quite convincingly argue that the pandemic has provided further reasons to rethink the pros and cons of gvcs, assessing the merits of cross-border production networks while highlighting the structural shortcomings of extreme specialization. we suggest leaving aside calls for hyper-globalization on one side and national self-sufficiency on the other, arguing for a european industrial policy plan targeted to the consolidation of resilient gvcs on a continental scale to pursue common development goals. such regional/continental level production networks-partially resembling attempts made by china to coordinate production within the south east asian region-could provide a less fragile basis for the development of more selective, strategically oriented, intercontinental networks. at the same time, a european industrial policy which goes in this direction would significantly increase the growth rate of capital formation and expand the matrix of technology used, two tendencies which appear at odds with the productive hyper-specialization induced by the modern global fragmentation of production. crisi di offerta e crescita senza investimenti, siepi.org the impact of trade on plant scale, production-run length, and diversification global supply chains: why they 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what policymakers should be doing about it big farms make big flu: dispatches on influenza, agribusiness, and the nature of science world development report : trading for development in the age of global value chains global value chain development report : technological innovation, supply chain trade, and workers in a globalized world fdi patterns and global value chains in the digital economy acknowledgements open access funding provided by università degli studi di urbino carlo bo within key: cord- -q zlgwy authors: pang, raymond ka-kay; granados, oscar; chhajer, harsh; legara, erika fille title: an analysis of network filtering methods to sovereign bond yields during covid- date: - - journal: nan doi: nan sha: doc_id: cord_uid: q zlgwy in this work, we investigate the impact of the covid- pandemic on sovereign bond yields amongst european countries. we consider the temporal changes from financial correlations using network filtering methods. these methods consider a subset of links within the correlation matrix, which gives rise to a network structure. we use sovereign bond yield data from european countries between the and period as an indicator of the economic health of countries. we find that the average correlation between sovereign bonds within the covid- period decreases, from the peak observed in the - period, where this trend is also reflected in all network filtering methods. we also find variations between the movements of different network filtering methods under various network measures. the novel coronavirus disease (covid- ) epidemic caused by sars-cov- began in china in december and rapidly spread around the world. the confirmed cases increased in different cities of china, japan, and south korea in a few days of early january , but spread globally with new cases in iran, spain, and italy within the middle of february. we focus on sovereign bonds during the covid- period to highlight the extent to which the pandemic has influenced the financial markets. in the last few years, bond yields across the euro-zone were decreasing under a range of european central bank (ecb) interventions, and overall remained stable compared with the german bund, a benchmark used for european sovereign bonds. these movements were disrupted during the covid- pandemic, which has affected the future trajectory of bond yields from highly impacted countries, e.g., spain and italy. however, in the last months, the european central banks intervened in financial and monetary markets to consolidate stability through an adequate supply of liquidity countering the possible margin calls and the risks of different markets and payment systems. these interventions played a specific role in sovereign bonds because, on the one side, supported the stability of financial markets and, on the other side, supported the governments' financial stability and developed a global reference interest rate scheme. understanding how correlations now differ and similarities observed in previous financial events are important in dealing with the future economic effects of covid- . we consider an analysis of sovereign bonds by using network filtering methods, which is part of a growing literature within the area of econophysics [ , , , , ] . the advantages in using filtering methods is the extraction of a network type structure from the financial correlations between sovereign bonds, which allows the properties of centrality and clustering to be considered. in consequence, the correlation-based networks and hierarchical clustering methodologies allow us to understand the nature of financial markets and some features of sovereign bonds. it is not clear which approach should be used in analyzing sovereign bond yields, and so within this paper, we implement various filtering methods to the sovereign bond yield data and compare the resulting structure of different networks. our analysis shows that over the last decade, the mean correlation peaks in october and then decreases during the period, when covid- is most active in europe. these dynamics are reflected across all network filtering methods and represent the wide impact of covid- towards the spectrum of correlations, compared to previous financial events. we consider the network centrality of sovereign bonds within the covid- period, which remains consistent with previous years. these trends are distinctive between filtering methods and stem from the nature of correlations towards economic factors e.g., positive correlations show a stable trend in the individual centrality, compared with the volatile trends for negative correlations, where central nodes within these networks are less integrated in the euro-area. although there is a change in the magnitude of correlations, the overall structure relative to the central node is maintained within the covid- period. previous studies have used different methods to analyze historic correlations as random matrix theory to identify the distribution of eigenvalues concerning financial correlations [ , , ] , the approaches from information theory in exploring the uncertainty within the financial system [ , ] , multilayer network methods [ , , , , , ] , and filtering methods. several authors have used network filtering methods to explain financial structures [ , ] , hierarchy and networks in financial markets [ ] , relations between financial markets and real economy [ ] , volatility [ ] , interest rates [ ] , stock markets [ , , , ] , future markets [ ] or topological dynamics [ ] to list a few. also, the comparison of filtering methods to market data has been used for financial instruments. birch, et al [ ] consider a comparison of filtering methods of the dax stocks. musmeci, et al [ ] propose a multiplex visual network approach and consider data of multiple stock indexes. kukreti, et al [ ] use the s&p market data and incorporate entropy measures with a range of network filtering methods. aste, et al [ ] apply a comparison of network filtering methods on the us equity market data and assess the dynamics using network measures. in order to evaluate the european sovereign bonds, based on filtering methods, this work is organized as follows. in section , we describe the network filtering methods and present the data sets with some preliminary empirical analyses. we apply in section the filtering methods to sovereign bond yields and analyze the trend of financial correlations over the last decade and consider aspects of the network topology. we construct plots in section representing the covid- period for all methods and analyze the clustering between countries. in section , we discuss the results and future directions. we introduce a range of network filtering methods and consider a framework as in [ ] for sovereign bond yields. we define n ∈ n to be the number of sovereign bonds and bond yields y i (t) of the ith sovereign bond at time-t, where i ∈ { , ..., n}. the correlation coefficients r ij (t) ∈ [− , ] are defined using pearson correlation as: with · denoting the average of yield values. the notion of distance d ij ∈ [ , ] considers the values of the entries r ij of the correlation matrix r ∈ [− , ] n×n , with d ij = ( − r ij ). a distance of d ij = represents perfectly positive correlations and d ij = represents bonds with negative correlations. the network filtering methods are then applied to the distance matrix d ∈ [ , ] n×n , where a subset of links (or edges) are chosen under each filtering method. the set of edges is indicated by {(i, j) ∈ e(t) : nodes i and j are connected} at time-t, defined for each filtering method. we define the time frames of financial correlations as x for the set of observations, with n different columns and t rows. from the set of observations x, we consider windows of length , which is equal to six months of data values. we then displace δ windows by data points, which is equal to two weeks of data values, and discard previous observations until all data points are used. by displacing the data in this way, we can examine a time series trend between each window x. we verify the statistical reliability of correlations by using a non-parametric bootstrapping approach as in efron [ ] , which is used in tumminello, et al [ , ] . we randomly choose rows equal in number to the window length t , allowing repeated rows to be chosen. we compute the correlation matrix for this window x * m and repeat the procedure until m samples are generated, which is chosen at , . the error between data points described in efron [ ] is equal to ( − ρ )/t , where highly positive and negative correlated values ρ have the smallest errors. the minimum spanning tree (mst) method is a widely known approach which has been used within currency markets [ ] , stocks markets [ , ] and sovereign bond yields [ ] . the mst from table considers the smallest edges and prioritizes connections of high correlation to form a connected and undirected tree network. this approach can be constructed from a greedy type algorithm e.g. kruskal's and prim's algorithm and satisfies the properties of subdominant ultrametric distance i.e, d ij ≤ max{d ik , d kj } ∀i, j, k ∈ { , ..., n}. a maximum spanning tree (mast) constructs a connected and undirected tree network with n − edges in maximizing the total edge weight. analyses involving mast have been used as comparisons to results seen within mst approaches [ , ] . an mast approach is informative for connections of perfectly anti-correlation between nodes, which are not observed within the mst. a network formed from asset graphs (ag) considers positive correlations between nodes of a given threshold. within the mst, some links of positive correlation are not considered in order to satisfy the properties of the tree network. all n − highest correlations are considered in an ag, allowing for the formation of cliques not observed within a mst or mast network. the use of ag has been considered in onnela, et al [ ] , which identifies clustering within stock market data. as the method only considers n − links, some nodes within the ag may not be connected minimum spanning tree (mst) n − [ ] a connected and undirected network for n nodes which minimizes the total edge weight. maximum spanning tree (mast) n − [ ] a connected and undirected network for n nodes which maximizes the total edge weight. asset graph (ag) n − [ ] choose the smallest n− edges from the distance matrix. triangulated maximal filtering graph (tmfg) [ ] a planar filtered graph under an assigned objective function. for the given threshold and therefore the connection of unconnected nodes is unknown, relative to connected components. the triangulated maximal filtering graph (tmfg) constructs a network of (n − ) fixed edges for n nodes, similar to the planar maximal filtered graph (pmfg) [ ] , which has been used to analyze us stock trends [ ] . the algorithm initially chooses a clique of nodes, where edges are then added sequentially, in order to optimize the objective function e.g., the total edge weight of the network, until all nodes are connected. this approach is non-greedy in choosing edges and incorporates the formation of cliques within the network structure. a tmfg is also an approximate solution to the weighted planar maximal graph problem, and is computationally faster than the pmfg. the resulting network includes more information about the correlation matrix compared with spanning tree approaches, while still maintaining a level of sparsity between nodes. the european sovereign debt has evolved in the last ten years, with some situations affecting the convergence between bond yields. after the crisis, european countries experienced a financial stress situation starting in that affected bond yields, thus the investors saw an excessive amount of sovereign debt and demanded higher interest rates in low economic growth situations and high fiscal deficit levels. during - , several european countries suffered downgrades in their bond ratings to junk status that affected investors' trust and fears of sovereign risk contagion resulting, in some cases, a differential of over , basis points in several sovereign bonds. after the introduction of austerity measures in giips countries, the bond markets returned to normality in . the european debt crisis particularly revealed spillover effects between different sovereign bonds, which have been studied using various time series models e.g. var [ , ] and garch [ ] . the results showed that portugal, greece, and ireland have a greater domestic effect, italy and spain contributed to the spillover effects to other european bond markets and a core group of abfn (austria, belgium, france, and netherlands) countries had a lower contribution to the spillover effects, with some of the least impacted countries residing outside of the euro zone. during the sovereign debt crisis, public indebtedness increased after greece had to correct the public finance falsified data, and other countries created schemes to solve their public finance problems, especially, bank bailouts. in consequence, the average debt-to-gdp ratio across the euro-zone countries rose from % in to . % in , as well as the increase in sovereign credit risk [ , ] . after the fiscal compact treaty went into effect at the start of , which defined that fiscal principles had to be embedded in the national legislation of each country that signed the treaty, the yield of sovereign bonds started a correction, although some investors and institutions . four of the listed countries are part of the g and g economic groups (germany, france, italy and the uk). we consider sovereign bond yields with a year maturity between january and june . this data is taken from the financial news platform . in total, there are , data values for each country with an average of data points within year. table provides summary statistics of the y bond yield data. the results show greek yields to have the highest values across all statistical measures compared with other countries yields, particularly within the - (max yield of . ). in contrast, swiss bond yields exhibit the smallest mean and variance, with a higher than average positive skewness compared with other sovereign bonds. under the jb test for the normality of data distributions, all bond yield trends have a negligible p-value with non-gaussian distributions. the left skewed yield distributions (except for iceland), which represent an average decrease in yield values each year are high for giips countries compared with the uk, france, and germany, with flattening yield trends. we compute the correlation matrix for each window x with a displacement of δ between windows, and consider the mean and variance for the correlation matrix. we define the mean correlation r(t) given the correlations r ij for n sovereign bonds from figure , we find that the mean correlation r(t) is highest at . in oct . this suggests that a covid- impact was a continuation on the decrease of the mean correlation, and throughout the punitive lock down measures introduced by the majority of european countries in feb-mar . the decreases in mean correlation are also observed within the in the period during the european debt crisis, in which several european countries received eu-imf bailouts to cope with government debt and in , under a combination of political events within the uk and the increased debt accumulation by italian banks. the variance u(t) also follows a trend similar to the mean correlation, with the smallest variance of . in october . within , the variance increases between sovereign bonds and reflects the differences between the correlations of low and high yield. we consider the normalized network length l(t), which is introduced in onnela, et al [ ] as the normalized tree length. we define the measure as the normalized network length, as this measure is considered for ag and tmfg non-tree networks. the network length is a measure of the mean link weights on the subset of links e(t), which are present within the filtered network on the distance matrix at time-t the plots in figure represent the mean and variance of the network length. as each filtering method considers a subset of weighted links, the normalized length l(t) is monotonic between all methods and decreases with the increased proportion of positive correlated links within the network. we highlight the movements in the normalized network length during the covid- period, which is reflected across all filtering methods. this movement is observed within , but only towards a subset of correlations, in which the network length of the mast and tmfg increases compared with the mst and ag. the relative difference between the normalized networks lengths is least evident in periods of low variance; this is observed in the - period, where the difference between all methods decreases. we find the variance is highest within the tmfg and lowest with the ag approach. the increased inclusion of links with a higher reliability error in the tmfg increases the variance, particularly within the - period. the variance of the mst on average is higher compared with the mast, but when considering only the highest correlated links in the ag, the variance decreases. we define the degree centrality for the node of maximum degree c(t) at time-t. this measure considers the number of direct links the mean occupation layer η(t) (mol) introduced in onnela, et al [ ] is a measure of the centrality of the network, relative to the central node υ(t). we define lev i (t) as the level of the node, which is the distance of the node relative to υ(t), where the central node and nodes unconnected relative to the central node have a level value of we use the betweenness centrality to define the central node υ(t) for the mol. introduced in freeman [ ] , the betweenness b(t) considers the number of shortest paths σ ij (k) between i and j which pass through the node k, relative to the total number of shortest paths σ ij between i and j, within the mst, the degree centrality ranges between to for euro-zone countries. the trend within the mst remains stable, where the central node under degree centrality is associated with multiple sovereign bonds e.g., netherlands %, portugal % and belgium % across all periods. the mast has the highest variation, with a centralized network structure in some periods e.g., c(t) of , forming a star shaped network structure. this is usually associated with greece, iceland and hungary, which are identified as the central node % of the time. the degree centrality on average is naturally highest with the tmfg, under a higher network density, where the central nodes are identified as hungary and romania sovereign bonds, similar to the mast. the ag identifies the netherlands and belgium within the degree centrality, under a higher proportion of % and % compared with the mst. within figure , the mol on average is smallest for the ag, because of the level values from unconnected nodes, in which an unconnected node is present within % of considered windows. we find that the nodes within the tmfg are closest within the network, where the central node is directly or indirectly connected for all nodes, with an average path length of . across all periods. between the mst and mast, the mol is higher within the mast, where nodes within the network have a higher degree centrality. we analyze the temporal changes of sovereign bond yields between october and june . the associated link weights on each filtering method for window x are the proportions in which the link appears within the correlation matrix, under the statistical reliability, across all samples m for the randomly sampled windows x * m . under the mst, austria has the highest degree centrality of . the network also exhibits clusters between southern european countries connected by spain, and the uk towards polish and german sovereign bond yields. within the network, there is a connection between all abfn countries, but countries within this group also facilitate the connecting component within giips countries, where belgium is connected with spain and irish sovereign bonds. the uk and eastern european countries remain on the periphery, with abfn countries occupying the core of the network structure. for the mast in figure , there exists a high degree centrality for polish sovereign bonds between western european countries e.g., france and netherlands. this contrasts to the observed regional hub structure within the mst, with the existence of several sovereign bonds with high degree centrality in the network. the uk remains within the periphery of the mast structure when considering anti-correlations, and shows uk bond yields fluctuate less with movements of other european bonds compared with previous years. this is also observed for sovereign bonds for other countries with non-euro currencies such as czech republic, hungary, and iceland. we find nodes within the tmfg to have the highest degree in iceland at and poland at . although the mst is embedded within the tmfg network structure, a high resemblance is observed to links from the mast, where % of links which are present within the mast are common in both networks. there is also the associated degree centrality of the mast, which is observed within the tmfg connected nodes. under the tmfg, nodes have a higher degree connectivity when considering an increased number of links, this is the case for the uk, which has links compared with other spanning tree approaches. the ag exhibits three connected components between western european countries, southern european countries and the uk with eastern european countries. these unconnected nodes within the ag are associated with non-euro adopting countries, with the remaining countries connected in an individual component. by solely considering the most positive correlations, we observe the formation of -cliques between countries, which is prevalent within the western european group of nodes. the average statistical reliability is highest at . within the mast and ag, . for the mst and . for the tmfg. under the tmfg, the increased inclusion of links with a lower magnitude in correlations decreases the reliability in link values. other filtering approaches which consider a smaller subset can still result in low reliability values between some nodes e.g. austria and romania at . in the mst, germany and netherlands at . in ag. under various constraints, we observe a commonality between sovereign bonds across network filtering methods. we find for tree networks, that euro-area countries have a high degree centrality and countries with non-euro currencies e.g. czech republic and the uk are predominately located within the periphery of the network. this is further observed within the ag, where cliques are formed between giips and abfn countries, which is distinctive during the covid- period compared with previous years. the anti-correlations within the mast inform the trends of the negative correlations between eastern european countries and other european countries. by considering the tmfg with an increased number of links for positive correlations, we find similarities with the mast degree centrality. as a response to the covid- pandemic, most countries implemented various socio-economic policies and business restrictions almost simultaneously. an immediate consequence was an increase in yield rates for these nations. the resulting upward co-movement and upward movements in other yield rates explain the decrease in the mean correlation in bond dynamics, coinciding with the pandemic outbreak. thus, understanding the dynamics of financial instruments in the euro area is important to assess the increased economic strain from events seen in the last decade. in this paper, we consider the movements of european sovereign bond yields for network 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sovereigns' cds market hierarchy: a correlation-filtering approach portfolio optimization based on network topology complex networks and minimal spanning trees in international trade network hierarchical structure in financial markets network filtering for big data: triangulated maximally filtered graph interest rates hierarchical structure relation between financial market structure and the real economy: comparison between clustering methods the multiplex dependency structure of financial markets dynamic asset trees and black monday asset trees and asset graphs in financial markets clustering and information in correlation based financial networks random matrix approach to cross correlations in financial data the multi-layer network nature of systemic risk and its implications for the costs of financial crises universal and nonuniversal allometric scaling behaviors in the visibility graphs of world stock market indices pruning a minimum spanning tree on stock market dynamics through ultrametricity of minimum spanning tree causality networks of financial assets complexities in financial network topological dynamics: modeling of emerging and developed stock markets cross-border interbank networks, banking risk and contagion a tool for filtering information in complex systems spanning trees and bootstrap reliability estimation in correlation-based networks hierarchically nested factor model from multivariate data correlation, hierarchies, and networks in financial markets a cluster driven log-volatility factor model: a deepening on the source of the volatility clustering multiscale correlation networks analysis of the us stock market: a wavelet analysis network formation in a multi-asset artificial stock market key: cord- -ea nbkh authors: mitzner, veera title: conclusion and further thoughts date: - - journal: european union research policy doi: . / - - - - _ sha: doc_id: cord_uid: ea nbkh the conclusion chapter not only summarizes the main results of the research conducted for this book but also connects the events and discussions between the s and s to later political developments. it shows striking ideational and institutional continuity and reveals a substantial character of european integration: by relying on powerful political framings and discourses, as well as on sturdy institutions, the european community/union was able to move into areas that were not sanctioned by the treaties. the chapter further argues that to stay relevant, the eu research policy must be capable of breaking with the past and dramatically expand its mission to embrace the social and environmental challenges of the twenty-first century. in fact, with the existential threat of climate change and other global challenges, and the urgent need for socio-technological transformation at scale, opportunities and imperatives for european level activity in research might be greater than ever. separation of europe from the technologically more advanced united states, the european political and economic elites appeared open to arguments for unifying european resources in science and innovation. with the postwar market liberalization, rapid technological change, and the gradual shift of production to the newly industrialized countries, the case for joint european research effort seemed plausible. maria nedeva and linda wedlin have divided the development of european union research policy in two main phases: "science in europe" and "european science." according to these two scholars, the first phase, "science in europe," which roughly covers the time period analyzed in this book, was characterized by the principle of subsidiarity-an idea that european-level action would only be appropriate if action at a national level was insufficient; focus on increasing collaboration amongst researchers from different european countries; and concerns of the technology gap, which nedeva and wedlin call the "european paradox," as europe as a world leader of science seemed to lag behind in the industrial and economic exploitation of scientific ideas. furthermore, "science in europe" witnessed three distinct waves: the establishment of large, transnational facilities, such as european organization for nuclear research (cern), euratom, european molecular biology organization (embo), and european molecular biology laboratory (embl); the creation of "diffuse" organizations, such as european cooperation in science and technology (cost) and the european science foundation (esf), including all fields of research and providing platforms for cross-national cooperation rather than supporting science at european level; and finally the creation of the ec/eu framework program for research, bringing the scattered research programs under one administrative umbrella. for nedeva and wedlin, "science in europe" was a period "during which a partial and fragmented science and research system was developed." it complemented rather than challenged national science, research, and innovation arrangement; it had a limited impact on universities, research institutes, and national funding agencies and landscapes; and it "didn't in a rule, include explicitly european-level research performing organizations." this book doesn't challenge nedeva's and wedlin's analysis. for a long time, the ec/eu research policy had a partial character-the first initiatives were not designed to directly challenge activity in national settings. the proponents of european level activities in the field, while often ambitious and visionary, were aware of the political realities. even the "european paradox" argument is solid, although the debate had its ebbs and flows and occurred in different variations depending on the context and advocates. fears of european technological retard did push initiatives forward, at different degrees of success, giving them rationale and a sense of urgency. also, european level research facilities were still few and far in between, and decisions in brussels barely influenced the daily operations of universities and research institutes in the ec member states. however, with a closer historical analysis, we also see an emerging policy that was constantly challenged, contested, and vetted by a number of groups, and in particular those who would mostly have been affected by it. and this is the vital "more to the story" that nedeva's and wedlin's and others' accounts do not fully tell. this book also unveils the bigger ambitions of policy-makers and dreamers by fletching out the protracted struggles for initiatives that either never quite made it to the fruition or were realized as truncated versions, to the disappointment of many. it sheds light to the already forgotten and fluid policy spaces where the feasible and unfeasible were defined, and where future pathways were sometimes painfully determined. crucially, these spaces were shaped by broader political, social, and economic circumstances within and beyond europe, and the prevalence and strength of specific ideational shifts and conditions, such as the fading of the naïve enthusiasm about science that had characterized the immediate postwar period; increasing uneasiness about growth policies and calls for better accountability of economic and scientific activity; new limitations imposed by economic austerity; persisting distrust in european level bureaucracy among academics and national policy-makers; and ongoing political battles in other policy areas within the ec context. yet one distinct thread runs through all these events and debates: a basic agreement on the beneficial role of research and research policy in achieving greater growth and competitiveness, and the need to create joint political mechanisms to that end. how this would eventually be achieved remained open for contestation. besides being a story of continuity and struggle, this book also is an account of the vulnerability of a political idea that survived, grew big, and eventually transformed european research. it was just as strong as its proponents, which at the beginning were not many. although the commission made sure that science policy experts would be involved in the elaboration of the various schemes from the start, these experts often represented governments and national science funding bodies rather than the broader science community. the lack of active participation of scientists and their advocates, as well as of industry and the wider public in the early discussion on the topic, is striking; until the s, most proposals were developed in technocratic milieus with little public attention. the backing of pro-european associations such as union des industries de la communauté européenne (unice), the european parliament, and some individuals from national research institutions and administrations remained too weak and fragmented to put significant pressure on key decision-makers. this is the tragedy of the early ec/eu research policy. politicians in the ec member states, whose main concern was often winning the support of the electorate, tended to focus on issues with more immediate political benefit. it was not the case that the citizens in the community member countries had opposed the idea of the europeanization of scientific and technological activities: in the community's surveys of public opinion, research has consistently been at the top of the list of policies that people think should not be managed exclusively at a national level. research was, nevertheless, not the kind of "high politics" that would be surrounded by glamour and drama and that would have reached the headlines. it remained distant to the lives of ordinary citizens who barely knew of the commissions initiatives. it is also exactly this technical, abstract, complex, and distant nature of this policy domain that at least in part explains the persisting gap between rhetoric and reality in european research policy from the mid- s to the mid- s. for national politicians, expressing catchy phrases and stirring rhetoric about science and its many promises for the future, could be politically advantageous but also relatively safe: sacrifices in this sector rarely had a direct impact on the lives of voters, and therefore, they seldom provoked significant public protests. the late s' protests at the ispra euratom joint research centre stand out as a notable exception, as multiple employees were threatened to lose their jobs as a result of decisions made in brussels. but in most cases, lip service in research policy came at a low political price. in the absence of a systematic backing of scientists or their representatives, the commission alone was too weak to induce the national governments to truly commit to developing a community research policy. moreover, commission's strategy was not always consistent: important tensions prevailed between the dgs, while each commissioner in charge of research pursued a slightly different policy. the existence of dg research, under various names since , however, became a central structure for guaranteeing institutional and ideational continuity. the dg constituted a center where bureaucrats and experts, sharing some important ideas and interests, could further elaborate schemes for new community activity. this finding supports katia seidel's research on dgs iv (agriculture) and vi (competition), which shows that the administrative cultures developed in those dgs and the subsequent socialization of officials into these cultures, "helped to bring about policies in relevant areas, and then to keep them on track." through the discussions initiated by dg research and others, the ec gradually formed an arena for "europeanization" in research where socialization of different experts and political actors could take place. intra-european connections were reinforced, and similarities increased, while "europe" was given new contents and contours. at the same time, it would be misleading to argue that all contacts within the realm of the research policy debate would automatically have led to greater cohesion. these exchanges also exposed to governments and experts the diverse character of european research systems and made them aware of the difficulty in creating a strongly integrated common policy. yet they prepared the ground for subsequent initiatives that would take off with a more favorable political momentum. in the early s, the various plans and scattered activities of the previous twenty years finally evolved into something that could be said to have an important scope and political and economic significance. the first step into this new direction was the launch of sizeable technological programs, the pioneer being european strategic program on research in information technology (esprit), an initiative that was intended to develop a european strategic scheme based on collaboration between major european companies, small-and medium-sized firms, universities, and research institutes. the project was successful: of the proposals received in response to the first call, less than percent could be funded. not surprisingly, after , esprit became the model for successive community programs, such as research in advanced communications in europe (race) and basic research in industrial technologies (brite). the expansion of these activities led to the creation of the first framework program for research ( ) ( ) ( ) ( ) in . the € . billion budget of the framework program, together with the budget for esprit of € million, signified a threefold increase in the community's research spending in . moreover, the program sparked off a steady and fast increase of the community's research budgets. peter tindemans has observed that after esprit, there were a growing number of scientists and industrialists involved in the consultations, projects, and evaluations, and thereby directly benefiting from the community's activity in the field. this created "an almost unstoppable dynamic of pressure [that] arose in national capitals and in the directorates-general responsible for the fp's [framework programs] to increase the budget." although the role of the ec as a focal point of common european research activities remained contested, as was demonstrated by the creation of eureka, an intergovernmental initiative intended to promote "near-market research," in , and by the difficult negotiations of the subsequent framework programs, the political context had changed. national governments were now more willing to see the ec take a central role in the field, and a horizontal and vertical growth of brussels-led activities ensued. all this was supported by the favorable conjuncture in european integration that ensued the single market program. the first framework programs were still focused on advancing applied, marketoriented research, but during the s, these programs became more comprehensive and versatile as they included new mechanisms and stretched into different disciplines and sectors. they also opened up for countries outside the ec/eu framework, developed collaboration with other european and international organizations, and finally became more closely integrated with the ec/eu core policies. an eu-centric european research space started to emerge. this is, indeed, where wedlin and nedeva see the shift to "european science," marked by a changing understanding of the "european added value." whereas the previous ec/eu activities had been primarily focused on fostering collaboration between researchers, now there was a widely shared desire to increase the level of integration in different aspects of european science and research. new tools were designed to strengthen the european knowledge base and to support basic research through competitive funding. a key concept here is the european research area (era). originally dreamed up by commissioner ralf dahrendorf in the s, the idea was revived by commissioner alberto ruberti in the s, and finally pushed to the center of the european political agenda by commissioner philippe busquin. endorsed as an ambitious effort to pool european scientific and technological resources, the initiative marked a clear break with the distributive approach that had characterized the eu activities in the preceding years. while the definition of what the era stands for has evolved over the years, it has brought new inclusiveness into the eu research policy and signaled a move toward a more europeanized research policy field, "where the era agenda provides justification for the adoption of new institutions and funding tools." era aimed to increase the commission's autonomy in initiating projects and programs with direct implications for national activities. in , the commission defined era as "a unified research area open to the world based on the internal market, in which researchers, scientific knowledge and technology circulate freely and through which the union and its member states strengthen their scientific and technological bases, their competitiveness and their capacity to collectively address grand challenges." era was launched in the political framework of the lisbon european council of march , where the eu set itself the new strategic goal of becoming "the most competitive and dynamic knowledge-based economy in the world capable of sustainable economic growth, with more and better jobs and greater social cohesion." in lisbon, research and development were drawn to the center of the eu's strategy for achieving its goal by . essentially, scientific research, technological development, and innovation were defined as key factors in growth, competitiveness, and employment in a knowledge-based european economy. this trend was continued in the eu's new growth strategy, europe , launched in . one of the seven flagship initiatives announced in the strategy was innovation union, aiming to "improve conditions and access to finance and innovation, to ensure that innovative ideas can be turned into products and services that create growth and jobs." era was considered a core element of the innovation union and the europe strategy. the eu's barcelona target to achieve r&d funding of percent of gdp-an objective largely motivated by the higher r&d investment levels in the united states and japan-gave era additional political support and visibility. according to wedlin and nedeva, "european science" has created "an organizational space for the support of research that is aligned, and in competition with national-level research spaces." a key invention in this space was the european research council (erc), created in as a part of the seventh framework program. by allowing european researchers to compete with all other researchers in the eu area on the basis of excellence, erc revolutionized the eu's approach to supporting scientific activity. with scientific excellence as the sole criterion for selection and sizeable pots of money to distribute (the first budget in the seventh framework program ( - ) was . billion euros, and it almost doubled for horizon , constituting percent of the overall program budget), erc soon became a powerful tool for boosting european knowledge space and shaping science in europe. the shifting agendas and the broadening scope of the ec/eu activities have been paralleled by the gradual strengthening of the community's legal status. after the still very restricted formulation of the single european act ( ), the treaty of maastricht ( ) included a new article , which required that "the community and the member states shall coordinate their research and technological development activities so as to ensure that national policies and community policy are mutually consistent." moreover, the commission was granted the explicit right to take any "useful initiative" to promote such consistency. the amsterdam treaty ( ) then abandoned the requirement for council unanimity for adoption of the co-decision on the framework program. from now on, the program could be adopted by a qualified majority vote. the union's competences in the domain were further widened by the article of lisbon treaty ( ), stating that: . the union shall have the objective of strengthening its scientific and technological bases by achieving a european research area in which researchers, scientific knowledge and technology circulate freely, and encouraging it to become more competitive, including in its industry, while promoting all the research activities deemed necessary by virtue of other chapters of the treaties. . for this purpose the union shall, throughout the union, encourage undertakings, including small and medium-sized undertakings, research centres and universities in their research and technological development activities of high quality; it shall support their efforts to cooperate with one another, aiming, notably, at permitting researchers to cooperate freely across borders and at enabling undertakings to exploit the internal market potential to the full, in particular through the opening-up of national public contracts, the definition of common standards and the removal of legal and fiscal obstacles to that cooperation. . all union activities under the treaties in the area of research and technological development, including demonstration projects, shall be decided on and implemented in accordance with the provisions of this title. within ten years, the ec/eu research policy activity had firmly become woven into the legal fabric of the union. while the ec's/eu's legal base strengthened and its activities broadened, the community/union gradually moved into territories traditionally occupied not only by european nation-states but also by various intergovernmental organizations promoting research. in this book we have seen how already in the s and s, the ec challenged more established organizations such as the oecd and the council of europe. in the subsequent years, this trend continued. indeed, as the eu is thus increasingly dominating the european research field, one can observe an incremental transformation of previously non-ec/eu projects and institutions into eu or quasi-eu projects and institutions, or alternatively, a launching of new eu operations practically identical to existing activities in other european organizations. a good example here is space research. the cooperation agreement signed between the eu and the european space agency in constituted a new framework for a comprehensive european space policy involving the coordination of the actions of the european commission and esa through a joint secretariat, a small team of eu administrators and esa executives, as well as ministerial-level meetings in the space council. in there was a further development when twenty-nine european countries expressed their support for a european space policy, prepared jointly by the european commission and the esa's director-general. moreover, the lisbon treaty gave the european union an explicit competence in space, calling for the development of a european space program. the implementation of this program has further increased cooperation between the eu and the esa. in the recent past, almost percent of the funds managed by esa have originated from the eu budget. the european science foundation, on the other hand, after almost nearing death in , has acknowledged the eu's growing role in promoting european science and wound down all its collaborative research instruments and networks. with a stated mission to serve the european research area for the sole benefit of science, it now focuses on running an expert division called science connect that provides services to the science community. this is a significant shift: since its creation in , esf has run over world-class science programs and networks, with support from eighty member organizations in thirty countries. through this work, it has made a solid contribution to the creation of the european research space. in the most recent years, its financial resources, however, have faced a dramatic downturn. between and , the esf member contributions diminished by . million euros. these budget cuts, together with the wishes of its members, led esf to radically reduce its activities and staff and to move policy-related work into newly formed science europe. cost, another intergovernmental institution supporting european research set up in the early s and analyzed in this book, has weathered the emergence of a comprehensive eu research policy somewhat better. it fills a specific niche in the european research funding landscape as an organization providing financial support for the creation of european-wide multi-stakeholder research networks, called cost actions. most of cost's funding, however, comes from the eu research framework program, which makes it highly dependent on the political developments in the union. the various impacts of the emergent eu research policy on the wider european research space still deserve more thorough scholarly attention. one interesting trend, however, is the emergence of stronger and more organized organizations and networks promoting the interests of researchers, research funders, and the higher education sector at the european level. these include euroscience, a grassroots non-profit organization of researchers in europe, created in to shape policies for science, technology, and innovation. with its individual members, euroscience aims to be "a direct and democratic way to influence the construction of the europe of science and technology, for the benefit of europe's international position in science, and to enhance science's contribution to society, opportunities and mobility for both young and experienced researchers." the european association of research and technology organizations (earto) was established two years later to promote research and technology organizations and represent their interest in europe. its current activities are geared toward influencing the eu policy. in , a merger between the association of european universities and the confederation of european union rectors' conferences led to the creation of the european university association (eua). eua represents over universities in forty-eight countries, and its activities are targeted at influencing the eu policies on higher education, research, and innovation. a year later, another belgium-based organization, the league of european research universities (leru), was formed with the objective to "advance the understanding and knowledge of decision makers, policy makers and opinion leaders about the role and activities of research-intensive universities." both institutions seek to promote the interests of higher education and research institutions in eu policymaking. in , twenty-three countries joined in the founding assembly of science europe, an organization representing research funders and other research organizations in the eu. science europe was built from two former advocacy groups, the esf and eurohorcs (an organization of the heads of the european research councils), aspiring to give its members a stronger and more united voice on european level policy. the list could be continued, in particular if one includes more informal networks and alliances. the rise of these various groups is largely a response to the gradual strengthening of the eu's research arm, which has made advocating for researchers' and research and higher education institutions' interests in brussels a worthwhile endeavor. undoubtedly, there is a new reality for research in europe. the current eu research funding program is exceptional in its size, long duration (seven years), budgetary framework stability, and scope: it encompasses research as well as innovation; grants as well as loans, equity, and procurement; it combines a broad top-down focus on grand societal challenges with bottom-up "frontier" research; it is crossborder and cross-sectoral, encouraging inter-disciplinary collaboration, mobility, and coordination. and it is attracting increasing attention and interest within research communities both within and beyond europe. the horizon mid-term evaluation accounted a percent increase in the annual number of applications compared to the program's predecessor, the seventh framework programme (fp ). with an . percent success rate, which was much lower than under fp ( . percent), the competition for horizon grants had become fierce. while . percent of funding went to participants from the eu member states, the program involved people from over countries. moreover, eu-funded projects are also producing top-notch research. between and alone, horizon , supporting approximately , researchers, produced more than peer-reviewed publications that were cited more than twice the world average. two-thirds of them were also openly accessible to the public. it is also noteworthy that within the european institutions, research has taken a prominent role. in , a total of of the commission's , officials worked for dg research, making this dg the fourth largest division. if this number were to include staff members in the eu joint research centre, research would occupy the first place. research also continues to be the third largest item in the eu budget. despite the prevailing uncertainties about the future of european integration and the different priorities of the member states, the trend of strengthening the role of the eu in supporting and shaping european research is likely to continue. in june , the commission published its proposal for the next eu funding program, horizon europe, which would cover the years of - , involve a budget of billion euros-an increase from horizon -and be "the most ambitious research and innovation funding programme ever." there was little modesty in the commission's statement that "'[b]eneficiaries' research capacities and scientific outputs would have significantly decreased had they received national funding instead. this decrease would have been especially large in terms of their ability to collaborate with industry and business, transfer of knowledge, the number of participations in scientific conferences and the knowledge in new areas." the covid- outbreak in , effectively highlighting the importance of research in tackling global crisis is likely to give a further boost to eu research and innovation activities. the eu acted relatively fast by providing emergency funding from horizon and processing proposals at a record speed. one could note emerging tensions between european and national research spaces, as the national funding bodies and the eu instruments might increasingly compete for resources, applicants, and legitimacy. the larger the eu research budget grows, and the more extensive the european level programs become, the more accentuated this competition will become. already now the eu research funding programs include almost all research themes and forms of research promotion that exist in national research and technology policies. linda wedlin and maria nedeva explain the shift from "science in europe" to "european science" by two important changes in policy rationales and framing. first, the understanding of "european added value" shifted from focusing on coordination to incorporating competition. epitomized in the creation of the european research area, which signaled a move away from collaboration toward integration, this new framing precipitated the implementation of policy instruments that stretched beyond simple support for collaboration between researchers and encouraged genuine competition between them. second, there was growing evidence and concern that europe was not doing so well in science compared with its main commercial competitors. this new version of the technology gap debate argued for a european retard that was more fundamental than failure in making the link between scientific research and commercial applications. "by ," wedlin and nedeva write, "the long-standing assumption of this 'european paradox' morphed into a full-blown 'gap' argument, that is, that europe was clearly lagging behind the usa and japan both in terms of science and its application." this concern led to a change of rationales and the target for policy intervention. it served as a justification for a powerful move to the realm of basic science and the creation of the european research council. tim flink, through an analysis on the creation of erc, supports this finding. a renewed concern of european global retard and a conviction of the need to strengthen the european knowledge base underpinned a political discussion that ushered the birth of erc. moreover, language played a critical role: with the concept "frontier research," the european commission avoided directly violating the principle of subsidiarity, which had curtailed its activity in basic research-an area traditionally belonging to the nation states. "frontier research," falling somewhere in between basic and applied research, aligned well with the eu's traditional mission of fostering european competitiveness and the geopolitical objective of the lisbon strategy to make europe the world's leading commercial power, drawing its strength from knowledge. "frontier research" projected an image of global competition, and it made a solid reference to vannevar bush's monumental report science, the endless frontier and the founding myth of the us national science foundation. "[i]n this policy process," concludes flink, "the narrative structure only seemed to be successful if it followed the chimera of a geostrategic security understanding (isolation by the european research area) and a prosperity identity (market imperative of the eu)." also, terttu luukkonen, while noting the critical role of stakeholder groups, such as the leading life sciences organizations, stresses the role of the gap debate in the formation of the erc. "there was a concern," she writes, "about funding being too low for basic research and about quality of science and its institutions in europe and, as in european research policy in general, usa provided a benchmark with which comparisons were made." furthermore, the focus on frontier research, provided "a strong argument that the erc is justified from the point of view of technological and economic competitiveness, not just of scientific competitiveness." this is confirmed by just a cursory reading of the commission's documents from that time period. the prominence of economic concerns and the gap narrative is striking, as well as the discourse that highlights the insufficiency of national initiatives. the commission's proposal for era, for example, argues that in europe, "the situation concerning research is worrying. without concerted action to rectify this the current trend could lead to a loss of growth and competitiveness in an increasingly global economy. the leeway to be made up on the other technological powers in the world will grow still further. and europe might not successfully achieve the transition to a knowledge-based economy." the discursive continuity is remarkable. throughout this book we have seen how european commission officials and other supporters of greater eu competences in the field of research used a specific language and political framing to advance their initiatives. this language presented scientific research as an engine for economic growth and material prosperity, which were widely accepted as favorable political goals by the european governing elites. combined with arguments about european technological delay vis-à-vis its main commercial competitors, it presented an increasingly convincing case for new european level activity. by the early s, these framings had become dominant truth claims in the european debates, and they started to essentially determine political agendas. at the same time, the scope and nature of the scientific enterprise changed, the pace of discovery and innovation accelerated, and as a result, not only did competition between research institutes, nations, and individual researchers accentuate, but science and technology became increasingly vital forces shaping societies and determining the geopolitical and economic success of countries. it could be argued that one reason for the continuous popularity of the instrumental-economic conception of research policy has been its malleability and extraordinary ability to adapt to different political conditions. as amenable as it seemed to the western european expansive welfare economies of the s and s, it has proved perfectly compatible with the intensifying drive for market liberalism since the early s. this is not only because of the prevailing appeal of growth. research policy has proved useful in facilitating and accelerating the shift to the world of open markets. in this study we have seen how research policy, especially at european level, was commonly pictured as a means to adapt to an exogenous and irreversible transformation that later became labeled as globalization. the eu research policy came into being with an explicit objective of enhancing european economic performance in an international framework in which competition was accelerating. this goal remains as one of the guiding principles of the current eu research policy. when research commissioner máire geoghegan-quinn in november said: "fundamentally, support through horizon for research, innovation and science is an economic policy," she reiterated the principle on which the union's activity had been based since the very beginning. the economic framing remains sturdily in place. the horizon mid-term evaluation report, for example, made a blatant calculation a gdp gain, concluding that "[e]very euro invested under horizon brings back to . euros." the explanatory memorandum included in the commission's proposal for establishing horizon europe included similar calculations. according to the memorandum, horizon europe was expected to generate positive effects on growth, with an increase of gdp on average by . percent to . percent over twenty-five years, "which means that each euro invested can potentially generate a return of up to euro of gdp over the same period"-better ratio than in horizon . the memorandum went on to calculate that discontinuing the union research and innovation program could result in a decline of competitiveness and growth up to billion euros of gdp loss over twenty-five years. to some degree, the eu is tied to this language. initially, the lack of a solid juridical basis for research policy forced the commission to frame research as a tool for achieving prosperity, which has traditionally been one of the union's core objectives. although the gradual strengthening of the juridical basis of research policy during the last three decades has granted the commission more room for maneuver, the eu's activity and ambitions are still limited by the principle of subsidiarity and the constant search for european added value. the eu is most likely to gain legitimacy by anchoring its initiatives and programs to the objectives of competitiveness and growth-a territory where eu activity is seldom questioned. but continuously liking research and growth and articulating science and innovation policy in economic terms, is more than a smart political strategy. it is a durable policy framing that was constructed in the early s and that has been broadly embraced and absorbed by national decisionmakers, european officials, and a number of other actors shaping research policy. it embodies a very specific understanding of the role of science and research in society and the objectives of public support for research and innovation communities. policy framings can and need to change if they become unsuited to responding to the society's most pressing needs and challenges. johan schot and w. edward steinmueller, for example, have argued for a new framing for innovation policy, "linked to contemporary social and environmental challenges such as the sustainable development goals and calling for transformative change." this framing should take priority over the two existing framings, which emerged in the s and s, and which, while still relevant, "offer little guidance for managing the substantial negative consequences of the socio-technical system of modern economic growth to which they have contributed and of which they are a part." the first framing, which dominated during the time period analyzed in this book and-as we have seen-still influences eu policy, emphasized the role of research and innovation for increasing economic growth, aimed at boosting the potential for science and technology for prosperity, and cultivated socio-technical systems supporting mass production and consumption. the second framing gained ground toward the late s and more explicitly aimed to address challenges related to intensifying international competition and globalization. with a focus on developing and analyzing national systems of innovation, enhancing capacities for learning, and supporting connectivity between different societal sectors within the system, it added a layer of sophistication and complexity to the first framing. however, it didn't depart from the premises of increasing growth through maintained and improved competitiveness. the third framing suggested by schot and steinmueller draws upon the recognition that a deep and rapid transformation of socio-technical systems is needed in the backbone systems of modern societies, including energy, mobility, food, water, healthcare, and communication, and that research and innovation policy has a vital role to play in supporting this transformation. that requires a dramatic change in thinking, broadening of perspectives and alliances, and setting entirely new goals and objectives. according to the two authors, this new framing "focuses on innovation as a search process on the system level, guided by social and environmental objectives, informed by experience and the learning that accompanies that experience, and a willingness to revisit existing arrangements to deroutinize them in order to address societal challenges." further, the innovation process in this framing "is likely to be effective in achieving these goals if it is inclusive, experimental and aimed at changing the direction of socio-technical systems in all its dimensions." essentially, this framing will propose building a new knowledge base, setting up mission-oriented policies with missions formulated in an open-ended way, encouraging new forms of engagement and networks between public, private, and third sector actors, and supporting intermediary actors to advocate competitive niches, as well as new visions and policies. it will mean a shift away from the old research and innovation policy proposition that social and economic goals can be achieved through economic growth if surpluses are redistributed and technocratic elites can regulate the externalities of growth. a more comprehensive and ambitious-if not radical-approach is needed to ensure the continued relevance of research and innovation policy in the contemporary world in rapid transition. schot and steinmueller are not alone with this proposition. in the last few years, there has been increasing scholarly interest in rethinking the premises of science and innovation policy and exploring ways of better aligning policy objectives with broader social and environmental challenges. a new policy paradigm is emerging, "layered upon but not fully replacing earlier paradigms of science and technology and innovation systems policy." according to gijs diercks, henrik laren, and fred steward, this "'normative turn' that is currently taking place insists that innovation policy must not only optimize the innovation system to improve economic competitiveness and growth, but also include strategic directionality and guide processes of transformative change towards desired societal objectives." further, this "emerging paradigm of transformative innovation policy is still a heavily contested discursive space" and "there is considerable uncertainty regarding the paths of this paradigm shift in the making." looking at the most recent eu language on research and innovation policy, one can discern elements of this shift. while competitiveness and growth retain a solid place in the framework programs, with the increasing focus on strengthening the european knowledge base and the objective of tackling grand societal challenges through research, the political framing of the eu research policy has widened. one of the earliest visible expressions of this ambition was the declaration, endorsed by over researchers, policy-makers, and representatives from industry and research funding institutions during the "new world -new solutions" conference in lund, sweden. the declaration stated that "european research must focus on the grand challenges of our time moving beyond current rigid thematic approaches. this calls for a new deal among european institutions and member states, in which european and national instruments are well aligned and cooperation builds on transparency and trust." the "grand challenges" listed in the declaration included global warming; tightening supplies of energy, water, and food; aging societies; public health; pandemics; and security. the lund declaration also states that the "european knowledge society must tackle the overarching challenge of turning europe into an eco-efficient economy." since then, broader social and global concerns have found a firmer footing in the eu's research policy agenda. between and , a total of percent of horizon funding went to reinforcing and extending the excellence of european science base and consolidating the era. the second biggest pot, percent, went to stimulating a critical mass of research and innovation efforts to help address grand societal challenges. promoting industrial leadership came only as third, with percent of funding. the remaining percent was dedicated to other priorities, such as widening participation, including society, euratom, and the pilot for fast-tracking innovation. the three pillars of horizon europe-excellent science, global challenges and european industrial competitiveness, and innovative europe-seem to continue this trend, although the commission notes that "[i]industrial leadership will be prominent in this [global challenges] pillar and throughout the program as a whole." indeed, the proposal for horizon europe was "framed by the premise that research and innovation (r&i) delivers on citizen's priorities, boosts the union's productivity and competitiveness, and is crucial for sustaining our socio-economic model and values, and enabling solutions that address challenges in a more systematic way." in other words, "horizon europe will strengthen the union's scientific and technological bases in order to help tackle major global challenges of our time and contribute to achieving the sustainable development goals (sdgs). at the same time, the programme will boost the union's competitiveness, including that of its industries." these quotes reveal a striking feature in the commission's blueprints for horizon europe: an effort to achieve the double objective of responding to global challenges and maintaining economic growth. overall, however, the program seems to be defined as serving the european economy. in march , carlos moedas, the research commissioner, noted that "ware now on track to launch the most ambitious ever european research and innovation programme in , shaping the future for a strong, sustainable and competitive european economy and benefiting all regions in europe." even the lund declaration underlines that "[m]eeting the grand challenges will be a prerequisite for continued economic growth." a vital task for the commission and the union more broadly in the future will be resolving the tension between its traditional policy goals of pursuing growth and competitiveness and its newer mission of tackling grand challenges and promoting sustainability transitions. can both objectives be successfully pursued in parallel, and how possible conflicts between growth and broader socio-environmental issues can be reconciliated, has not explicitly been addressed in any of the key documents or declarations-until very recently. ursula von der leyen's commission's proposal for a european green new deal contains explicit language, suggesting that the challenge of climate change can be addressed without compromising economic growth and competitiveness. it presents the european green new deal as "a new growth strategy that aims to transform the eu into a fair and prosperous society, with a modern, resource-efficient and competitive economy where there are no net emissions of greenhouse gases in and where economic growth is decoupled from resource use." moreover, "careful attention will have to be paid when there are potential trade-offs between economic, environmental and social objectives." the language stressing the eu's "collective ability to transform its economy and society to put it on a more sustainable path," outlines an ambitious and transformative vision that departs from narrow sectorial propositions. in the european new green deal, science and innovation also has a solid place. the document calls for "new technologies, sustainable solutions and disruptive innovation" and notes that "conventional approaches will not be sufficient." the eu's new research and innovation agenda will emphasize experimentation, and working across sectors and disciplines, thereby taking the "systemic approach needed to achieve the aims of the green deal." for now it seems that four of horizon europe's five research missions would be closely linked to the green deal objectives. also, how the covid- pandemic transforms the european research landscape, remains to be seen. in the last few decades, the eu has shown an important capacity of selfcriticism and learning. horizon europe is being built on lessons learned in horizon , such as the need for increasing support for breakthrough innovation; adopting mission-orientation and encouraging citizen involvement; strengthening international cooperation; reinforcing openness; rationalizing the funding landscape; and encouraging participation. new initiatives in horizon europe, such as european innovation council, address these objectives. ursula von der leyen's commission has set a new level of ambition for achieving a pressing transformative change that transcends several sectors of european society. the rapid response to covid- also shows proactivity and agility from an institution often seen as stiff and bureaucratic. these serve as demonstrations that the eu does not need to or even cannot be constrained by its past policy framings and discourses. and this is a real opportunity. in today's europe, torn by nationalism and distrust in governing elites, preserving peace and producing prosperity no longer suffice to provide the legitimacy that carried the process of european integration this far. it is time for a more powerful and compelling mission, aiming at a profound socio-technological transformation that a union-wide science and research effort is well positioned to shape and realize. despite its contested origins, research policy has become one of the eu's core instruments for advancing its policies, promoting european integration, and achieving change. it has won the support of researchers, both in europe and beyond, gained prestige, and found a solid place on the eu political agenda. with its extensive programs and significant funding, it has transformed the european research landscape for good. however, to remain relevant, the eu research policy needs to be tailored to serve the most compelling needs of the european societies. if designed right, it can be a powerful force, enabling critical and truly transformative effort for which none of the individual member states would be capable alone. from 'science in europe' to 'european science qualitative study on the image of science and the research policy of the european union european commission, qualitative study on the image of science and the research policy of the european union the process of politics in europe. the rise of european elites and supranational institutions europeanization in the twentieth century: historical approaches post-war research, education and innovation policy-making in europe history and memory of an institution towards european science. dynamics and policy of an evolving european research space (cheltenham and political dynamics of the era," in changing governance of research and technology policy − the european research area explaining changes and continuity in eu technology policy: the politics of ideas european research area: an evolving policy agenda political dynamics european commission communication: a reinforced european research area partnership for excellence and growth the 'european research area' idea in the history of community policy-making european research area european commission communication: europe flagship initiative innovation union european commission communication: a reinforced european research area partnership for excellence and growth broadening aims and building support in science, technology and innovation policy: the case of the european research area towards european science die entstehung des europäischen forschungsrates: marktimperative -geostrategie -frontier research (weiserswist: velbrück wissenschaft consolidated version of the treaty on the functioning of the european union-part three: union policies and internal actions, title xix: research and technological development and space-article (ex article tec) esa website the european science foundation; death or mid-life crisis? the science connect website the european science foundation from 'science in europe science europe lobby group hit by sudden exodus. brussels-based advocacy group aimed to provide single voice for scientists in the eu-but is losing members key findings from the horizon eu-forschungspolitik-von der industrieförderung zu einer pan-europäischen wissenschaftspolitik? proposal for a decision of the european parliament and of the council on establishing the specific programme implementing horizon europe-the framework programme for research and innovation eu funding for research and innovation key findings from the horizon from 'science in europe eu-forschungspolitik from 'science in europe vannevar bush, science, the endless frontier. a report to the president by vannevar bush, director of the office of scientific research and development - ; - . the author's translation from german european research area european commission: communication from the commission to the council, the european parliament, the economic and social committee, and the committee of the regions: towards a european research area innovation and science: remarks at press conference launching horizon press conference brussels key findings from the horizon interim evaluation european commission: proposal for a regulation of the european parliament and of the council establishing horizon europe-the framework programme for research and innovation, laying down its rules for participation and dissemination three frames for innovation policy: r&d, systems of innovation and transformative change goal conflicts and goal alignment in science, technology and innovation policy discourse combining insights from innovation systems and multi-level perspective in a comprehensive 'failures' framework next generation innovation policy and grand challenges in addition to scholars in sts and sustainability transitions studies, there have also been historians arguing for a change. for instance, andrew jamison wrote in that "as in the s, there is a need for fundamentally rethinking the relations between science, technology, and society, in europe as well as internationally. in particular, there needs to be much more coordination between policies for science and technology and all the other policies that national governments, as well as local authorities and intergovernmental bodies pursue. in order to meet the challenge of climate change and sustainable development, science and technology need to be reconfigured so that the 'solutions' they provide can be relevant for the problems that humanity faces. and in order to provide appropriate solutions, scientists and engineers will need to be better educated about the problems that need to be solved transformative innovation policy european research area," ; ulnicane swedish presidency: research must focus on grand challenges european commission: the lund declaration: europe must focus on the grand challenges of our time european research area key findings from the horizon interim evaluation european commission: proposal for a regulation of the european parliament and of the council establishing horizon europe-the framework programme for research and innovation, laying down its rules for participation and dissemination the next eu research and innovation investment programme european commission, the lund declaration european commission: communication from the commission to the european parliament, the european council, the european economic and social committee, and the committee of the regions, the european green deal the next eu research and innovation investment programme key: cord- -i vhj authors: simha, ashutosh; prasad, r. venkatesha; narayana, sujay title: a simple stochastic sir model for covid infection dynamics for karnataka: learning from europe date: - - journal: nan doi: nan sha: doc_id: cord_uid: i vhj in this short note we model the region-wise trends of the evolution to covid- infections using a stochastic sir model. the sir dynamics are expressed using textit{it^o-stochastic differential equations}. we first derive the parameters of the model from the available daily data from european regions based on a -day history of infections, recoveries and deaths. the derived parameters have been aggregated to project future trends for the indian subcontinent, which is currently at an early stage in the infection cycle. the projections are meant to serve as a guideline for strategizing the socio-political counter measures to mitigate covid- . covid- has brought misery to many regions in the world, especially in europe. however, it has just taken off in other parts of the world. the remaining regions, including india hosts huge population approximately more than % of the world population. there is an eminent danger and most of these population are under the poverty-line or the countries do not have huge resources for medical interventions for affected patients. the options are limited -either go for herd immunity which can backfire and it is too risky or lockdown and restrict the population from moving around. thus we try to factor in the percentage of population allowed to move around (some are essential services and some not obeying the orders) and assuming the mobility factors appropriately , we try to predict what can happen during the lockdown. we specifically take italian case for karnataka because of the population resemblance. the dynamics governing the evolution of the covid- infections have been modeled using a stochastic differential equation sir model [ ] . the parameters of this model have been initially optimized for a set of european regions, individually. an aggregate of these parameters have then been used for projecting the future trends for the indian region, specifically for karnataka state. multiple projections have been generated by varying the exposure factor that influences the growth rate of infections. this is done in order to provide insights for selective regional quarantining and lock-downs. the evolution of covid- infections in each region, has been modeled via the stochastic susceptible-infected-recovered (sir) model [ ] which is given as, model states ) t is a daily-time parameter. ) c(t) denotes the number of active infections at time t. ) s(t) denotes the total susceptible population at time t. ) r(t) denotes the total number of recoveries and deaths at time t. ) dc(t), ds(t), dr(t) denotes the change in the states at time t. ) dw t is an incremental weiner process (brownian motion), which models the randomness in the evolution. • growth rate: the constant β denotes the growth rate, which factors the rise in the number of infections, due to interactions between susceptible and infected population. this parameter is a lumped constants which is meant to account for: (a) the population size, (b) reproduction number r of covid- , and (c) exposure-factor (which depends on mobility, precautionary measures, etc.). • γ is the rate of outcomes, i.e., the rate at which the infections are neutralized, which may be due to recovery or death. it is assumed that recovered persons would not spread the infections again (at least for a window of a month). • σ is a parameter used to model the stochasticity or randomness in the evolution, which may cause local deviations from the typical (exponential) trends. • p total is the population of the region, c and s are initial number of infections and susceptible individuals. the parameters of the sir model were optimized based on the data obtained for different european regions and india. the criterion for optimization was to simultaneously minimize the square integral error, terminal error and terminal rate error, between the actual data and daily samples of the simulated data. further, because we have data for more number of days for european countries, we try to use the parameters from those countries and appropriate it on the indian data which is for lesser number of days. the assumption is that india may be in the catch game (which, of course, we do not want) if the behaviour of people is taken to be similar. we have to base our predictions based on some gross assumptions under the given circumstances. note: we have taken the italian and german data days earlier than france and spain in order to reflect the earlier trend before their respective lockdown conditions. this will enable us to simulate for various levels of lockdown percentages (exposure factor). the sir model has been simulated and the parameters have been optimized based on the infection trends obtained for european countries for days, and india for days (after th march, when the infections started to show an exponential trend). the stochastic differential equations have been simulated using the euler-maruyama numerical integration method [ ] . we present below how our model is with respect to various european countries. the sir model has been simulated beyond the days of considered data for india, with parameters averaged over the european regions considered above, as well as with the parameters obtained for the indian region. this is done in order to demonstrate the possible growth of infections in india, based on the current trend so far (which is an optimistic case), or based on the european trend (which is the case to be prepared for). an exposure factor has been introduced to scale down the rate of infections β. this factor illustrates the additional reduction in mobility due to state-wide lock-down and quarantining measures, which are essential for maintaining the infections within reasonable limits. for india we have two sets, (i) using the available data only from india and (ii) taking the model parameters from considered european countries and projecting on indian case. note: we have four graphs in each set. for example, when we use % exposure, that means there is no lockdown. when we use % exposure half of the country is lockdown. this is to show the gravity of the situation. the italy and average european parameters have been used to project the future trends for the state of karnataka. the italy parameters have been chosen to predict the worst-case scenario, and the european average parameters. the initial number of infections is assumed to be (as of today) to accounted for additional unreported or undetected infections, over reported ones. we have made the assumption here that lockdown is in place and we have taken varied percentage of people obeying the rules. we only consider %- % of the population mobile. let us summerize very briefly what we observe. • below % exposure required for avoiding exponential growth, with european average trend. • below % exposure required for avoiding exponential growth, with italian trend. this is a very significant aspect in the whole of this work because, karnataka has less infrastructure and similar population. only advantage is that the mobility is impeded by lack of high speed transportation, which is a blessing in disguise compared to italy. • below % exposure required for decline in infections, with european average trend. • below % exposure required for decline in infections, with italian trend. that means we need to have less than lakh people to be on the street to be well within the limit because of heavy assumptions. • fig. suggests the inevitable need for an extended lockdow period for several months for karnataka, or until a vaccine is available. • by 'lockdown' we mean restricted mixing i.e. fewer susceptible individuals s(t) interact with fewer infected individuals c(t), thereby introducing a factor in the prod- this is a continuous work in which we are trying to find the model parameters everyday and project the possible scenarios, by varying the exposure factor for the rate of infection, as a result of evolving levels of quarantining. while this is not a completely verifiable projection, the model parameters look quite consistent, however they may reflect an over estimation of the projected number of infections in order to compensate for the unreported or undetected infections. thus, we may take the numbers in this note to be a guide for further action by the law enforcing authorities. infectious disease spread analysis using stochastic differential equations for sir model the numerical solution of stochastic differential equations key: cord- -altqn l authors: fernández-díaz, elena; iglesias-sánchez, patricia p.; jambrino-maldonado, carmen title: exploring who communication during the covid pandemic through the who website based on w c guidelines: accessible for all? date: - - journal: int j environ res public health doi: . /ijerph sha: doc_id: cord_uid: altqn l health crisis situations generate greater attention and dependence on reliable and truthful information from citizens, especially from those organisations that represent authority on the subject, such as the world health organization (who). in times of global pandemics such as covid- , the who message “health for all” takes on great communicative importance, especially from the point of view of the prevention of the disease and recommendations for action. therefore, any communication must be understandable and accessible by all types of people, regardless of their technology, language, culture or disability (physical or mental), according to the world wide web consortium (w c), taking on special relevance for public health content. this study analysed whether the who is accessible in its digital version for all groups of citizens according to the widely accepted standards in the field of the internet. the conclusion reached was that not all the information is accessible in accordance with the web content accessibility guidelines . , which implies that there are groups that are, to some extent, left out, especially affecting the elderly. this study can contribute to the development of proposals and suggest ways in which to improve the accessibility of health content to groups especially vulnerable in this pandemic. the use of the internet has increased in the last decade, as approximately % of the world's population now uses this medium. in , there are more than . billion people using the internet [ ]. therefore, not only has the capacity of access to the internet increased but more and more users have access to internet content, also as a source of information. in the face of a pandemic such as covid- , access to reliable information by citizens is crucial as a means of preventing the disease and enabling citizens to take action in certain everyday situations, as demonstrated in a study by the european parliament's european science-media hub [ ] . the world health organization (who) website is one of the main sources of information for the public, providing daily updates and interactive maps showing the evolution of the pandemic and offering credibility and security of information [ ] . in addition to the who, other international sources of information stand out, such as the european centre for disease prevention and control and the european commission's coronavirus and coronavirus response page, as well as the european medicines agency, among others. as has happened in the world with the zika and ebola pandemics, the internet is not only a source of communication for prevention and action in the face of health crises but also becomes a channel for misinformation [ , ] , making the need to share accurate and accessible information even more significant. the objective reasons for carrying out this study are mainly based on analysing the web accessibility of the world health organization (who) during the covid- pandemic. thus, the objective will be to determine whether the content offered to inform about the disease is prepared so that any person can access it, regardless of their technology (hardware, software, or network infrastructure), language, culture, or disability, whether physical or mental, as determined by the world wide web consortium (w c). this institution is an international community that aims to ensure that anyone can access the content offered by a website, promoting the social value it provides for all citizens [ ] . a lack of equity of access to health information can generate a digital divide [ , ] and affect the ability to deal the disease [ ] . consequently, it represents a social determinant of health. in addition, the elderly are an important group of citizens who benefit from web accessibility, since their skills are weaker as a result of age [ ] . bearing in mind that older people are considered one of the main groups at risk in this pandemic, it is especially important that they are the ones who have the most access to the content offered by information sources such as the who [ ] . from the point of view of web accessibility, it should be emphasised how few studies have been carried out on international and official organisations with important social responsibilities such as the who, as they are more oriented towards the education sector such as universities and local corporations such as town councils [ ] [ ] [ ] [ ] [ ] . most of these studies do not focus on recent recommendations but only on some that have been detected in the area of education [ , ] ; likewise, studies have shown that citizens' dependence on the media is more intense in crisis situations [ ] . however, traditional media and the internet play different roles for people in these public health crises, and the internet's dependence on individuals is greater than that of traditional media [ ] . therefore, one of the major contributions of this research is based on the originality of the analysis of universally accessible public health risk communication in times of global pandemic. likewise, the consideration that access to information is a health determinant is also relevant. this research is structured in the following blocks. after the introduction, the theoretical framework shows the who as the main health agency and information reference in the covid- pandemic. the methodology of the research is of an exploratory type, contemplating an analysis of the who' page using the wave tool and a manual review of each of the criteria of the content accessibility guidelines. the results obtained reflect the level of accessibility of the who according to international standards. the practical implications of this study highlight the need to raise awareness among health organisations such as the who about the importance of accessibility to their digital content in times of health crisis with the social objective of universal health information accessibility. the main contribution is the analysis of accessibility, taking particular account of elderly people, as one of the most vulnerable groups in this pandemic, and consequently, it provides practical proposals for addressing this challenge for health institutions. the who is an international organisation founded in , and since then, the world has undergone great political and economic changes, not only from the point of view of health [ ] . its headquarters are located in geneva, with offices in different countries and six regional offices [ ] . any country that is a member of the united nations can become a member of this organisation [ ] . in order to become a member, countries must agree to its constitution, which currently includes member states [ ] . among the different activities addressed by this organisation, one of the main objectives is to provide universal health coverage by supporting integrated health services for citizens. they offer prevention, surveillance and a response to possible risks that may threaten citizen security, for example, in pandemics such as covid- [ ] . this underlines the importance of this organisation as the importance of this organisation as the main source of information on this disease, which has become a pandemic caused by the coronavirus, the outbreak of which began in december in wuhan, china, and has affected the whole world [ ] . the who acts as an agency that promotes accessibility in all its fields, including digital, and provides recommendations on its website [ ] . for its part, the spanish association of scientific communication (aecc) [ ] highlights the who as one of the main sources of consultation for the citizens of affected countries. in the united states, . % of the population seeks health information on the internet [ ] . in order to offer as much information as possible on covid- , the who has created specific pages on prevention, symptoms and action protocols and even a solidarity page for donations to help research and detect the spread of the virus. these donations also go to unicef partners to support their work in communities that are the most vulnerable, such as children [ ] . it is important to note that an outbreak such as covid- causes important social consequences, affecting social distance in the most affected countries and generating more anxiety [ ] . all of the above have resulted in an increase in google searches for the keywords "world health organization" and "coronavirus world health organization" worldwide, which began to grow in early march, as can be seen in figure . this search result confirms that the who has a social responsibility to provide quality content and information that is accessible to all types of people, since as the network evolves, different challenges are being addressed, resulting in a continuous need for relationships and trust [ ] . moreover, it is a way of ensuring equity, eliminating disparities and improving the health of all groups [ ] . it is important to highlight the responsibility that the management of pandemics entails for the who, since depending on the type, it involves a problem of uncertainty when the end of the pandemic is declared, as occurred in the case of influenza a (h n ) in [ ] . lamb-white [ ] refers to the who's commitment to improving communicable diseases through the international health regulations (ihr) to improve public health, and this would therefore help countries to strengthen their capacity to achieve this. figure shows the significant increase in visits, unique visitors and pages per visit in the last months for the who website. moreover, it is surprising that the average duration of visits has also increased, so it can be said that the who website has been and is a reference for consultation on public health on a global level, especially in times of pandemics this search result confirms that the who has a social responsibility to provide quality content and information that is accessible to all types of people, since as the network evolves, different challenges are being addressed, resulting in a continuous need for relationships and trust [ ] . moreover, it is a way of ensuring equity, eliminating disparities and improving the health of all groups [ ] . it is important to highlight the responsibility that the management of pandemics entails for the who, since depending on the type, it involves a problem of uncertainty when the end of the pandemic is declared, as occurred in the case of influenza a (h n ) in [ ] . lamb-white [ ] refers to the who's commitment to improving communicable diseases through the international health regulations (ihr) to improve public health, and this would therefore help countries to strengthen their capacity to achieve this. figure shows the significant increase in visits, unique visitors and pages per visit in the last months for the who website. moreover, it is surprising that the average duration of visits has also increased, so it can be said that the who website has been and is a reference for consultation on public health on a global level, especially in times of pandemics such as covid- , as can be seen in the data. in addition, the semrush tool (semrush.com), comparing the who's traffic data with those of the website of the european centre for disease prevention and control (the second most popular website after the who's according to the european parliament's european science-media hub [ ] ), shows that the who website visits increased from the end of february to april. prevention and control (the second most popular website after the who's according to the european parliament's european science-media hub [ ] ), shows that the who website visits increased from the end of february to april. figure shows that the who pages related to the covid- pandemic were the most visited in the last months according to a study by the european science-media hub of the european parliament [ ] . specifically, this tool has shown that in countries such as spain, the keyword "coronavirus" has increased organic traffic on the who website, accounting for more than % of organic web traffic, as can be seen in figure , in addition to terms such as covid- being incorporated in the top positions, showing that citizens have real public health concerns through the search for these keywords that are incorporated into the ranking of new searches related to the who. apart from the organic traffic referenced above, it should be noted that the who is making great communication efforts to reach all citizens through the paid searches of search engines such as google, specifically through ads in different languages, depending on the search keyword. people with disabilities have many difficulties in becoming independent as a result of the lack of commitment of the different public policies in force. a report by the spanish committee of representatives of people with disabilities (cermi) states: "universal accessibility is the great failure of public policies in our country" [ ] (p. ). this causes people with disabilities to figure shows that the who pages related to the covid- pandemic were the most visited in the last months according to a study by the european science-media hub of the european parliament [ ] . specifically, this tool has shown that in countries such as spain, the keyword "coronavirus" has increased organic traffic on the who website, accounting for more than % of organic web traffic, as can be seen in figure , in addition to terms such as covid- being incorporated in the top positions, showing that citizens have real public health concerns through the search for these keywords that are incorporated into the ranking of new searches related to the who. [ ] ), shows that the who website visits increased from the end of february to april. figure shows that the who pages related to the covid- pandemic were the most visited in the last months according to a study by the european science-media hub of the european parliament [ ] . specifically, this tool has shown that in countries such as spain, the keyword "coronavirus" has increased organic traffic on the who website, accounting for more than % of organic web traffic, as can be seen in figure , in addition to terms such as covid- being incorporated in the top positions, showing that citizens have real public health concerns through the search for these keywords that are incorporated into the ranking of new searches related to the who. apart from the organic traffic referenced above, it should be noted that the who is making great communication efforts to reach all citizens through the paid searches of search engines such as google, specifically through ads in different languages, depending on the search keyword. people with disabilities have many difficulties in becoming independent as a result of the lack of commitment of the different public policies in force. a report by the spanish committee of representatives of people with disabilities (cermi) states: "universal accessibility is the great failure of public policies in our country" [ ] (p. ). this causes people with disabilities to apart from the organic traffic referenced above, it should be noted that the who is making great communication efforts to reach all citizens through the paid searches of search engines such as google, specifically through ads in different languages, depending on the search keyword. people with disabilities have many difficulties in becoming independent as a result of the lack of commitment of the different public policies in force. a report by the spanish committee of representatives of people with disabilities (cermi) states: "universal accessibility is the great failure of public policies in our country" [ ] (p. ). this causes people with disabilities to encounter physical and technological barriers in their daily lives. consequently, it suggests that institutions with competences could address this matter and pay attention to accessibility to minimise the digital divide [ ] , achieve health equity, and allow all groups to better face the disease [ ] . according to webaim, the internet is an opportunity for people who have some kind of disability, since it allows them to access information through diverse content quickly and by means of different devices and software, for example, screen readers for people with vision problems. however, these opportunities that the world wide web (www) should offer through websites are not sufficiently optimised and adapted to the different needs of citizens according to their disability [ ] . according to who data, the aging of the population and the increase in chronic diseases are one of the main reasons for the increase in disability rates, which is about % for the world's population [ ] . in fact, age is a physical and social determinant directly correlated with health [ ] . in times of crisis, communication through written messages is remembered more than those transmitted through other formats, so it must be not only accessible but also accurate so that it is understood by the majority of the population [ ] . studies have confirmed that because of the speed with which these types of diseases such as covid- are transmitted, citizens and different countries need to increase their vigilance and prepare themselves through preventive responses [ ] . communication is particularly important in this regard, as is access to equal opportunities for all. however, if information tends to be complex and ambiguous in terms of the interpretations that citizens may make, situations of panic and anxiety may arise [ ] . exceptional crisis situations such as the covid- pandemic generate greater attention or dependence on information, especially reliable and accurate information. in addition to this maxim, which can be contrasted with previous literature, internet penetration makes it easy to access information and increases the level of information that each person has, which is why the following research questions are posed: rq : does the who make itself accessible to all groups of citizens according to accepted standards in the field of the internet? rq : what aspects of web content analysis can be improved, and which audiences are affected? this research analyses the web accessibility of the who website based on the web content accessibility guidelines . in an exploratory way. the analysis was carried out during the covid- pandemic in march-may , coinciding with one of the world's most popular periods for citizens to search for information (figure ) . the methodology was combined using a web accessibility evaluation tool and manual analysis carried out by an evaluator [ , ] . the tool used for accessibility evaluation was the wave tool [ ] , developed by the webaim organisation. the website accessibility conformity assessment methodology (wcag-em) was used, which is considered in the web content accessibility guidelines . but is applicable to wcag . [ ] . as for the variables analysed, they belong to the web content accessibility guidelines (wcag), which explain how to make content more accessible to developers and other professional profiles related to web accessibility authoring and evaluation tools, including mobile accessibility [ ] . it should be recalled in historical retrospect that wcag . was a recommendation in may . it consists of a total of guidelines and priority , and checkpoints depending on the level of compliance [ ] . wcag . was recommended in december . unlike the previous ones, it is composed of guidelines and four principles-perceptible, operable, understandable and robust-with criteria for success [ ] . however, the latest guidelines recommended in june are wcag . , with a total of guidelines and compliance criteria; in this case, the w c has included new criteria, maintaining the four principles mentioned above [ ] . the web content accessibility guidelines present different conformance levels-a, aa and aaa [ ] [ ] [ ] [ ] . in the case of wcag . , the levels depend on satisfying the priority levels to ; for example, it is determined that level a is met when all the priority checkpoints are satisfied [ ] . however, in wcag . and . , the levels do not refer to priorities to ; for example, it is determined that level a is met when all the level a compliance criteria are satisfied [ , ] . in order to carry out a more in-depth analysis, six representative pages from the entire website were analysed (table ). the sample was selected by taking representative pages that allow the checking of each of the analysed criteria; for example, a page with forms must be checked to determine the compliance of the labels in the fields, or a page with tables must be checked to ensure that the conent is in an accessible form; additionally, videos must be checked for their audiovisual accessibility. the methodology (wcag-em) suggested by w c [ ] recommends selecting representative urls for each criterion: standard page: a second-level reference page of the website that describes the structure of the website. page with tables: a page that shows content laid out using tables. page with forms: registration forms, application forms, information forms, etc. result of a search: the information necessary for the location of contents is extracted and checked by means of a keyword search; in this case of analysis, the word "covid- " is used as an example. . page containing video: to analyse compliance with the guidelines in the case of videos. once the representative urls of the rest of the who website were selected, as shown in table , the compliance with each of the variables to wcag . was analysed. these are shown in tables and , divided into levels of compliance a or double a respectively: after analysing each of the criteria, the results obtained were collected with the data analysis tool (table ); the variables of the wcag . analysed are shown in the upper part of the table, facilitating manual data collection and the checking of compliance. the first step was to check if the success criterion could be applied to the analysed url and how many times it was applied to (a). the second step was to check whether the success criterion was approved (b) or not (m). the symbols have the following meanings (table ) : p: pages analysed for each service a: pages to which the criterion applies b: pages that are correct according to the criterion m: pages that breach the criterion as can be seen in table , the total number of pages analysed (tp) was calculated, the correct (tb) and incorrect (tm) pages were counted, and as a result, a percentage was obtained of the correct who pages, obtaining an average that represents the percentage of web accessibility compliance of each page analysed (%b). the formula is as follows: (%b) = (tb × /tp). source: author's elaboration based on the infoaccessibility observatory of discapnet [ ] for the manual assessment phase, table details the tools that enabled the level of compliance to be checked according to the wcag . guidelines on those points that required more in-depth review, apart from the wave tool, in the first phase. the analysis shows that the who website is % compliant regarding web accessibility based on the pages analysed; however, at the double-a level, the figure is slightly less than a % level of compliance. from the point of view of the four principles (appendix a, table a ) that underpin the wcag . -perceptible, operable, understandable and robust-it can be seen that the principle that is most complied with on the who website is understandable at both levels, with % and . % compliance, respectively. therefore, the who's digital health information is readable and understandable based on this principle [ ] . however, robust is the worst performer at both the a and aa levels, at % and %, respectively. it is precisely this principle that focuses on adapting the content to user applications and providing technical aids [ ] . the perceivable principle, directly related to the alternative text of the images, although it does not present outstanding values at level a with respect to the rest, is the second principle that best meets the double-a conformity criteria, with regard to the size and contrast of the text, benefiting those with vision problems derived from both age and sensory disabilities [ ] . finally, it should be noted that the operable principle shows significant differences upon comparison at both levels, worsening at the double-a level, and therefore, navigation aspects have to be improved [ ] . generally, the principles that are most closely adhered to are found in level a, so it is concluded that those in level aa are the ones that need to be improved for each compliance criterion analysed in the wcag . . with respect to the total number of errors detected by the wave tool and later analysed manually, it is worth highlighting in figure that the home page is the one with the most errors, followed by the form page and the page with tables. generally, the principles that are most closely adhered to are found in level a, so it is concluded that those in level aa are the ones that need to be improved for each compliance criterion analysed in the wcag . . with respect to the total number of errors detected by the wave tool and later analysed manually, it is worth highlighting in figure that the home page is the one with the most errors, followed by the form page and the page with tables. if one analyses it from the point of view of contrast errors, one will find that the page with the highest number of contrast errors is the table page, followed by the type page and the page with video ( figures and ). contrast errors are based on the fact that the visual presentation of the text and the images of the text must be sufficiently differentiated so that users with some type of visual disability can differentiate the text when reading, especially in the case of older age groups [ ] . if one analyses it from the point of view of contrast errors, one will find that the page with the highest number of contrast errors is the table page, followed by the type page and the page with video ( figures and ). contrast errors are based on the fact that the visual presentation of the text and the images of the text must be sufficiently differentiated so that users with some type of visual disability can differentiate the text when reading, especially in the case of older age groups [ ] . if one analyses it from the point of view of contrast errors, one will find that the page with the highest number of contrast errors is the table page, followed by the type page and the page with video ( figures and ). contrast errors are based on the fact that the visual presentation of the text and the images of the text must be sufficiently differentiated so that users with some type of visual disability can differentiate the text when reading, especially in the case of older age groups [ ] . regarding the non-text content errors detected, it can be seen that they also happen on all pages, especially form pages (figure ) , so people who need a screen reader will be especially affected because there is no alternative text for the images. regarding the non-text content errors detected, it can be seen that they also happen on all pages, especially form pages (figure ) , so people who need a screen reader will be especially affected because there is no alternative text for the images. from the point of view of the type of errors detected, figures and show each of them in detail. it is worth noting from the comparison between the level a and double a errors that they coincide in both cases, and there are a total of seven success criteria with % error rates for the pages analysed. of the total errors detected, the most significant are those referring to non-text content within level a, as they are directly related to the alternative text of the image by means of the alt tag, preventing screen readers from accessing the content by means of images for visually impaired citizens and, in the case of level aa, the visible focus, since if the user cannot clearly see where the keyboard tab is when browsing the page, it is difficult for them to conduct proper and understandable navigation through the content. regarding the non-text content errors detected, it can be seen that they also happen on all pages, especially form pages (figure ) , so people who need a screen reader will be especially affected because there is no alternative text for the images. from the point of view of the type of errors detected, figures and show each of them in detail. it is worth noting from the comparison between the level a and double a errors that they coincide in both cases, and there are a total of seven success criteria with % error rates for the pages analysed. of the total errors detected, the most significant are those referring to non-text content within level a, as they are directly related to the alternative text of the image by means of the alt tag, preventing screen readers from accessing the content by means of images for visually it is therefore determined that each of these errors detected requires a complete review that, in turn, contemplates alternative solutions based on guidelines established by the w c to comply with the requirements set by the wcag . . it is therefore determined that each of these errors detected requires a complete review that, in turn, contemplates alternative solutions based on guidelines established by the w c to comply with the requirements set by the wcag . . as a main point of discussion, it should be noted that the who [ ] states on its website that one of its objectives as an international organisation is based on improving universal accessibility to health services, both in internet media and in physical media; however, its website does not show any kind of statement on accessibility with which they currently comply. it should be emphasised that the home page is one of the pages with the most errors; taking into account the fact that it is usually the first page that a user consults before proceeding to browse the rest of the web, it should be a principal target for improvement. in addition, the who recommends adopting digital media for health education [ ] , so it could be considered a special committer to digital media. as is seen in previous research work, assuming this responsibility is necessary to ensure health equity and to reduce the digital divide, which can affect the ability to face the disease for some population groups [ , , ] . considering the studies that have been carried out on citizens' dependence on the media in times of public health crisis, most are based on analysing the differences between dependence on traditional media and that on the internet [ , ] , but they do not focus on analysing the information offered from the point of view of the accessibility of a particular agency and the social responsibility they have towards citizens facing a pandemic and seeking information. therefore, this study provides originality based on a specific case of accessibility in a health agency such as the who and provides points of improvement to make the content universal, at a crucial time of global pandemic, such as that presented by covid- . furthermore, this article reinforces the conclusions reached by other studies in which it is highlighted that the population seeks information on public health mainly through the internet [ , ] . from a technical point of view, more associated with web accessibility studies, it is worth mentioning that due to the recent approval of the wcag . , most studies have focused on the previous guidelines, so there is a shortage of studies based on the new guidelines, among which [ , ] , focusing on the education sector, stand out. based on the results obtained, it is considered that the who is not accessible to all groups of citizens according to the web content accessibility guidelines . , being less than % accessible at one of the levels analysed. it is concluded that many aspects need to be improved in order to make it fully accessible. one of the main online messages transmitted by the who [ ] is "health for all", and therefore, this research calls for "web accessibility for all" as the main aim and contribution to ensure that citizens have access to accurate, understandable and direct information; in short, there should be universal accessibility. it is also one of the overarching goals of the healthy people initiative that specifically pays attention to achieving health equity and improving the health of all groups [ ] . hence, the concept of accessibility in times of crisis such as the covid- pandemic is especially relevant, regarding the social value of the web. among the most notable errors are those concentrated largely within the principles understandable and perceivable, which shows that they are essential variables of communication with citizens, since they are directly related to content that is easy to understand in the first case and offer text alternatives for non-text content in the second case, especially for people with vision problems who use screen readers and even groups of elderly citizens who have vision problems as a result of physical aging. it is therefore determined that one of the aspects that most needs to be improved in terms of accessibility parameters is directly related to these two principles. with regard to the limitations, it should be mentioned that the analysis could be completed with a heuristic study that would include manual checking as well as checking over time. on the other hand, comparison with other key information sources and the incorporation of who web users or those involved in this health crisis could offer a more complete vision of the phenomenon of web accessibility during a pandemic as well as the evaluation of the websites of other institutions. with regard to future lines of research, since wcag . must also be implemented in the mobile applications of public administrations, the analysis of the mobile applications of international organisations with social implications such as the who is proposed. it has also been considered as a future line of research to carry out user tests, a practice recommended by the w c [ ] . there are also different systems for groups with physical disabilities, where information can be collected from sources other than websites-for example, with bots-so it is proposed for these lines of research be considered in the future to make this research work more substantial. the practical implications of this study are mainly based on the fact that international organisations with competence in the matter should review the structure and texts as well as everything related to the content in order to approach this challenge in an equitable way and provide the interested public with the same options of access to information. this study allows us to consider the accesibility of the who web resources with a special focus on elderly groups. the diagnosis performed will help health organisations to make decisions and to pay attention to critical points. the absence of text in images (non-text content) and errors in the html code (parsing) should be stressed. this study acts as a first attempt to analyse accessibility for the most representative health institution, the who. this is the main social value that this research aims to convey, that the main sources of information-international organisations, whose responsibility for health is crucial in times of global pandemics such as the one we are experiencing-can be given solutions that provide greater visibility to the information. global digital overview european parliament's european science-media hub. esmh selection: sources of information about coronavirus twitter, and misinformation: a dangerous combination? zika 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organisation world health organisation comité español de representantes de personas con discapacidad. derechos humanos y discapacidad an introduction to web accessibility, web standards, and web standards makers passing crisis and emergency risk communications: the effects of communication channel, information type, and repetition coronavirus disease (covid- ): a literature review public health communication in time of crisis: readability of on-line covid- information a comparative test of web accessibility evaluation methods applying heuristics to perform a rigorous accessibility inspection in a commercial context evaluating web accessibility of educational websites website accessibility conformance evaluation methodology. wcag-em web content accessibility guidelines (wcag) overview web content accessibility guidelines . web content accessibility guidelines (wcag) . accesibilidad web. wcag . de forma sencilla world wide web consortium (w c) world wide web consortium (w c) accesibilidad web en los portales de ayuntamientos de capitales de provincia who guidance for digital health: what it means for researchers involving users in evaluating web accessibility this article is an open access article distributed under the terms and conditions of the creative commons attribution (cc by) license the authors declare no conflict of interest. key: cord- -dyjpfvvf authors: gardner, anthony luzzatto title: foreign aid and humanitarian assistance date: - - journal: stars with stripes doi: . / - - - - _ sha: doc_id: cord_uid: dyjpfvvf together the us and eu provide two-thirds of global humanitarian assistance for the alleviation of emergencies arising from natural and man-made disasters and % of global foreign aid for longer-term development assistance programs. it is therefore vital that they continue their close partnership to ensure their dollars and euros are spent as effectively as possible in an era of increasingly tight budgetary constraints. the outbreak of ebola in west africa in is a good example of how the us and the eu successfully addressed (albeit belatedly) a major health crisis that could have turned into a global pandemic. in many areas in africa, they are collaborating closely on the foundation of shared priorities, including on food security, resilience, and electrification. they are also among the largest donors to the global fund to fight hiv/aids, tuberculosis, and malaria and to the global alliance for vaccines and immunisation. us and the eu to undertake repeatedly costly emergency measures to respond to disaster. indeed, some short-term solutions can be counterproductive: the dumping of food aid in local markets to address urgent food needs, for example, can undermine the ability of poor countries to develop sustainable farms and functioning markets. during my diplomatic post, i witnessed first-hand how the us and the eu work together in several areas of both humanitarian assistance and foreign aid. there are notable differences in how they pursue their agenda, including their degree of willingness to partner with the private sector and the military, but these have not prevented frequent and deep collaboration. in the fall of , my wife and i found ourselves in a large, abandoned field in downtown brussels to visit the training site of médecins sans frontières (msf), also known as doctors without borders, a non-profit and non-governmental international medical organization of french origin best known for its projects in conflict zones and in countries affected by endemic diseases. we were impressed by the explanations about how the modular kits in the large containers lying in a storeroom could be rapidly shipped to humanitarian disaster zones around the world and assembled on site within hours to provide functioning power generators, water purification, lodging for aid workers, medical treatment, and waste disposal. the logistical know-how and deep experience of msf made it the first, and sometimes most significant, responder to major medical emergencies. although the logistics were interesting, what really grabbed our attention was the sight of several of the organization's instructors training staff on how to put on and take off protective clothing to avoid contamination. the eight-piece clothing, entirely covering the human body, looked like a space suit but was more ominous because it was being worn on land to respond to the outbreak of the ebola virus in west africa in march . the scene reminded me of outbreak, a film released in starring many hollywood icons, including dustin hoffman, kevin spacey and morgan freeman. the film was about the public panic and response by military and civilian agencies in the wake of a fictional outbreak of an ebola-like virus in zaire, and later in a small town in california. a reallife outbreak of the ebola virus was ongoing in zaire when the film was released. the film contained exaggerations to enhance its shock value but wasn't too far from fact. we were transfixed by the demonstration. our guide told us that medical staff could only wear the space suit for several hours in real-world conditions on the ground because temperatures inside the suits can easily reach up to degrees centigrade (nearly degrees fahrenheit). the suit takes a long time to put on in order to ensure that the body is entirely protected from contamination. taking off soiled suits requires even longer (up to twenty minutes) because of a meticulous and rigorous twelve-step process that healthcare workers must repeat three or more times per day. it was clear that there is no margin for error. we were also shown disinfection routines, sample containment wards for infected patients, and the incineration units for soiled clothing and used equipment. during the first months following the detection of an ebola outbreak in march, msf was the most effective response of the developed world. msf remained on the ground throughout the crisis, even when staff members contracted the virus and died. on top of this risk of infection, msf workers also faced angry opposition from villagers suspecting that the treatment facilities were spreading the disease. msf had to close one facility in southern guinea after being attacked in april by a stone-throwing mob. villagers later killed eight african members of an msf team trying to raise awareness of ebola in the region. despite these risks, msf had nearly international workers and local employees fighting the outbreak by the time my wife and i were touring the facility in brussels. by contrast, the eu and its member states struggled to come up with a coherent and decisive response in the early months of the crisis. this was disappointing in light of the eu's deep partnership with africa, europe's proximity to the continent and greater vulnerability to a spread of ebola, and the history of france and britain as former colonial powers in guinea and sierra leone. similarly, the initial us response was slow and focused on domestic preparations. it took until the fall of for the us and the eu to respond in a coordinated fashion. our joint response should have been more effective. as i toured the msf facility i recalled how, as a young director for european affairs in the white house in - , i had played a role in the us-eu new transatlantic agenda that had specifically flagged the importance of transatlantic cooperation on infectious diseases: we are committed to develop and implement an effective global early warning system and response network for new and re-emerging communicable diseases such as aids and the ebola virus, and to increase training and professional exchanges in this area. together, we call on other nations to join us in more effectively combating such diseases. we did not make as much progress on this key challenge as we would have liked in the subsequent twenty years. our msf guide explained that an ebola outbreak starts when a human has direct contact with the blood, body fluids, or organs of infected animals, such as bats, chimpanzees, monkeys, or gorillas. the transmission of the virus from that human to other humans occurs through personal direct contact either with the patient's blood or other body fluids (including sweat and saliva) or through contact with objects, such as needles and syringes, that contain these fluids. transmission of the virus often occurs through caregivers such as family members or medical personnel in homes or healthcare environments where infection-control practices are weak. ebola crises, such as the one in zaire that occurred when outbreak was playing in movie theaters, can be amplified by the transmission of the virus in overcrowded hospitals and burial practices in which highly infectious corpses are washed or touched by family and members of the community in a sign of love for the deceased. ebola is a disease from one of the deeper circles of dante's hell. fortunately, it is not transmitted through the air, like influenza or tuberculosis, or transmitted before symptoms appear, like measles or hiv. nonetheless, it can result in terrible consequences for its victims and can spread quickly if not brought under control. in its early phase, victims of ebola can be misdiagnosed because it is relatively rare and because some of the symptoms, including fever, fatigue, muscle pain, and headache, are difficult to distinguish from those of other infectious diseases, such as malaria and typhoid fever. victims often subsequently experience vomiting, diarrhea, rashes, and hemorrhages resulting in internal and/or external bleeding. the dysfunction or collapse of multiple body organs leads to severe injury and often death. the fatality rate varies from to %, depending on the strain. the ebola outbreak in west africa in was first reported by the world health organization on march in a remote, forested region of south-eastern guinea bordering liberia and sierra leone. multiple chains of transmission of the virus had gone unrecognized for months. by the end of the year, the virus had claimed lives and had infected , persons; by march , the numbers were dead and , sick; and by the time the crisis had been brought under control in march , the dead numbered , . the number of infections and deaths far exceeded those in approximately twenty previous outbreaks since the s in central and eastern africa; in each of those outbreaks, the number of reported cases never exceeded . there were many reasons for the severity of the crisis. like the remote areas of central and eastern africa where prior outbreaks occurred, the virus appeared in a remote area of guinea. normally, this might have helped contain the disease, but in an unfortunate twist of geographic fate the region lay at the junction of guinea with sierra leone and liberia where people regularly move across borders. the region's lack of rudimentary public health infrastructure, partly because of its recent history of civil war and violence, and its lack of experience with a previous outbreak of the virus compounded the problem. as the virus spread to urban areas and expanded into an epidemic, the number of cases quickly overwhelmed the capacity of diagnostic and other healthcare facilities. at the onset of the outbreak, there was a very small number of experienced healthcare workers to deal with it, including in europe and the united states. even at msf, involved in most of the prior outbreaks, there were only "veterans." medical teams were simply not prepared to deal with a disease that kills at least % of its patients and for which no treatments existed. had ebola been a first world disease, there would have been a vaccine. but there wasn't because, in the eyes of the major pharmaceutical companies, the numbers of patients are small, and nearly exclusively in the third world. the business case for the recovery of significant upfront research and production costs had simply not been present. some ebola treatment units (etus) were filled beyond capacity, requiring the facilities to turn away people suspected of having the disease, thereby fostering new chains of transmission. one of the critical factors in bringing an ebola outbreak under control-an exhaustive tracing of contacts between victims and others with whom they had been in contact-was absent. moreover, poor infection control in hospitals led to many infections and deaths among healthcare workers and a rapid collapse of the region's healthcare system. the control of other devastating viruses, such as malaria, and even the provision of routine medical services declined. i had seen reports from our embassies that people who collapsed from heart attacks were sometimes left to die because no one wanted to touch them for fear that they had contracted ebola. children started missing out on education because of school closures. generalized fear and even panic threatened to devastate the region's economies, especially by reducing agricultural output and trade, while driving up prices. my friend and former colleague samantha power, us ambassador to the united nations during president obama's second term, visited brussels several times to brief and coordinate with eu officials on the ebola crisis after her trips to the infected region. on one of those trips, she relayed reports that farming communities were "eating their seeds," indicating not only that current harvests were poor but warning that future harvests and even food security were in danger. a total collapse of civil society was imminent as governments lost control of the situation. in the early days of the crisis, the eu allocated extra emergency funding to msf and other humanitarian organizations, such as the red cross and red crescent, the international medical corps, save the children and the international rescue committee. the aid contributed to the faster deployment of doctors and nurses and the purchase of diagnostic equipment and medical supplies. disaster assistance response teams of the us agency for international development (usaid) and teams from the atlanta-based centers for disease control and protection (cdc) were deployed to the region to carry out an assessment of what needed to be done. the us airlifted significant amounts of personal protective gear, generators, and medical equipment. by early summer, it seemed that wiring funds and providing assistance from a distance would be enough as the number of reported cases leveled off and then dropped, suggesting the outbreak could be contained as in the previously reported outbreaks in africa since . instead, the virus spread. by late july it reached, for the first time in history, densely populated metropolitan areas, not only in the three countries of origin but further afield. a traveler with ebola had flown from monrovia, liberia, to lagos, nigeria, africa's most populous city (with million inhabitants) where he had contact with multiple people who later contracted the illness. a massive effort by the nigerian government, assisted by the cdc, managed to contain the outbreak to just cases in two cities. by early august, the world health organization categorized the outbreak as a "public health emergency of international concern," a declaration that caught media headlines and unlocked significant new funding. claus sorensen, an old friend and the director-general of european civil protection and humanitarian aid operations (echo), conceded that in an ebola crisis "speed is of the essence, and there is a feeling that all of us have been behind the curve." the decisive factor in galvanizing action in the eu and the us was a series of shocking announcements regarding the ebola infections outside africa. in late july and early august, two us citizens, including a doctor with samaritan's purse, were repatriated to atlanta, where they (successfully) underwent treatment for ebola in a specialized isolation ward in emory university hospital. they were the first two patients ever to receive such treatment in the united states. in early october, two spanish priests died in madrid after contracting the virus in sierra leone; a nurse who had treated them also tested positive (but later recovered), the first person to have been infected outside of west africa. shortly thereafter a liberian national who had recently returned from liberia to the united states died of ebola in a dallas hospital. the nurses who had cared for him contracted the virus but recovered. on top of the shock of ebola infections appearing in europe and the united states, public health authorities on both sides of the atlantic conducted modeling about the potential spread of the virus. the results were sobering: in september, the cdc estimated that approximately , ebola cases ( . million cases when corrected for under-reporting) could occur in west africa by january , , if approximately % of all persons with new cases were not effectively isolated. the world health organization projected that new ebola cases could reach , per week by december. when respondents to a telephone poll were asked in october whether they were concerned that there would be a large outbreak of ebola in the united states within the next months, % reported that they were "very" or "somewhat" concerned. the initial us response (before october) was largely domestic because the country was not prepared for an epidemic of this magnitude. when the crisis broke out, only one facility in the united states (the cdc laboratory in atlanta) was qualified to test for ebola and there were only three facilities that could treat ebola patients; by january there were laboratories in states that could do so and by october there were treatment centers in states. during that period, , healthcare workers received instruction on how to identify, isolate, diagnose, and care for patients under investigation for ebola. the cdc and customs and border protection implemented intensive screening of air passengers arriving from west africa. there was significant public support for cutting off all air links with west africa and quarantining anyone who had recently been in the region. while generalized measures of this kind were avoided, the defense department did impose a -day quarantine for all personnel returning from ebola-affected areas, regardless of risk, because of political rather than scientific considerations. the presidential commission established to review the government's response to the ebola crisis was scathing in its final report. it criticized federal, state, and local unpreparedness to cope with the threat of an epidemic and the government's focus on the political implications of public reactions rather than on the underlying health concerns. by the end of the year, the us government's response started shifting decisively toward attacking ebola at its source. president obama was clear about the stakes: this is an epidemic that is not just a threat to regional security -it's a potential threat to global security if these countries break down, if their economies break down, if people panic. that has profound effects on all of us, even if we are not directly contracting the disease. the united states and the eu (as well as its member states), together and in parallel, supported by the african union and several african countries, finally undertook a major and highly successful effort to bring the crisis under control. washington focused on liberia, while paris focused on guinea and london on sierra leone. speaking at the cdc in mid-september, president obama announced that the us military had recently dispatched personnel to monrovia to establish a base under the control of us africa command. its main objectives were to build etus, including new isolation spaces and more than beds, and to recruit and train more than medical personnel to staff them. the department of defense built seven mobile laboratories in west africa that cut turn-around times for testing blood samples from five to seven days to between three and five hours, thereby freeing up bed space in overcrowded clinics and hospitals. the uk and france quickly followed the us example with similar military missions of their own to sierra leone and guinea to build hospitals and diagnostic centers; our experts often singled out the uk effort as being rapid and effective in bringing down new infection rates. in addition to these efforts, thousands of cdc employees and government-supported civilians were deployed in all of west africa, partnering with national governments to train healthcare workers, treat patients, staff field laboratories, trace contacts of patients to identify chains of transmission, develop border and airport-screening programs, promote safe burials, and educate communities. the united states was to airlift more than metric tons of personal protective equipment and other medical and relief supplies during the subsequent months. in the fall of , the white house announced major efforts to accelerate the development of vaccines (to prevent new infections) and therapeutics (to treat those already infected). and in december the us congress overwhelmingly supported legislation providing $ . billion in emergency funding for the cdc and other health services, the state department, and usaid; much of this funding was earmarked for the prevention, detection, and response to the ebola crisis in west africa, as well for efforts to assist in the region's recovery. the eu made an important contribution as a coordinator and donor. the european centre for disease prevention and control, headquartered in sweden, coordinates the work of health experts in different countries but does not have its own emergency-response teams. similarly, eu member states, rather than the european commission, dispose of their own medical personnel, hospitals, labs, and stock of specialized equipment. but as the only european body with a global picture of the fast-moving epidemic, the european commission successfully played the role of "traffic cop" to ensure that europe's response was consistent and effective. that role included the identification of the type and destination of emergency supplies for west africa, providing a clearinghouse of information about the crisis and the disease, and the creation of a list of available member state assets relevant for the treatment of ebola in europe. the european commission also played a key role in identifying european assets and trained personnel that could be deployed for the medical evacuation of patients back to europe and in negotiating a us-eu agreement about when us and european patients could call on their respective emergency medivac "air bridges." ensuring that international healthcare workers could be airlifted to equipped facilities in europe within hours was critical to the ability of the us and eu to recruit such workers. in october, the eu appointed christos stylianides, commissioner for humanitarian aid and crisis management, as eu ebola coordinator to ensure that the eu institutions and member states acted in a coordinated manner with each other and with international partners. in addition to the directorate-general for humanitarian aid, other commission departments were involved in the response to the ebola crisis: these were principally the directorate-general for international cooperation and development (devco), the counterpart to usaid and responsible for foreign aid, and the european external action service (the eu's diplomatic service with delegations in most countries around the world). the role of other departments also had to be coordinated: the directorate-general for health identified facilities in member states that were willing and able to accept ebola patients; the directorate-general for internal affairs (including justice and law enforcement) coordinated entry and exit procedures at airports in case of travelers suspected of having ebola; and the directorategeneral for research worked to promote vaccines and therapies. the eu was also an important donor to help combat ebola in west africa. the european commission and eu member states contributed almost e billion (without counting the value of in-kind contributions from many member states such as personal protective equipment, vehicles, and field hospitals). of this total, the european commission contributed e million out of the eu budget for emergency measures, financial support for the african union's own medical mission to the region, and long-term relief (such as budgetary support for the restoration of vital public services and the strengthening of food security). moreover, the european commission announced substantial funding from the eu budget to promote projects on ebola research, including immediate largescale clinical trials of potential vaccines and tests of existing and novel compounds to treat ebola. the european commission also partnered with the european pharmaceutical industry in launching a e million longer-term research program involving clinical trials of new ebola vaccines, the development of fast diagnostic tests, and new approaches to manufacture, store and transport vaccines. in summary, while the us and eu were both slow in responding to the ebola crisis, by the fall of they had significantly scaled up their efforts and were working very closely together to provide an effective series of measures that brought the crisis under control by the summer of . the lessons learned from that dramatic experience-including european commission coordination of eu member state activities and intensive us-eu coordination to combat epidemics-are important for coping with future humanitarian disasters. the lessons enabled the us and eu to respond to an outbreak of ebola in the democratic republic of congo (drc) in may , the second worst in history and the longest and deadliest of the nation's nine previous outbreaks. us-eu cooperation will be equally important in dealing with the outbreak of coronavirus. while ebola and other epidemics are just one area where the us and eu have worked well in humanitarian assistance, they are emblematic of the many other examples of how they are indispensable partners in alleviating suffering around the globe. one example is their delivery of aid to those suffering from the syrian civil war that has displaced over million people and killed , . the war has been catastrophic, but it would have been far worse without us and eu efforts. under the eu's - multi-annual financial plan, the european commission's annual humanitarian assistance budget averaged e billion per year and is projected to rise under the - budget cycle. in addition to the formal budget, the eu has drawn from other sources to spend hundreds of millions of euros annually to respond to unforeseen events and major crises, including the humanitarian disaster caused by the syrian civil war and the refugee crisis in - . several eu member states-especially the uk, germany, and sweden-are generous donors of humanitarian assistance as well. together with the eu, they provide roughly the same amount of funding as the united states ($ billion per year as of ). us funding, largely administered through a specific bureau within usaid (the office of us foreign disaster assistance), has responded to the same emergencies as the eu-not only the ebola outbreak, but also the syrian civil war and many other crises concentrated in the middle east and africa. both the us and the eu contribute significant amounts through united nations agencies such as the un high commission for refugees, unicef, and other non-governmental organizations such as the world food program and the international red cross. the predominance of the us and the eu as humanitarian assistance actors means that their practices shape those of other donors, including states, ngos, and multilateral organizations. when we join forces to minimize overlaps or inconsistent approaches, we ensure that our dollars and euros have maximum impact, leading to real improvements in the lives of millions of people affected by humanitarian assistance. as one study rightly pointed out: failure by these two parties to enhance their cooperation…would result in additional, yet avoidable, human death and suffering…and could lead to increased insecurity and instability across the globe., threatening us and eu strategic interests. us administrations of both political parties have recognized the importance of this partnership and for good reason. the george h. w. bush administration launched an annual strategic dialogue on humanitarian assistance with the eu at senior levels in usaid, the state department and echo, supplemented with regular contacts on the ground among field officers. in the new transatlantic agenda concluded in under the clinton administration, the us and the eu set forth an extensive list of areas where they should work closely, including improving the effectiveness of international humanitarian relief agencies, and urged the creation of joint missions whenever possible, greater operational coordination, staff exchanges, and information sharing. close dialogue has continued since then, despite the turbulence of us-eu relations during the presidency of donald trump. the us and the eu provide an even larger share of total foreign aid than they provide of total humanitarian assistance. the united states is the largest single provider of foreign aid, accounting for one-quarter of the $ billion disbursed worldwide every year. but the eu, together with its member states (especially germany, the united kingdom, and france), provides over half the total foreign aid disbursed. while the us contribution appears generous, it represents only slightly more than % of the us federal budget and about . % of us gdp, far below the equivalent percentages of many eu member states. the us-eu partnership on foreign aid has worked well, in part because the two are the biggest players globally. an even bigger reason for the successful partnership is that they share values and objectives such as the promotion of human rights, democracy, good governance, gender equality, and open markets. nonetheless, their policies occasionally reflect different outlooks and priorities. us foreign aid policy is often shaped by national security concerns, especially during major wars, and devotes significant resources to military and non-military security assistance (concentrated in afghanistan, israel, egypt, and iraq). moreover, us foreign aid is sometimes used as a tool to open global markets to us exports and is often tied to the purchase of us the united nations has urged countries to spend at least . % of their gdp on foreign aid, a target met by sweden, luxembourg, norway, denmark, the netherlands, and the united kingdom. according to an opinion poll conducted in the united states in , americans on average think that % of the federal budget is spent on foreign aid; given this misconception, many believe that the us should reduce its spending (unlike their european counterparts who support giving generously). see bianca dijulio, jamie firth, and mollyann brodie, "data note: americans' views on the u.s. role in global health," kaiser family foundation, january , . https://www.kff.org/globalhealth-policy/poll-finding/data-note-americans-views-on-the-u-s-role-in-global-health/. goods and services (especially food). some us aid is explicitly made conditional on the recipients' agreement to take certain actions. for example, the millennium challenge corporation, a foreign aid agency established by but independent from the us government, provides large five-year grants to countries that meet certain political and economic criteria and sign up to "compacts" detailing the domestic policies they will pursue. by contrast, the eu's commitment to development assistance is grounded in a widely shared feeling among the european public that the eradication of extreme poverty is a moral obligation and an investment in europe's long-term security. unlike the united states, that has the luxury of large oceans on either side, europe is far more exposed to instability on its borders. without development assistance, significant migration flows into europe from northern africa and the eastern mediterranean are certain. while the eu's development assistance is not shaped by military considerations or the desire to promote exports, the eu has recently moved closer to the us view that aid should be subject to strict conditions about the behavior of recipients, especially their willingness to undertake economic reforms. most us presidents have considered foreign aid as an investment in global and us security and prosperity, and a significant pillar of us foreign policy, rather than a gift to undeserving foreign countries. president trump has departed from that consensus by considering foreign aid wasteful and ineffective unless given to allies. in his speech before the un general assembly, president trump made the latter point in stark terms: "moving forward, we are only going to give foreign aid to those who respect us and, frankly, are our friends." his white house now appears to see africa largely as a playground of big-power rivalry, where chinese and russian influence is on the rise. other than as a destination for growing us exports and investment, africa appears to be of little inherent interest. the president's budgets have regularly proposed massive cuts in foreign aid. his budget has called for % cuts in the foreign aid budget-a target that is not only immoral but geo-politically nonsensical in light of the growing influence of china and russia in africa and other parts of the developing world. fortunately, congress has maintained most of the programs, partly in sympathy with the argument that deploying diplomats and development experts today is cheaper than deploying troops tomorrow. in an open letter to congress in , more than retired admirals and generals argued cogently: we know from our service in uniform that many of the crises our nation faces do not have military solutions alone…[the] state department, usaid…and other development agencies are critical to preventing conflict and reducing the need to put our men and women in uniform in harm's way…the military will lead the fight against terrorism on the battlefield, but it needs strong civilian partners in the battle against the drivers of extremism -lack of opportunity, insecurity, injustice and hopelessness. as with humanitarian assistance, the new transatlantic agenda also tried to introduce greater structure around us-eu cooperation on foreign aid, especially in their joint efforts to "help developing countries by all appropriate means in their efforts towards political and economic reforms." but efforts at a structured dialogue suffered from disagreements in other areas and were only revived when the us and eu launched a development dialogue in . although the annual meetings at ministerial level have not always occurred as envisioned by the dialogue, regular meetings at senior levels and continuous technical exchanges between staff, at headquarters and especially in the field, have enabled the parties to exchange information on policies and programs, as well as to promote greater policy consensus and coordination. the us-eu annual summit in that i attended several weeks after i arrived in brussels issued an ambitious set of targets for the parties' development agenda. some of the objectives were aspirational, such as delivering on the "unfinished business" of the millennium development goals, a set of eight extremely ambitious international development goals for (including the eradication of extreme poverty and hunger) that had been established by the united nations in . these goals have been replaced by the un's global goals for sustainable development, an agenda of social and economic development objectives for that the us and the eu support. us-eu cooperation on foreign aid is not always easy because each party has different budgetary cycles, implementation systems, and measures to ensure accountability. much of the "real" day-to-day work occurs in the field in dozens of countries, making coordination from headquarters in washington and brussels rather complex. even when coordination is successful, moreover, it can be overtaken by fast-moving events. representatives of usaid often observed to me that they preferred to deal with eu member states-such as the uk, the netherlands, sweden, and denmark-because they were less bureaucratic and nimbler than the eu. nonetheless, the constant dialogue between the us and the eu has enhanced their mutual trust and the effectiveness of their foreign aid programs. one example is how each party has increasingly relied on the other's geographic expertise: usaid relies on france and the eu in francophone africa where the us has a relatively more modest presence; the eu relies on the united states in the horn of africa where the former lacks the latter's resources and expertise. during my diplomatic mission, usaid and devco evidenced their mutual trust and intent to specialize by signing an agreement enabling each to fund the other's projects. an important by-product of the us-eu dialogue on development is that it has forced each side to coordinate better among its own government departments. in the case of the eu, that means devco, the european external action service and also the directorate-general for neighbourhood and enlargement negotiations (that implements assistance programs in the western balkans, turkey, the former soviet union, and the maghreb). in the case of the united states, that means not only usaid, but also includes the state department and even the defense department, the department of health and human services, the us treasury, and the department of agriculture. this internal coordination can sometimes be more challenging than transatlantic coordination, as i witnessed many times. the us and the eu development dialogue has covered a wide range of topics. one of the areas of focus has been the challenge of how to improve the "resilience" of developing countries. in the development context "resilience" means the ability of people, households, communities, countries, and systems to mitigate, adapt to and recover from shocks and stresses in a manner that reduces chronic vulnerability and facilitates economic growth that is fairly distributed across society. resilience can be strengthened in many ways: for example, with cash transfer programs to provide a safety net to the poorest households in drought-prone areas; by vaccinating livestock and planting crops that are more resistant to pests and drought; through early warning systems and insurance against extreme weather, plagues, and earthquakes; and with budget support for countries to maintain vital state functions, including policing and health care services. the dialogue has also covered topics such as the effectiveness of aid in achieving economic or human development, adaptation to climate change (especially developing countries' implementation of low-carbon growth strategies and adaptation to harsher weather), improving the availability and accessibility of food ("food security"), the interplay between security and development, electrification (with a focus on rural areas in sub-saharan africa), and health. the last four areas merit further elaboration. at the g- summit in l'aquila, italy, members of the g- and other donors, including the eu, pledged $ billion to support food security over a three-year period. that initiative led to the launch at the g- summit at camp david in may of a new alliance for food security and nutrition between the donors and ten african countries suffering chronic food shortages. the purpose of the initiative is to attract private investment in agriculture, to complement public investment and create the right conditions for the recipient countries to increase agricultural productivity, adopt improved production technologies (including improved seed varieties), and improve their post-harvest management practices to reduce their dependency on food imports and food aid. under the "cooperation frameworks" signed with the donor countries, the recipients agree to implement reforms in a wide variety of areas, including infrastructure improvements, regulatory and tax reforms, and easier conditions for the marketing and trade in farm products. another important example of us-eu cooperation to promote food security in the developing world was the agreement by the bill and melinda gates foundation and the european commission to provide $ million each to fund agricultural and climate-change research during - to assist farmers with crop improvement, protection, and management. the us and the eu have both focused on the importance of providing security as a precondition for effective long-term development. their common views have translated into practical consequences on the ground. for example, the eu and the us ambassadors to south sudan, working with the united nations, averted a military confrontation between two tribes a few years after the country achieved its independence in . the lou nuer and murle tribes had been fighting each other for decades over cattle, with revenge killings occurring frequently. the ambassadors traveled together by helicopter to remote and dangerous areas to negotiate with the tribe's leadership and local elders to mediate an end to the impending conflict. as major providers of aid, the eu and especially the us were successful because they insisted that peace was a precondition for continued aid. electrification is another focus of us-eu cooperation on development. two-thirds of the population of sub-saharan africa, around million people, lacks access to power. that number is growing as rapid population growth creates demand that outstrips increased supply from investments in electrification. the remaining one-third cannot consume as much power as it would like because of blackouts and brownouts. in rural areas, the average electrification rate is only %. the main reasons for this situation include droughts that affect hydropower capacity, aging infrastructure and poor maintenance, unreliable fuel supply and inadequate transmission and distribution capacity. lack of electricity has numerous dramatic effects: for example, it stunts industrial growth and agricultural yields, hurts healthcare services (such as hospital care and the delivery of drugs requiring refrigeration), impedes digital connectivity that is increasingly essential to participate in the knowledge economy, and increases the number of premature deaths, especially among women and children, because of household air pollution caused by the use of solid biomass for cooking and of candles and kerosene lamps for indoor lighting. electrifying africa, especially sub-saharan africa, is therefore crucial to progress; at the same time, electrification using cleaner fuel sources, such as natural gas and renewable energies, will be key to avoid major harm to the environment from meeting the energy needs of a rapidly growing population with dirty coal or oil. ensuring that all people in sub-saharan africa have access to electricity by , one of the un's sustainable development goals, will require a major effort by the region's governments and the international community, above all the united states and the european union. according to various estimates, the region will need to increase its electrical capacity by about gigawatts and invest at least $ billion per year to achieve this goal. attracting that investment from the private sector is a huge challenge because almost none of sub-saharan electric utilities are currently financially sustainable due to artificially low tariffs, low operational efficiency due to losses during transmission and distribution, and poor bill collection. wasteful subsidies incentivize inefficient forms of energy, disincentivize maintenance and investment, and overwhelmingly benefit higher income groups. political patronage, corruption, and a poor regulatory environment present further challenges. investments in the electricity sector are overwhelmingly in the traditional fossil fuel sector generating power on the grid rather than in the renewables sector generating power off the grid. the latter, especially in the form of solar photovoltaic, small hydropower and small wind turbines, are especially relevant for the three-fifths of the population that lives in rural areas. even despite ongoing technological improvements that increase efficiency, renewable energy projects require significant upfront capital commitments and high transaction costs relative to the amount of power produced and the return on investment. sub-saharan africa will only be able to substantially increase electrification rates, especially with renewable energy projects in rural areas, through energy sector reform and international public-private partnerships that mobilize private capital. there are dozens of international initiatives originating in asia, the middle east, europe, and the americas to improve access to power in africa. china is increasingly active, including in sub-saharan africa where chinese contractors (the vast majority of them state-owned) were responsible for % of new electrical capacity between and . the us and the eu are working closely to align their initiatives to promote electrification in sub-saharan africa. in , they signed a memorandum of understanding that outlined their cooperation to reduce energy poverty and increase energy access in sub-saharan africa. although the mou is non-binding and does not require either party to make financial commitments, it establishes a structure for cooperation in several areas, including joint financial support, stimulating private sector investment, and the alignment of technical assistance and reform efforts. as of , there were separate initiatives originating from the member states and the eu institutions. while it is understandable that various member states wish to have separate initiatives to promote national political and commercial interests, it appears rather inefficient for the eu institutions (the european commission and the european investment bank) to have numerous ones as well. nonetheless, during the five years ending in , the eu budget alone allocated more than e . billion in grants to support sustainable energy in sub-saharan africa; those grants enabled the private sector to commit several times that amount in equity and debt capital as well. together with the member states, the eu has supported projects that have brought electricity to more than million people in the region. one of the key projects is the european commission's electrification financing initiative (electrifi) to support the adoption of renewable energy, with an emphasis on decentralized energy solutions in rural areas around the developing world, principally in sub-saharan africa. usaid not only assisted the european commission to structure the program but also approved a us investment of e million in electrifi. that investment represented a crucial "seal of approval" that enabled the european commission to access a far larger pool of capital than would otherwise have been possible. electrifi provides financing and technical support, even at an early stage and in partnership with other funders, to enable projects to overcome gaps in available market financing and achieve maturity in order to attract private long-term capital. the united states has also been active in promoting the electrification of sub-saharan africa. power africa, announced by president obama in , is the largest public-private partnership in history, involving many agencies of the us government, african governments, more than memorandum of understanding between the united states and the european union for reducing energy poverty and increasing energy access in sub-saharan africa, signed july , . https://www.usaid.gov/sites/default/files/documents/ /eu% signed% mou% from% july% % .pdf. private sector partners and international organizations like the african development bank and the world bank. power africa was underpinned by the us electrify africa act of , passed with overwhelming bipartisan support to promote african developments, as well as to assist us exports and counter chinese influence. power africa was one of the few obama-era executive decisions that president trump did not cancel upon entering office; indeed, his administration supported it as a model for how governments can leverage private capital to build infrastructure. the aim of the initiative has been to finance by gigawatts of electricity capacity and million new domestic electricity connections, especially from renewable projects in rural areas, by unlocking sub-saharan africa's substantial wind, solar, hydropower, natural gas, and geothermal resources. by the end of , power africa had attracted over $ billion in commitments and had catalyzed about $ billion in investment into power projects and over gigawatts of capacity. these projects connected about million homes and million people. most of these connections are from solar lanterns that power a single light and enable the charging of a mobile phone. as basic as that may sound, even such connections can result in dramatic improvements in livelihood. power africa is moving beyond these connections to include larger, more on-grid power projects using non-renewable sources. in addition to the power sector, both the us and the eu are actively engaged in the promotion of global health. for example, they are the main contributors to the global fund to fight hiv/aids, tuberculosis and malaria and to the global alliance for vaccines and immunisation (gavi). they sit on the governing boards and closely align their policies. founded in as a partnership among governments, nongovernmental organizations, and the private sector, the global fund raises and invests the world's money-about $ billion per year-to support programs in more than countries that combat the three deadliest infectious diseases. the eu (the european commission and eu member states combined) and the united states provide roughly and % of the global fund's financing, respectively. the health programs supported with this money have reduced the number of deaths caused by hiv/aids, tb, and malaria by one-third since and have saved million lives, the majority of these in sub-saharan africa, as of the end of . the global fund has enabled millions to be on antiretroviral therapy and therefore to be spared the death sentence that hiv/aids used to represent. of the million people living with hiv, million are on antiretroviral therapy ( . million of these thanks to the global fund). improved access to hiv treatment has cut the number of aids-related deaths in half since the peak in , from . million to under million in . however, hiv infections remain very high and especially among adolescent girls and young women who are up to eight times more likely to be hiv positive than young men in some african countries. on the current trajectory, the global fund is unlikely to meet its goal of reducing new infections to , globally by . in addition to the terrible human cost of the disease, the economic impact of hiv/aids is estimated to be over $ billion in lost earnings in . the global fund provides more than % of all international financing to combat tb. the global fund has disbursed about $ billion in the fight against tb by the end of . much of this has focused on expanding molecular diagnostic technology which delivers faster and more accurate results, supporting programs that identify those living with the disease without treatment, and enabling millions to be treated. progress is being made: the mortality rate for tb fell by % between and . but tb remains a serious threat to global health security because it is highly contagious, airborne and increasingly drug resistant. it remains the leading cause of death from infectious disease, with . million deaths per year, not including hiv co-infections. deaths from drug-resistant tb are responsible for about one-third of all deaths due to antimicrobial resistance worldwide; if trends continue, . million people will die of drug-resistant tb by , costing the global economy trillions of dollars in lost output. the global fund is also the leading provider of funding to combat malaria, a disease transmitted to humans by mosquitoes. malaria is a major killer: in , there were million infections and , deaths from malaria (most of them children under age ). in africa alone the economic impact of malaria is estimated to be $ billion per year, including the costs of healthcare, absenteeism, days lost in education, decreased productivity, and loss of investment. but thanks to the support of the global fund, hundreds of millions of insecticide-treated mosquito nets have been distributed and over million cases of malaria have been treated by . as a result, global malaria deaths have dropped by % since . unfortunately, progress has stalled in the past few years due to drug and insecticide resistance. some countries are even losing ground to the disease. launched in with the help of a $ million five-year pledge from the bill and melinda gates foundation, gavi is an international organization that brings together the public and private sectors in the shared goal of creating access to new and underused vaccines for children living in the world's poorest countries. the us and the eu are among gavi's six original donor countries; as with the global fund, they are the largest donors, providing roughly $ million each per year. gavi estimates that it has helped treat over million children and has prevented more than million future deaths in the first years of its existence. looking to the future demands on foreign aid and humanitarian assistance are certain to grow in the future, principally because of population growth and climate change, causing extreme weather patterns (including heat and drought), pests, disease, and rising oceans. some studies predict that of the . billion increase in world population between and . billion will be in africa. the oecd estimates that by half a billion people may be living in "fragile states," defined as countries that are incapable of exercising basic functions, because of climate change and conflict. every year hundreds of millions of people require humanitarian assistance, largely because of natural disasters and conflicts. hunger is one of the main urgent challenges: the food and agriculture organization estimates that over million people suffer from food insecurity, of which over million (roughly half of them children) face acute hunger, even starvation. only one-fifth of children affected by severely acute malnutrition receive adequate care, with the result that many become ill and suffer impaired growth and cognitive development. there are nearly million people around the globe-principally in syria, turkey, lebanon, palestine, yemen, afghanistan, south sudan, somalia, and myanmar-requiring protection, shelter, food, and other basic services due to forced displacement, often lasting a decade or more. very often these people lack access to water, sanitation, and hygiene, resulting in heightened risk of epidemic outbreaks. the world bank estimates that % of land area worldwide, home to approximately % of the world's population, is exposed to drought. at the same time, rapid population growth and urbanization are contributing to a steady increase in the demand for water. as a result, the number of people without access to safe drinking water is expected to double by to billion. in light of increasing demand for urgent humanitarian assistance and longer-term development aid, the us and the eu, including its member states, need to build on their cooperation as the world's leading donors to coordinate more frequently and deeply than ever before. this coordination is not only necessary to make the dollars and euros stretch further, but also to ensure that their common values shape the global development agenda despite the rapid rise of new state donors (especially china) that are focused almost exclusively on the expansion of political power and economic ties, rather than the promotion of democracy, human rights, and good governance. the us and the eu also need to work together to ensure that their activities in foreign aid and humanitarian assistance are consistent with the growing role of private development assistance coming from ngos, foundations, and corporations in the oecd. it will be more challenging, but important, for the us and eu to reconsider some of their policies that undermine their joint objectives to promote more stable economic and political conditions in the poorest countries. in the case of the us, that means its practice of tying aid to the purchase of us agricultural commodities. in the case of the eu, that means its practice of dumping into african markets the cheap surplus food that results from generous european production subsidies; and it also means its opposition to genetically modified food and feed that prevents african countries from accepting some food aid and planting more resilient crops. the signature of a cooperation agreement between the us and the eu in on the sharing of data received from the eu's copernicus constellation of earth observation satellites will assist joint efforts to manage and mitigate natural disasters the new alliance for food security and nutrition in africa the invaluable assistance of dr. emmanuel de groof in the preparation of this chapter is gratefully acknowledged. key: cord- -ev pvr s authors: werth, annette; gravino, pietro; prevedello, giulio title: impact analysis of covid- responses on energy grid dynamics in europe date: - - journal: appl energy doi: . /j.apenergy. . sha: doc_id: cord_uid: ev pvr s when covid- pandemic spread in europe, governments imposed unprecedented confinement measures with mostly unknown repercussions on contemporary societies. in some cases, a considerable drop in energy consumption was observed, anticipating a scenario of sizable low-cost energy generation, from renewable sources, expected only for years later. in this paper, the impact of governmental restrictions on electrical load, generation and transmission was investigated in european countries. using the indices provided by the oxford covid- government response tracker, precise restriction types were found to correlate with the load drop. then the european grid was analysed to assess how the load drop was balanced by the change in generation and transmission patterns. the same restriction period from was compared to previous years, accounting for yearly variability with ad-hoc statistical technique. as a result, generation was found to be heavily impacted in most countries with significant load drop. overall, generation from nuclear, and fossil coal and gas sources was reduced, in favour of renewables and, in some countries, fossil gas. moreover, intermittent renewables generation increased in most countries without indicating an exceptional amount of curtailments. finally, the european grid helped balance those changes with an increase in both energy exports and imports, with some net exporting countries becoming net importers, notably germany, and vice versa. together, these findings show the far reaching implications of the covid- crisis, and contribute to the understanding and planning of higher renewables share scenarios, which will become more prevalent in the battle against climate change. the covid- emergency established an unprecedented challenge for modern societies. governments had to face a novel disease with exponential spread and considerable death rate. while intensive care cases were overwhelming healthcare systems, scientific experts tried to determine new behavioural models with little scientific knowledge, due to the substantial novelty of the situation. despite the economic and political interconnections between european countries, the countermeasures against the epidemic were diverse in severity and temporal implementation. the efficacy of these interventions and their economic impact on industries [ ] , stock markets [ ] , environment [ , ] and energy markets are still unclear. as governments try relaunching economies and softening restrictions, it is crucial to understand the short and long term consequences of the adopted measures, not only to be better prepared for future crises, but also to unveil potential opportunities [ ] , in particular for sustainability research on electricity [ , ] . with the analysis of electricity data, available at almost real time, it was possible to provide a fine-grained view on the economic and environmental impact of covid- . indeed, as fezzi and colleagues [ ] demonstrated for the case of italy, high frequency electricity data could be used to estimate the effect of covid- on the national gdp. previous studies reported that national lockdowns had a strong influence on the electricity consumption in many countries heavily affected by the epidemic [ , , ] . norouzi and colleagues [ ] used an artificial neural network model to evaluate the elasticity of oil and electricity demand in china, based on parameters including number of infections and gross domestic product without actual electricity data in input. ruiz and colleagues [ ] found that electricity consumption in china increased due to large quarantine and health service demand. other studies ascribed the decrease in energy consumption to governmental interventions in the usa and canada [ , , , ] , in brazil [ ] and in europe [ , , , ] . the present work aims at understanding the impact that restrictions, associated to covid- , had on the electrical energy consumption in europe, and the downstream consequences to generation and international exchange of electricity. leveraging the work from [ ] , the types of intervention whose severity correlated with consumption loss were identified, and their time of implementation was determined. following this data driven approach, such period of active restrictions in was analysed comparing it the same time interval from past years, using the percent deviation statistic, and accounting for yearly variability with ad hoc statistical procedures. with this methodology, energy load, generation and international transmission were studied for european countries, for which data were available, to understand how europe's electric grid was affected by the epidemic. the data were obtained from the transparency platform of the european network of transmission system operators for electricity (entsoe [ ] ). the available data consist of time-series with a , or minute resolution depending on the country. following time series for european countries from / / to / / were used: • generation data for each type of source • generation capacities for each type of source (annually) • cross-country power flows for neighbouring countries (imports and exports) because of inconsistencies or omissions in the data, the following were removed: • netherlands generation data, since they disagreed with load data, particularly for the years before ; • italy and switzerland generation data, because largely incomplete. all other non-substantial missing data points were imputed by linear interpolation. in the evolving scenario of covid- emergency, quantifying in a comparable way governments responses is a very complex task. several attempts have been done to systematize institutional intervention data [ , ] . the oxford covid- government response tracker [ ] (oxcgrt in the following) provides a systematic cross-national, cross-temporal measure to understand how governmental responses have evolved during the epidemic. this reference was identified as most suitable to compare the intensity of interventions in the covid- emergency with different nations' electricity load. the oxcgrt includes indices sorted into main categories: "containment and closure", "economic response", "health systems". each indicator quantifies, on a daily basis, the intensity of a governmental restriction executed from a country included in their data-set. for the present analysis, all the indices of the "containment and closure" were used, that most obviously have a more direct relation with the energy load. "economic response" indices were instead excluded for the lack of a direct relation with the electric load and also for the timescale of their action. economic responses, in fact, do have consequences on a longer time scale while the present study takes into account only the first months of . "health systems" indices were also excluded because of their negligible impact on the energy load, with the exception of the "public info campaigns", which, in principle, could affect people behaviours and thus energy consumption on a large scale. the list of the indices taken in consideration can be found in table . for each of these indices, the time of action was determined for each country. the time of action was defined as the days when the relative index is greater than zero. summing an index over its time of action provided the cumulative extent of severity for the governmental actions associated with it, in that country, for the first months of . this quantity was named cumulative index. in order to study the relation between intervention intensity and load variations, also the cumulative energy load was calculated in the time of action of each index in each country. the mean of the cumulative load of the five previous years was then used as a forecast reference for cumulative load, and the cumulative forecast excess (cfe) was calculated as where cl y is the cumulative load in the time of action of a given index for the year y. e.g. cl is the cumulative load observed in in the same days corresponding to the time of action in of the given index. from this procedure, two values were obtained for each combination of one oxcgrt index and one country: • the cumulative index, representing the cumulative severity of the intervention; • the cumulative forecast excess of the load, representing the total decrease in the load in the observed period compared to the same period in previous years. to understand and measure the impact of governmental restrictions on the electrical power system, at a country level, it is necessary to assess the data recorded during covid- emergency against a counterfactual scenario where the epidemic did not occur. therefore, defined the real number x y as the total energy transmitted, generated or consumed, over a fixed time period of year y (for y ∈ { , ..., }), the fluctuation of x y was estimated by the percent deviation from years before , namely restrictions as the deviation of x from the average of previous years, which estimates year without epidemic. to quantify how largely s deviated from past years, taking yearly variability into account, the number of years that were more extreme than were compared against the others averaged, using the value r = #{y < : p y ≥ s }, and #a being the number of elements in a set a. this approach follows the same rationale as a test for statistical significance by permutations [ ] assessing the null hypothesis that x y , for y = , . . . , , are independent observations from the same distribution. when r = , the test leads to the rejection of the null hypothesis, with a level of significance equal to /#{y: y ≤ } (i.e. . or about . for or yearly data points, respectively). when r > , instead, the null hypothesis cannot be rejected. note that this measure does not take into account special weather conditions. after the spread of covid- in europe, lockdowns were imposed in several countries to oppose the epidemic. simultaneously, large drops in electric load were reported ( fig. ) , which is mostly due to a reduction in industrial and commercial activities with a possible increase in residential sector [ ] . for instance, for the month of april, all countries combined had a mean load drop of gw, or % if compared to the previous years. yet, load levels did not revert to normality after the confinements ended, as observed in fig. , and other limitations persisted throughout the pandemic and after strictest restrictions were lifted. first, the oxcgrt indicators whose impact resulted in reduced load were identified. for every index, using the cumulative index and the cumulative forecast excess from each country, the spearman's correlation coefficient and the relative p-value were calculated. the indices with significant correlations j o u r n a l p r e -p r o o f (p-value < %) were selected. results are reported in table . only indices showed positive and significant correlations: "school closing", "workplace closing", "restriction on internal movements", and "stay at home". these four indices were then combined into a novel macro index named "energy stringency index ". this new index was then used to calculate its time of action, i.e. the time range where the index is greater than zero. the days in when the energy stringency index is not null, determine a period of active restrictions with a strong relation on a country's energy load. thus, this temporal arc will be hereafter referred to as the "restrictions period". this procedure enabled a systematic determination of the interval of time to study how european countries modulated electrical energy generation, and transmission in order to balance for the reduced consumption. each nation had its own time range, based on when the energy stringency index was greater than zero in that nation. they had different starting dates but the ending date is the same for every country, the st of july , because the index happened to be greater than zero at least until that date. finally, the energy stringency index was compared against the load drops, resulting in a strong and significant positive correlation ( . with p-value of . ), as outlined in fig. reported results suggest that the quantification of severity for governmental restrictions could be exploited to improve the forecast of the load, which became loads [ , ] , that cancelled out upon aggregation. therefore, a more in depth study (with finer data) is required to pin down the actual causes and impact. overall, out of countries present null r-value (in brackets next to each country's name), indicating as the most extreme year, with many loads reducing by % from past years average. j o u r n a l p r e -p r o o f journal pre-proof as energy load dropped in the restrictions period, significant changes were expected at the level of electricity generation. observing the weekly rolling mean of generation from january to june , sorted by energy source, many countries presented a generation decrease, particularly in concomitance with march and april when most severe lockdown measures were imposed (fig. ) . for some cases, such as italy, the drop steepness suggested causes other than the changing of weather or seasons, when transitioning from winter to spring time energy production typically reduces with the increasing of atmospheric temperatures. for instance, during the month of april, the total mean generation, from all countries combined, decreased by gw (- %) compared to the previous years averaged. during that same month, fossil fuel generation dropped by gw (- %), nuclear by gw (- %) while combined renewable generation increased by gw (+ %). also, more in general, the load comparisons presented in the previous section suggested that seasonality could not be responsible for all the variation observed. in order to account for the seasonality, the data from the restrictions period for the generation (total and split by different source groups) were analysed using the percent deviation statistic. this procedure enabled the study of how generation was regulated among different energy sources, and which of these were reduced or curtailed. the difference between imported and exported energy, namely the net energy balance, was finally investigated to achieve a complete overview of the energy composition for each country. total generation. as hypothesised, most of the countries with significant load decrease also showed a significant generation decrease (fig. a , r-value = ), with two exceptions: austria and belgium. austria, however, presented a remarkable change of trend with respect to the previous three years. inference for belgium, instead, was hindered by the high variability in its generation pattern, thus no conclusion could be drawn. nuclear and fossil generation. the non-renewable energy sources, such as nuclear and fossil (which includes gas, oil and coal) that typically cover the baseline load, were observed decreasing in generation for most countries, although yearly productions were quite variable (fig. b,c) . still, nuclear output reduced significantly in france and the uk, two eu countries with large nuclear generation. each point is a cumulative sum over a country's restriction period. baseline value (at %) corresponds to the average from years before (excluded), by which all points relative to a given country are centered. next to a country's name, within parentheses, the r-value depicts how many years are more extreme, compared to others, than . to reduce their output than nuclear. this agrees with the evidence that energy generation from fossil fuels was affected by largest percent deviation drops (fig. c ). in particular, coal and oil decreased over the past years (fig. ) , replaced by cleaner alternatives (see supplementary material) . still, germany, poland and czechia, countries whose production share of coal is still relatively high, shifted to more fossil gas generation over the years (fig. e) , a trend continuing also through . renewable generation. renewable energy has no fuel cost and should therefore be prioritised for both environmental and economic reasons. the variability and intermittency in generation from wind and solar, however, challenges the matching of supply and demand, and may result in curtailments. in particular, generation from renewables -specifically intermittent renewables-was queried about its full utilization even during the load drop. to analyse renewable sources, these were grouped into intermittent renewables (solar, wind onshore and wind offshore), and non-intermittent renewables (hydro, biomass, geothermal, waste), on the basis that the output from intermittent renewables cannot be shifted in time, but only curtailed. indeed, as low load and favourable weather conditions co-occurred in april , an unprecedented number of negative wholesale market prices was witnessed across europe (especially in germany) [ , ] , thus raising the question whether intermittent renewable energy was extraordinarily curtailed due to covid- restrictions. as new wind and solar generation capacity was progressively added over the years (fig. a) , generation from intermittent renewables steadily increased as well, and this trend seemed to continue in , for all european countries but portugal, spain, germany and austria (fig. f) . a more in depth study of weather conditions and markets would be needed to determine reasons for this. for another viewpoint, generation was divided by the installed capacity, which measures the proportion of utilization from the installed assets. for the analyzed countries, no significant percentage deviation in generation/capacity was observed for the lockdown phase of , except for portugal and spain (fig. b ). this could suggest that the other countries did not recur to exceptional curtailments in , and intermittent renewable output was consumed or exported similarly as previous years. as for portugal (and to a lesser about non-intermittent renewables (mostly hydro), their output considerably increased in the united kingdom and slightly decreased in czechia from previous years, but did not present outstanding deviations overall. in fact, being hydro a mature and flexible technology, its utilisation did not change significantly over the years (fig. g ). the unusual situation of , showed that the european grid helps balancing between countries with often greatly different generation mixes or -in this case-government interventions leading to load drops. in general, both exports and imports were higher compared to previous years. to inspect the international exchange of electrical energy, countries were first separated into net importers and net exporters, respectively depending on the positive or negative sign of the net energy balance summed over the restrictions periods from years to . in figure (fig. ) , net balance naturally presented a larger percentage variations than load or generation, because of smaller amount of energy that is imported and exported, relative to the load (with portugal being the most extreme example -see supplementary material). during the restriction period of , italy, traditionally a big net-importing country, diminished the net balance by %, indicating that imports were much reduced as the load dropped. for belgium and austria, instead, the declining trend, over the past years trade, slowed and inverted, respectively, and exports overcame imports by . in this year, a significant increase in net balance occurred for poland and czechia, which are fossil based, and for germany. strikingly, the latter country incremented so much as to become a net importer during the restrictions period related to the covid- . the understanding of covid- emergency is an important case study to prepare for scenarios of large load reduction, and high renewable output. the present paper investigates the impact of restrictions, related to the epidemic, on the electricity flows in european countries. using the oxcgrt indices, four limitation types, that significantly correlated with the load reduction observed during the crisis, were identified: "stay at home", "school closing", "restriction on internal movements", and "workplace closing", ordered by increasing correlation. these findings highlighted how the daily severity of these interventions could be accounted for in energy consumption forecast models, to obtain more accurate predictions in the eventuality of successive waves or in future emergencies. in the period of active restrictions, most of europe was characterized by a remarkable load drop, except for scandinavia and switzerland, whose consumption did not decrease significantly (possibly due to less restrictive limitations, peculiarities concerning the load compartments, or a country's industrial activity). concurrently, energy generation by coal, oil and nuclear was reduced considerably, in favour of intermittent renewable sources and, in some countries, fossil gas. in most countries, no extraordinary curtailment was found concerning intermittent renewables, confirming the general trend of increasing exploitation of these sustainable sources, even in critical times. the energy transmissions between countries was also explored, showing a general increase of both imports and exports. coal-based countries, such as poland, czechia and germany, highly increased their net incoming energy, with germany even becoming a net importer in . italy, instead, halved its net imports, which is significant for such a big net-importing country. future studies are required to precisely model the impact of restrictions on generation, net balance and load, ideally split the latter by sector (residential, industrial, commercial). such models will allow for a better long-term forecast, which could be strategic for policy-makers. also short-term forecast will be improved by these models, supporting the activity of the electric grid stake-holders. the results outlined above provide an overview on how energy dynamics in the european electric system played out, adapting to rapidly changing conditions as a consequence of the covid- restrictions imposed by governments. understanding the ramifications of covid- responses provides unique insights not only on how different societies manage critical situations but also on how higher shares of intermittent renewables will impact the grid infrastructure, the energy markets and related investments. as gillingham and colleagues pointed out [ ], pushing back renewable investments would outweigh emission reductions from march to june of and only an energy policy response could change that. the path undertaken through this crisis will be incorporated into j o u r n a l p r e -p r o o f new policies and determine long-term consequences towards a 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climate change: exploring the impacts of covid- on oil and electricity demand in china analysis of the electricity demand trends amidst the covid- coronavirus pandemic canadian electricity markets during the covid- pandemic:an initial assessment effects of the covid- pandemic on the brazilian electricity consumption patterns the impact of different covid- containment measures on electricity consumption in europe variation in government responses to covid- -blavatnik school of government the csg covid- control strategies list (cccsl testing statistical hypotheses emergency measures to protect energy consumers during the covid- pandemic: a global review and critical analysis commercial down vs. residential up: covid- 's electricity impact eu day-ahead markets see th week of negative hourly prices the authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. key: cord- - ugoxns authors: jensen, bent borg title: extensive literature search on the ‘effects of copper intake levels in the gut microbiota profile of target animals, in particular piglets’ date: - - journal: nan doi: . /sp.efsa. .en- sha: doc_id: cord_uid: ugoxns nan background and terms of reference as provided by the requestor . . in the context of the development of the scientific opinion linked to the mandate related to the revision of maximum levels of copper in feed, the experts of the ad-hoc working group (wg) of the feedap panel have considered relevant to evaluate the effects of copper intake levels in the gut microbiota profile of target animals. published scientific papers have addressed this issue and it seems there is a relationship between levels of dietary copper in animals and differences in their gastrointestinal microbiota, which in turn may result e.g. in more nutrients becoming available for absorption; the mode of actions have also been recently studied. a possible outcome of the opinion might be a recommendation of different maximum levels compared to those currently authorised in feed (table ) . therefore it is necessary to know the potential effects of a variation in copper intake levels on the gut microbiota profile of the target animals which might be accounted in the feedap panel opinion. piglets should be particularly considered in this literature search, as this is animal category for which the maximum copper in feed is allowed by commission regulation (ec) no / ; at least, two other relevant animal species/categories (chickens for fattening, dairy cows, fish, cat, dog) shall be included. the project should be accomplished in a structured manner, that is following an extensive literature search (els) process. the steps listed below should be followed and duly documented under the present call for tender: -development of an els protocol, which will be subject to efsa´s approval the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. -databases to be consulted -criteria to select the papers -criteria for data extraction -assessment of the quality of the data -synthesis of the data the purpose of the contract was to provide scientific assistance to the feed unit of efsa in the completion of an extensive literature search on the effects of copper intake levels in the gut microbiota profile of target animals, in particular piglets. the procedure includes the provision of a report collecting, collating, analysing and synthesising the scientific data and information on the els topic. the specific objectives of the contract resulting from the present procurement procedure are as follows: the searches in web of science resulted in hits (shown in appendix a ). the search in cab abstracts resulted in hits of which one (p in appendix a and appendix b) was not among the hits from the search in web of science. consulting the reference list of relevant papers from the search another studies/papers were retrieved (p to p in appendix a and appendix b). search terms and boolean operators (strings): ts=(copper or cu or copper sulphate) and ts=(gut or intestinal or intestine or ileal or digestive tract or gastrointestinal tract or git) and ts=(bacteria or microbiota or microflora or microorganism or microbial community) and ts=(chickens or layer or hen or broiler) the searches in web of science resulted in hits (shown in appendix a ). the search in cab abstracts resulted in hits of which one (ch in appendix a and appendix b) was not among the hits from the search in web of science. no further studies/papers were found by consulting the reference lists from relevant papers from the search. search terms and boolean operators (strings): ts=(copper or cu or copper sulphate) and ts=(gut or intestinal or intestine or ileal or digestive tract or gastrointestinal tract or git) and ts=(bacteria or microbiota or microflora or microorganism or microbial community) and ts=(cow or cattle or rumen or ruminant or bovine). the searches in web of science resulted in hits (shown in appendix a ). the search in cab abstracts resulted in hits of which all were included in the hits from the search in web of science. no further studies/papers were found by consulting the reference lists from relevant papers from the search. effect of copper intake on gut microbiota (pigs/piglets, chickens, dairy cows) www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. for the general approach to complete this els the contractor considered the technical manual for performing electronic literature searches in food and feed safety and the efsa guidance: application of systematic review methodology to food and feed safety assessments to support decision making. the inclusion criteria for the scientific papers were no limitation on timespan, no limitation on language and inclusion of primary research and review articles (the later to get a comprehensive view on the subject). the els protocol was primarily based on primary research. papers on foreign languages, where translation was not possible, were excluded.  the study had to contain a control diet and diet supplemented with one or several copper concentrations in the range of to mg/kg.  studies including the effect of any sources of dietary copper on any aspect of the microbiota in the gastrointestinal tract or faeces (profile, activity, metabolites and ph) were selected. the total number of references retrieved, the references rejected at the title and abstract screening and the references from which data was extracted are shown as a flow chart in annexes a to a . the list of included and excluded references are shown in appendix b. data collected was the effect of dietary copper in the range from to mg/kg on any aspect on the microbiota in the gastrointestinal tract or faeces. copper content: background content of feed, supplemented copper, concentration and source (data given in table for piglet/pigs and in table for broilers/layers). animals: species/category and age (weight) of the animals (data given in table for piglet/pigs and in table for broilers/layers). number of animals; animals per treatment; replicates (data given in table for piglet/pigs and in table for broilers/layers). composition and activity of the microbiota in the gastrointestinal tract or faeces (profile, activity, metabolites and ph). an overview of the data obtained is given in table for piglets/pigs and in table for broilers/layers. the data obtained is narratively described in the text and an overview of the effect of copper concentrations and sources on the concentrations on bacterial populations is shown in tables , and for piglets/pigs and in table for broilers/layer. if available, performance parameters (data given in table for piglet/pigs and in table for broilers/layers). the results are narratively described in the text and shown in tables , , , and , for piglets/pigs and in tables , and for broilers/layers. www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the study design (incl. e.g. the duration of the study), the conduction of the study and the description of the study was evaluated. it was checked if the number of animals in each group in the studies were adequate. it was checked if the methods used to describe the microbiota profiles were adequate. it was checked if the statistical methods (incl. e.g. randomisation of the assignment of animals in treatment groups) used were adequate. the methodological quality of the selected studies is captured in table for piglets/pigs and in table for broilers/layers. of the references found for pigs and piglets by the literature search, were considered appropriate to be included in the els (annex a ). references were duplicates. of the excluded references, were excluded when screening the title and abstract and after consulting the full paper. the list of included and excluded references is provided in appendix b. of the references found for chickens by the literature search, were considered appropriate to be included in the els (annex a ). references were duplicates. of the excluded references, one full paper could not be retrieved, were excluded when screening the title and abstract and after consulting the full paper. it should be mentioned that out of the included papers were written in foreign languages ( in korean and in chinese) with english abstracts and table and figure legends. the list of included and excluded references is provided in appendix b. of the references found for cows by the literature search, were considered appropriate to be included in the els (annex a ). references were duplicates. of the excluded references, were excluded when screening the title and abstract and after consulting the full paper. a closer examination of the five studies first selected to be included in the els study on cows, revealed the papers not to be appropriate to be included in the els. one of the studies (cow ) was published in polish, two of the studies were dealing with antibiotic resistance (cow and cow ), one was dealing with the effect of copper on parasite control (cow ) and the last one on the effect of copper on abomasa ulcers (cow ). the list of included and excluded references is provided in appendix b. effect of copper intake on gut microbiota (pigs/piglets, chickens, dairy cows) www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. pigs and piglets . . assessment of the methodological quality of the studies table : assessment of the methodological quality of the piglet/pig studies as shown in table the quality of the papers retrieved from the search on piglet/pig was generally judged to be good although some of them were rather old. five out of papers were written in foreign languages ( in german and in chinese) with english abstracts and table and figure legends four types of copper sources were used in the research with pigs (table ) : ) inorganic copper, ) organic bound copper, ) clay bound copper and ) copper-loaded nanoparticles. inorganic copper was used in of the studies. the sources were cuso (concentrations - mg cu/kg feed, in studies p , p , p , p , p , p , p , p , p , p , p , p , p , p , p , p , p , p , p , p , p , p and p ), cuo (concentrations - mg cu/kg feed, in studies p and p ) and elemental copper (concentration mg cu/kg feed, in study p ). organic bound copper was used in of the studies. the sources were cu-aa complex (cupric amino acid complex, concentration mg cu/kg feed, in study p ), cu-cit (cupric citrate, concentrations , and mg cu/kg feed, in studies p and p ), cu-met (copper the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. bbj methionate, concentrations and mg cu/kg feed, in study p ) and cu-gly (copper glycinate, concentrations , and mg cu/kg feed, in study p ). clay bound copper was used in of the studies. the sources were cu-mm (copper-bearing montmorillonite from aluminosilicate clay). it was used in studies p , p , p , p and p at concentrations from to mg cu/kg. copper-loaded nanoparticles were used in of the studies. the source was copper loaded chitosan nanoparticles, it was used at a concentration of . mg cu/kg feed in study p . the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. usa ----- ppm cuso . experiment ----- ppm cuso . ----- ppm cucit . ----- ppm cucit . ----- ppm cucit . ----- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the microbial characteristics measured in each of the selected studies with piglets/pigs are shown table . eighteen of the studies were done with piglets (p , p , p , p , p , p , p , p , p , p , p , p , p , p , p , p , p , p and p ) and were done with growing pigs (p , p , p , p , p , p , p , p and p ). the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. bbj pigs (hu et al., ) the population of clostridium was significantly reduced in as well si as caecum content in pigs fed mg/kg cu as cu-montmorillonite compared to the control fed pigs; however, although numerically lower, the effect was not significantly different from pigs fed copper free montmorillonite. in study p (kroger et al., ) addition of mg/kg cu as cuo or cuso to diets of growing pigs were shown significantly to reduce pigs containing costridium perfringens in gut content from the small intestine and colon. no effect was found of elementary copper. the effects of cuo and cuso addition were only seen if cu was supplemented until the pigs were slaughtered at kg live weight. if it was withdrawn from the feed at kg live weight no effect was detected when the pigs were slaughtered at kg. in conclusion low concentrations of copper as clay bound copper seem to reduce the population of clostridium both in piglets and in growing pigs. further high concentrations of inorganic copper as either cuso or cuo may reduce the population of clostridium in growing pigs. coliform bacteria. the effect of copper on the population on coliform bacteria was investigated in studies with piglets (p , p , p , p , p , p , p , p , p , p , p , p , p , p ) and four studies with growing pigs (p , p , p , p ). the results from the studies are summarised in table . it may be concluded from the experiments that supplementing piglet (and growing pigs) diet with low additional copper amounts (below mg/kg cu) seems to inhibit the population of coliform bacteria in the gastrointestinal tract. especially clay bound copper seems to be effective. when supplemented to the feed in amounts above mg/kg copper it does not seem to have any effect on the population of coliform bacteria in the gastrointestinal tract of pigs. www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. enterococci. the effect of copper on the population density enterococci was investigated in one study with piglets. dietary doses of mg/kg cu as cuso significantly reduced the counts of enterococci in stomach content of piglets while no effect was seen in content from the si, caecum or colon (study p ). lactic acid bacteria. the effect of copper on the population density of lactic acid bacteria was investigated in one study with piglets. dietary doses of mg/kg cu as cuso significantly reduced the counts of enterococci in stomach content of piglets while no effect was seen in content from the si, caecum or colon (study p ). the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. lactobacilli. the effect of copper on the population density and/or the composition of lactobacilli was investigated in studies with piglets (p , p , p , p , p , p , p , p , p , p , p and p ) and in four studies with growing pigs (p , p , p and p ). the results from the studies are summarised in table . in study p with piglets (højberg et al., ) dietary doses of mg/kg cu as cuso significantly reduce the counts of lactobacilli in stomach content while no effect was seen in content from the si, caecum or colon. in study p (jensen, ) the population of lactobacilli was reduced throughout the gastrointestinal tract in piglets fed mg/kg cu as cuso . in study p (ma et al., ) no effect of addition of mg/kg cu as cumm was detected on the population of lactobacilli in content from the si or colon. in study p (mei et al., ) a numeric reduction in the population of lactobacilli was detected by addition of , and mg/kg cu as cuso to piglet diets when the population density were enumerated using selectively cultivation, however if the population of lactobacilli were quantified by use of qpcr the reduction found for and mg/kg cu as cuso were significant different from the results found in the control fed piglets. in study p (mei et al., ) no significant effect of addition of mg/kg cu as cuso was found on the population of lactobacilli in cecal content. in study p (namkung et al., ) the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. bbj from piglets was found by supplementing the diet with mg/kg in study p (kellog et al., ) . in the study with growing pigs (p , hawbaker et al., ) no significant effect on the population of staphylococci in fecal samples were detected in any of the three experiments carried out with mg/kg cu as cuso . in conclusion the effect of dietary copper on the gastrointestinal populations of staphylococci in pigs seems to be non-existent or very week. streptococci. the effect of copper on the population density and composition of streptococci was investigated in two studies with piglets (p and p ) and four experiments with growing pigs (p , p , p and p ). with piglets no significant effect in the fecal counts of streptococci were found in study p experiment and study p experiment (bunch et al., ) neither by feeding the piglets , or mg/kg cu as cuo or by feeding them mg/kg cu as cuso . there was, however, a numerical reduction in streptococci in both experiments by feeding mg/kg cu as cuso . no significant reduction in the population of streptococci was found by feeding piglets with mg/kg cu as cuso in study p (kellog et al., ) . in study p (varel et al., ) with growing pigs, mg/kg cu as cuso reduced the number of ureoluytic organisms in fecal samples with a marked decrease in the streptococcus spp., which made up % of the ureolytic bacteria in fecal samples from pigs fed the control diet. a marked fall in the number of streptococci in fecal samples was found in study p (fuller et al., ) when growing pigs were fed mg/kg cu as cuso compared to pigs fed a control diet. the extent of the reduction varied but the counts of streptococci in the copper treated pigs were always lower than in the control pigs and the reduction could be as much as from to cfu/g faeces. in study p (experiments , and ), a significant reduction of streptococci in fecal samples of growing pigs was found in experiment and while only a numerical reduction was found in experiment by supplementing the diet of growing pigs with mg/kg cuso . in study p (hendericks et al., ) , it was concluded that cuso (concentration of cu not given) strongly inhibited the growth of streptococci in an ex vivo experiment with si content from growing pigs. in conclusion the results strongly indicate that supplementation with mg cu/kg, or higher, as cuso significantly reduces the population of streptococci in the gastrointestinal tract of growing pigs. the effect of cuso on the population of streptococci in piglets is more questionable. total anaerobe bacteria. the effect of supplementary copper on the population of total anaerobic bacteria is shown in table . no effect on the anaerobic population was observed if the amount of copper added to the diet was below mg/kg copper. however, addition of mg/kg cu as cuso to the diet may reduce the population of total anaerobic bacteria in the colon, while copper as cuo does not seem to have any effect even at cu concentrations of mg/kg. www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. table : effect of copper source and copper concentration on the population of anaerobic bacteria in the gastrointestinal tract of piglets and growing pigs. total aerobe bacteria. the effect of copper on the population of total aerobe bacteria was investigated in two studies with piglets (p and p ) and in one study with growing pigs (p ). in study p experimet (bunch et al., ), piglets were fed either a control diet or diets supplemented with , or mg/kg cu as cuo and a diet supplemented with mg/kg cu as cuso . a significant reduction in the population of total aerobe bacteria in faecal samples was found in the piglets fed mg/kg cu as cuso . in contrast cu as cuo resulted in a significant linear increase with increasing cuo supplementation in the population of total aerobic bacteria in faecal samples. in study p experiment , no significant effect of either mg/kg cuo or mg/kg cuso was found on the population of total aerobic bacteria in piglet faecal samples. in agreement with that no effect on the population of aerobic bacteria in fecal samples from piglets was found in study p (kellog et al., ) by supplementing the diet with mg/kg cu as cuso . in the study with growing pigs (p , hawbaker et al., ) a significant reduction in the population of total aerobic bacteria in fecal samples was detected in all three experiments carried out with mg/kg cu as cuso . in conclusion the effect of copper concentration and source on the population on total aerobe bacteria seems to be very variable. ureolytic bacteria. the number of ureolytic bacteria was investigated in study p (varel et al., ) . addition of copper sulfate to the diet at a concentration of mg/kg copper significantly reduced the number of ureolytic bacteria compared to control fed pigs, in fecal samples collected after , and weeks of feeding the experimental diets, with a marked decrease occurring in streptococcus spp., which make up % of the ureolytic isolates. quantitative real-time polymerase chain reaction (qpcr) has been used in one investigation with piglets (p , mei et al., ) to detect the effect of copper supplementation on bacterial populations (lactobacilli, enterobacteria or total anaerobic bacteria) in gastrointestinal content. it was shown that the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. bbj the population of lactobacilli in caecum content from piglets was significantly decreased in piglets fed diets supplemented with or mg/kg cu as cuso while no effect was seen by a diet supplemented with mg/kg cu as cuso . no effect was found of any of the cuso supplemented diets on the population of enterobacteria or total anaerobic bacteria. use of traditional microbiological culturing methods did also show a reduction in the population of lactobacilli with increasing cuso addition; however these results were not significant. the microbial community was investigated in four studies with piglets (p , p , p and p ). in study p (namkung et al., ) , study p ( the results obtained from the microbial community structure studies are summarized in table . . . in all studies the number of bands per sample was measured and in study p and p the similarity indices for bands between samples were calculated. the number of bands is indicative for the number of bacterial species within a sample; the similarity coefficient is indicative of how similar the bacteria within a treatment group are. table : effect of copper sources and copper concentrations on microbial diversity and microbial similarity obtained from microbial community structure studies. no effect of copper concentration or sources was found in study p neither on the number of species nor on the similarity of the microbiota in faecal samples. however in studies p , p and p it was shown that supplementing piglet diets with high amounts of copper ( to mg/kg) as cuso affect the number of bacterial species in ileal (p ), in caecal (p and p ) and in colonic (p ) digesta. furthermore, in study p addition of mg/kg cu as cuso significantly reduced the degree of similarity of the microbiota in ileal samples compared to pigs fed a diet without supplementary copper addition. as also pointed out by the authors in study p , the combination of faecal samples and the microbial methods used in their study may not have been sensitive enough to detect differences among the dietary treatments. in study p , the similarity analysis further shows that the piglets did restore a diverse microbiota weeks after withdrawal from the high cu diet, but the community structure still seems to be different from that in the control fed pigs. in general, the community structure studies strongly suggest that supplementing piglet diets with to mg/kg cu as cuso change the microbial community in the small intestine, the caecum and the colon. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. digesta content. the amount of gastrointestinal content was investigated in study p (højberg et al., ) . the addition of high concentration of cuso ( mg/kg cu as cuso ) to piglet diets had no significant influence on digesta content in any part of the gastrointestinal tract compared to piglets fed a control diet. dry matter. the dry matter content in gastrointestinal digesta was investigated in piglets in study p (højberg et al., ) . in contrast to high concentrations of zno (addition of mg/kg zn as zno to the diet) -which significantly increase the dry matter content in the stomach and decrease the dry matter content in caecum and colon-, high concentration of cuso ( mg/kg cu as cuso ) had no significant influence on the dry matter content in any part of the gastrointestinal tract (stomach, si, caecum and colon). ph. the effect of copper on faecal ph and ph of digesta were investigated in three studies with piglets (p , p and p ) and one study with growing pigs (p ). in study p (højberg et al., ) and p (jensen ) no effect of feeding piglets diets supplemented with either mg/kg (p ) or mg/kg of cu (p ) as cuso was found on ph in any segments of the gastrointestinal tract (stomach, si, caecum and colon) compared to piglets fed control diets. in study p (zhu et al., ) a significant effect on ph in caecum and colon content was detected by feeding copper as cu-chitosan ( and mg/kg cu) or as cuso ( mg/kg cu) to piglets compared to piglets fed a control diet. in study p (huang et al., ) the effect of feeding copper supplemented diets to growing pigs on ph in faecal samples taken at the beginning of the experiment and after three and five weeks on the experimental diets were investigated. four different copper supplemented diets were used: either or mg/kg cu as cuso or or mg/kg cu as cucitrate. apart from a small but significant decrease in ph in faecal samples taken after weeks from the pigs fed the diet supplemented with mg/kg cu as cuso (ph . in faecal samples from the control fed pigs versus . in the copper fed pigs), no effect on ph was detected. in general the effect of supplementing pig diet with copper on faecal or gastrointestinal ph seems to be limited. diarrhoea. the effect of copper on diarrhoea incidence was investigated in three studies with piglets (p , p and p ) and in one study with growing pigs (p one where an antibiotic was included in the all diets tested ( mg/kg mecadox) and one without antibiotic in the diets. in both, experiment faecal samples were taken at day for measurement of the odour characteristics. in the study with antibiotic the pigs were kept on the experimental diets through the growing-finishing phase and samples taken after days for measurements of odour characteristics. in samples taken after days from the piglets fed the diets with antibiotics addition effect of copper intake on gut microbiota (pigs/piglets, chickens, dairy cows) www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. of mg/kg cu as cuso and and mg/kg cu as cu citrate significantly reduced two of the all three odour characteristics measured (odour intensity and irritation intensity) while no significant effect were found on the third characteristic (odour quality). no significant effect of diets with mg/kg cu as cuso or mg/kg cu as cuso was found on any of the odour characteristics measured. in samples taken after days from the piglets fed the diets without antibiotics, odour intensity was significantly reduced in piglets fed , and mg/kg cu as cu citrate while no significant effects were found for piglets fed or mg/kg cu as cuso . none of the experimental diets affected irritation intensity. however, mg/kg cu as cuso and , and mg/kg cu as cu citrate all significantly improved odour quality. no effect of mg/kg cuso was found on odour quality. in samples taken after days from the pig fed the diets with antibiotics odour intensity and irritation intensity were significantly reduced by all experimental diets except the diet supplemented with mg/kg cu as cu citrate which had no effect on odour intensity and irritation intensity. odour quality was significantly improved in faecal samples from all pigs fed copper supplemented diets compared to faecal samples from the control pigs. in contrast to the above mentioned results no effect of adding copper ( or mg/kg as cu as cuso or , , or mg/kg cu as cu citrate) to antibiotic supplemented diets were found in study p (armstrong et al., ) on either odour intensity, irritation intensity or odour quality of faecal samples from piglets (weaned at to days) after days feeding the experimental diets. in conclusion supplementing pig diet with or mg/kg as cu citrate or mg/kg as cuso may improve odour characteristics of swine waste. the mechanism of action was believed to be attributed to an antibiotic-like function of copper on the microbiota in the gastrointestinal tract. short chain fatty acids (scfa) concentration. lactate. microbial production of lactate was investigated in two studies with piglets (p and p ). addition of mg/kg zn as zno to the diets in study p significantly reduced the concentration of lactate in digesta in the stomach and si. however in the caecum and colon, where lactate is usually not detectable, a significant accumulation of lactate was observed in the piglets receiving the high zno diet. in contrast, the addition of mg/kg cu as cuso to the feed had no significance on the concentration of lactate in any segments of the gastrointestinal tract, but counteracted the accumulation of lactate found in the high zno fed piglets in caecum and colon. in study p (song et al., ) , the concentration of d-lactate was measured in plasma samples. d-lactate is produced by the microbiota in the gastrointestinal tract and absorbed to the blood. mammals do not have the enzymatic system to metabolise d-lactate so plasma d-lactate may be a measurement of the microbial activity in the gastrointestinal tract. a significant reduction in plasma d-lactate was found in study p in pigs fed mg/kg cu as either cu-camontmorillonite or cu-namontmorillonite compared to the control fed pigs, indicating a reduced fermentation in the gastrointestinal tract in the piglets fed the copper supplemented diets. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. bbj gastrointestinal tract, but counteracted the accumulation of succinate in caecum and colon found in the high zno fed piglets. absorption of ammonia. effect of copper on ammonia absorption was investigated in growing pigs in study p (yen and nienaber, ) . the addition of mg/kg cu as cuso to the diet significantly reduced the net absorption of ammonia to the portal vein compared to pigs fed a diet without the addition of extra copper. as concluded by the author, that may reflect decreased urease activity and ammonia production by the gastrointestinal microbiota in the pigs receiving the copper supplemented diet. deamination and decarboxylation of amino acids. the effect of copper on microbial formation of ammonia by deamination of amino acids and production of amines (histamine, putrescine, cadaverine, tyramine and phenylethylamine) by decarboxylation of amino acids in the si has been investigated using ex vivo incubations in study p (dierick et al., ) using si content from growing pigs. both processes were severely decreased if mg/kg cu as cuso was added to the incubations, indicating that copper has a sparing action on microbial degradation of amino acids in the small intestine, leaving more amino acids available for absorption. urease activity. effect of copper on urease activity has been investigated in two studies, one study with piglets (study p ) and one study with growing pigs (study p ). in the study with piglets (p , højberg et al., ) urease activity was investigated in content from the stomach, si, caecum and colon in piglets fed diets either added mg/kg cu as cuso or control diets without the addition of extra cuso both diets were fed with or without addition of mg/kg zno. the urease activity was below the detection limit in stomach and si content, highest urease activity was found in the colon. addition of cuso to the diets had no significant effect of the urease activity. in contrast to piglets, a significant effect (p< . ) of mg/kg cu as cuso was found on urease activity in fecal samples from growing pigs in study p (varel et al., ) . atp concentrations. bacterial activity as determined by the concentration of adenosine triphosphate (atp) was measured in study p and study p . in study p (højberg et al., ) no significant effects were found on atp concentration in any of the gastrointestinal segment investigated (stomach, si caecum and colon) by addition of mg/kg cu as cuso to piglet diets compared to a control diet without additional addition of cuso . similar results were found in study p (jensen, ) , where no effect of the addition of mg/kg cuso to a diet containing mg/kg cu was found on the atp concentrations in content from the stomach, three segments from the si, the caecum and three segments from the colon. microbial enzyme activity. the effect of copper on microbial enzyme activity of β-glycosidase and βglucuronidase has been investigated in one study (p , xia et al., ) . no significant effect was found by the addition of mg/kg cu as cuso to the diets on either β-glycosidase or βglucuronidase activity in si or colon content. addition of mg/kg cu as cu-montmorillonite reduced the activity of both enzymes in si as well as colon content compared to the control fed pigs. however apart from the activity of β-glucuronidase in the colon content, the effects were not significantly different from the activities found in pigs fed copper free montmorillonite. feeding high copper diets to germ free pigs has been investigated in one study with growing pigs (p , shurson et al., ) . germ free pigs tended to have higher average daily gain (adg) and average daily feed intake (adfi) than conventionally reared pigs. addition of mg/kg cu as cuso to the feed tended to reduce adg and adfi in germ-free pigs while it increases adg and adfi in conventionally reared pigs. the high copper diets did not appear to make organ weight or intestinal characteristics in conventionally reared pigs different than those of the germ free pigs. high copper diets seem to reduce intestinal cell turnover in germ free pigs while it tended to accelerate intestinal turnover in conventionally reared pigs, perhaps via an interaction with the gut microbiota. www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. coliform bacteria. it may be concluded from the experiments that supplementing piglet and growing pigs diet with low additional copper amounts (below mg/kg cu) seems to inhibit the population of coliform bacteria in the gastrointestinal tract. especially clay bound copper seems to be effective. when supplemented to the feed in amounts above mg/kg copper, it doesn't seem to have any effect on the population of coliform bacteria in the gastrointestinal tract of pigs. lactobacilli. from the results it may be concluded that supplementing pig diets with copper streptococci: in conclusion the results with growing pigs strongly indicate that the addition of mg cu/kg, or higher, as cuso significantly reduces the population of streptococci in the gastrointestinal tract of growing pigs. the effect of cuso on the population of streptococci in piglets is more questionable. in general, the studies on community structure strongly suggest that supplementing piglet diets with to mg/kg cu as cuso change the microbial community in the small intestine, as well as in the caecum and the colon. in conclusion, supplementing pig diets with or mg/kg as cu citrate or mg/kg as cuso may improve odour characteristics of swine waste. the mechanism of action was believed to be attributed to an antibiotic-like function of copper on the microbiota in the gastrointestinal tract. in general, the effect of supplementing pig diets with copper on faecal or gastrointestinal ph seems to be limited. the overall conclusion from the studies with piglets and growing pigs is that copper even at low concentrations (< mg/kg feed) may affect the microbiota in the gastrointestinal tract. especially the population of clostridia and coliform bacteria seems to be affected by low concentrations of copper. in particular, copper bound clay minerals seem to have an effect on the populations of coliform bacteria and clostridia. at higher concentrations (> mg/kg feed) cu as cuso reduce the population of lactobacilli in piglet as well as growing pigs. in growing pigs, the addition of cu as cuso reduces the population of streptococci in colonic and fecal samples, the population of ureolytic bacteria in the colon of which streptococci make up %, the urease activity in the colon, and decarboxylation and deamination of amino acids in the small intestine. however, no effect of cu as cuso on the population of streptococci and on urease activity was seen in piglets. supplementing piglet diets with to mg/kg cu as cuso significantly affects the community structure of the microbiota in the small intestine, the caecum and the colon. effect of copper intake on gut microbiota (pigs/piglets, chickens, dairy cows) www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. bbj chickens . . assessment of the methodological quality of the studies table : assessment of the methodological quality of the chicken studies. as shown in table the quality of the papers retrieved from the search on chickens was generally judged to be low. ten out of papers were written in foreign languages ( in korean and in chinese) with english abstracts and table and figure four types of copper sources were used in the research with chickens (table ) : ) inorganic copper, ) organic bound copper, ) clay bound copper and ) copper-loaded nanoparticles. inorganic copper was used in of the studies. the sources were cuso (concentrations - mg cu/kg feed, in studies). organic bound copper was used in of the studies. the sources were cu-soy proteinate (concentrations - mg cu/kg feed, in studies ch , ch , ch , ch , and ch ), cu-met (concentrations - mg cu/kg feed, in studies ch , ch and ch ), cu-fish meal (concentration mg cu/kg feed, in study ch ), starch cu complex (concentration mg cu/kg feed, in study ch ), starch methionine cu complex (concentration mg cu/kg feed, in study ch ), and na-alginate cu complex (concentration mg cu/kg feed, in study ch ). clay bound copper was used in of the studies. the source was copper-bearing montmorillonite from aluminosilicate clay (cu-mmt). it was used in study ch and ch at concentrations from to mg cu/kg feed. copper-loaded nanoparticles were used in of the studies. the source was copper-montmorillonite nanomaterial (concentrations - mg cu/kg feed, in study ch ) and copper silicate nanoparticles (concentration mg cu/kg feed, in study ch ). the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the microbial characteristics measured in each of in the selected studies with chickens are shown in table . fifteen of the studies were done with broilers (ch , ch , ch , ch , ch , ch , ch , ch , ch , ch , ch , ch , ch , ch , ch and ch ) and were done with laying hens (ch and ch ). the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. ), a significant increase in the population of bifidobacteria in content from the cecum was found by addition of or mg/kg copper as cuso to the diet both in broilers slaughtered after and weeks feeding the treatment diets. in contrast to that, addition of mg/kg copper to the diet as cuso resulted in a significant reduction in the population of bifidobacteria in cecal content in broilers slaughtered after weeks while no effect was found in broilers slaughtered after weeks. in all studies the population density of bifidobacteria were determined by counting the bacteria on selective media but several studies has shown that no really good selective media exist for the enumeration of bifidobacteria. campylobacter. the effect of copper on the population density of campylobacter was investigated in study ch (aydin et al., ) . no significant effect of an addition of mg/kg copper as cuso to the diet was found on the population density of campylobacter in ileal content. clostridia. the effect of copper on the population density on clostridia in content from the small intestine was investigated in studies with broilers (ch , ch , ch , ch , ch and ch ) and in two studies with layers (ch and ch ). the effect of copper concentration and copper sources on populations of clostridia in chicken gut content is shown in table . a significant reduction in the population of clostridia in si content from broilers were found by the addition of mg/kg cu as cuproteinate in study ch (kim et al., a) the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. cu-proteinate. similarly, no effect on the population of clostridia was found in study ch (min et al., ) by the addition of mg/kg cu as either cuso , cu-methionine or cu-fish meal to the diet; or in study ch by the addition of mg/kg cu as cuso . in the studies with layers a significant reduction of the population of clostridia were found in both investigations (ch and ch ). in study ch (choi et al., ) the population of clostridia was reduced by the addition of and mg/kg cu whatever it was added to the diet as cuso or cu-proteinate, and in study ch (paik et al., ) , by addition of and mg/kg cu whatever it was added as cu-methionine or cu-proteinate. the effect of copper on the density of clostridia in content from the caecum was investigated in two studies (ch and ch ). in study ch (xia et al., ) a significant reduction in the population of clostridia was found by the addition of mg/kg cu as cu-montmorillonite to the diet while no effect was found by the addition of mg/kg cu as cuso to the diet. in study ch (xu et al., ) no effect on the population of clostridia in caecum content was found by the addition or mg/kg cu-montmorillonite to the diet. supplementing chicken (broilers and layers) diets with additional copper seems to reduce the population of clostridia in the gastrointestinal tract of the birds, even at low copper concentrations. in particular, clay bound copper seems to be effective in reducing the population of clostridia. www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. coliform bacteria. the effect of copper on the population density of coliform bacteria in content from the gizzard was investigated in three investigations (ch , kim et al., and ch experiments and , min et al., ) . as shown in the effect of copper on the population density of coliform bacteria in si content was investigated in experiments with broilers and in experiments with layers (table ) . a significant reduction in the population of coliform bacteria was found in study ch (choi et al., ) by the addition of , or mg/kg cu as cuso to the diet, while no effect of an addition of mg/kg cu as cuso was found in studies ch (kim et al., ) , ch (min et al., ) , ch experiments and (min et al., ) ; and of mg/kg cu as cuso in study ch . furthermore, no effect was found in layers (ch , kim et al., b) fed diets containing or mg/kg cu as cuso . in study ch (kim et al., ) a significant reduction in the population of coliform bacteria in broiler si content was found by the addition of mg/kg cu as cu-methionine to the diet while it had no effect at a concentration of mg/kg. no effect was found by the addition of mg/kg cu as cu-methionine in study ch (min et al., ) and no effects were found in layers at concentrations of and mg/kg (ch , paik et al., ) . copper as cu-protionate was shown to reduce the population of coliform bacteria in study ch (kim et al., ) when it was added to the diet at a concentration of mg/kg cu while it had no effect at a concentration of mg/kg. no effect of cu-protionate was found in study ch (aydin et al., ) at a concentration of mg/kg cu or in study ch (kim et al., a) at a concentration of mg/kg cu. cu-propionate at concentrations of and mg/kg cu had no effect on the density of coliform bacteria in si content from layers neither in study ch nor in study ch (paik et al., ) . a significant reduction in the population of coliform bacteria in si content from broilers were found in study ch (ma et al., ) conducted where small intestinal content was incubated for hours with or without addition of copper. in both experiments it was found that mg/kg as cuso reduced the growth of coliform bacteria, while mg/kg cu as cuso or tribasic copper chloride at concentrations of mg/kg and mg/kg had no effect on the growth of coliform bacteria. the effect of copper on the population of coliform bacteria in caecal content has been investigated in seven studies with broilers (ch , ch experiment , ch experiment , ch , ch , ch and ch ). a summary of results from the experiments are shown in table . no effect on the population of coliform bacteria were found by feed concentrations of mg/kg cu as cuso in study ch (kim et al., ) the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. bbj (ch experiment , min et al., ) , or with mg/kg cu as a cu-starch/methionate complex (ch experiment and ch experiment ) or with a na-alginate cu complex (ch experiment ). in general the effect of copper on the gastrointestinal populations of coliform bacteria seems to be rather weak, in most cases where an effect was seen, the concentrations were higher than mg/kg cu; the exception may be copper bound clay particles, especially copper bound nanoparticles that seems to have an effect at concentrations below mg/kg cu (ch and ch ). that high concentrations of copper may reduce the population of coliform bacteria in gut content from chickens is in agreement with the ex vivo experiment studies (ch and ch ) that show that cu as cuso inhibit the growth of coliform bacteria at a concentration of mg/kg cu but not at a concentration of mg/kg cu. enterococci. the effect of copper on the population of enterococci has only been investigated in a single experiment (ch , aydin et al., ) . supplementation of the diet with cu-proteinate corresponding to a cu concentration at mg/kg cu had no significant effect on the population of enterococci in broiler ileal content. lactobacilli. the effect of copper concentration and copper sources on populations of lactobacilli in gut content is shown in table . the population lactic acid bacteria (lab) in content from the gizzard was investigated in three experiments with broilers: ch (kim et al., ) , ch experiment and ch experiment (min et al., ) . no effect of either copper concentration or copper source was found (table ). the effect of copper on the population density of lactobacilli in si content was investigated in experiments with broilers and in experiments with layers. a significant reduction in the population of lactobacilli was found in study ch (choi et al., ) by addition of or mg/kg cu as cuso to the diet, while no effect was found by addition of mg/kg cu as cuso . no effect of the addition of mg/kg cu as cuso was found in studies ch (kim et al ) , ch (min et al., ) , ch experiments and (min et al., ) ; and of mg/kg cu as cuso in study ch (xia et al., ) . in the studies with layers (ch , kim et al., b ) the population of lactobacilli was significantly increased when the diet was supplemented with either or mg/kg cu as cuso . cu-methionat at a cu concentration of mg/kg cu had no effect on the population of lactobacilli (ch , min et al., ) while at cu concentrations of mg/kg, it had a significant stimulating effect on the population of lactobacilli in broiler si content (ch , kim et al., ) . only a tendency to an increase (ch ) was found when it was added to a diet at a concentration of mg/kg cu. in layers fed a diet supplemented with mg/kg cu as cu-methionate a significant increase in the population of lactobacilli was observed (ch , paik et al., ) , while at a concentration of mg/kg it resulted in a tendency to an increase (ch ). addition of cu-propionate corresponding to cu amount of mg/kg to broiler diets resulted in an increase in the population in study ch while no effect was seen in study ch (kim et al., a) . at an addition corresponding to a cu concentration of mg/kg no effect was found of cu-propionate on the population of lactobacilli. in layers addition of cu-propionate corresponding to an addition of mg/kg cu resulted either in a significant reduction in the population of lactobacilli (study ch ) or to a tendency to an increase (study ch ). both in study ch and ch , an increase was detected in the population of lactobacilli when layer diets were supplemented with ppm cu as cu-propionate. no effect on the population of lactobacilli in si content from broilers were found in study ch by addition of or mg/kg cu as cu- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the population lactobacilli in content from the caecum of broilers was investigated in seven experiments (table ) : ch , ch experiment , ch experiment , ch , ch , ch and ch ). in experiment ch , no effects of supplementing feed with mg/kg cu as cuso were found on the population of lactobacilli in cecal content. the same was the case in study experiments and when feed was supplemented with mg/kg cu-starch, cu starch/met and cu-naalginate; in experiment ch , for supplementation with mg/kg cuso ; and in experiments ch and ch for supplementation with and mg/kg cu-montmorillonite. in study ch , addition of mg/kg cu as cu-montmorillonite-nanomaterial to broiler fed stimulated the population of lactobacilli in cecal content. the same was found in study ch when it was added at concentrations of mg/kg and mg/kg cu as cuso to broiler diets, while cu as cuso at a concentration of mg/kg resulted in a significant reduction in the population of lactobacilli. from the investigation of copper concentrations and sources on the population of lactobacilli in gastrointestinal content it may be concluded that copper concentrations at or below mg/kg seems to stimulate the population of lactobacilli in chicken gut content, while it does not seem to have any effect at concentrations from to mg/kg but has an inhibitory effect at concentrations above streptococci. the effect of copper on the population of streptococci was investigated in three experiments with broilers from two studies (ch and ch experiment and ). in study ch (choi et al., ) the population of streptococci were higher after week but lower after weeks in content from the si of broilers fed either , or mg/kg cu as cuso compared to the control fed birds, while no effect were seen of any of the cu concentrations after or weeks on the experimental diets. in study ch experiment (min et al., ) the effect of the addition of mg/kg cu as cuso , cu-starch or cu-starch/methionine to diets to broilers on the population of streptococci in content from the gizzard, upper and lower si and cecum was investigated. no effect of any of the added copper sources was found on the population of streptococci in any of the investigated git segments. in study ch experiment , the effect of the addition of mg/kg cu as cuso , a cu-starch /methionine complex, a cu-starch/methionine complex (added to a diet where the methionine was reduced with the amount of methionine added with the cu-starch/methionine complex) and a na-alginate cu complex to diets of broilers on the population of staphylococci in content from the gizzard, upper and lower si and cecum was investigated. no effect of any of the added copper sources was found on the population of streptococci in any of the investigated git segments. in general supplementing broiler diets with additional copper dose not seems to have any effect on the population of streptococci in the gastrointestinal tract. total aerobe bacteria. the effect of copper on the population of total anaerobic bacteria was investigated in three studies (ch , ch and ch ). no effect was found in study ch (ma et al., ) the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the microbial community was investigated in two studies (ch and ch ). in both studies the microbial community was determined by isolating bacterial dna, amplifying the v region of the s ribosomal dna and performing denaturing gradient gel electrophoreses (dgge). in both studies the number of bands per sample and the similarity indices for bands between samples were calculated. the number of bands is indicative for the number of bacterial species within a sample whereas the similarity coefficients indicate how similar the bacteria within a treatment group are. in study ch (thompsen et al., ) broilers were fed either a control diet, the control diet supplemented with mg/kg bacitracin or the control diet supplemented with mg/kg cu as cuso from day to day . at day , six birds per treatment group were slaughtered and the bacterial community structure analysed in samples of ileal content and samples of ileal mucosa. amplicon profiles from the ileal content revealed that dietary treatment had no effect on band numbers or similarity coefficients. likewise, band numbers of the mucosa-associated bacteria were not affected by dietary treatments. the comparison similarity coefficients, however, revealed that the mucosa-associated bacterial communities of birds fed mg/kg cu as cuso had higher similarity indices than either the control or bacitracin fed broilers. the high similarity coefficient observed in birds fed copper might indicate that addition of high concentrations of copper may select for copper resistant bacteria. in study ch (pang et al., a) broilers were fed either a control diet, a diet supplemented with mg/kg cu as cuso or a diet supplemented with mg/kg cu as tribasic copper chloride (tbcc) from hatch to day . on day eight birds per treatment were slaughtered and the bacterial community structure analysed in samples of ileal content and samples of ileal mucosa. neither cu supplementation at . mg/kg nor cu sources affected the number of predominant bacterial species (dgge band number) in ileal content (mean = . ) or ileal mucosa (mean = . ). furthermore, neither cu supplementation at . mg/kg nor cu sources (cuso or tbcc) affected the similarity coefficient of the microbial community structure in ileal content. however supplementation of the diet with . mg/kg cu as tbcc increased the similarity coefficient of ileal mucosal bacteria, while supplementing the diet with . mg/kg cu as cuso did not affect the similarity coefficient of ileal mucosal bacteria. this shows that the mucosa associated microbiota was more similar between individual birds for the tbcc fed broilers, compared to the control and the cuso fed birds. in conclusion the effect of copper supplementation of broiler diets seems to have little effect on the community structure of the microbiota in the small intestine. no effect of supplementing broiler diets with either or mg/kg cu as cuso or mg/kg cu as tbcc were found on band numbers (number of bacterial species) neither in digesta samples nor in mucosal samples. moreover, no effect of copper www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. bbj supplementation was found on population similarity in digesta samples, while mg/kg cu as cuso affected the population similarity in mucosa samples in study ch but no effect of mg/kg cu as cuso was found on the population similarity in mucosal samples in study ch . in study ch , however, an effect of mg/kg cu as tbcc was found on the population similarity in mucosal samples. fecal moisture content. fecal moisture content was investigated in one study (ch , choi et al., ) . it was measured each week during the six weeks of the experiment. the moisture content was significantly higher at week in the chickens fed and mg/kg cu as cuso and numerically higher in the group fed mg/kg cu as cuso compared to the control group ( . , . , . and . % for the control group and the groups fed diets supplemented with , and mg/kg cu as cuso , respectively). no effect on the moisture content of any of the copper concentrations investigated was found from week to . ph. the effect of copper on ph of gastrointestinal or fecal content was investigated in five experiments with broilers (ch , ch , ch experiment , ch experiment and ch ). in study ch (aydin et al., ), no effect of mg/kg cu as cu-proteinate was found on ph of ileal content in broilers slaughtered at day . in study ch experiment (min et al., ) , the addition of mg/kg cu as either cuso , a cu-starch complex or a cu-starch/methionine complex to broiler diets had no effect on ph of content from gizzard, upper si, lower si or cecum taken from broilers slaughtered at day . in study ch (min et al., ) , supplementation of broiler diets with mg/kg cu as cuso , cu-methionine or cu-fish meal significantly reduced the ph in content from the gizzard ( . , . , . and . for the control group and the three cu supplemented groups, respectively) from broilers slaughtered at day . no effects of any of the copper sources were found on ph in content from the upper or lower si or the caecum. in study ch experiment (pang et al., ), supplementation of broiler diets with mg/kg cu as either cuso , a cu-starch /methionine complex, a cu-starch/methionine complex (added to a diet where the methionine was reduced with the amount of methionine added with the cu-starch/methionine complex) and a na-alginate cu complex significantly reduced the ph in content from the gizzard ( . , . , . , . , . for the control group and the four cu supplemented groups respectively) from broilers slaughtered at day . no effect of any of the copper sources was found on ph in content from the upper or lower si, while the cu starch complex and the na-alginate cu complex significantly reduced the ph in content from the caecum ( . , . and . for the control group and the two cu supplemented groups, respectively). in study ch (shao et al., ) , supplementation of broiler diets with mg/kg cu as cu silicate nanoparticles significantly reduce the ph of content from the caecum from birds slaughtered at day ( . and . for the control fed and the copper fed birds, respectively). no effect of copper supplementation were found on ph of caecal content from birds slaughtered at days or on ph of fecal samples taken throughout the experimental period of days. in conclusion the effect of supplementing broiler diets seems to reduce ph in gizzard content while the effect on the rest of the gastrointestinal tract seems to be marginal. ammonia emission. effect of copper on ammonia emission was investigated in study ch (shi et al., ) . supplementation of broiler diets with mg/kg cu as cu silicate nanoparticles significantly reduce the ammonia emission at day , , , , and to about half of the emission found for the control fed birds. gizzard erosion. the effect of copper on gizzard erosion was investigated in five experiments with broilers (ch , ch , ch , ch experiment , and ch experiment ). in study ch (choi et al., ) , gizzard erosion was investigated in broilers fed four different diets (a control and three diets supplemented with mg/kg, mg/kg and mg/kg cu as cuso ). gizzard erosion was measured in three birds per treatment after , and weeks on the experimental diet. gizzard erosion indices were scored from (normal) to (severe). the incidence of gizzard erosion increased as the level of copper in the diet increased and with time on the experimental diets. after weeks on the experimental diet the gizzard erosion indices were . , . , . and . for the birds fed the www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. bbj control diet and the birds fed , and mg/kg cu as cuso , respectively. in study ch (kim et al., ,) gizzard erosion was investigated in broilers in a x factorial experiment including levels of added fat ( % and %) and levels of added copper (no addition, mg/kg cu as cuso .h o and mg/kg cu as cuso . h o). again increasing gizzard erosion indices were evident in birds fed the copper supplemented diets; both in the birds fed the low and the high fat diet. in experiment ch (min et al., ) , gizzard erosion was investigated in birds fed a control diet and three diets supplemented with three different copper sources ( mg/kg cu as cuso , mg/kg cu as cu-methionine and mg/kg cu as cu-fish meal). gizzard erosion was measured as described for study ch in four birds per treatment after days on the experimental diets. again severe gizzard erosion was observed in the birds fed the copper supplemented diets ( . , . , . and . for the control fed birds and the birds fed mg/kg cu as cuso , cu-methionine and cu-fish meal, respectively). in study ch experiments and (min et al., ) , gizzard erosion was investigated for five sources of copper all added to the diets to increase the cu concentration with mg/kg. the five copper sources were cuso , a cu-starch complex, a cu-starch/methionine complex, a cumethionine complex and a na-alginate cu complex. regardless of copper sources, severe gizzard erosion was observed in all broilers fed copper supplemented diets in all four studies where it was investigated. in conclusion, supplementation of broiler diets with copper at concentrations above mg/kg strongly increases gizzard erosion. ammonia concentration. effect of copper on ammonia concentration in fecal samples was investigated in study ch (shi et al., ) . supplementation of broiler diets with mg/kg cu as cu silicate nanoparticles had no effect on ammonia concentration in fecal samples neither on day nor on day . microbial enzyme activity. the effect of copper on microbial enzyme activity of β-glycosidase and βglucuronidase has been investigated on two studies with broilers (ch and ch ). in study ch , broilers were fed a control diet and two experimental diets supplemented with either or mg/kg cu as cu-montmorillonite-nanomaterial (cu-mmn). the diets were fed to the broilers for days. eight birds from each treatment were slaughtered at day and day and the enzyme activity measured in content from the si and the caecum. the activities of β-glucosidase and β-glucuronidase in the small intestine of birds fed on the diet supplemented with both concentrations cu-mmn were significantly lower than the control, both for the birds slaughtered at day and for the birds slaughtered at day . similarly both enzyme activities were reduced in the cu-mmn fed birds compared to the control fed birds in caecal content at day but not at day . in study ch (xu et al., ) , broilers were fed a control diet and two experimental diets supplemented with either g/kg montomorillonite (mm) or g/kg cu-montomorillonite (cu-mm) (resulting in an addition of mg/kg cu to the feed). the diets were fed to the broilers for days. eight birds from each treatment were slaughtered at day and day and the enzyme activity measured in content from the si and the caecum. the activities of β-glucosidase and β-glucuronidase in the small intestine of birds fed on the diet supplemented with cu-mm were significantly lower than the control both for the birds slaughtered at day and for the birds slaughtered at day . β-glucosidase activities were reduced in the cu-mm fed birds compared to the control fed birds in caecal content at day but not at day . the caecal β-glucuronidase was not significantly affected by cu-mm. the enzyme activities in the mm fed birds were somehow between the control fed birds and the birds fed the copper enriched mm (cu-mm). clostridia: supplementing chicken (broilers and layers) diets with additional copper seems to reduce the population of clostridia in the gastrointestinal tract of the birds, even at low concentrations of copper. especially clay bound copper seems to be effective in reducing the population of clostridia. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. coliform bacteria: in general the effect of copper on the gastrointestinal populations of coliform bacteria seems to be rather weak; in most cases where an effect was seen, the concentrations was higher than mg/kg cu, the explanation may be copper bound clay particles, especially copper bound nanoparticles seem to have an effect at concentrations below mg/kg cu (ch and ch ). that high concentrations of copper may reduce the population of coliform bacteria in gut content from chickens is in agreement with the ex vivo experiment studies (ch and ch ) that show that cu as cuso inhibit the growth of coliform bacteria at a concentration of mg/kg cu but not at a concentration of mg/kg cu. lactobacilli: from the investigation of copper concentrations and sources on the population of lactobacilli in gastrointestinal content it may be concluded that copper concentrations at or below mg/kg seem to stimulate the population of lactobacilli in chicken gut content, while it does not seem to have any effect at concentrations from to mg/kg but has an inhibitory effect at concentrations above mg/kg. in agreement with that, the ex vivo experiments (ch and ) show that mg/kg cu as cuso stimulated the growth of lactobacilli in broiler si content. however no inhibitory effect on the growth was seen by a copper concentration of mg/kg cuso . that no effect of tribasic copper chloride (tbcc) was seen in the ex vivo experiment is properly due to the low solubility of tbcc in water (ch ). streptococci. supplementing broiler diets with additional copper does not seem to have any effect on the population of streptococci in the gastrointestinal tract. aerobe bacteria: only three studies could be found and copper supplementation showed no effect (the effect in study ch may be an effect of the nanoparticles rather than an effect of copper). total anaerobe bacteria: copper supplementation had no effect in most studies (only in one study with high copper concentration). ex vivo studies. mg/kg cu as cuso inhibit the growth of coliform bacteria in chicken si content. no effect of mg/kg cuso on growth of lactobacilli. no effect on growth of either coliform bacteria or lactobacilli by tbcc (properly due to low water solubility). ph. supplementing broiler diets with copper seems to reduce ph in gizzard content while the effect on the rest of the gastrointestinal tract seems to be marginal. community structure. the effect of copper supplementation of broiler diets seems to have little effect on the community structure of the microbiota in the small intestine. neither cu supplementation at . mg/kg or mg/kg nor cu sources (cuso or tbcc) affected the number of predominant bacterial species (dgge band number) in ileal content or ileal mucosa or affected the similarity coefficient of the microbial community structure in ileal content. in study ch , it was found that supplementation of the diet with . mg/kg cu as tbcc increase the similarity coefficient of ileal mucosal bacteria while supplementing the diet with . mg/kg cu as cuso had no effect. in contrast to that, it was found in study ch that supplementing broiler diets with mg/kg cu as cuso increase the similarity coefficient of the mucosa associated microbiota. gizzard erosion. supplementation of broiler diets with copper at concentrations above mg/kg strongly increase gizzard erosion. of the references found for cows by the literature search, were in the first phase considered appropriate to be included in the els. however, a closer examination of the five studies revealed the papers not to be appropriate to be included in the els. one of the studies was published in polish, two of the studies were dealing with antibiotic resistance, one was dealing with the effect of copper on parasite control and the last one on the effect of copper on abomasal ulcers. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. of the references found for pigs and piglet by the literature search, were considered appropriate to be included in the els. in total different characteristics related to the gastrointestinal microbiota were investigated in the selected studies. the number of characteristics investigated in the individual studies varied from to . the number of studies in which each characteristic was studied varied from to . the characteristic studied could be divided in themes: plate count of bacteria ( studies), qpcr of bacterial populations ( study), microscopic counts of bacteria ( study), community structure analyses ( studies), gut content characteristics ( studies), microbial metabolites ( studies), microbial activity ( studies) and studies in germ free pigs ( study). the copper concentrations used in the studies varied from . to mg/kg feed. four types of copper sources were used in the studies: inorganic bound copper ( studies at concentrations from - mg/kg feed), organic bound copper ( studies at concentrations from - mg/kg feed), copper bound to clay minerals ( studies at concentrations from - mg/kg feed) and copper-loaded nanoparticles ( study at a concentration of . mg/kg). it may be concluded from the experiments that supplementing pig diets with low additional copper amounts (below mg/kg cu) seems to inhibit the population of coliform bacteria in the gastrointestinal tract of as well piglets as growing pigs. in particular, clay bound copper seems to be effective. when supplemented to the feed in amounts above mg/kg copper, the effect on the population of coliform bacteria in the gastrointestinal tract of piglets as well as pigs was marginal. supplementing pig diets with copper concentrations above mg/kg as cuso has a reducing effect on the population of lactobacilli in the gastrointestinal tract of as well piglets as growing pigs. the effect of cu as cuo did not affect the population of lactobacilli at high concentrations ( and mg/kg feed). diets with copper concentration below mg/kg feed seem to have marginal effect on the population of lactobacilli in the gastrointestinal tract both in piglets and in growing pigs. the results with growing pigs strongly indicate that the addition of mg cu/kg, or higher, as cuso significantly reduces the population of streptococci in the gastrointestinal tract of growing pigs. the effect of cuso on the population of streptococci in piglets is more questionable. community structure studies strongly suggest that supplementing piglet diets with to mg/kg cu as cuso affects the community structure of the microbiota in the small intestine, the caecum and the colon. the overall conclusion from the studies with piglets and growing pig is that copper, even at low concentrations (< mg/kg feed), may affect the microbiota in the gastrointestinal tract. especially the population of clostridia and coliform bacteria seems to be affected of low concentrations of copper, and especially copper bound clay minerals seem to have an effect. at higher concentrations (> mg/kg feed) cu as cuso reduces the population of lactobacilli in piglets as well as in growing pigs. in growing pigs cuso reduces the population of streptococci in colonic and fecal samples, the population of ureolytic bacteria in the colon of which streptococci make up %, the urease activity in the colon, and decarboxylation and deamination of amino acids in the small intestine. no effect of cu as cuso on the population of streptococci and on urease activity was seen in piglets. supplementing piglet diets with to mg/kg cu as cuso significantly affected the community structure of the microbiota in the small intestine, the caecum and the colon. www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. of the references found for chickens by the literature search, were considered appropriate to be included in the els. in total different characteristics related to the gastrointestinal microbiota were investigated in the selected studies. the number of characteristics investigated in the individual studies varied from to . the number of studies in which each characteristic was studied varied from to . the characteristic studied could be divided in themes: plate count of bacteria ( studies), community structure analyses ( studies), gut content characteristics ( studies), microbial metabolites ( study) and microbial activity ( studies). the copper concentrations used in the studies varied from to mg/kg feed. four types of copper sources were used in the studies: inorganic bound copper ( studies at concentrations from - mg/kg feed), organic bound copper ( studies at concentrations from - mg/kg feed), copper bound to clay minerals ( studies at concentrations from - mg/kg feed) and copper-loaded nanoparticles ( study at a concentration from - mg/kg). from the plate count investigations of bacterial population it can be concluded that supplementing chicken (broilers and layers) diets with additional copper seems to reduce the population of clostridia in the gastrointestinal tract of the birds, even at low concentrations of copper. especially clay bound copper seems to be effective in reducing the population of clostridia. in general the effect of copper on the gastrointestinal populations of coliform bacteria seems to be rather week, in most cases where an effect were seen the concentrations was higher than mg/kg cu, the exception may be copper bound clay particles, especially copper bound nanoparticles seems to have an effect at concentrations below mg/kg cu. that high concentrations of copper may reduce the population of coliform bacteria in gut content from chickens is in agreement with the ex vivo experiment studies that show that cu as cuso inhibits the growth of coliform bacteria at a concentration of mg/kg cu but not at a concentration of mg/kg cu. from the investigation of copper concentrations and sources on the population of lactobacilli in gastrointestinal content it may be concluded that copper concentrations at or below mg/kg seems to stimulate the population of lactobacilli in chicken gut content, while it does not seem to have any effect at concentrations from to mg/kg. however, it has an inhibitory effect at concentrations above mg/kg. in agreement with that, the ex vivo experiments show that mg/kg cu as cuso stimulated the growth of lactobacilli in broiler si content. however, no inhibitory effect on the growth was seen by a copper concentration of mg/kg cuso . in conclusion the effect of supplementing broiler diets with copper at concentrations of mg/kg either as inorganic or organic bound copper seems to reduce ph in gizzard content while the effect in the rest of the gastrointestinal tract seems to be marginal. in conclusion the effect of copper supplementation of broiler diets seems to have little effect on the community structure of the microbiota in the small intestine. neither cu supplementation at . mg/kg or mg/kg nor cu sources (cuso or tbcc) affected the number of predominant bacterial species (dgge band number) in ileal content or ileal mucosa or affected the similarity coefficient of the microbial community structure in ileal content. in study ch , it was found that supplementation of the diet with mg/kg cu as tbcc increase the similarity coefficient of ileal mucosal bacteria while supplementing the diet with mg/kg cu as cuso had no effect. in contrast to that, it was found in study ch that supplementing broiler diets with mg/kg cu as cuso increase the similarity coefficient of the mucosa associated microbiota. supplementation of broiler diets with copper at concentrations above mg/kg strongly increases gizzard erosion. the overall conclusion from the studies with broilers is that copper even at low concentrations (< mg/kg feed) may affect the microbiota in the gastrointestinal tract. especially the population of clostridia seems to be affected by low concentrations of copper. in particular, copper bound clay minerals seem to have an effect. at higher concentrations (> mg/kg feed) inorganic or organic effect of copper intake on gut microbiota (pigs/piglets, chickens, dairy cows) www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. bbj bound copper also seems to affect the population of lactobacilli, and coliform bacteria, to reduce the ph in gizzard content and to produce severe gizzard erosion. of the references found for cows by the literature search, were in the first phase considered appropriate to be included in the els. however, a closer examination of the five studies revealed the papers not to be appropriate to be included in the els. one of the studies was published in polish, two of the studies were dealing with antibiotic resistance, one was dealing with the effect of copper on parasite control and the last one on the effect of copper on abomasal ulcers. while implementing this extensive literature search some suggestions and remarks were raised:  the quality of the papers retrieved from the search on chickens was generally judged to be low. out of papers were written in foreign languages ( in korean and in chinese) with english abstracts and table legends .  a closer examination of the five studies first selected to be included in the els study on cows, revealed the papers not to be appropriate to be included in the els. one of the studies (cow ) was published in polish, two of the studies were dealing with antibiotic resistance (cow and cow ), one was dealing with the effect of copper on parasite control (cow ) and the last one on the effect of copper on abomasa ulcers (cow ). www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. schole j, szasz e, peters j, eikemeyer j and sickel e, . the total anabolic activity of the blood as a connecting link between the intestinal-flora theory and the intermediary action of growth the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. new bacterium comprising lactic acid bacterium and bacterium of genus bifidobacterium comprising metal nanoparticle bound to its surface, useful e.g. as contrast agent for mri of digestive tract of subject, and for treating cancer contribution to the study of gut hypersensitivity reactions to soybean proteins in preruminant calves and early-weaned piglets ddesaturation of organ lipids and tissue-lipids and influence on depot triglyceride structure by feeding high levels of different copper-compounds composite preblend material for pig capable of improving immunologic function includes ferrous sulfate, copper sulfate, zinc sulfate, manganese sulfate, composite microbial ecological agent, ethoxyquinoline, choline chloride and bran nutritional alternatives for antimicrobial control in pigs scientific opinion on the safety and efficacy of copper compounds (e ) as feed additives for all animal species: cupric sulphate pentahydrate based on a dossier submitted by manica s swine. proceedings of the rd western nutrition conference, optimizing efficiency of animal production how to avoid making mistakes when choosing alternatives to feed antibiotics. svinovodstvo (moskva) silver nanoparticles as a potential antimicrobial additive for weaned pigs utilization of fermented flocculated poultry sludge as a feed constituent for pigs nutritional compound additive alleviates jejunal mucosal structure disruption of piglets challenged with feed containing corn contaminated with mycotoxins this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document antibacterial effects of the cu(ii)-exchanged montmorillonite on escherichia coli k and salmonella choleraesuis antibacterial effects of the cu(ii)-loaded montmorillonite in vitro and its on mechanisms effect of supplementing tylosin and olaquindox or copper on growth, tissue mineral contents and enteric bacteria counts of weanling pigs bacterial resistance to copper in animal feeds feed without antibiotic by microbial fermentation for later period of growing/fattening pigs comprises batch mixture and compound fermented material e.g. lactobacillus acidophilus, aspergillus niger in specific weight ratio microbially fermented piglet early growth feed without antibiotic comprises e.g. bacillus subtilis, aspergillus based fermentation material and composite material action of antimicrobial agents on intestinal bacteria of pigs in vitro gastrointestinal health and 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published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document copper and manganese in hay samples from scrapie-free, scrapie-prone and scrapie-afflicted farms in iceland icelandic agricultural sciences microbial decomposition of organic cpds.|comprises a mixt. of magnetic field-treated gastrointestinal microorganism culture and micronutrients rumen bacterial metabolism as affected by extracellular redox potential antibiotic growth promoters in animal nutrition assessment of sustainable vermiconversion of water hyacinth by eudrilus eugeniae and eisenia fetida alterations in ruminal utilization of magnesium and zinc in lambs fed different ratios of concentrate -forage chronic botulism in a saxony dairy farm: sources, predisposing factors, development of the disease and treatment possibilities influence of experimentally induced theileriosis (theileria annulata) on the pharmacokinetics of a long-acting formulation of oxytetracycline (otc-la) in calves effects of experimentally induced theileria annulata infection on the pharmacokinetics of oxytetracycline in cross-bred calves effects of freeze-dried mitsuokella jalaludinii culture and natuphos (r) phytase supplementation on the performance and nutrient utilisation of broiler chickens effects of altered dietary iron intake in mycobacterium-paratuberculosis-infected dairy-cattle -sequential observations on growth, iron and copper-metabolism and development of para-tuberculosis intracellular iron storage and the pathogenesis of para-tuberculosis -comparative studies with other mycobacterial, parasitic or infectious conditions of veterinary importance new biologically pure culture of a probiotic bacteria strain for producing bacteriocins useful in animal feed formulation for promoting growth and feed conversion in monogastric animal e.g. avian, dog, rabbit, mink effect of copper deficiency upon erythro kinetics in growing cattle preliminary results praseodymium and copper cation-exchange capacities of neutral-detergent fibers relative to composition and fermentation kinetics this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author purification and characterization of bacterial cm-cellulase from camel rumen fluid ( camelus dromedarius) studies on the mechanism of toxicity of the mycotoxin sporidesmin . . inhibition by copper-chelating agents of the generation of superoxide radical by sporidesmin protective efficacy of azomite enriched diet in oreochromis mossambicus against aeromonas hydrophila the diagnosis of adult bovine diarrhea -trial to put forward a new technique acute sheep poisoning from a copper sulfate footbath combined control of the principal diseases of sheep in the spring khimioprofilaktika, patogenez i epizootologiya parazitozov sel'skokhozyaistvennykh zhivotnykh phosphorus digestion and metabolism in ruminants: applications to production disease and environmental considerations. production diseses in farm animals th international conferences east lansing copper deficiency in an infant with giardiasis fed with cows milk biogenic amines in animal nutrition: origin, metabolism and physiological aspects uses of plants, animal and mineral substances in mediterranean ethno-veterinary practices for the care of small ruminants prevalence of strains with multiple resistance to antibiotics, and the transferability in vitro of the r factor of strains of escherichia coli and salmonella of various origins. lucrari stiintifice, institutul agronomic isolation of clostridiumperfringens from neonatal calves with ruminal and abomasal tympany, abomasitis, and abomasal ulceration annual report of studies in animal nutrition and allied sciences annual report of studies in animal nutrition and allied sciences. volume parasitic coenoses in the intestine of sheep and their control. ussr, akademiya nauk ukrainskoi ssr, institut zoologii the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. the present document has been produced and adopted by the bodies identified above as author. this task has been carried out exclusively by the author in the context of a contract between the european food safety authority and the author, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author. key: cord- - w fkd authors: nan title: abstract date: - - journal: eur j epidemiol doi: . /s - - - sha: doc_id: cord_uid: w fkd nan the organisers of the european congress of epidemiology , the board of the netherlands epidemiological society, and the board of the european epidemiology federation of the international epidemiological association (iea-eef) welcome you to utrecht, the netherlands, for this iea-eef congress. epidemiology is a medical discipline that is focussed on principles and methods of research on causes, diagnosis and prognosis of disease, and establishing the benefits and risks of treatment and prevention. epidemiological research has proven its importance by contributions to the understanding the origins and consequences of diseases, and has made major contributions to the management diseases and improvement of health in-patients and populations. this meeting provides a major opportunity to affirm the scientific and societal contributions of epidemiological research in health care practice, both in clinical medicine and in public health. during this meeting major current health care problems are addressed alongside methodological issues, and the opportunities and challenges in approaching them are explored. the exchange of ideas will foster existing co-operation and stimulate new collaborations across countries and cultures. the goal of this meeting is to promote the highest scientific quality of the presentations and display advanced achievements in epidemiological research. our aim is to offer a comprehensive and educational programme in the field of epidemiological research in health care and public health, and create room for discussions on contemporary methods and innovations from the perspective of policy makers, professionals and patients. above all, we want to stimulate open interaction among the congress participants. your presence in utrecht is key to an outstanding scientific meeting. the european congress of epidemiology is organised by epidemiologists of utrecht university, under the auspices of the iea-eef, and in collaboration with the netherlands epidemiological society. utrecht university, founded in , is the largest university in the netherlands and harbours the largest academic teaching hospital in the netherlands. the epidemiologists from utrecht university work in the faculties of medicine, veterinary medicine, pharmacy and biology. the current meeting was announced through national societies, taking advantage of their newsletters and of the iea-eef newsletter. in addition, avoiding the costs and disadvantages of the traditional journal advertisements and leaflets, information about the congress was disseminated via an internet mailing list of epidemiologists, which was compiled from, among other, the meeting in porto in , the european young epidemiologist network (http://higiene.med.up.pt/eye/) and several institutions and departments. many of the procedures followed this year were based on or directly borrowed from the stimulating iea-eef congress in porto in . publication in an international journal of large circulation of the congress programme and abstracts selected for oral and poster presentation, signifies the commitment of the organisers towards all colleagues that decided to present their original work at our meeting, and is intended to promote our discipline and to further stimulate the quality of the scientific work of european epidemiologists. and methods to the objectives and quality of its description; presentation of results; importance of the topic; and originality. a final rating was given on a - point scale. the two junior epidemiologists independently evaluated each abstract. based on ratings of the juniors, the senior epidemiologist gave a final abstract rating. the senior reviewer decided when juniors disagreed, and harnessed against untoward extreme judgements of the juniors. based on the judgement by the seniors abstracts with a final rating of or higher were accepted for presentation. next, in order to shape the scientific programme according to scientific and professional topics and issues of interest for epidemiologists, members of the scientific programme committee grouped the accepted abstracts in major thematic clusters. for these, topics, keywords and title words were used. within each cluster, abstracts with a final rating of or higher, as well as abstracts featuring innovative epidemiological approaches were prioritised to be programmed as an oral presentation. the submitted abstracts had an average final rating of (sd= ). in total abstracts ( %) with a final rating of or lower were rejected. because of the thematic programming some abstracts with a final rating of or higher will be presented as posters, while few with a final rating of are programmed as oral presentation. there were abstracts ( %) accepted and programmed for poster presentation; each poster will be displayed for a full day. in total abstracts ( %) are accepted for oral presentation. these are programmed in parallel sessions. based on the topics of their abstracts the oral sessions were arranged into themes, notably epidemiology of diseases, methods clinical & population research, burden of disease, high risk populations, growth and development, public health strategies, translational epidemiology. sessions from one theme never are programmed parallel. in table we present the submitted and accepted abstracts (oral or poster) according to the distribution of country of origin. in table submitted abstracts are displayed according to their topic, as classified by the authors using the topic long list of the submission form. the scientific programme committee convened in a telephone meeting by the end of the summer of and decided on the above programming process. the scientific programme committee was informed on the result of the programming process by the end of april . fifteen abstracts were submitted for the eye session work in progress. of these abstracts were selected for oral presentation and thereby nominated for the eye award. in total, abstracts were submitted in relation to the student award, of which were programmed for oral presentation and thus nominated. during the congress authors of poster presentations may name themselves as candidate for the poster award. during the closing ceremony the winners of the student award and the poster award will be announced. these awards are an initiative of the netherlands epidemiological society that will fund them in . according to the iea rules expenses of congress participation for applicants from low-income countries will be covered. the board of the iea-eef will select a maximum of candidates; their travel and registration expenses will be (partly) covered from the congress budget. in order to stimulate participation form as many as possible junior researchers and young epidemiologists the congress budget covers registration fee reduction for undergraduate (msc) participants and eye members. this also holds for the registration fee reduction of iea-eef members and nes members. it is years ago ( ) that the iea regional european meeting was held in the hague, the marcon wei leray gehring leray spallek greving laan brussee brussee may- de groot hoefman goettsch posters ordered by abstract number olawuyi de vries uotinen smits gimeno houben de vries de vries ten berg diaconu diaconu streppel belo sauvaget koopman kilsztajn pembrey schmidt kretzschmar scholtens defraye medronho eljedi van de garde mello hosper feleus bayingana de wit stolk teixeira teixeira verhoef capon de boer lazarevska terschu¨ren khosravi boroujeni molag mikolajczyk luijsterburg stolk mirabelli barreto curzio pereira van gageldonk-lafeber van nispen roobol mokkink rava haukka jansen de kraker bogers donalisio behrens borders melis pac muller van den hooven van der sande van den berg novoa van den boogaard vannoord koedijk kuczerowska giorgi rossi vannoord baussano agabiti bierma-zeinstra van wier faustini jarrin juhl miguel maira lindert van den berg mierzejewska fonseca cardoso martens cotton corte ursoniu vernic boer ruskamp szurkowska bijkerk fonseca cardoso mazur ahrens dijkstra ajdacic-gross ajdacic-gross lucas santos gielkens-sijstermans tobi de kraker proteomics and genomics are supposed to be related to epidemiology and clinical medicine, among other because of the putative diagnostic usefulness of proteomics and genomics tests. hence, clinical and sometimes even public health applications are promised by basic sciences. it is debated whether such promises and subsequent expectation are fulfilled. what are at meaningful and consequential examples of current findings in proteomics, genomics and similar approaches in biomedical research? are they different from the ''classic'' tools and frameworks of clinical epidemiology? in the context of proteomics and genomics, etiologic studies, primary prevention, epidemiological surveillance and public health are concerned with the influence of environmental exposures on gene expression and on the accumulation of genetic alterations. proponents and advocates of proteomics and genomics have suggested that their products can yield clinically useful findings, e.g., for early diagnosis, for prognosis, for therapeutic monitoring, without always needing to identify the proteins, peptides or other 'biomarkers' at stake. do we feel comfortable with this ''black-box'' reasoning, i.e. do we question the role of pathophysiological and mechanistic reasoning in clinical medicine? how much sense does it make for epidemiology to play with and scrutinize proteomics and genomics approaches in epidemiology and clinical medicine? what are at present (and in the near future) the main biological, clinical and public health implications of current findings in these research fields? in this plenary session these and other questions regarding the place and role of proteomics and genomics in clinical epidemiological research are discussed from different perspectives. infection diseases: beneficial or disaster for man? infectious diseases pose an increasing risk to human and animal health. they lead to increasing mortality, in contrast to the situation fifty years ago when new control measures still provided hope of overcoming many problems in the future. improved hygiene, better socio-economic circumstances, vaccination and use of antibiotics has led to a gradual decline of tuberculosis, rheumatic fever, measles and mumps in western societies over the last five decades. paradoxically, absence of exposure to infectious agents has a major impact as well. the decline in infectious disease risk is accompanied by a gradual increase of allergic and autoimmune diseases and this association is believed to be causal. exposure to infectious agents from early on in life can markedly boost an individual's natural resistance and hence influence the individual's reaction to future exposure to both biological and non-biological antigens. in this plenary session we want to emphasise both aspects of the effect of infectious agents on human and animal health. evidence based medicine in health care practice and epidemiological research p. glasziou & l. bonneux evidence-based medicine is defined as the conscientious, explicit and judicious use of current best evidence in making decisions about the care of individual patients. proponents of evidence-based medicine maintain that coming form a tradition of pathophysiological rationale and rather unsystematic clinical experience, clinical disciplines should summarize and use evidence concerning their practices, by using principles of critical appraisal and quantitative clinical reasoning. for this they should convert clinical information needs into answerable questions, locate the available evidence, critically appraise that evidence for its validity and usefulness, and apply the results of the best available evidence in practice. applying the principles of evidence-based medicine implies improvement of the effectiveness and efficiency of health care. therefore, evidence-based medicine has commonalties with clinical medical and epidemiological research. for integration of evidence-based medicine into health care practice the challenge is to translate knowledge from clinical medical and epidemiological research, for example in up to date practice guidelines. the limitations of using evidence alone to make decisions are evident. the importance of the values and preference judgments that are implicit in every clinical management decision are also evident. critics of evidence based medicine argue that applying best available research evidence in practice in order to improve the effectiveness and efficiency of health care contradicts with the importance of the values and preference judgments in clinical management decisions. in this plenary session we want to contrast these and other viewpoints on evidence based medicine in health care practice. statistical topics i: missing data prof. dr. t. stijnen this workshop will be an educational lecture on missing data by professor stijnen from the department of epidemiology and biostatistics of the erasmus mc rotterdam, the netherlands. every clinical or epidemiological study suffers from the problem of missing data. in practice mostly simple solutions are chosen, such as restricting the analysis to individuals without missing data or imputing missing values by the mean value of the individuals with observed data. it is not always realised that these simple approaches can lead to considerable bias in the estimates and standard errors of the parameters of interest. in the last to years much research has been done to better methods for dealing with missing values. in this workshop first an overview will be given of the methods that are available and their advantages and disadvantages will be discussed. most attention will be given to multiple imputation, to date generally considered as the best method for dealing with missing values. also the available software in important statistical packages such as spss and sas will be shortly discussed. prof. dr. b. van hout suppose that one wants to know how often individuals have a certain characteristic, and suppose that one doesn't have any knowledge -any knowledge at all -how often this is the case. now, suppose that one starts by checking individuals and only finds individual with this characteristic. than the probability that the ' th individual has the characteristic is / . the fact that this is not / (although it will be close to that if the numbers of observations increase) may be counter-intuitive. it will become less so, when realising how it is obtained from the formal integration of the new information with the complete uncertainty beforehand. this formal integration, with a prior indicating that it is as likely to be / as it is to be / as it is / , and with negative and positive observation -is by way of bayes rule. the italian mathematician, actuary, and bayesian, bruno de finetti ( - , estimated that it would take until the year for the bayesian view of statistics to completely prevail. the purpose of this workshop is to not only convince the attendants that this is appealing outlook but also to aid the workshop participants in realising this prediction. after a first introduction of the work of reverend bayes, a number of practical examples are presented and the attendant is introduced in the use of win-bugs. a first example -introducing the notion of noninformative priors -concerns a random effects logistic regression analysis. second, the use of informative priors, is illustrated (in contrast with non-informative priors) using an analysis of differences in quality of life as observed in a randomised clinical trial. it will be shown how taking account of such prior information changes the results as well as showing how such information may increase the power of the study. in a third example, it will be shown how winbugs offers a powerful and flexible tool to estimate rather complex multi-level models in a relatively easy way and how to discriminate between various models. within this presentation some survival techniques (or stress control techniques) will be presented for when winbugs starts to spit out obscure error-codes without giving the researcher any clue where to search for the reason behind these errors. communicating research to the public h. van maanen, prof. dr. ir. d. kromhout, a. aarts most researchers will at some point in their career face difficulties in communicating research results to the public. whereas most scientific publications will pass by the larger public in silence, now and then a publication provokes profound interest of popular press. interest from the general public should be regarded as positive. after all, public money is put into research and a researcher has a societal responsibility of spreading new knowledge. however, often, general press interest is regarded upon as negative by the researcher. the messages get shortened, distorted or ridiculed. whose responsibility is this misunderstanding between press and researchers? should a researcher foresee press reaction and what can be done to prevent negative consequences? is the press responsible background and relevance: patients with a carotid artery stenosis, including those with an asymptomatic or moderate stenosis, have a considerable risk of ischemic stroke. identification of risk factors for cerebrovascular disease in these patients may improve risk profiling and guide new treatment strategies. objectives and question: we cross-sectionally investigated whether carotid stiffness is associated with previous ischemic stroke or tia in patients with a carotid artery stenosis of at least %. design and methods: patients were selected from the second manifestations of arterial disease (smart) study, a cohort study among patients with manifest vascular disease or vascular risk factors. arterial stiffness, measured as change in lumen diameter of the common carotid arteries during the cardiac cycle, forms part of the vascular screening performed at baseline. the first participants with a stenosis of minimally % in at least one of the internal carotid arteries measured by duplex scanning were included in this study. logistic regression analysis was used to determine the relation between arterial stiffness and previous ischemic stroke or tia. results: the risk of ischemic stroke or tia in the highest quartile (stiffest arteries) relative to the lowest quartile was . ( % ci . - . ). these findings were adjusted for age, sex, systolic blood pressure, minimal diameter of the carotid artery and degree of carotid artery stenosis. conclusion and discussion: in-patients with a ‡ % carotid artery stenosis, increased common carotid stiffness is associated with previous ischemic stroke and tia. measurement of carotid stiffness may improve selection of high-risk patients eligible for carotid endarterectomy and may guide new treatment strategies. background (and relevance): patients with advanced renal insufficiency are at increased risk for adverse cardiovascular disease (cvd) outcomes. objectives and question: the aim was to establish whether impaired renal function is an independent predictor of cvd and death in an unselected high-risk population with cvd. design and methods: the study was performed in patients with cvd. primary outcomes were all vascular events and all cause death. during a median follow-up of months, patients had a vascular event ( %) and patients died ( . %). results: the adjusted hazard ratio (hr) of an estimated glomerular filtration rate < vs > ml/min per . m was . ( % ci . - . ) for vascular events and . ( % ci . - . ) for all cause death. for stroke as a separate outcome it was . ( % ci . - . ) . subgroup analysis according to vascular disease at presentation or the risk factors hypertension, diabetes and albuminuria had no influence on the hr's. conclusion and discussion: renal insufficiency is an independent risk factor for adverse cvd events in patients with a history of vascular disease. renal function was a particularly important factor in predicting stroke. the presence of other risk factors hypertension, diabetes or albuminuria had no influence on the impact of renal function alone. background and relevance: patients with hypertension have an increased case-fatality during acute mi. coronary collateral (cc) circulation has been proposed to reduce the risk of death during acute ischemia. objectives and question: we determined whether and to which degree high blood pressure (bp) affects the presence and extent of cc-circulation. design and methods: cross-sectional study in patients ( % males), admitted for elective coronary angioplasty between january and july . collaterals were graded with rentrop's classification (grade - ). ccpresence was defined as rentrop-grade ‡ . bp was measured twice with an inflatable cuff-manometer in seated position. pulse pressure was calculated by systolic blood pressure (sbp) )diastolic blood pressure (dbp). mean arterial pressure was calculated by dbp + / *(sbp-dbp). systolic hypertension was defined by a reading ‡ mmhg. we used logistic regression with adjustment for putative confounders. results: sbp (odds ratio (or) . per mmhg; % confidence-interval (ci) . - . ), dbp (or . per mmhg; % ci . - . ), mean arterial pressure (or . per mmhg; % ci . - . ), systolic hypertension (or . ; % ci . - . ), and antihypertensive treatment (or . ; % ci . - . ), each were inversely associated with the presence of cc's. also, among patients with cc's, there was a graded, significant inverse relation between levels of sbp, levels of pulse pressure, and collateral extent. conclusion and discussion: there is an inverse relationship between bp and the presence and extent of cc-circulation in patients with ischemic heart disease. background and relevance: silent brain infarcts are associated with decreased cognitive function in the general population. objectives and question: we examined whether this relation also exists in patients with symptomatic arterial disease. furthermore, we compared cognitive function of patients with stroke or tia, with cognitive function of patients with symptomatic arterial disease at other sites in the arterial tree. design and methods: an extensive screening was done in consecutive patients participating in the second manifestations of arterial disease (smart) study, including a neuropsychological test. inclusion diagnoses were cerebrovascular disease, symptomatic coronary artery disease, peripheral arterial disease, or abdominal aortic aneurysm. mri examination was performed to assess the presence of silent infarcts in patients without symptomatic cerebrovascular disease. the patients were assigned to one of three categories according to their patient history and inclusion diagnosis: no stroke or tia, no silent infarcts (n = ; mean age years); no stroke or tia, but silent infarcts present (n = ; mean age years); stroke or tia at background and relevance: patients with manifest vascular disease are at high risk of a new vascular event or death. modification of classical risk factors is often not successful. objectives and question: we determined whether the extra care of a nurse practitioner (np) could be beneficial to the cardiovascular risk profile of high-risk patients. design and methods: randomised controlled trial based on the zelen design. patients with manifestations of a vascular disease and who had ‡ modifiable vascular risk factors were prerandomised to receive treatment by a np plus usual care or usual care alone. after year, risk factors were re-measured. primary endpoint was achievement of treatment goals for risk factors. results: of the pre-randomised patients, of ( %) in the intervention group and of ( %) in the control group participated in the study. after a mean follow-up of months, the patients in the intervention group achieved significantly more treatment goals than did the patients in the control group (systolic blood pressure % versus %, total cholesterol % vs %, ldl-cholesterol % vs %, and bmi % vs %). medication use was increased in both groups and no differences were found in patients' quality of life (sf- ) at followup. conclusion and discussion: treatment delivered by nps, in addition to a vascular risk factor screening and prevention program, resulted in a better management of vascular risk factors compared to usual care alone in vascular patients after year follow-up. were used as non-invasive markers of vascular damage and adjusted for age and sex if appropriate. results: the prevalence of the metabolic syndrome in the study population was %. in pad patients this was %; in chd patients %, in stroke patients % and in aaa patients %. patients with the metabolic syndrome had an increased mean imt ( . vs. . mm, p-value < . ), more often a decreased abpi ( % vs. %, p-value . ) and increased prevalence of albuminuria ( % vs. %, p-value . ) compared to patients without this syndrome. an increase in the number of components of the metabolic syndrome was associated with an increase in mean imt (p-value for trend < . ), lower abpi (p-value for trend < . ) and higher prevalence albuminuria (p-value for trend < . ). conclusion and discussion: in patients with manifest vascular disease the presence of the metabolic syndrome is associated with advanced vascular damage. background (and relevance): in patients with type diabetes the progression of atherosclerosis is accelerated, as observed by the high incidence of cardiovascular events. objectives (and question): to estimate the influence of location and extent of vascular disease on new cardiovascular events in type diabetes patients. design and methods: diabetes patients (n = ), mean age years, with and without prior vascular disease were followed through - (mean follow-up years). patients with vascular disease (n = ) were classified according to symptomatic vascular location, and number (extent) of locations. we analyzed occurrence of new (non)-fatal cardiovascular events using cox proportional hazards models and kaplan-meier analysis. results: multivariate-adjusted hazard ratios (hrs) were comparable in diabetes patients with cerebrovascular disease (hr . ; % ci . - . ), coronary heart disease (hr . ; . - . ) and peripheral arterial disease (hr . ; . - . ), compared to those without vascular disease. multivariate-adjusted hr was . ; ( . - . ) in patients with one vascular location and . ; ( . - . ) in those with ‡ locations. the -year risks were respectively . % ( . - . ) and . % ( . - . ) . conclusion and discussion: diabetes patients with prior vascular disease have an increased risk of cardiovascular events, irrespective of symptomatic vascular location. cardiovascular risk increased with the number of locations. data emphasize the necessity of early and aggressive treatment of cardiovascular risk factors in diabetes patients. background (and relevance): despite recent advances in medical treatment, cardiovascular disease (cvd) is still health problem number one in western societies. a multifactorial approach with the aid of nurse practitioners (nps) is beneficial for achieving treatment goals and reducing events in patients with manifest cvd. objectives (and question): in the self-management of vascular patients activated by internet and nurses (spain) pilot study, we want to implement and test a secure personalized website with additional treatment and coaching of a np for hypertension, hyperlipidemia, diabetes mellitus, smoking and obesity in patients with clinical manifestations of cvd. design and methods: interesting patients are going to use the secure patient-specific website. before the use of the web-application, risk factors are measured. realistic treatment goal(s) for elevated risk factors based on current guidelines are made and appointments how to achieve the treatment goal(s) are determined between the patients and the np in a face to face contact. patients can enter his/ her own weight or a new blood pressure measurement for instance, besides the regular exchange information with the responding np through e-mail messages. the np personally replies as quick as possible and gives regular but protocol driven feedback and support to the patient. the risk factors are remeasured after six months. conclusion and discussion: the spain study is aimed to implement and test a patient specific website. secondary outcome is the change in cardiovascular risk profile. the pre-post measurements of risk factors and the amount of corresponding e-mail messages between the patient and the np enhances the feasibility of this innovative way of risk factor management. background (and relevance): modification of vascular risk factors has been shown to be effective in reducing mortality and morbidity in patients with symptomatic atherosclerosis. nevertheless, reduction of risk factors in clinical practice is difficult to achieve and maintain. objectives (and question): in the risk management in utrecht and leiden evaluation (rule) study, a prospective, comparative study, we assess the effects of a multidisciplinary vascular screening program on improvement of the cardiovascular risk profile and to compare this to a setting without such a program that provides current standard practice in patients referred for cardiovascular disorders. design and methods: patients with diabetes mellitus, coronary artery disease, cerebrovascular disease, or peripheral arterial disease ( per disease category in each hospital) referred by the general practitioner will be enrolled, started january . at the umcu, patients need to be enrolled in the vascular screening program or will be identified through the hospital registration system. at the lumc patients will be identified through the hospital registration system. risk factors will be measured in the two hospitals at baseline and one year after their initial visit. a risk function will be developed for this population based on data of the whole cohort. analysis will be performed on the two comparison groups as a whole, and on subgroups per disease category. changes in risk factors will be assessed with linear or logistic regression procedures, adjusting for differences in baseline characteristics between groups. conclusion and discussion: the rule study is aimed to evaluate the added value of a systematic hospital based vascular screening program on risk factor management in patients at high risk for vascular diseases. background: signs of early cerebral damage are frequently seen on mri scans in elderly people. they are related to future manifest cerebrovascular disease and cognitive deterioration. cardiovascular risk factors can only partially explain their presence and progression. evidence that inflammation is involved in atherogenesis continues to accumulate. chronic infections can act as an inflammatory stimulus. it is possible that subclinical inflammation and chronic infections play a role in the pathogenesis of early cerebral damage. objectives (and question): to unravel the role of inflammation and chronic infection in the occurrence and progression of early cerebral damage in patients with manifest vascular disease. design and methods: participants of the smart study with manifest vascular disease underwent an mr investigation of the brain between may and december . starting in january of all patients are invited for a second mr of the brain after an average follow-up period of four years. both at baseline and after follow-up all cardiovascular risk factors are measured and blood samples are stored to assess levels of inflammatory biomarkers and antibodies against several pathogens. occurrence and progression of early cerebral damage is assessed by measuring the volume of white matter lesions, the number of silent brain infarctions, cerebral atrophy, aberrant metabolic ratios measured with mr spectroscopy and cognitive function at baseline and after follow-up. the relation between inflammation, chronic infection and the occurrence and progression of early cerebral damage will be investigated using both crosssectional and longitudinal analysis. abstract monocyte chemoattractant protein (mcp- ) polymorphism and susceptibility for coro-nary collaterals j.j. regieli , , j. koerselman , ng sunanto , , m. entius , p.p. de jaegere , y. van der graaf , d.e. grobbee , p.a. doevendans heart lung institute, dept of cardiology clinical epidemiology, julius center for health sciences and primary care, utrecht, netherlands background (and relevance): collateral formation is an important beneficial condition during an acute ischemic event. a marked interindividual variability in high risk patients is seen, but at present the basis for this variability is unclear and can not be explained solely by environmental factors. a genetic factor might be present that could influence coronary collateral formation. objectives (and question): we have analyzed the association between a single nucleotide polymorphism in mcp- and the formation of coronary collaterals in patients admitted for angioplasty. mcp- has been suggested to play an important role in collateral development. design and methods: this study involved caucasian patients who were admitted for coronary angioplasty. coronary collateral development was defined angiographically as rentropgrade ‡ . polymorphisms in the promoter region of mcp- () ) were identified by pcr and allele specific restriction digestion. this method allows identification of individuals with either aa, ag or gg at mcp- position ) . statistical analysis was performed using a x -test, unconditional logistic regression, likelihood ration and a wald's test. results: we could genotype of the patients. coronary collaterals (rentropgrade > ) were found in patients. the allele frequency for aa, ag and gg was . %, . % and . %, respectively. the dis-tribution of mcp- genotypes in subjects without collaterals was in hardy weinberg equilibrium. we found that individuals with g allele ( %) were more likely to have collaterals than those with homozygous aa (or . , % ci . to . ) adjusted for potential confounders. linear regression shows that the allele g increased the likelihood for collateral presence with a factor . . conclusion and discussion: this study provides evidence for a role for genetic variation of mcp- gene in the occurrence of coronary collaterals in high risk patients. until september patients with recently established clinically manifest atherosclerotic disease with > modifiable vascular risk factors were selected for the study. the mean self-efficacy scores were calculated for vascular risk factors (age, sex, vascular disease, weight, diabetes mellitus, smoking behavior, hypercholesterolemia, hypertension, and hyperhomocysteinemia). results: diabetes, overweight, and smoking, but none of the other risk factors, were significantly associated with the level of self-efficacy in these patients. patients with diabetes had lower self-efficacy scores ( . ) for exercise and controlling weight ( . ) than patients without diabetes ( . p = . ) and ( . p = . ) respectively. overweight patients scored low on controlling weight ( . and . p< . ) and choosing healthy food ( . and . p = . ) than patients who were on a healthy weight ( . and . ). conclusion and discussion: patients with vascular diseases appear to have high levels of self-efficacy regarding medication use ( . ), exercise ( . ), and controlling weight ( . ). in patients with diabetes, overweight and in smokers, self efficacy levels were lower. practice implications: in nursing care and research on developing self-efficacy based interventions, lower self-efficacy levels can be taken into account for specific vascular patient groups. background (and relevance): little is known about the role of serum uric acid in the metabolic syndrome and increased risk of cardiovascular disease. we investigated the association between uric acid levels and the metabolic syndrome in a population of patients with manifest vascular diseases and whether serum uric acid levels conveyed an increased risk for cardiovascular disease in patients with the metabolic syndrome. design and methods: this is a nested case-cohort study of patients originating from the second manifestations of arterial disease (smart) study. all patients had manifest vascular diseases, constituting peripheral artery disease, cerebral ischemia, coronary artery disease and abdominal aortic aneurysm. analyzing the relationship of serum uric acid with the metabolic syndrome, age, sex, creatinine clearance, alcohol and diuretics were considered as confounders. investigating the relationship of serum uric acid levels with the risk for cardiovascular disease, values were adjusted for age and sex. results: the metabolic syndrome was present in % of the patients. serum uric acid levels in patients with metabolic syndrome were higher compared to patients without ( . ± . mmol/l vs. . ± . mmol/l). serum uric acid concentrations increased with the number of components of the metabolic syndrome ( . mmol/l to . mmol/l) adjusted for age, sex, creatinine clearance, alcohol and use of diuretics. increased serum uric acid concentrations showed to be independently associated with the occurrence of cardiovascular events in patients without the metabolic syndrome (age en sex adjusted hr: . , % ci . - . ) , contrary to patients with the metabolic syndrome (adjusted hr: . , % ci . - . ). conclusion: elevated serum uric acid levels are strongly associated with the metabolic syndrome, yet are not linked to an increased risk for cardiovascular disease in patients with the metabolic syndrome. however, in patients without the metabolic syndrome elevated serum uric acid levels are associated with increased risk for cardiovascular disease. the objective of this study is to investigate the overall and combined role of late-life depression, prolonged psychosocial stress exposure, and stress hormones in the etiology of hippocampal atrophy and cognitive decline. design and methods: as part of the smart study, participants with manifest vascular disease underwent an mri of the brain between may and december . in a subsample of subjects, cognitive function and depressed mood were assessed. starting in january , all patients are invited for a follow-up mri of the brain. at this follow-up measurement, minor and major depression, hypothalamic-pituitary-adrenal (hpa) axis function indicated by salivary cortisol, psychosocial stress exposure indicated by stressful life events early and later in life, and cognitive functioning will also be assessed. the independent and combined effects of late-life depression, (change in) hpa-axis activity, and psychosocial stress exposure on risk of hippocampal atrophy and cognitive decline will be estimated with regression analysis techniques adjusting for potential confounders. introduction the netherlands epidemiological society advocates according to good epidemiological practice, that research with sound research questions and good methodology should be adequately published independent of the research outcomes. although reporting bias in clinical trials is fully acknowledged, failure to report outcomes or selective reporting of outcomes in non-clinical trial epidemiological studies is less well known, but most likely occurs as well. in this mini-symposium the netherlands epidemiological society wants to give attention to this phenomenon of not publishing research outcomes, to encourage publication of all outcomes of adequate research. different scopes to this subject will be addressed: the background, an example of occurrence, initiatives to possibly avoid it and an editor's point of view. selective reporting of outcomes in clinical studies (reporting bias) has been described to occur frequently. therefore a registration of clinical trials is started which enables to address this problem in the future since occurrence of not publishing negative or adverse outcomes can be investigated with this registration. in non-clinical epidemiological studies the failure to report outcomes or selective reporting of outcomes most likely occurs as well, but is less studied and reported. again studies with negative outcomes or no associations are the ones most likely not to be reported. the most important obstacles for not publishing no or negative associations are tradition and priorities of researchers and journals. the reviewers might play a role in this as well. the netherlands epidemiological society advocates according to good epidemiological practice, that research with sound research questions and good methodology should be adequately published independent of the research outcomes. however, reality occurs not to be accordingly. therefore we would like to give attention to this phenomenon of not publishing research outcomes in non-trial-based epidemiological studies, to encourage publication of all outcomes of adequate research. in this mini-symposium, firstly the effects of failure or selective publishing of outcomes on subsequent meta-analysis in a non-clinical research setting will be demonstrated. afterwards, initiatives to promote and improve publication of observational epidemiological research will be addressed, the editor's point of view on this phenomenon will be given and finally concluding remarks will be given. background: there are several reasons for suspecting reporting bias in time-series studies of air pollution. such bias could lead to false conclusions concerning causal associations or inflate estimates of health impact. objectives: to examine time-series results for evidence of publication and lag selection bias. design and methods: all published time-series studies were identified and relevant data extracted into a relational database. effect estimates were adjusted to an increment of lg/m . publication bias was investigated using funnel plots and two statistical methods (begg, egger). adjusted summary estimates were calculated using the ''trim and fill'' method. the effect of lag selection was investigated using data on mortality from us cities and from a european multi-centre panel study of children. results: there was evidence of publication bias in a number of pollutant-outcome analyses. adjustment reduced the summary estimates by up to %. selection of the most significant lag increased estimates by over % compared with a fixed lag. conclusion and discussion: publication and lag selection bias occurs in these studies but significant associations remain. presentation and publication of time-series results should be standardised. background: selective non-publication of study outcomes hampers the critical appraisal and appropriate interpretation of available evidence. its existence could be shown empirically in clinical trials. observational research often uses an exploratory approach rather than testing specific hypotheses. results of multiple data analyses may be selected based on their direction and significance. objectives: to improve the quality of reporting of observational studies. to help avoid selective non-publication of study outcomes. methods: ''strengthening the reporting of observational studies in epidemiology (strobe)'' is an international multidisciplinary initiative that currently develops a checklist of items recommended for the reporting of observational studies (http:// www.strobe-statement.org). results: strobe recommends to avoid selective reporting of 'positive' or 'significant' study results and to base the interpretation on main results rather than on results of secondary analyses. discussion: strobe cannot prevent data dredging, but it promotes transparency at the publication stage. for instance, if multiple statistical analyses were performed in a large dataset to identify new exposure-outcome associations, authors should give details and not only report significant associations. strobe could have a ''feedback effect'' on study quality since, ideally, researchers think ahead when a study is planned and consider points that are essential for later publication. good publishing practice begins with researchers considering ( ) whether an intended study can bring added value, irrespective its result, ( ) and whether its methodology is valid to pick up positive and negative outcomes equally well. when reporting ( ) they should adequately discuss the significance of a negative result ( ) and be as eager to publish negative results as positive ones. as to editors, intentional bias in relation to study results is considered editorial malpractice, whatever its motivation. unintentional bias may be more frequent but will not easily be noticed, also by editors. editorial responsibility implies several levels (accepting for review, choice of reviewers, assess their reviews, decision making, and a repeated process in case of resubmission). various designs for process evaluation can be considered. evaluation will be more difficult for journals with few professional support. collaboration between journals can help, and may also avoid 'self evaluation bias'. in line with registering of randomized trials, registers for observational study protocols could facilitate monitoring for bias and searching unpublished results. but practicalities, methodological requirements, and bureaucratic burden should not be underestimated. in principle, in an era of electronic publishing every study can be made widely accessible widely also if not 'accepted', by editors or authors themselves. however, this would need huge changes in culture of authoring and reading, editorial practice, publishing business, and scientific openness. background: high circulating levels of insulin-like growth factor-i (igf-i), a mitogenic and anti-apoptotic peptide, have been associated with increased risk of several cancer types. objective: to study circulating levels of igf-i and igf binding protein- (igfbp- ) in relation to ovarian cancer risk. design and methods: within the european prospective investigation into cancer and nutrition (epic), we compared levels of igf-i and igfbp- measured in blood samples collected at baseline in women who subsequently developed ovarian cancer ( women diagnosed before age ) and controls. results: the risk of developing ovarian cancer before age ('premenopausal' was increased among women in the middle or top tertiles of igf-i, compared to the lowest tertile: or = . [ % ci: . - . ], and or = . [ % ci: . - . ], respectively (p trend = . ). results were adjusted for bmi, previous hormone use, fertility problems and parity. adjustment for igfbp- levels slightly attenuated relative risks. in older women we observed no association between igf-i, igfbp- and ovarian cancer risk. discussion and conclusion: in agreement with the only other prospective study in this field (lukanova et al, int j cancer, ) , our results indicate that high circulating igf-i levels may increase the risk of premenopausal ovarian cancer. background: the proportion of glandular and stromal tissue in the breast (percent breast density) is a strong breast cancer risk factor. insulin-like growth factor (igf- ) is hypothesized to influence breast cancer risk by increasing breast density. objectives: we studied the relation between premenopausal circulating igf- levels and changes in breast density over menopause. design and methods: mammograms and blood samples of premenopausal participants of the prospect-epic cohort were collected at baseline. a second mammogram was collected after these women became postmenopausal. we determined serum igf- levels. mammographic density was assessed using a computer-assisted method. changes in percent density over menopause were calculated for quartiles of igf- , using linear regression, adjusted for age and bmi. results: premenopausal percent density was not associated with igf- levels (mean percent density . in all quartiles). however, women in the highest igf- quartile showed less decrease in percent density over menopause ( st quartile: ) . vs th quartile: ) . , p-trend = . ). this was mostly explained by a stronger decrease of total breast size in women with high igf- levels. conclusion and discussion: women with high igf- levels show a lower decrease of percent density over menopause than those with low igf- levels. background: body mass index (bmi) has been found to be associated with risk of colon cancer in men, whereas weaker associations have been reported for women. reasons for this discrepancy are unclear but may be related to fat distribution or use of hormone replacement therapy (hrt) in women. objective: to examine the association between anthropometry and risk of colon cancer in men and women. design and methods: during . years of followup, we identified cases of colon cancer among , subjects free of cancer at baseline from european countries. results: bmi was significantly related to colon cancer risk in men (rr per kg/ m , . ; %-ci . - . ) but not in women (rr . ; . - . ; p interaction = . ), whereas waist-hip-ratio (whr) was equally strong related to risk in both genders (rr per . , men, . ; %-ci . - . ; women, . ; . - . ; p interaction = . ). the positive association for whr was not apparent among postmenopausal women who used hrt. conclusions: abdominal obesity is an equally strong risk factor for colon cancer in both sexes and whr is a disease predictor superior to bmi in women. the association may vary depending on hrt use in postmenopausal women; however, these findings require confirmation in future studies. background: fruits and vegetables are thought to protect against colorectal cancer. recent cohort studies, however, have not been able to show a protective effect. patients & methods: the relationship between consumption of vegetables and fruit and the incidence of colorectal cancer within epic was examined among , subjects of whom developed colorectal cancer. a multivariate cox proportional hazard model was used to determine adjusted cancer risk estimates. a calibration method based on standardized -hour dietary recalls was used to correct for measurement errors. results: after adjustment for potential confounding and exclusion of the first two years of follow-up, the results suggest that consumption of vegetables and fruits is weakly, inversely associated with risk of colorectal cancer (hr . , . , . , . , . , for quintiles of intake, % ci upper quintile . - . , p-trend . ), with each gram daily increase in vegetables and fruit associated with a statistically borderline significant % reduction in colorectal cancer risk (hr . ; . - . ). linear calibration strengthened this effect. further subgroup analyses will be presented. conclusion: findings within epic support the hypothesis that increased consumption of fruits and vegetables may protect against colorectal cancer risk. a diverse consumption of vegetables and fruit may influence the risk of gastric and oesophageal cancer. diet diversity scores (dds) were calculated within the epic cohort data from > , subjects in european countries. four scores, counting the number of ffq-based food-items usually eaten at least once in two weeks, were calculated to represent the diversity in the overall vegetable and/or fruit consumption. after an average follow-up of . years, incident cases of gastric and oesophageal cancer were observed. cox proportional hazard models were used to compute tertile specific risks, stratified by follow-up duration, gender and centre and adjusted for total consumption of vegetables and fruit and potential confounders.preliminary findings suggest that, compared to individuals who eat from only or less vegetable sub-groups, individuals who usually eat from eight different subgroups, have a reduced gastric cancer risk (hr . ; % ci . - . ). in comparison to all others, individuals who usually eat only the same fruit may experience an elevated risk (hr . ; % ci . - . ). these findings from the epic study suggest that a diverse consumption of vegetables may reduce gastric and oesophageal cancer risk. subjects with a very low diversity in fruit consumption may experience higher risk. g. steindorf , l. friedenreich , j. linseisen , p. vineis , e. riboli for the epic group german cancer research center, heidelberg, germany alberta cancer board, alberta, canada imperial college london, great-britain background: previous research on physical activity and lung cancer risk, conducted predominantly in males, has yielded inconsistent results. objectives: we examined this relationship among , men and women from the epic-cohort. design and methods: during . years of follow-up we identified men and women with incident primary lung cancer. detailed information on recreational, household and occupational physical activity, smoking habits, and diet was assessed. relative risks (rr) were estimated using cox regression. results: we did not observe an inverse association between occupational, recreational or household physical activity and lung cancer risk either in males or in females. we found a modest reduction in lung cancer risk associated with sports in males and cycling in females. for occupational physical activity, lung cancer risk was increased for unemployed men (rr = . ; % confidence interval . - . ) and men with standing occupations (rr = . ; . - . ) compared with sitting professions. conclusion: our study shows no convincing protective associations of physical activity with lung cancer risk. discussion: it may be speculated that the elevated risks for occupational physical activity could reflect the higher probability that manual workers are exposed to industrial carcinogens compared to workers having sitting/office jobs. purposes: epidemiological research almost always means using data and, increasingly, human tissue as well. the use of these resources is not free but is subject to various regulations, which differ in the european countries on several important aspects. usually these regulations have been determined without involvement of active epidemiological researchers or patient organisations. this workshop will address the issues involved in these regulations in the european context. it will serve the following purposes: -to provide arguments and tools and to exchange best practices for a way out of the regulatory labyrinths especially in cross european research projects; -to provide a platform for epidemiologists and patient groups to discuss their concerns about impediments for epidemiological research with other parties, like data protection authorities. targeted audience: the mini symposium is primarily meant for epidemiologists, but provides an excellent opportunity to meet and discuss with other stakeholders, like from patient groups, data protection authorities, the european commission etc. as well. therefore program allows for extra time for discussion. the other stakeholders will be explicitly invited. a special 'day ticket' is available to satellite symposium epidemiology and the seventh eu research framework over the last few years the seventh eu research framework has been drafted. it is now rapidly moving towards the first calls for proposals. previous eu research programmes and frameworks have been criticised because they are considered to include too few possibilities for epidemiological research and public health research. this satellite-symposium will provide an outline of the research framework and inform researchers about the current state of affairs of the seventh eu research framework. special focus will be on the possibilities for epidemiology and public health research. - . welcome by our host prof. jan willem coebergh, rotterdam, introduction, international and national regulations on the use of data and tissue or research in europe, different approaches to: evert-ben van veen l.l.m. (medlawconsult, the netherlands) -'identifiability' of data -consent for using data and tissue for research the tubafrost code of conduct to exchange data and tissue across europe. . - . data and tissuebanking for research in denmark: a liberal approach the danish approach to use patient data for epidemiological research, cooperation of the danish data protection authority, the danish act of to use anonymous but coded tissue for research based on an opt-out system, first experiences hans storm ph.d. (copenhagen, denmark) . - . estonian data protection act: a disaster for epidemiology the story of the birth of the act, implementing the european data protection directive and of its consequences reveal political and administrative incapability resulting in gradual vanishing of register-based epidemiological research. background: non-invasive assessment of atherosclerosis is important. most of the evidence of coronary calcium has been based on images obtained by electron beam ct (ebct). current data suggest that ebct and multi-slice ct (msct) give comparable results. since msct is more widely available than ebct, information on its reproducibility is relevant. objective: to assess inter-scan reproducibility of msct and to evaluate whether reproducibility is affected by different measurement protocols, slice thickness, cardiovascular risk factors and technical variables. design: cross-sectional study. materials and methods: the study population comprised healthy postmenopausal women. coronary calcium was assessed in these women twice at two separate visits using msct (philips mx idt ). images were made using . and . mm slice thickness. the agatston, volume and mass scores were assessed. reproducibility was determined by mean differences, absolute mean differences and intra-class correlation coefficients (iccc). results: the reproducibility of coronary calcium measurements between scans was excellent with iccc of > . , and small mean and absolute mean differences. reproducibilility was similar for . as for . mm slices, and equal for agatston, volume and mass measurements. conclusion: inter-scan reproducibilility of msct is excellent, irrespective of slice thickness and type of calcium parameter. background: it has been suggested that the incidence of colorectal cancer is associated with socioeconomic status (ses). the major part of this association may be explained by known lifestyle risk factors such as dietary habits. objective: to explore the association between diet and ses measured at area-based level. methods: the data for this analysis were taken from a multi-centre case-control study conducted to investigate the association between some environmental, genetic factors and colorectal cancer incidence. the townsend scores (as deprivation index) were categorized into fifths. a linear regression analysis was used to estimate difference in mean of each continuous variable of diet by deprivation fifth. results: the mean of processed meat consumption in the most deprived area was higher compared to the mean of that in the most affluent areas (mean difference = . , % ci: . , . ). by contrast, the mean of vegetables and fruits consumption in the most deprived areas was lower than that in the affluent areas. conclusion: our findings suggest that lifestyle factors are likely to be related to ses. thus any relation between ses and colorectal cancer may direct us to seek for the role of different life style factors in aetiology of this cancer. background: the reason for the apparent decline in semen quality during the past years is still unexplained. objective: to investigate the effect of exposure to cigarette smoke in utero on the semen quality in the male offspring. design and methods: in this prospective follow-up study, adult sons of mothers, who during pregnancy provided information about smoking and other lifestyle factors, are sampled in six strata according to prenatal tobacco smoke exposure. each man provides a semen sample, a blood sample, and answers a questionnaire, which is collected in a mobile laboratory. external quality assessment of semen analysis is performed twice a year. results: until now, a total of men have been included. the participation rate is %. the percentage of men with decreased sperm concentration (< mill/ml) is %. the unadjusted median ( - % percentile) sperm concentration in the non-exposed group (n = ) is ( - ) mill/ml compared to ( - ) mill/ml among men exposed to > cigarettes per day in fetal life (n = aim: to estimate the prevalence of overweight and obesity, and their effects in physical activity (pa) levels of portuguese children and adolescents aged - years. methods: the sample comprises subjects ( females- males) attending basic/secondary schools. the prevalence of overweight and obesity was calculated using body mass index (bmi), and the cut-off points suggested by cole et al. ( ) . pa was assessed with the baecke et al. ( ) questionnaire. proportions were compared using chi-square tests and means by anova. results and conclusions: overall, . % were overweight (females = . %; males = . %) and . % were obese (females = . %; males = . %). prevalence was similar across age and gender. bmi changed with age (p< . ), and a significant interaction between age and gender was found (p = . ): whereas bmi in males increased with aging, in females increased up to years and stabilized onwards. males showed significantly higher values of pa (p< . ). both genders had a tendency to increase their pa until - years. a significant interaction between age and gender (p = . ) points out different gender patterns across age: pa increased with aging in males but in females started to decline after years. no significant differences in pa were found between normal weight, overweight and obese subjects (p = . ). background: atherosclerosis is an inflammatory process. however, the relation between inflammatory markers and extent and progression of atherosclerosis remains unclear. objectives: we studied the association between c-reactive protein (crp) and established measures of atherosclerosis. design and methods: within the rotterdam study, a population-based cohort of , persons over age , we measured crp, carotid plaque and intima-media thickness (imt), abdominal artery calcification, ankle-brachial index (abi) and coronary calcification. using ancova, we investigated the relation between crp and extent of atherosclerosis. we studied the association between progression of extra coronary atherosclerosis (mean follow-up period: . years) and crp using multinomial regression analysis. results: crp levels were positively related to all measures of atherosclerosis, but the relation was weaker for measures based on detection of calcification only. crp levels were associated with severe progression of carotid plaque (multivariable adjusted odds ratio: . , % ci: . - . ), imt ( . , . - . ) and abi ( . , . - . ). no relation was observed with progression of abdominal artery calcification. conclusion and discussion: crp is related to extent and progression of atherosclerosis. the relation seems weaker for measures based on detection of calcification only, indicating that calcification of plaques might attenuate the inflammatory process. background: maternal stress during pregnancy has been reported to have an adverse influence on fetal growth. the terrorist attacks of september , on the united states have provoked feelings of insecurity and stress worldwide. objective: our aim was to test the hypothesis that maternal exposure to these acts of terrorism via the media had an unfavourable influence on mean birth weight in the netherlands. design and methods: in a prospective cohort study, we compared birth weights of dutch neonates who were in utero during the attacks with those of neonates who were in utero exactly year later. results: in the exposed group, birth weight was lower than in the non-exposed group (difference, g, %ci . , . , p = . ). the difference in birth weight could not be explained by tobacco use, maternal age, parity or other potential confounders, nor by shorter pregnancy durations. conclusion: these results provide evidence supporting the hypothesis that exposure of dutch pregnant women to the september events via the media has had an adverse effect on the birth weight of their offspring. objective: asian studies suggested potential reduction in the risk of pneumonia among patients with stroke on ace-inhibitor therapy. because of the high risk of pneumonia in patients with diabetes we aimed to assess the effects of ace-inhibitors on the occurrence of pneumonia in a general, ambulatory population of diabetic patients. methods: a case-control study was performed nested in , patients with diabetes. cases were defined as patients with a first diagnosis of pneumonia. for each case, up to controls were matched by age, gender, practice, and index date. current ace-inhibitor use was defined within a time-window encompassing the index date. results: ace-inhibitors were used in . % of , cases and in , % of , matched controls (crude or: . , % ci . to . ). after adjusting for potential confounders, ace-inhibitor therapy was associated with a reduction in pneumonia risk (adjusted or: . , % ci . to . ). the association was consistent among different relevant subgroups (stroke, heart failure, and pulmonary diseases) and showed a strong dose-effect relationship (p< . ). conclusions: use of ace-inhibitors was significantly associated with reduced pneumonia risk and may apart from blood pressure lowering properties be useful in prevention of respiratory infections in patients with diabetes. background: progressive decline in serum levels of testosterone occurs with normal aging in both men and women. this is paralleled by a decrease in physical performance and muscle strength, which may lead to disability, institutionalization and mortality. objective. we examined whether low levels of testosterone were associated with three-year decline in physical performance and muscle strength in two population-based samples of older men and women. methods: data were available for men in the longitudinal aging study amsterdam (lasa) and men and women in the health, aging, and body composition (health abc) study. levels of total testosterone and free testosterone were determined at baseline. physical performance and grip strength were measured at baseline and after three years. results: total and free testosterone were not associated with change in physical performance or muscle strength in men. in women, low levels of total testosterone (<= ng/dl) increased the risk of decline in physical performance (p = . ), and low levels of free testosterone (< pg/ ml) of decline in muscle strength (p = . ). conclusion: low levels of total and free testosterone were associated with decline in physical performance and muscle strength in older women, but not in older men. background: obesity and physical inactivity are key determinants of insulin resistance, and chronic hyperinsulinemia may mediate their effects on endometrial cancer (ec) risk. aim: to examine the relationships between prediagnostic serum concentrations of cpeptide, igf binding protein (igfbp)- and igfbp- , and ec risk. methods: we conducted a case-control study nested within the epic prospective cohort study, including incident cases of ec, in pre-and post-menopausal women, and matched control subjects. odds ratios (or) and % confidence intervals (ci) were calculated using conditional logistic regression models. results: in fasting women (> h since last meal), serum levels of c-peptide, igfbp- and igfbp- were not related to risk. however, in nonfasting women ( h or less since last meal), ec risk increased with increasing serum levels of c-peptide ( background: tobacco is the single most preventable cause of death in the world today. tobacco use primarily begins in early adolescent. objective: to estimate the prevalence and evaluate factors associated with smoking among high school going adolescents in karachi, pakistan. methods: a school based cross sectional survey was conducted in three towns of karachi from january through may . two-stage cluster sampling stratified on school types was employed to select schools and students. self-reported smoking status of school going adolescents was our main outcome in analysis. results: prevalence of smoking ( days) among adolescents was . %. multiple logistic regression model showed that after adjustment for age, ethnicity and place of residence, being student of a government school (or= . ; % ci: . - . ), parental smoking (or = . ; % ci: . - . ), uncle (or = . ; % ci: . - . ) , peer smoking (or = . ; % ci: . - . ) and spending leisure time outside home (or = . ; % ci . - . ) were significantly associated with adolescents smoking. conclusion: a . % prevalence of smoking among school going adolescents and influence of parents and peers in initiating smoking in this age group warrant the need for effective tobacco control in the country especially among the adolescents. background: individual patient data meta-analyses (ipd-ma) have been proposed to improve subgroup analyses that may provide clinically relevant information. nevertheles, comparison of the effect estimates of ipd-ma and meta-analyses of published data (map) are lacking. objective: to compare main and subgroup effect estimates of ipd-ma and map. methods: an extended literature search was performed to identify all ipd-ma of randomized controlled trials, followed by a related article search to identify maps with a similar domain, objective, and outcome. data were extracted regarding number of trials, number of subgroups, effect measure, effect estimate and their confidence intervals. results: in total ipd-ma and map could be included in the analysis. twentyfive main effect estimates could be compared; of which were in the same direction. although over subgroups were studied in both ipd-ma and map, only effect estimates could be compared; were in the same direction. subgroup analyses in map most often related to trial characteristics, whereas subgroup analyses in ipd-ma were related to patient characteristics. conclusion: comparable ipd-ma and map report similar main and subgroup effect estimates. however, ipd-ma more often study subgroups based on patient characteristics, and thus provide more clinically relevant information. patients with diabetes have an increased risk of a complicated course of community-acquired lower respiratory tract infections. although influenza vaccination is recommended for these persons, vaccination levels remain too low because of conflicting evidence regarding potential benefits. as part of the prisma nested casecontrol study among , persons recommended for vaccination, we studied the effectiveness of single and repeat influenza vaccination in the subgroup of adult diabetic population ( , ) during the - influenza a epidemic. case patients were hospitalized for diabetes, acute respiratory or cardiovascular events, or died and controls were sampled from the baseline cohort. after control for age, gender, health insurance, prior health care, medication use and co-morbid conditions logistic regression analysis showed that the occurrence of any complication ( hospitalizations, deaths) was reduced by % ( % confidence interval % to %). vaccine effectiveness was similar for those who received the vaccine for the first time and for those who received an earlier influenza vaccination. although we did not perform virological analysis or distinguish type i from type ii diabetes we conclude that patients with diabetes benefit substantially from influenza vaccination independent of whether they received the vaccine for the first time or received earlier influenza vaccinations. background: construction workers are at risk of developing silicosis. regular medical evaluations to detect silicosis preferably in the pre-clinical phase are needed. objectives: to identify the presence or absence of silicosis by developing an easy to use diagnostic model for pneumoconiosis from simple questionnaires and spirometry. design and methods: multiple logistic regression analysis was done in dutch construction workers, using chest x-ray indicative for pneumoconiosis (ilo profusion category > / ) as the reference standard (prevalence . %). model calibration was assessed with graph and the hoshmer-lemeshow goodness of fit test; discriminative ability using area under receiver operating characteristic curve (auc); and internal validity using bootstrapping procedure. results: age > years, current smoking, high exposure job title, working > years in the construction industry, 'feeling unhealthy', and standardized residual fev below ) . were selected as predictors. the diagnostic model showed a good calibration (p = . ) and discriminative ability (auc . ; % ci . to . ). internal validity was reasonable (correction factor of . and optimism corrected auc of . ). conclusions: and discussion: our diagnostic model for silicosis showed reasonable performance and internal validity. to apply the model with confidence, external validation before application in a new working population is recommended. background: artemisinin based combination therapy (act) reduces microscopic gametocytaemia, the malaria parasite stage responsible for transmission from man to mosquito. as a result, act is expected to reduce the burden of disease in african populations. however, molecular techniques recently revealed high prevalences of gametocytaemia below the microscopic threshold. our objective was to determine the importance of sub-microscopic gametocytaemia after act treatment. methods: kenyan children (n= ) aged months - years were randomised to four treatment regimens. gametocytaemia was determined by microscopy and pfs real-time nucleic acid sequence-based amplification (qt-nasba). transmission was determined by membrane feedings. findings: gametocyte prevalence at enrolment was . % ( / ) as determined by pfs qt-nasba and decreased after treatment with act. membrane feedings in randomly selected children revealed that the proportion of infectious children was up to fourfold higher than expected when based on microscopy. act did not significantly reduce the proportion of infectious children but merely the proportion of infected mosquitoes. interpretation: sub-microscopic gametocyte densities are common after treatment and contribute considerably to mosquito infection. our novel approach indicates that the effect of act on malaria transmission is much smaller than previously suggested. these findings are sobering for future interventions aiming to reduce malaria transmission. background: adequate folate intake may be important in the prevention of breast cancer. factors linked to folate metabolism may be relevant to its protective role. objectives: to investigate the association between folate intake and breast cancer risk among postmenopausal women and evaluate the interaction with alcohol and vitamin b intake. methods: a prospective cohort analysis of folate intake among , postmenopausal women from the e n french cohort who completed a validated food frequency questionnaire in was conducted. during years follow-up , cases of pathology-confirmed breast cancer were documented through followup questionnaires. nutrient intakes were categorized in quintiles and energy-adjusted using the regression-residual method. cox modelderived relative risks (rr) were adjusted for known risk factors for breast cancer. results: the multivariate rr comparing the extreme quintiles of folate intake was . ( % ci . - . ; p-trend= . ). after stratification, the association was observed only among women whose alcohol consumption was above the median (= . g/day) and among women who consumed = . lg/day of vitamin b . however, tests for interaction were not significant. conclusions: in this population, high intakes of folate were associated with decreased breast cancer risk; alcohol and vitamin b intake may modify the observed inverse association. background: the simultaneous rise in the prevalence of obesity and atopy in children has prompted suggestions that obesity might be a causal factor in the inception of atopic diseases. objective: we investigated the possible role of ponderal index (kg/m ) as marker for fatness at birth in early childhood atopic dermatitis (ad) in a prospective birth cohort study. methods: between november and november , mothers and their newborns were recruited after delivery at the university of ulm, germany. active follow-up was performed at the age of months. results: for ( %) of the children included at baseline, information on physician reported diagnosis of ad was obtained during follow-up. incidence of ad was . % at the age of one year. mean ponderal index at birth was . kg/m . risk for ad was higher among children with high ponderal index at birth (adjusted or for children within the third and fourth compared to children within the second quartile of ponderal index: . ; % ci . respectively) background: the relationship between duration of breastfeeding and risk of childhood overweight remains inconclusive, possibly in part caused by using never breastfeeding mothers as the reference category. objectives: we assessed the association between duration of breastfeeding and childhood overweight among ever breastfed children within a prospective birth cohort study. methods: between november and november all mothers and their newborns were recruited after delivery at the university of ulm, germany. active follow-up was performed at age months. results: among children ( % of children included at baseline) with available body mass index at age two ( . %) were overweight. whereas children ( . %) were never breastfed, ( . %) were breastfed for at least six months, and ( . %) were exclusively breastfed for at least six months. compared to children who were exclusively breastfed less than three months, the adjusted or for overweight was . ( % ci . ; . ) in children who were exclusively breastfed for at least three but less than six months and . ( % ci . ; . ) in children who were exclusively breastfed for at least six months. conclusion: these results highlight the importance of prolonged breastfeeding in the prevention of overweight in children. background: in africa, hiv and feeding practices influence child mortality. exclusive breastfeeding for months (bf ) and formula feeding (ff) when affordable are two who recommendations for safe feeding. objective: we estimated the proportion and the number of children saved with each recommendation at population level. design and methods: data on sub-saharan countries were analysed. we considered saved a child remaining hiv-free and alive after two years of life. a spreadsheet model based on a decision tree for risk assessment was used to calculate this number according to six scenarios that combine the two recommendations without and with promotion then with promotion and group education. results: whatever the country, the number of children saved with bf would be higher than with ff. overall, without promotion, ( background: farming has been associated with respiratory symptoms as well as protection against atopy. effects of different farming practices on respiratory health in adults have rarely been studied. objectives: we studied associations between farming practices and hay fever and current asthma in organic and conventional farmers. design and methods this cross-sectional study evaluated questionnaire data of conventional and organic farmers. associations between health effects and farm exposures were assessed by logistic regression. results: organic farmers reported slightly more hay fever than conventional farmers ( . % versus . %, p = . ). however, organic farming was no independent determinant for hay fever in multivariate models including farming practices and potential confounders. livestock farmers who grew up on a farm had a five-fold lower prevalence of hay fever than crop farmers without farm childhood (or . , % ci . - . ). use of disinfectants containing quaternary ammonium compounds was positively related to hay fever (or . , % ci . - . ). no effects of farming practices were found for asthma. conclusion and discussion: our study adds to the evidence that a farm childhood in combination with current livestock farming protects against allergic disorders. this effect was found for both organic and conventional farmers. background: although a body mass index (bmi) above kg/m is clearly associated with an increase in mortality in the general population, the meaning of high levels of bmi among physically heavily working men is less clear. methods: we assessed the association between bmi and mortality in a cohort of male construction workers, aged - years, who underwent an occupational health examination in wu¨rttemberg (germany) during - and who were followed over a years period. covariates considered in the proportional hazard regression analysis included age, nationality, smoking status, alcohol consumption, and comorbidity. results: during the follow-up deaths occurred. there was a strong u-shaped association between bmi and all-cause mortality, which was lowest for bmi levels between and kg/m . this pattern persisted after exclusion of the first years of follow-up and control for multiple covariates. compared with men with a bmi < . kg/m , the relative mortality was . ( % confidence interval: , - , ), . ( . - . ) and . ( . - . ) for bmi ranges - . , - . and = . kg/m . conclusion and discussion: bmi levels commonly considered to reflect overweight or moderate obesity in the general population may be associated with reduced mortality in physically heavily working men. background: colonoscopy with removal of polyps may strongly reduce colorectal cancer (crc) incidence and mortality. empirical evidence for optimal schedules for surveillance is limited. objective. to assess risk of proximal and distal crc after colonoscopy with polypectomy. design and methods: history and results of colonoscopies were obtained from cases and controls in a population-based case-control study in germany. risk of proximal and distal crc according to time since colonoscopy was compared to risk of subjects without previous colonoscopy. results: subjects with previous detection and removal of polyps had a much lower risk of crc within four years after colonoscopy (adjusted odds ratio . , % confidence interval . - . ), and a similar risk as those without colonoscopy in the long run. within four years after colonoscopy, risk was particularly low if only single or small adenomas were detected. most cancers occurring after polypectomy were located in the proximal colon, even if polyps were found in the sigma or rectum only. conclusion and discussion: our results support suggestions that surveillance colonoscopy after removal of single and small adenomas may be deferred to five years and that surveillance should include the entire colorectum even if only distal polyps are detected. background: a population-based early detection programme for breast cancer has been in progress in finland since . recently, detailed information about actual screening invitation schemes in - has become available in electronic form, which enables more specific modeling of breast cancer incidence. objectives: to present a methodology for taking into account historical municipality-specific schemes of mass screening when constructing predictions for breast cancer incidence. to provide predictions for numbers of new cancer cases and incidence rates according to alternative future screening policies. methods: observed municipality-specific screening invitation schemes in finland during - were linked together with breast cancer data. the incidence rate during the observation period was analyzed using poisson regression, and this was done separately for localized and nonlocalized cancers. for modeling, the screening programme was divided into seven different phases. alternative screening scenarios for future mass-screening practices in finland were created and an appropriate model for incidence prediction was defined. results and conclusion: expanding the screening programme would increase the incidence of localized breast cancers; the biggest increase would be obtained by expanding from women aged - to - . the impacts of changes in the screening practices on predictions for non-localized cancers would be minor. background: new screening technologies are implemented to routine screening in increasing numbers, with limited evidence on their effectiveness. randomised evaluation of new technologies is encouraged but rarely done. objective: to evaluate in a randomised design whether the effectiveness of an organised cervical screening programme can be improved by means of new technologies. methods: since , - , women have been invited annually to a randomised multi-arm trial ran within the finnish organised cervical screening programme. the invited women are randomly allocated to three study arms of different primary screening tests: conventional cytology, automation-assisted cytology and, since , human papillomavirus (hpv) testing. up to , we have gathered information on , screening visits in the automation-assisted arm and , in the hpv arm, and we have compared the results to conventional screening. results: automation-assistance resulted in a slightly increased detection of precancers, but the efficacy based on interval cancers is not known. results on hpv screening suggest higher detection of precancers and cancers compared to conventional screening. conclusion: evidence of higher effectiveness of new screening technologies is needed, especially when changing the existing screening programmes. the multi-arm trial shows how these technologies can be implemented to routine in a controlled manner. introduction: nodules and goitres are important risk factors for thyroid cancer. as the number of diagnosed cases of thyroid cancer is increasing, the incidence of such risk factors has been assessed in a french cohort of adults. methods: the su.vi.max (supple´mentation en vitamines et mine´raux antioxydants) cohort study included middle-aged adults followed-up during eight years. incident cases of goitres and nodules have been identified retrospectively by scheduled clinical examinations and spontaneous consultations by the participants. cox proportional hazards modeling was used to identify factors associated to thyroid diseases. results: finally, incident cases of nodules and goitres were identified among , subjects free of thyroid diseases at inclusion. after an average follow-up of years, the incidence of goitres and nodules was . % in - year old men, . % in - year old women and . % in - year old women. identified associated factors were age, low urinary thiocyanate level and oral contraceptive use in women, and high urinary thiocyanate level and low urinary iodine level in men. conclusion: estimated incidences are consistent with those observed in other countries. the protective role of urinary thiocyanate in both men and women and, in women, oral contraceptives deserve further investigation. background: various statistical methods for outbreak detection in hospital settings have been proposed in the literature. usually validation of those methods is difficult, because the long time series of data needed for testing the methods are not available. modeling is a tool to overcome that difficulty. objectives: to use model generated data for testing sensitivity and specificity of different outbreak detection methods. methods: we developed a simple stochastic model for a process of importation and transmission of infection in small populations (hospital wards). we applied different statistical outbreak detection methods described in the literature to the generated time series of diagnosis data and calculated and the sensitivity and specificity of different methods. results: we present roc curves for the different methods and show how they depend on the underlying model parameters. we discuss how sensitivity and specificity measures depend on the degree of underdiagnosis, on the ratio of admitted colonised patients to colonisation resulting from transmission in the hospital, and on the frequency of testing patients for colonisation. conclusions: modeling can be a useful tool for evaluating statistical methods of outbreak detection especially in situation where real data is scarce or its quality questionable. associated with higher mammographic density and breast pain, has been increased which has bearing on screening performance. objective: we compared the screening performance for women aged - years with dense and lucent breast patterns in two time periods and studied the possible interaction with use of hrt. methods: data were collected from a dutch regional screening programme for women referred in - (n = ) and - (n = ) . in addition, we sampled controls for both periods that were not referred (n = and n = resp.) and women diagnosed with an interval cancer. mammograms were digitised and computer-assisted methods used to measure mammographic density. among other parameters, sensitivity was calculated to describe screening performance. results: screening performance has improved slightly, but the difference between dense and lucent breast patterns still exists (e.g. sensitivity % vs. %). hrt use has increased; sensitivity was particularly low ( %) in the group of women with dense breast patterns on hrt. discussion: in conclusion, the detrimental effect of breast density and the interaction with hrt on screening performance warrants further research with enlargement of the catchment area, more referred women, interval cancers and controls. background: population based association studies might lead to false-positive results if possibly underlying population structure is not adequately accounted for. to assess the nature of the population structure some kind of cluster analysis has to be carried out. we investigated the use of self-organizing maps (soms) for this purpose. objectives: the two main questions concern identification of an either discrete or an admixed population structure and identification of the number of subpopulations involved in forming the structured population under investigation. design and methods: we simulated data sets with different population models and included varying informative marker and map sizes. sample sizes ranged from to individuals. results: we found that a discrete structure can easily be accessed by soms. a near to perfect assignment of individuals to their population of origin can be obtained. for an admixed population structure though, soms do not lead to reasonable results. here, even the correct number of subpopulations involved can not be identified. conclusion: in conclusion, soms can be an alternative to a model-based cluster analysis if the researcher assumes a discrete structure but should not be applied if an admixed structure is likely. background: little is known about the combined effect of duration of breastfeeding, sucking habits and malocclusion in the primary dentition. objectives: we studied the association of breastfeeding and non-nutritive sucking habits on malocclusion on the primary dentition. design and methods: a cross-sectional study nested in a birth cohort was carried out in pelotas, brazil. a random sample of children aged was examined and their mothers interviewed. the foster and hamilton criteria were used to define anterior open bite (aob) and posterior cross bite (pcb). information regarding breastfeeding and non-nutritive sucking habits was collected from birth to years-old. poisson's regression analysis was used. results: non-nutritive sucking habits between months and years of age (pr . [ . ; . ] ) and digital sucking at years of age (pr . [ . ; . ]) were risk factors for aob. breastfeeding for less than months (pr . [ . ; . ] ) and the regular use of a pacifier between months and years of age (pr . [ . ; . ]) were the risk factors for pcb. for pcb an interaction was identified between lack of breastfeeding and the use of a pacifier. conclusion: lack of breastfeeding and longer non-nutritive sucking habits during early childhood were the main risk factors for malocclusion in primary dentition. background: recent, dramatic coronary heart disease (chd) mortality increases in beijing, can be mostly explained by adverse changes in risk factors, particularly total cholesterol and diabetes. it is important for policy making to predict the impact of future changes in risk factors on chd mortality trends. objective: to assess the potential impact of changes in risk factors on numbers of chd deaths in beijing from to , to provide evidence for future chd strategies. design: the previously validated impact model was used to estimate the chd deaths expected in a) if recent risk factor trends continue or b) if levels of risk factors reduce. results: continuation of current risk factor trends will result in a % increase in chd deaths by , (almost half being attributable to increases in total cholesterol levels). even optimistically assuming a % annual decrease in risk factors, chd deaths would still rise by % because of population ageing. conclusion: a substantial increase in chd deaths in beijing may be expected by . this will reflect worsening risk factors compounded by demographic trends. population ageing in china will play an important role in the future, irrespective of any improvements in risk factor profiles. background: since smoking cessation is more likely during pregnancy than at other times, interventions to maintain quitting postpartum may give the best opportunity for a long-time abstinence. it is still not clear what kind of advice or counseling should be given to help prevent the relapse postpartum. objectives: to identify the factors, which predispose women to smoking relapse after delivery. design and methods: the cohort study was conducted in and in public maternity units in lodz, poland. the study population consisted of pregnant women between - weeks of pregnancy who have quit smoking no later than three months prior to participation in the study. smoking status was verified using saliva cotinine level. women were interviewed twice: during pregnancy and three months after delivery. results: within three months after delivery about half of women relapsed into smoking. the final model identified the following risk factors for smoking relapse: having partner and friends who smoke, quitting smoking in late pregnancy, and negative experiences after quitting smoking such as dissatisfaction with weight, nervousness, irritation, loosing pleasure. conclusion. this study advanced the knowledge of the factors that determine smoking relapse after delivery and provided preliminary data for future interventions. introduction: it remains difficult to predict the effect of an particular antihypertensive drug in an individual patient and pharmacogenetics might optimise this. objective: to investigate whether the association between use of angiotensin converting enzyme (ace)-inhibitors or ß-blockers and the risk of stroke or myocardial infarction (mi) is modified by the t-allele of the angiotensinogen m t polymorphism. methods: data were used from the rotterdam study, a population-based prospective cohort study. in total, subjects with hypertension were included from july st, onwards. follow-up ended at the diagnosis of mi or stroke, death, or the end of study period (january st, ) . the drug-gene interaction and the risk of mi/stroke was determined with a cox proportional hazard model (adjusted for each drug class as time-dependent covariates). results: the interaction between current use of ace-inhibitors and the angiotensinogen m t polymorphism increased the risk of mi (synergy index (si) = . ; % ci: . - . ) and non-significant increased risk of stroke (si = . ; % ci: . - . ). no interaction was found between current use of ß-blockers and the agt m t polymorphism on the risk of mi or stroke. conclusion: subjects with at least one copy of the t allele of the agt gene might have less benefit from ace-inhibitor therapy. [ . - . ] to . [ . - . ] in those without ms-idf and . [ . - . ] with ms-idf. ms-ncep had no effect. conclusion and discussion: although cardiovascular disease was self-reported, we conclude that the higher prevalence of cardiovascular disease is partly accounted for by marked differences in the prevalence of metabolic syndrome. the ms-idf criteria seem better for defining metabolic syndrome in ethnic groups than the ms-ncep criteria. background: selenium is an essential trace mineral with antioxidant properties. objective: to perform meta-analyses of the association of selenium levels with coronary heart disease (chd) endpoints in observational studies and the efficacy of selenium supplements in preventing chd in randomized controlled trials. methods: we searched medline and the cochrane library from through . relative risk (rr) estimates were pooled using an inversevariance weighted random-effects model. for observational studies reporting three or more categories of exposure we conducted a dose-response meta-analysis. results: twenty-five observational studies and clinical trials met our inclusion criteria. the pooled rr comparing the highest to the lowest categories of selenium levels was . ( % confidence interval . - . ) in cohort studies and . ( . - . ) in case-control studies. in dose-response models, a % increase in selenium levels was associated with a % ( - %) reduced risk of coronary events. in randomized trials, the rr comparing participants taking selenium supplements to those taking placebo was . ( . - . ). conclusion: selenium levels were inversely associated with the risk of chd in observational studies. the randomized trials findings are still inconclusive. these results require confirmation in randomised controlled trials. currently, selenium supplements should not be recommended for cardiovascular prevention. background propensity score analysis (psa) can be used to reduce confounding bias in pharmacoepidemiologic studies of the effectiveness and safety of drugs. however, confidence intervals may be falsely precise because psa ignores uncertainty in the estimated propensity scores. objectives: we propose a new statistical analysis technique called bayesian propensity score analysis (bpsa). the method uses bayesian modelling with the propensity score as a latent variable. our question is: does bpsa yield improved interval estimation of treatment effects compared to psa? our objective is: to implement bpsa using computer programs and investigate the performance of bpsa compared to psa. design and methods: we investigated bpsa using monte carlo simulations. synthetic datasets, of sample size n = , , , were simulated by computer. the datasets were analyzed using bpsa and psa and we estimated the coverage probability of % credible intervals. results the estimated coverage probabilities ranged from % to % for bpsa, and from % to % for psa, with simulation standard errors less than %. background: several factors associated with low birth weight, such as smoking and body mass index (bmi) do not explain all ethnic differences. this study investigates the effects of working conditions on birth weight among different ethnic groups. methods: questionnaire data, filled in weeks after prenatal screening, was used from the amsterdam born children and their development (abcd) study (all pregnant women in amsterdam [ / / - / / (n = . ), response ( %)]. ethnicity (country of birth). was dichotomised into dutch and non-dutch. working conditions were: weekly working hours, weekly hours standing/walking, physical load and job-strain (karasek-model). only singleton deliveries with pregnancy duration = weeks were included. results: although only . % of the non-dutch women worked during first trimester ( . % of the dutch women), they reported significantly more physical load ( . % vs . %), more hours standing/walking ( . % vs . %) and more high job-strain ( . vs . ). linear regression revealed that only high job-strain lowered significantly birth weight (non-dutch: gram and dutch: gram). after adjusting for confounders (gender, parity, smoking, maternal length, maternal bmi and education), this was only significant in the non-dutch group ( vs. gram). conclusion: job-strain has more effect on birth weight in non-dutch compared to dutch women. background: in panama population was estimated in . million habitants, from which three millions lived in malaria endemic areas. until january malaria control activities were accomplished under a vertical structure. objective: to evaluate the evolution of malaria control in panama, before and after the decentralization of the malaria program. design and methods: average (standard deviation) of the program indexes are described for the last decades. the correlation between positive smears index and per capita cost of the program is analyze. results: in the 's the average (standard deviation) positive smears index per habitants was . % ( . ); in the 's: . % ( . ); in the 's: . % ( . ); in the 's: . % ( . ); and in the first five years of : . % ( . ). after the decentralization of the program was accomplished in , the positive smears index increased . fold. the average per capita cost involved in malaria control activities per decade ranged between . y . us dollars and presented a determination coefficient of . in the reduction of the positive smears index. discussion: the decentralization had significant detrimental implications in the control program capabilities. background: notification rates of new smear-positive tuberculosis in the central mountainous provinces ( / , population) are considerably lower than in vietnam in general ( / , population). this study assessed whether this is explained by low case detection. objective: to assess the prevalence and case detection of new smear-positive pulmonary tuberculosis among adults with a prolonged cough in central mountainous vietnam. design and methods: a house-to-house survey of adults years or older was carried out in randomly selected districts in three mountainous provinces in central vietnam in . three sputum specimens were microscopically examined of persons reporting a cough of weeks or longer. results: the survey included , persons with a response of %. a cough of weeks or longer was reported by , ( . % % ci . - . ) persons. of these, were sputum smear-positive of whom had had anti-tuberculosis treatment. the prevalence of new smear-positive tuberculosis was / , population ( % ci - / , population). the patient diagnostic rate was . per person-year, suggesting that the case notification rate as defined by who was %. conclusion: low tuberculosis notification rates in mountainous vietnam are probably due to low tuberculosis incidence. explanations for low incidence at high altitude need to be studied. background: although patients with type diabetes (dm ) have an increased risk of urinary tract infections (utis), not much is known about predictors of a complicated course. objective: this study aims to develop a prediction rule for complicated utis in dm patients in primary care. design and methods: we conducted a -month prospective cohort study, including dm patients aged years or older from the second dutch national survey of general practice. the combined outcome measure was defined as the occurrence of recurrent cystitis, or an episode of acute pyelonephritis or prostatitis. results: of the , dm patients % was male and mean age was years (sd ). incidence of the outcome was per patient years (n = ). predictors were age, male sex, number of physician contacts, incontinence of urine, cerebro vascular disease or dementia and renal disease. the area under the receiver-operating curve (auc) was . ( % ci . to . ). subgroup analyses for gender showed no differences. there is an increased early postoperative mortality (operation risk) after elective surgery. this mortality is normally associated with cardiovascular events, such as deep venous thrombosis, pulmonary embolism, and ischemic heart diseases. our objective was to quantify the magnitude of the increased mortality and how long the mortality after an operation persists. we focused on the early postoperative mortality after surgery for total knee and total hip replacements from the national registries in australia and norway, which cover more than % of all operations in the two nations. only osteoarthritis patients between and years of age were included. a total of . patients remained for analyses. smoothed intensity curves were calculated for the early postoperative period. effects of risk factors were studied using a nonparametric proportional hazards model. the mortality was highest immediately after the operation ($ deaths per . patients per day), and it decreased until the rd postoperative week. the mortality was virtually the same for both nations and both joints. mortality increased with age and was higher for males than for females. a possible reduction of early postoperative mortality is plausible for the immediate postoperative period, and no longer than the rd postoperative week. background/objectives: single, modifiable risk factors for stroke have been extensively studied before, but their combined effects were rarely investigated. aim of the present study was to assess single and joint effects of risk factors on stroke and transitoric ischemic attack (tia) incidence in the european prospective investigation into cancer and nutrition (epic)-potsdam study. methods: among participants aged - years at baseline total stroke cases and tia cases occurred during . years of follow-up. relative risks (rr) for stroke and tia related to risk factors were estimated using cox proportional hazard models. results: after adjustment for potential confounders rr for ischemic stroke associated with hypertension was . ( % ci, . - . ) and for tia . ( % ci . - . ). the highest rr for ischemic stroke (rr . , % ci . - . , p trend< . ) and tia (rr . , % ci . - . , p trend= . ) were observed among participants with or modifiable risk factors. . % of ischemic strokes and . % of tia cases were attributable to hypertension, diabetes mellitus, high alcohol consumption, hyperlipidemia, and smoking. conclusion: almost % of ischemic stroke cases could be explained by classical modifiable risk factors. however, only one in four tia cases was attributable to those risk factors. background: the investigation of genetic factors is gaining importance in epidemi-ology. most relevant from a public health perspective are complex diseases that are characterised by complex pathways involving gene-gene-and gene-environment-interactions. the identification of such pathways requires sophisticated statistical methods that are still in their infancy. due to their ability in describing complex association structures, directed graphs may represent a suitable means for modelling complex causal pathways. objectives: we present a study plan to investigate the appropriateness for using directed graphs for modelling complex pathways in association stud-ies. design and methods: graphical models and artificial neural networks will be investigated using simulation studies and real data and their advantages and disadvantages of the respective ap-proaches summed up. furthermore, it is planned to construct a hybrid model exploiting the strengths of either model type. results and conclusions: the part of the project that concerns graphical models is being funded and ongoing. first results of a simulation study have been obtained and will be presented and discussed. a second project is currently being applied for. this shall cover the investigation of neural networks and the construction of the hybrid model. this study investigates variations in mortality from 'avoidable' causes among migrants in the netherlands in comparison with the native dutch population. data were obtained from population and mortality registries in the period - . we compared mortality rates for selected 'avoidable' conditions for turkish, moroccan, surinamese and antillean/aruban groups to native dutch. we found slightly elevated risk in total 'avoidable' mortality for migrant populations (rr = . ). higher risks of death among migrants were observed from almost all infectious diseases (most rr> . ) and several chronic conditions including asthma, diabetes and cerebro-vascular disorders (most rr> . ). migrant women experienced a higher risk of death from maternity-related conditions (rr = . ). surinamese and antillean/ aruban population had a higher mortality risk (rr = . and . respectively), while turkish and moroccans experienced a lower risk of death (rr = . and . respectively) from all 'avoidable' conditions compared to native dutch. control for demographic and socioeconomic factors explained a substantial part of ethnic differences in 'avoidable' mortality. conclusion: compared to native dutch, total 'avoidable' mortality was slightly elevated for all migrants combined. mortality risks varied greatly by cause of death and ethnicity. the substantial differences in mortality for a few 'avoidable' conditions suggest opportunities for improvement within specific areas of the healthcare system. warmblood horses scored by the jury as having uneven feet will never pass yearly selection sales of the royal dutch warmblood studbook (kwpn).to evaluate whether the undesired trait 'uneven feet' influences performance, databases of kwpn (n = horses) and knhs (n = show jumpers, n = dressage horses) were linked through the unique number of each registered horse. using a proc glm model of sas was investigated whether uneven feet had effects on age at first start and highest performance level. elite show jumpers with uneven feet start at . years and dressage horses . years of age, which is a significant difference (p< . ) with elite even feet horses ( . respectively . years). at their maximum level of performance horses with even feet linearly scored in show jumping . at regular and . at elite level ( . resp. . with uneven feet), while in dressage horses scores were . at regular and . at elite level ( . resp. . with uneven feet).the conformational trait 'uneven feet' appears to have a significant effect on age at first start, while horses with even feet demonstrate a higher maximal performance than horses with uneven feet. objectives: to identify children with acute otitis media (aom) who might benefit more from treatment with antibiotics. methods: an individual patient data meta-analysis (ipdma) on six randomized trials (n = children). to preclude multiple testing, we first performed a prognostic study in which predictors of poor outcome were identified. subsequently, interactions between these predictors and treatment were studied by fixed effect logistic regression analyses. only if a significant interaction term was found, stratified analyses were performed to quantify the effect in each subgroup. results: interactions were found for: age and bilateral aom, and otorrhea. in children less than years with bilateral aom, a rate difference (rd) of ) % ( % ci ) ; ) %) was found, whereas in children aged years or older with unilateral aom the rd was ) % ( % ci ) ; ) %). in children with and without otorrhea the rd were ) % ( % ci ) ; ) %), and ) % ( % ci ) %; ) %). conclusion: although there still are many areas in which ipdma can be improved, using individual patient data appear to have many advantages especially in identifying subgroups. in our example, antibiotics are beneficial in children aged less than years with bilateral aom, and in children with otorrhea. major injuries, such as fractures, are known to increase the risk of venous thrombosis (vt). however, little is known of the risk caused by minor injuries, such as ankle sprains. we studied the risk of vt after minor injury in a population-based case-control study of risk factors for vt, the mega-study. consecutive patients, enrolled via anticoagulation clinics, and control subjects, consisting both of partners of patients and randomly selected control subjects, were asked to participate and filled in a questionnaire. participants with cancer, recent plastercasts, surgery or bedrest were excluded from the current analyses. out of patients ( . %) and out of controls ( . %) had suffered from a minor injury resulting in a three-fold increased risk of vt (odds ratio adjusted for age and sex . ; % confidence interval . - . ) compared to those without injury. the risk was highest in the first month after injury and was no longer increased after months. injuries located in the leg increased the risk five-fold, while those located in other body parts did not increase the risk. these results show that minor injuries in the leg increase the risk of vt. this effect appears to be temporary and mainly local. introduction: in southeast asia, dengue was considered a childhood disease. in the americas, this disease occurs predominantly in older age groups, indicating the need for studies to investigate the immune status of the child population, since the presence of antibodies against a serotype of this virus is a risk factor for dengue hemorrhagic fever (dhf). objective: to evaluate the seroprevalence and seroincidence of dengue in children living in salvador, bahia, brazil. methods: a prospective study was carried out in a sample of children of - years by performing sequential serological surveys (igg/ dengue). results: seroprevalence in children was . %. a second survey (n = seronegative children) detected an incidence of . % and no difference was found between males and females or according to factors socioeconomic analyzed. conclusion and discussion: these results show that, in brazil, the dengue virus circulates actively in the initial years of life, indicating that children are also at great risk of developing dhf. it is possible that in this age group, dengue infections are mistaken for other febrile conditions, and that there are more inapparent infections in this age group. therefore, epidemiological surveillance and medical care services should be aware of the risk of dhf in children. since , in the comprehensive cancer centre limburg (cccl) region, all women aged - years are invited to participate in the cervical cancer screening programme once every five years. we had the unique opportunity to link data from the cervical screening programme and the cancer registry. we studied individual pap smear testing and participation in the screening programme preceding the diagnosis of cervical cancer. all invasive cases of cervical cancer of women aged - years in the period - were selected. subgroups were based on results of the pap smear and invitation and participation in the screening programme. time interval between screening and detection of tumours was calculated. in - , the non-response rate was %. in total, invasive cervical cancer cases were detected of which were screening and interval carcinomas. in the group of women who were invited but did not participate and women who were not invited, respectively and tumours were detected. these tumours had a higher stage compared to screening carcinomas. in the cccl region, more and higher stage tumours were found in women who did not participate in the screening compared to women with screening tumours. background: pcr for mycobacterium tuberculosis (mtb) has already proved to be a useful tool for the diagnosis and investigation of molecular epidemiology. objectives: evaluation of pcr assay for detection of mycobacterium tuberculosis dna as a diagnostic aid in cutaneous tuberculosis. design and methods: thirty paraffinembedded samples belonging to patients were analyzed for acid fast bacilli. dna was extracted from tissue sections and pcr was performed using specific primers based on is repeated gene sequence of mtb. results: two of the tissue samples were positive for acid fast bacilli (afb). pcr was positive in eight samples from six patients. amongst them, two were suspected of having lupus vulgaris confirmed histopathologically, whom their entire tests were positive. accounting histopathology as gold standard, the sensitivity of pcr in this study was determined as %. conclusion: from cases of skin tuberculosis diagnosed by histopathology, were positive by pcr technique, which shows the priority of previous method to molecular technique. discussion: pcr assay can be used for rapid detection of mtb from cutaneous tuberculosis cases, particularly when staining for afb is negative and there is a lack of growth on culture or when fresh material has not been collected for culture. background: recent epidemiological studies used automated oscillometric blood pressure (aod) devices that systematically measure higher blood pressure values than random zero sphygmomanometer devices (rzs) hampering the comparability of the blood pressure values between these studies. we applied both a random zero and an automated oscillometric blood pressure device in a randomized order in an ongoing cohort study. objectives: the aim of this analysis was to compare the blood pressure values by device and to develop a conversion algorithm for the estimation of blood pressure values from one device to the other. methods: within a randomized subset of subjects aged - years, each subject was measured three times by each device (hawskley random zero and omron hem- cp) in a randomized order. results: the mean difference (aod-rzs) between the devices was . mmhg and . mmhg for the systolic and diastolic blood pressure respectively. linear regression models including age, sex, and blood pressure level can be used to convert rzs blood pressure values to aod blood pressure values and vice versa. conclusions: the results may help to better compare blood pressure values of epidemiological studies that used different blood pressure devices. a form was used to collect relevant perinatal clinical data, as part of a european (mosaic) and italian (action) project. the main outcomes were mortality and a variable combining mortality and severe morbidity at discharge. the cox proportional hazards and logistic regression models were used, respectively, for the two outcomes. results: twenty-two of percent of vpbs were among fbms. comparing to control group, the percentage of babies below weeks and plurality was statistically significant higher among babies of fbms: % vs. . and . % vs. . %. adjusting for potential confounders, no association for mortality among immigrant group was found, whereas a slightly excess of morbidity-mortality was observed (odd ratio, . ; % cis . - . ). conclusions: the high proportion of vpbs among fbms and the slight excess observed in morbidity and mortality indicate the need to improve the health care delivery for the immigrant population. background: high-risk newborns have excess mortality, morbidity and use of health services. objectives: to describe re-hospitalizations after discharge from an italian region. design and methods: the population study consisted of all births with < weeks' gestation discharged alive from the twelve neonatal intensive care units in lazio region during . the perinatal clinical data was collected as part of a european project (mosaic). we used the regional hospital discharge database to find hospital admissions within months, using tax code for record linkage. data were analyzed through logistic regression for re-hospitalization. results: the study group included children; among these, ( . %) were re-hospitalized; overall, readmission were observed. the median total length of stay for re-admissions was d. the two most common reasons for re-hospitalization were respiratory ( . %) and gastrointestinal ( . %) disorders. the presence of a severe morbidity at discharge (odd ratio . : % cis . - . ) and male sex (odd ratio . ; % cis . - . ) predicted re-hospitalization in multivariate model. conclusions: almost one out three preterm infants was re-hospitalized in the first months. readmissions after initial hospitalization for a very preterm birth could be a sensitive indicator of quality of follow-up strategies in high risk newborns. background: self-medication with antibiotics may lead to inappropriate use and increase the risk of selection of resistant bacteria. in europe the prevalence varies from / to / respondents. self-medication may be triggered by experience with prescribed antibiotics. we investigated whether in european countries prescribed use was associated with self-medication with antibiotics. methods: a population survey was conducted in european countries with respondents completing the questionnaire. multivariate logistic regression analysis was used to study the relationship between prescribed use and self-medication (both actual and intended) in general, for a specific symptom/disease or a specific antibiotic. results: prescribed use was associated with selfmedication, with stronger effect in northern/western europe (odds ratio . , % ci . - . ) than in southern ( . , . - . ) and eastern europe ( . , . - . ). prescribing of a specific antibiotic increased the probability of self-medication with the same antibiotic. prescribing for a specific symptom/disease increased the likelihood of self-medication for the same symptom/disease. the use of prescribed antibiotics and actual self-medication were both determinants of intended self-medication in general and for specific symptoms/diseases. conclusions: routine prescribing of antibiotics increases the risk of self-medication with antibiotics for similar ailments, both through the use of leftovers and buying antibiotics directly from pharmacies. background: in the american national kidney foundation published a guideline based on opinion and observational studies which recommends tight control of serum calcium, phosphorus and calcium-phosphorus product levels in dialysis patients. objectives: within the context of this guideline, we explored associations of these plasma concentrations with cardiovascular mortality risk in incident dialysis patients. design and methods: in necosad, a prospective multi-centre cohort study in the netherlands, we included consecutive patients new on haemodialysis or peritoneal dialysis between and . risks were estimated using adjusted time-dependent cox regression models. results: mean age was ± years, % was male, and % was treated with haemodialysis. cardiovascular mortality risk was significantly higher in haemodialysis patients (hr: . ; % ci: . to . ) and in peritoneal dialysis patients (hr: . ; . to . ) with elevated plasma phosphorus levels when compared to patients who met the target. in addition, having elevated plasma calcium-phosphorus product concentrations increased cardiovascular mortality risk in haemodialysis (hr: . ; . to . ) and in peritoneal dialysis patients (hr: . ; . to . ). conclusion: application of the current guideline in clinical practice is warranted since it reduces cardiovascular mortality risk in haemodialysis and peritoneal dialysis patients in the netherlands. background: urologists are increasingly confronted with requests for early detection of prostate cancer in men from hereditary prostate cancer (hpc) families. however, little is known about the benefit of early detection among men at increased risk. objectives: we studied the effect of biennial screening with psa in unaffected men from hpc families, aged - years, on surrogate endpoints (test and tumour characteristics). methods: the netherlands foundation for the detection of hereditary tumours holds information on approximately hpc families. here, nonaffected men from these families were included and invited for psa testing every years. we collected data on screening history and complications related to screening. results: in the first round, serum psa was elevated ( ng/ml or greater) in of men screened ( %). further diagnostic assessment revealed patients with prostate cancer ( . %). compared to population-based prostate cancer screening trials, the referral rate is equal but the detection rate is twice as high. discussion: in conclusion, the results of prostate cancer screening trials will not be directly applicable to screening in hpc families. the balance between costs, side-effects and potential benefits of screening when applied to a high-risk population will have to be assessed separately. background: in industrialized countries occupational tuberculosis among health care workers (hcws) is re-emerging as a public health priority. to prevent and control tuberculosis transmission in nosocomial settings, public health agencies have issued specific guidelines. turin, the capital of the piedmont region in italy, is experiencing a worrying rise of tuberculosis incidence. here, hcws are increasingly exposed to the risk of nosocomial tuberculosis transmission. objectives: a) to estimate the sex-and age-adjusted annual rate of tuberculosis infection (arti) (per person-years [%py]) among the hcws, as indicated by tuberculin skin test conversion (tst) conversion, b) to identify occupational factors associated with significant variations in the arti, c) to investigate the efficacy of the regional preventive guidelines. design and methods: multivariate survival analysis on tst conversion data from a dynamic cohort of hcws in turin, between and . results: the overall estimated arti was . ( % ci: . - . ) %py. the risk of tst conversion significantly differed among workplaces, occupations, and age of hcws. the guidelines implementation was associated with an arti reductions of . ( % ci: . - . ) %py. conclusions: we identify occupational risk categories for targeting surveillance and prevention measures and assessed the efficacy of the local guidelines. background: a positive family history (fh) of breast cancer is an established risk factor for the disease. screening for breast cancer in israel is recommended annually for positive-fh women aged = y and biennially for average-risk women aged - y. objective: to assess the effect of having a positive breast cancer fh on performing screening mammography in israeli women. methods: a cross-sectional survey based on a random sample of the israeli population. the study population consists of , women aged - y and telephone interviews were used. logistic regression models identified variables associated with mammography performance. results: a positive fh for breast cancer was reported by ( . %) participants. performing a mammogram in the previous year was reported by . % and . % of the positive and negative fh subgroups, respectively (p< . ). rates increased with age. among positive fh participants, being married was the only significant correlate for a mammogram in the previous year. conclusions: over % and around % of high-risk women aged - y and = y, respectively, are inadequately screened for breast cancer. screening rates are suboptimal in average-risk women too. discussion: national efforts should concentrate on increasing awareness and breast cancer screening rates. to evaluate the association between infertility, infertility treatments and breast cancer risk. methods: a historical prospective cohort with , women who attended israeli infertility centers between and . their medical charts were abstracted. breast cancer incidence was determined through linkage with the national cancer registry. standardized incidence ratios (sirs) and % confidence intervals were computed by comparing observed cancer rates to those expected in the general population. additionally, in order to control for known risk factors, a casecontrol study nested within the cohort was carried out as well based on telephone interviews with breast cancer cases and controls matched by : ratio. results: compared to . expected breast cancer cases, were observed (sir = . ;non-significant). risk for breast cancer was higher for women treated with clomiphene citrate (sir = . ; % ci . - . ). similar results were noted when treated and untreated women were compared, and when multivariate models were applied. in the nested case-control study, higher cycle index and treatment with clomiphene citrate were associated with significantly higher risk for breast cancer. conclusions: clomiphene citrate may be associated with higher breast cancer risk. smoking is a strong risk factor for arterial disease. some consider smoking also as a risk factor for venous thrombosis, while the results of studies investigating the relationship are inconsistent. therefore, we evaluated smoking as a risk factor for venous thrombosis in the multiple environmental and genetic assessment of risk factors for venous thrombosis (mega) study, a large population-based case-control study. consecutive patients with a first venous thrombosis were included from six anticoagulation clinics. using a random-digit-dialing method a control group was recruited in the same geographical area. all participants completed a questionnaire including questions on smoking habits. persons with known malignancies were excluded from the analyses, leading to a total of patients and controls. current and former smoking resulted in a small increased risk of venous thrombosis (ors adjusted for age, sex and bmi) (or-current: . ci : . - . , or-former: . ci : . - . ). an increasing amount and duration of smoking was associated with an increase in risk. the highest risk was found among young (lowest tertile: to yrs) current smokers; twenty or more pack-years resulted in a . -fold increased risk (ci : . - . ). in conclusion, smoking results in a small increased risk of venous thrombosis, with the greatest relative effect among young heavy smokers. objective: to explore whether the observed association between silica exposure and lung cancer was confounded by exposure to other occupational carcinogens, we conducted a nested case-control-study among a cohort of male workers in chinese mines and potteries. methods: lung cancer cases and matched controls were selected. exposure to respirable silica as well as relevant occupational confounders were evaluated quantitatively based on historical industrial hygiene data. the relationship between silica exposure and lung cancer mortality was analyzed by conditional logistic regression analysis adjusted for exposure to arsenic, polycyclic aromatic hydrocarbons (pahs), radon, and smoking habit. results: in a crude analysis adjusted for smoking only, a significant trend of increasing risk of lung cancer with exposure to silica was found for tin, copper/iron miners, and pottery workers. however, after the relevant occupational confounders were adjusted, no association can be observed between silica exposure and lung cancer mortality (pro mg/m -year increase of silica exposure: or = . , % ci: . - . ). conclusion: our results suggest that, the observed excess risk of lung cancer among silica exposed chinese workers is more likely due to exposure to other occupational carcinogens such as arsenic and pahs rather than due to exposure to respirable silica. background: modelling studies have shown that lifestyle interventions for adults with a high risk of developing diabetes are costeffective. objective: to explore the cost-effectiveness of lifestyle interventions for adults with low or moderate risk of developing diabetes. design and methods: the short-term effects of both a community-based lifestyle program for the general population and a lifestyle intervention for obese adults on diabetes risk factors were estimated from international literature. intervention costs were based on dutch projects. the rivm chronic diseases model was used to estimate long-term health effects and disease costs. costeffectiveness was evaluated from a health care perspective with a time horizon of years. results: intervention costs needed to prevent one case of diabetes in years range from , to , euro for the community program and from , to , euro for the intervention for obese adults. cost-effectiveness was , to , euro per quality adjusted life-year for the community program and , to , for the lifestyle intervention. conclusion: a lifestyle intervention for obese adults produces larger individual health benefits then a community program but, on a population level, health gains are more expensively achieved. both lifestyle interventions are cost-effective. background: in barcelona, the proportion of foreign-born patients with tuberculosis (tb) raised from . % in to , % in . objective: to determine differences in infection by country of origin among contacts investigated by the tb programme in barcelona from - . design and methods: data were collected on cases and their contacts. generalized estimating equations were used to obtain the risk of infection (or and % ci) to account for potential correlation among contacts. results: contacts of foreign born cases were more infected than contacts of natives patients ( % vs %, p< . ) factors related to infection among contacts of foreign cases were inner city residency (or: . , % ci: . - . ) and sputum smear positivity of the case (or: . , % ci: . - . ) and male contact (or: . , % ci: . - . ), but not daily contact (or: . , % ci: . - . ) among natives cases, inner city residency (or: . , % ci: . - . ), sputum smear positivity (or: . , % ci: . - . ) and daily exposure (or: . , % ci: . - . ) increased risk of infection. conclusion: contacts immigrant tb cases have a higher risk of infection than contacts of natives cases, however daily exposure to an immigrant case was not associated with a greater risk of infection. this could be explained by the higher prevalence of tb infection in their country of origin. background: an inverse association between birthweight and subsequent coronary heart disease (chd) has been widely reported but has not been formally quantified. we therefore conducted a systematic review of the association between birthweight and chd. design and methods: seventeen studies including a total of , singletons that had reported quantitative or qualitative estimates of the association between birthweight and chd by october were identified. additional data from two unpublished studies of individuals were also included. in total, the analyses included data on non-fatal and fatal coronary heart disease events in , individuals. results: the mean weighted estimate for the association between birthweight and chd incidence was . ( % ci . - . ) per kg of birthweight. overall, there was no significant heterogeneity between studies (p = . ) or evidence of publication bias (begg test p = . ). fifteen studies were able to adjust for some measure of socioeconomic position, but such adjustment did not materially influence the association: . ( % ci . - . ). discussion: these findings are consistent with one kilogram higher birthweight being associated with - % lower risk of subsequent chd, but the causal nature of this association remains uncertain and its direct relevance to public health is likely to be small. objective: diabetes has been reported to be associated with a greater coronary hazard among women compared with men with diabetes. we quantified the coronary risk associated with diabetes by sex by conducting a meta-analysis of prospective cohort studies. methods: studies reporting estimates of the relative risk for fatal coronary heart disease (chd) comparing those with and without diabetes, for both men and women were included. results: studies of type- diabetes and chd among , individuals were identified. the summary relative risk for fatal chd, diabetes versus not, was significantly greater among women than men: . ( % ci . to . ) versus . ( % ci . to . ), p< . . after excluding eight studies that had only adjusted for age, the sex risk difference was substantially reduced, but still highly significant (p = . ). the pooled ratio of the relative risks (female: male) from the multiple-adjusted studies was . ( % ci . to . ). conclusions: the relative risk for fatal chd associated with diabetes is % higher in women than in men. more adverse cardiovascular risk profiles among women with diabetes, combined with possible treatment disparities that favour men, may explain the greater excess coronary risk associated with diabetes in women. background: malaria in sri lanka is strongly seasonal and often of epidemic nature. the incidence has lowered in recent years which increased the relevance of epidemic forecasting for better targeting control resources. objectives: to establish the spatio/temporal correlation of precipitation and malaria incidence for use in forecasting. design and methods: de-trended long term ( de-trended long term ( - monthly time series of malaria incidence at district level were regressed in a poisson regression against rainfall and temperature at several lags. results: in the north and east of sri lanka, malaria seasonality is strongly positively correlated to rainfall seasonality (malaria lagging one or two months behind rainfall). however, in the south west, no significant (negative) correlation was found. also in the hill country, no significant correlation was observed. conclusion and discussion: despite high correlations, it still remains to be explored to what extent rainfall can be a used as a predictor (in time) of malaria. observed correlation could simply be due to two cyclical seasonal patterns running in parallel, without causal relationship. e.g. similarly, strong correlations were found between temperature and malaria seasonality at months time lag in northern districts, but causality is biologically implausible. background: few studies assessed the excess burden of acute respiratory tract infections (rti) among preschool children in primary care during viral seasons. objective: to determine the excess of rti in preschool children in primary care attributable to influenza and respiratory syncytial virus (rsv). methods: we performed a retrospective cohort study including all children aged - years registered in the database of the utrecht general practitioner (gp) network. during during - , gps recorded episodes of acute rti. surveillance data of influenza and rsv were obtained from the weekly sentinel system of the dutch working group on clinical virology. viral seasons and base-line period were defined as the weeks with respectively more than % and less than % of the yearly number of isolates of influenza or rsv. results: on average episodes of rti were recorded per , child years ( % ci: - ). notably more consults for rti occurred during influenza-season (rr . , % ci: . - . ) and rsv-season (rr . , % ci: . - . ) as compared to base-line period, especially in children younger than two years of age. conclusion: substantial excess rates of rti were demonstrated among preschool children in primary care during influenza-season and particularly during rsvseason, notably in the younger age group. background: many cancer patients who have already survived some time want to know about their prognosis, given the precondition that they are still alive. objective: we described and interpreted population-based conditional -year relative survival rates for cancer patients. methods: the longstanding eindhoven cancer registry collects data on all patients with newly diagnosed cancer in the southeastern part of the netherlands ( . million inhabitants). patients aged - years, diagnosed between and and followed up until january , were included. conditional -year relative survival was computed for every additional year survived. results: for many tumours conditional -year relative survival approached - % after having survived - years. however, for stomach cancer and hodgkin's lymphoma conditional -year relative survival increased to only - % and for lung cancer and non-hodgkin's lymphoma it did not exceed - %. initial differences in survival at diagnosis between age and stage groups disappeared after having survived for - years. conclusion: prognosis for patients with cancer changes with each year survived and for most tumours patients can considered to be cured after a certain period of time. however, for stomach cancer, lymphoma's and lung cancer the odds for death remains elevated compared to the general population. background: systematic review with meta-analysis, now regarded as 'best evidence', depends on availability of primary trials and on completeness of review. whilst reviewers have attempted to assess publication bias, relatively little attention has been given to selection bias by reviewers. method: systematic reviews of three cardiology treatments, that used common search terms, were compared for inclusion/exclusion of primary trials, pooled measures of principal outcomes and conclusions. results: in one treatment, reviews included , , , , and trials. there was little overlap: of trials in the last review only , , , and were included by others. reported summary effects ranged from (most effective to least significant); mortality relative risk . ( . , . ) in trials to . ( . , . ) in , and in one morbidity measure; standardised mean difference from . ( . , . ) in trials ( patients) to . () . , . ) in ( patients). reviewers' conclusions ranged from 'highly effective' to 'no evidence of effect'. conclusions: these examples illustrate strong selection bias in published meta-analyses. post hoc review contravenes one important principal of science 'first the hypothesis, then the test'. selection bias by reviewers may affect 'evidence' more than does publication bias. in the context of a large population based german case control study examining the effects of hormone therapy (ht) on breast cancer risk, we conducted a validation study comparing ht prescription data with participants' self-reports for data quality assurance. included were cases and controls aged - years, stratified by age and hormone use. study participants provided detailed information on ht use to trained interviewers, while gynecologists provided prescription data via telephone or fax. data were compared using proportion of agreement, kappa, intraclass correlation coefficient (icc), and descriptive statistics. overall agreement for ever/never use was . %, while agreement for ever/never use by type of ht was . %, . %, and . % for mono-estrogen, cyclical, and continuous combined therapy, respectively. icc for duration was high ( . ( % ci: . - . )), as were the iccs for age at first and last use ( . ( % ci: . - . ) and . ( % ci: . - . ), respectively). comparison of exact brand name resulted in perfect agreement for . % of participants, partial agreement for . %, and no agreement for . %. higher education and shorter length of recall were associated with better agreement. agreement was not differential by disease status. in conclusion, these self-reported ht data corresponded well with gynecologists' reports. background: legionnaires' disease (ld) is a pneumonia of low incidence. however, the impact of an outbreak can be substantial. objective: to stop a possible outbreak at an early stage, an outbreak detection programme was installed in the netherlands and evaluated after two years. design: the programme was installed nationally and consisted of sampling and controlling of potential sources to which ld patients had been exposed during their incubation period. potential sources were considered to be true sources of infection if two or more ld patients (cluster) had visited them, or if available patients' and environmental strains were indistinguishable by amplified fragment length polymorphism genotyping. all municipal health services of the netherlands participated in the study. the regional public health laboratory kennemerland sampled potential sources and cultured samples for legionella spp. results: rapid sampling and genotyping as well as cluster recognition helped to target control measures. despite these measures, two small outbreaks were only stopped after renewal of the water system. the combination of genotyping and cluster recognition lead to of ( %) patient-source associations. conclusion and discussion: systematic sampling and cluster recognition can contribute to ld outbreak detection and control. this programme can cost-effectively lead to secondary prevention. -up ( - ) , primary invasive breast cancers occurred. results: compared with hrt never-use, use of estrogen alone was associated with a significant . -fold increased risk. the association of estrogen-progestagen combinations with breast cancer risk varied significantly according to the type of progestagen: while there was no increase in risk with estrogen-progesterone (rr . [ . - . ]), estrogen-dydrogesterone was associated with a significant . -fold increase, and estrogen combined with other synthetic progestins with a significant . -fold increase. although the latter type of hrt involves a variety of different progestins, their associations with breast cancer risk did not differ significantly from one another. rrs did not vary significantly according to the route of estrogen administration (oral or transdermal/percutaneous). conclusion and discussion: progesterone rather than synthetic progestins may be preferred when an opposed estrogen therapy is to be prescribed. additional results on estrogen-progesterone are needed. background: although survival of hodgkin's lymphoma (hl) is high (> %), treatment may cause long-term side-effects like premature menopause. objectives: to assess therapy-related risk factors for premature menopause (age < ) following hl. design and methods: we conducted a cohort-study among female year hl-survivors, aged < at diagnose and treated between and . patients were followed from first treatment until june , menopause, death, or age . cumulative dose of various chemotherapeutic agents as well as radiation fields were studied as risk factors for premature menopause. cox-regression was used to adjust for age, year of treatment, smoking, bmi, and oral contraceptive-use. results: after a median follow-up of . years, ( %) women reached premature menopause. overall women ( %) were treated with chemotherapy only, ( %) with radiotherapy only and ( %) with both radio-and chemotherapy. exposure to procarbazine ), cyclophosphamide (hr . [ . - . ] ) and irradiation of the ovaries ]) were associated with significant increased risks for premature menopause. for procarbazine a dose-response relation was observed. procarbazine-use has decreased over time. conclusion: to decrease the risk for premature menopause after hl, procarbazine and cyclophosphamide exposure should be minimized and ovarian irradiation should be avoided. background: casale is an italian town where a large asbestos cement plant was active for decades. previous studies found increased risk for mesothelioma in residents, suggesting a decreasing spatial trend with distance from the plant. objective: to analyse the spatial variation of risk in casale and the surrounding area ($ , inhabitants) focussing on non-occupationally exposed individuals. design/methods: population-based case-control study including pleural mesotheliomas diagnosed between and . information on the cases and controls comprised lifelong residential and occupational history of subjects and their relatives. nonparametric tests of clustering were used to evaluate spatial aggregation. parametric spatial models based on distance between the longest-lasting subject residence (excluding the last years before diagnosis) and the source enabled estimation of risk gradient. results: mesothelioma risk appeared higher in an area of roughly - km radius from the source. spatial clustering was statistically significant (p = . ) and several clusters of cases were identified within casale. risk was highly related to the distance from the source; the best fitting model was the exponential decay with threshold. conclusion/discussion: asbestos pollution has increased dramatically the risk of mesothelioma in the area around casale. risk decreases slowly with the square of distance from the plant. malaria control programmes targeting malaria transmission from man to mosquito can have a large impact of malaria morbidity and mortality. to successfully interrupt transmission, a thorough understanding of disease and transmission parameters is essential. our objective was to map malaria transmission and analyse microenvironmental factors influencing this transmission in order to select high risk areas where transmission reducing interventions can be introduced. each house in the village msitu-wa-tembo was mapped and censused. transmission intensity was estimated from weekly mosquito catches. malaria cases identified through passive case detection were mapped by residence using gis software and the incidence of cases by season and distance to river were calculated. the distribution of malaria cases showed a clear seasonal pattern with the majority of cases during the rainy season (chisquare = . , p< . ). living further away from the river (p = . ) was the most notable independent protective factor for malaria infection. transmission intensity was estimated at . ( % ci . - . ) infectious bites per person per year. we show that malaria in the study area is restricted to a short transmission season. spatial clustering of cases indicates that interventions should be planned in the area closest to the river, prior and during the rainy season. background: the effectiveness of influenza vaccination of elders has been subject of some dispute. its impact on health inequalities also demands epidemiological assessments, as health interventions may affect early and most intensely better-off social strata. objectives: to compare pneumonia and influenza (p&i) mortality of elders (aged or more years old) before and after the onset of a largescale scheme of vaccination in sao paulo, brazil. methods: official information on deaths and population allowed the study of p&i mortality at the inner-city area level. rates related to the period to , during which vaccination coverage ranked higher than % of elders were compared with figures related to the precedent period ( ) ( ) ( ) ( ) ( ) . the appraisal of mortality decrease used a geo-referred model for regression analysis. results: overall p&i mortality reduced . % after vaccination. also the number of outbreaks, the excess of deaths during epidemic weeks, and the proportional p&i mortality ratio reduced significantly after vaccination. besides having higher prior levels of p&i deaths, deprived areas of the city presented a higher proportional decrease of mortality. conclusion: influenza vaccination contributed for an overall reduction of p&i mortality, while reducing the gap in the experience of disease among social strata. background: alcohol's first metabolite, acetaldehyde, may trigger aberrations in dna which predispose to developing colorectal cancer (crc) through several distinct pathways. our objective was to study associations between alcohol consumption and the risk of crc, according to two pathways characterized by mutations in apc and k-ras genes, and absence of hmlh expression. methods: in the netherlands cohort study, , men and women, aged - years, completed a questionnaire on risk factors for cancer in . case-cohort analyses were conducted using crc cases with complete data after . years of follow-up, excluding the first . years. gender-specific adjusted incidence rate ratios (rr) and % confidence intervals (ci) were estimated. results: neither total alcohol, nor beer, wine or liquor consumption was clearly associated with the risk of colorectal tumors lacking hmlh expression or harboring a truncating apc mutation and/or an activating k-ras mutation. in men and women, total alcohol consumption above g/day was associated with an increased risk of crc harboring a truncating apc and/or activating k-ras mutation, though not statistically significant. (rr: . ( % ci: . - . ) in men, rr: . ( % ci: . - . ) in women). in conclusion, alcohol consumption is not involved in the studied pathways leading to crc. background: educational level is commonly used to identify social groups with increased prevalence of smoking. other indicators of socioeconomic status (ses) might however be more discriminatory. objective: this study examines to what extent smoking behaviour is related to other ses indicators, such as labour market position and financial situation. methods: data derived from the european household panel, which includes data on smoking for european countries. we selected data for , respondents aged - years. the association between ses indicators and smoking prevalence was examined through logistic regression analyses. results: preliminary results show that, in univariate analysis, all selected ses indicators were associated with smoking. higher rates of smoking in lower social groups were observed in all countries, except for women in some mediterranean countries. in multivariate analyses, education retained an independent effect on smoking. no strong effect was observed for labour market position (occupational class, employment status) or for income. however, smoking prevalence was strongly related to economic deprivation and housing tenure. conclusion: these results suggest that different aspects of people's ses affect their smoking behaviour. interventions that aim to tackle smoking among high-risk groups should identify risk groups in terms of both education and material deprivation. objective: we investigated time trends in overweight and leisure time physical activities (ltpa) in the netherlands since . intra-national differences were examined stratified for sex, age and urbanisation degree. design and methods: we used a random sample from the health interview survey of about respondents, aged -to- years. self-reported data on weight, height and demographic characteristics were gathered through interviews (yearly) and data on ltpa were collected by selfadministered questionnaires . linear regression was performed for trend analyses. results: during - , mean body mass index (bmi) increased by . kg/m (p = . ). trends were similar across sex and urbanisation degrees. in -to- year old women, mean bmi increased more ( . kg/m ; p = . ) than in older women. concerning ltpa, no clear trend was observed during observed during - and observed during - . however, in year old women spent $ min/wk less on ltpa compared to older women, while this difference was smaller during - . conclusions: mean bmi increased more in younger women, which is consistent with the observation that this group spent less time on ltpa during recent years. although the overall increase in overweight could no´t be explained by trends in ltpa, physical activity interventions should target the younger women. background: prediction rules combine patient characteristics and test results to predict the presence of an outcome (diagnosis) or the occurrence of an outcome (prognosis) for individual patients. when prediction rules show poor performance in new patients, investigators often develop a new rule, ignoring the prior information available in the original rule. recently, several updating methods have been proposed that consider both prior information and information of the new patients. objectives: to compare five updating methods (that vary in extensiveness) for an existing prediction rule that preoperatively predicts the risk of severe postoperative pain (spp). design and methods: the rule was tested and updated on a validation set of new surgical patients ( ( %) with spp). we estimated the discrimination (the ability to discriminate between patients with and without spp) and calibration (the agreement between the predicted risks and observed frequencies of spp) of the five updated rules in other patients ( ( %) with spp). results: simple updating methods showed similar effects on calibration and discrimination as the more complex methods. discussion and conclusion: when the performance of a prediction rule in new patients is poor, a simple updating method can be applied to improve the predictive accuracy. about two million ethnic germans (aussiedler) have resettled in germany since . analyses with a yet incomplete follow-up of a cohort of aussiedler showed a different mortality compared to russia and germany. objectives: we investigated whether the mortality pattern changed after a complete follow-up and whether residential mobility after resettlement has an effect on mortality. we established a cohort of aussiedler who moved to germany between and . we calculated smr for all causes, external causes, cardiovascular deaths and cancer in comparison to german rates. results: with a complete follow-up, the cohort accumulated person years. overall, deaths were observed (smr . , % ci: . - . ). smr for all external causes, all cancer and cardiovascular deaths were . , . and . , respectively. increased number of moves within germany was associated with increased mortality. conclusion and discussion: the mortality in the cohort is surprisingly low, in particular for cardiovascular deaths. there is a mortality disadvantage from external causes and for some selected cancers. this disadvantage is however not as large as would be expected if aussiedler were representative of the general population in fsu countries. mobility as an indicator for a lesser integration will be discussed. background: breast cancer screening (bcs) provides an opportunity to analyze the relationship between lymph node involvement (ln), the most important prognostic factor, and biological and time dependent characteristics. objective: our aim was to assess those characteristics that are associated with ln in a cohort of screen-detected breast cancers. methods: observational population study of all invasive cancers within stage i-iiia detected from to through a bcs program in catalonia (spain). age, tumor size, histological grade, ln status and screening episode (prevalent or incident) were analyzed. pearson chi-square or fisher's exact test, mann-whitney test and stratified analyses were applied, as well as multiple logistic regression techniques. results: twenty nine percent ( % ci . - . %) out of invasive cancers had ln and . % were prevalent cancers. in the bivariate analysis, tumor size and age were strongly associated to ln (p< . ) while grade was related to ln only in incident cancers (p = . ). grade was associated with tumor size (p = . ) and with screening episode (p = . ). adjusting for screening episode and grade, age and tumor size were independent predictors of ln. conclusion: in conclusion, age and tumor size are independent predictors of ln. grade emerges as an important biological factor in incident cancers. background: the evidence regarding the association between smoking and cognitive function in the elderly is inconsistent. objectives: to examine the association between smoking and cognitive function in the elderly. design and methods: in , all participants of a population-based cohort study aged years or older were eligible for a telephone interview on cognitive function using validated instruments, such as the telephone interview of cognitive status (tics). information on smoking history was available from questionnaires administered in . we estimated the odds ratios (or) of cognitive impairment (below th percentile) and the corresponding % confidence intervals (ci) by means of logistic regression adjusting for age, sex, alcohol consumption, body mass index, physical exercise, educational level, depressive symptoms and co-morbidity. results: in total, participants were interviewed and had complete information on smoking history. former smokers had a lower prevalence of cognitive impairment (oradjusted = . ; % ci: . - . ) compared with never smokers, but not current smokers (oradjusted = . ; % ci: . - . ). conclusion: there is no association between current smoking and cognitive impairment in the elderly. discussion: the lack of association between current smoking and cognitive impairment is in line with previous non-prospective studies. the inverse association with former smoking might be due to smoking cessation associated with co-morbidities. background: many studies have reported late effects of treatment in childhood cancer survivors. most studies, however, focused on only one late effect or suffered from incomplete follow-up. objectives: we assessed the total burden of adverse events (ae), and determined treatment-related risk factors for the development of various aes. methods: the study cohort included -year survivors, treated in the emma childrens hospital amc in the netherlands between - . aes were graded for severity by one reviewer according to the common terminology criteria adverse events version . . results: medical follow-up data were complete for . % -year survivors. median follow-up time was years. almost % of survivors had one or more aes, and . % had even or more aes. of patients treated with rt alone, % had a high or severe burden of aes, while this was only % in patients treated with ct alone. radiotherapy (rt) was associated with the highest risk to develop an ae of at least grade , and was also associated with a greater risk to develop a medium to severe ae burden. conclusions: survivors are at increased risk for many health problems that may adversely affect their quality of life and long-term survival. background: studies in the past demonstrated that multifaceted interventions could enhance the quality of diabetes care. however many of these studies showed methodological flaws as no corrections were made for patient case-mix and clustering or a nonrandomised design was used. objective: to assess the efficacy of a multifaceted intervention to implement diabetes shared care guidelines. methods: a cluster randomised controlled trial of patients with type diabetes was conducted at general practises (n = ) and one outpatient clinic (n = ). in primary care, facilitators analysed barriers to change, introduced structured care, gave feedback and trained practice staff. at the outpatient clinic, an abstract of the guidelines was distributed. case-mix differences such as duration of diabetes, education, co-morbidity and quality of life were taken into account. results: in primary care, more patients in the intervention group were seen on a three monthly basis ( . % versus . %, p< . ) and their hba c was statistically significant lower ( . ± . versus . ± . , p< . ). however, significance was lost after correction for case-mix (p = . ). change in blood pressure and total cholesterol was not significant. we were unable to demonstrate any change in secondary care. conclusion: multifaceted implementation did improve the process of care but left cardiovascular risk unchanged. background: we have performed a meta-analysis including studies on the diagnostic accuracy of mr-mammography in patients referred for further characterization of suspicious breast lesions. using the bivariate diagnostic meta-analysis approach we found an overall sensitivity and specificity of . and . , respectively. the aim of the present analysis was to detect heterogeneity between studies. materials and methods: seventeen study and population characteristics were separately included in the bivariate model to compare sensitivity and specificity between strata of the characteristics. results: both sensitivity and specificity were higher in studies performed in the united states compared to non-united states studies. both estimates were also higher if two criteria for malignancy were used instead of one or three. only specificity was affected by the prevalence of cancer: specificity was highest in studies with the lowest prevalence of cancer in the study population. furthermore, specificity was affected by whether the radiologist was blinded for clinical information: specificity was higher if there was no blinding. conclusions: variation between studies was notably present across studies in country of publication, the number of criteria for malignancy, the prevalence of cancer and whether the observers were blinded for clinical information. objective: the aim of this project is to explore variation in three candidate genes involved in cholesterol metabolism in relation to risk of acute coronary syndrome (acs), and to investigate whether dietary fat intake modifies inherent genetic risks. study population: a case-cohort study is designed within the danish 'diet, cancer and health' study population. a total of cases of acs have been identified among , men and women who participated in a baseline examination between - when they were aged - years. a random sample of participants will serve as 'control' population. exposures: all participants have filled out a detailed -item food frequency questionnaire and a questionnaire concerning lifestyle factors. participants were asked to provide a blood sample. candidate genes for acs have been selected among those involved in cholesterol transport (atp-binding cassette transporter a , cholesterol-ester transfer protein, and acyl-coa:cholesterol acyltransferase ). five single nucleotide polymorphisms (snps) will be genotyped within each gene. snps will be selected among those with demonstrated functional importance, as assessed in public databases. methods: statistical analyses of association between genetic variation in the three chosen genes and risk of acs. explorations of methods to evaluate biological interaction will be of particular focus. background: c-reactive protein (crp) has been shown to be associated with type diabetes mellitus. it is unclear whether the association is completely explained by obesity. objective: to examine whether crp is associated with diabetes independent of obesity. design and methods: we measured baseline characteristics and serum crp in non-diabetic participants of the rotterdam study and followed them for a mean of . years. cox regression was used to estimate the hazard ratio. in addition, we performed a meta-analysis on published studies. results: during follow-up, participants developed diabetes. serum crp was significantly and positively associated with the risk to develop diabetes. the risk estimates attenuated but remained statistically significant after adjustment for obesity indexes. age, sex and body mass index (bmi) adjusted hazard ratios ( % ci) were . ( . - . ) for the fourth quartile, . ( . - . ) for the third quartile, and . ( . - . ) for the second quartile of serum crp compared to the first quartile. in the meta-analysis, weighed age, sex, and bmi adjusted risk ratio was . ( . - . ), for the highest crp category (> . mg/l) compared to the reference category (< . mg/l). conclusion: our findings shows that the association of serum crp with diabetes is independent of obesity. background: effectiveness of screening can be predicted by episode sensitivity, which is estimated by interval cancers following a screen. full-field digital or cr plate mammography are increasingly introduced into mammography screening. objectives: to develop a design to compare performance and validity between screen-film and digital mammography in a breast cancer screening program. methods: interval cancer incidence was estimated by linking screening visits from - at an individual level to the files of the cancer registry in finland. these data was used to estimate the study size requirements for analyzing differences in episode sensitivity between screen-film and digital mammography in a randomized setting. results: the two-year cumulative incidence of interval cancers per screening visits was estimated to be . to allow the maximum acceptable difference in the episode sensitivity between screenfilm and digital arm to be % ( % power, . significance level, : randomization ratio, % attendance rate), approximately women need to be invited. conclusion: only fairly large differences in the episode sensitivity can be explored within a single randomized study. in order to reduce the degree of non-inferiority between the screen-film and digital mammography, meta-analyses or pooled analyses with other randomized data are needed. session: socio-economic status and migrants presentation: oral. background: tackling urban/rural inequalities in health has been identified as a substantial challenge in reforming health system in lithuania. objectives: to assess mortality trends from major causes of death of the lithuanian urban and rural populations throughout the period of - . methods: information on major causes of death (cardiovascular diseases, cancers, external causes, and respiratory diseases) of lithuanian urban and rural populations from to was obtained from lithuanian department of statistics. mortality rates were age-standardized, using european standard. mortality trends were explored using the logarithmic regression analysis. results: overall mortality of lithuanian urban and rural populations was decreasing statistically significantly during - . more considerable decrease was observed in urban areas where mortality declined by . % per year in males and . % in females, compare to the decline by . % in males and . % in females in rural areas. the most notable urban/rural differences in mortality trends with unfavourable situation in rural areas were estimated in mortality from stoke, breast cancer in females, and external causes of death (traffic accidents and suicides). background: recent studies indicate that depression plays an important role in the occurrence of cardiovascular diseases (cvd). underlying mechanisms are not well understood. objectives: we investigated whether low intake of omega- fatty acids (fas) is a common cause for depression and cvd. methods: the zutphen elderly study is a prospective cohort study conducted in the netherlands. depressive symptoms were measured with the zung scale in men, aged - years, and free from cvd and diabetes in . dietary factors were assessed with a cross-check dietary history method. results: compared to high intake (= . mg/d), low intake (< . mg/d) of omega- fas, adjusted for demographics and cvd risk factors, was associated with an increased risk of depressive symptoms (or . ; % ci . - . ) at baseline, and non-significantly with -year cvd mortality (hr . ; % ci . - . ). the adjusted hr for a -point increase in depressive symptoms for cvd mortality was . ( % ci . - . ), and did not change after additional adjustment for omega- fas. conclusion: low intake of omega- fas may increase the risk of depression. our results, however, do not support the hypothesis that low intake of omega- fas explains the relation between depression and increased risk of cvd. background: during the last decades the incidence of metabolic syndrome has risen dramatically. several studies have shown beneficial effects of nut and seed intake on components of this syndrome. the relationship with prevalence of metabolic syndrome has not yet been examined. objectives: we studied the relation between nut and seed intake and metabolic syndrome in coronary patients. design and methods: presence of metabolic syndrome (according to international diabetes federation definition) was assessed in stable myocardial infarction patients ( % men) aged - years, as part of the alpha omega trial. dietary data were collected by food-frequency questionnaire. results: the prevalence of metabolic syndrome was %. median nut and seed intake was . g/day (interquartile range, . - . g/day). intake of nuts and seeds was inversely associated with the metabolic syndrome (prevalence ratio: . ; % confidence interval: . - . , for > g/day versus < g/day), after adjustment for age, gender, dietary and lifestyle factors. the prevalence of metabolic syndrome was % lower (p = . ) in men with a high nut and seed intake compared to men with a low intake, after adjustment for confounders. conclusion and discussion: intake of nuts and seeds may reduce the risk of metabolic syndrome in stable coronary patients. background: in epidemiology, interaction is often assessed by adding a product term to the regression model. in linear regression the regression coefficient of the product term refers to additive interaction. however, in logistic regression it refers to multiplicative interaction. rothman has argued that interaction as departure from additivity better reflects biological interaction. hosmer and lemeshow presented a method to quantify additive interaction and its confidence interval (ci) between two dichotomous determinants using logistic regression. objective: our objective was to provide an estimation method for additive interaction between continuous determinants. methods and results: from the abovementioned literature we derived the formulas to quantify additive interaction and its ci between one continuous and one dichotomous determinant and between two continuous determinants using logistic regression. to illustrate the theory, data of the utrecht health project were used, with age and body mass index as risk factors for diastolic blood pressure. conclusions: this paper will help epidemiologists to estimate interaction as departure from additivity. to facilitate its use, we developed a spreadsheet, which will become freely available at our website. background: the incidences of acute myocardial infarction (ami) and ischemic stroke (is) in finland have been among highest in the world. accurate geo-referenced epidemiological data in finland provides unique possibilities for ecological studies using bayesian spatial models. objectives: examine sex-specific geographical patterns and temporal variation of ami and is. design and methods: ami (n = , ) and is (n = , ) cases in - in finland, localized at the time of diagnosis according to the place of residence address using map coordinates. cases and population were aggregated to km x km grids. full bayesian conditional autoregressive models (car) were used for studying the geographic incidence patterns. results: the incidence patterns of ami and is showed on average % ( % ci - %) common geographic variation and significantly the rest of the variation was disease specific. there was no significant difference between sexes. the patterns of high-risk areas have persisted over the years and the pattern of is showed more annual random fluctuations. conclusions: although ami and is showed rather similar and temporally stable patterns, significant part of the spatial variation was disease specific. further studies are needed for finding the reasons for disease specific geographical variation. most studies addressing socio-economic inequalities in health services use fail to take into account the disease the patient is suffering from. the objective of this study is to compare possible socioeconomic differences in the use of ambulatory care between distinct patient groups: diabetics and patients with migraine. data was obtained from the belgian health interview surveys , and . in total patients with self reported diabetes or migraine were identified. in a multilevel analysis the probability of a contact and the volume of contacts with the general practitioner and/or the specialist were studied for both groups in relation to educational attainment and income. adjustment was made for age, sex, subjective health and comorbidity at the individual level, and doctors' density and region at district level. no socio-economic differences were observed among diabetic patients. among patients with migraine we observed a higher probability of specialist contacts in higher income groups (or , ; % ci , - , ) and higher educated persons (or , ; % ci , - , ), while lower educated persons tend to report more visits with the general practitioner. to correctly interpret socio-economic differences in the use of health services there is need to take into account information on the patient's type of disease. background: the suitability of non-randomised studies to assess effects of interventions has been debated for a long time, mainly because of the risk of confounding by indication. choices in the design and analytical phase of non-randomised studies determine the ability to control for such confounding, but have not been systematically compared yet. objective: the aim of the study will be to quantify the role of design and analytical factors on confounding in non-randomised studies. design and methods: a meta-regression analysis will be conducted, based on cohort and case-control studies analysed in a recent cochrane review on influenza vaccine effectiveness against hospitalisation or death in the elderly. primary outcome will be the degree of confounding as measured by the difference between the reported effect estimate (odds ratios or relative risks) and the best available estimate (nichol, unpublished data) . design factors that will be considered include study design, matching, restriction and availability of confounders. statistical techniques that will be evaluated include multivariate regression analysis with adjustment for confounders, stratification and, if available, propensity scores. results the rsults will be used to develop a generic guideline with recommendations how to prevent confounding by indication in non-randomised effect studies. the wreckage of the oil tanker prestige in november produced a heavy contamination of the coast of galicia (spain). we studied relationships between participation in clean-up work and respiratory symptoms in local fishermen. questionnaires including details of clean-up activities and respiratory symptoms were distributed among associates of fishermen's cooperatives, with postal and telephone follow-up. statistical associations were evaluated using multiple logistic regression analyses, adjusted for sex, age, and smoking status. between january and february , information was obtained from , fishermen (response rate %). sixty-three percent had participated in clean-up operations. lower respiratory tract symptoms were more prevalent in clean-up workers (odds ratio (or) . ; % confidence interval . - . ). the risk increased when the number of exposed days, number of hours per day, or number of activities increased (p for linear trend < . ). the excess risk decreased when more time had elapsed since last exposure (or . ( . - . ) and . ( . - . ) for more and less than months, respectively; p for interaction < . ). in conclusion, fishermen who participated in the clean-up work of the prestige oil spill show a prolonged dosedependent increased prevalence of respiratory symptoms one to two years after the beginning of the spill. background. hpv testing has been proposed for cervical cancer screening. objectives: evaluating the protection provided by hpv testing at long intervals vs. cytology every third year. methods: randomised controlled trial conventional arm: conventional cytology. experimental arm: in phase hpv and liquid-based cytology. hpv-positive cytology-negatives referred for colposcopy if age - , for repeat after one year if age - . in phase hpv alone. positives referred for colposcopy independently of age. endpoint: histologically confirmed cervical intraepithelial neoplasia (cin) grade or more. results: overall , women were randomised. preliminary data at recruitment are presented. overall, among women aged - years relative sensitivity of hpv versus conventional cytology was . ( % c.i. . - . ) and relative positive predictive (ppv) value was . ( % c.i. . - . ). among women aged - relative sensitivity of hpv vs. conventional cytology was . ( % c.i. . - . ) during phase but . ( % c.i. . - . ) during phase . conclusions: hpv testing increased cross-sectional sensitivity, but reduced ppv. in younger women data suggest that direct referral of hpv-positives to colposcopy results in relevant overdiagnosis of regressive lesions. measuring detection rate of cin at the following screening round will allow studying overdiagnosis and the possibility of longer screening intervals. background: plant lignans are present in foods such as whole grains, seeds, fruits and vegetables, and beverages. they are converted by intestinal bacteria into the enterolignans, enterodiol and enterolactone. enterolignans possess several biological activities whereby they may influence carcinogenesis. objective: to study the association between plasma enterolignans and the risk of colorectal adenomas. colorectal adenomas are considered to be precursors of colorectal cancer. design and method: the case-control study included cases with at least one histologically confirmed colorectal adenoma and controls with no history of any type of adenoma. associations between plasma enterolignans and colorectal adenomas were analyzed by logistic regression. results: associations were stronger for incident than for prevalent cases. when only incident cases (n = ) were included, high compared to low enterodiol plasma concentrations were associated with a reduction in colorectal adenoma risk after adjustment for confounding variables. enterodiol odds ratios ( % ci) were . , . ( . - . ), . ( . - . ), . ( . - . ) with a significant trend (p = . ) through the quartiles. although enterolactone plasma concentrations were fold higher, enterolactone's reduction in risk was not statistically significant (p for trend = . ). conclusion: we observed a substantial reduction in colorectal adenoma risk among subjects with high plasma concentrations of enterolignans, in particular enterodiol. background: smoking is a risk factor for tuberculosis diseases. recently the question was raised if smoking also increases the risk of tuberculosis infection. objective: to assess the influence of environmental tobacco smoke (ets) exposure in the household on tuberculosis infection in children. design and methods: a crosssectional community survey was done and information on children was obtained. tuberculosis infection was determined with a tuberculin skin test (tst) (cut-off mm) and information on smoking habits was obtained from all adult household members. univariate and multivariate analyses were performed, and odds ratio (or) was adjusted for the presence of a tb contact in the household, crowding and age of the child. results: ets was a risk factor for tuberculosis infection (or: . , % ci: . - . ) when all children with a tst read between two and five days were included. the adjusted or was . ( % ci: . - . ). in dwellings were a tuberculosis case had lived the association was strongest (adjusted or . , % ci: . - . ). conclusion and discussion: ets exposure seems to be a risk factor for tuberculosis infection in children. this is of great concern considering the high prevalence of smoking and tuberculosis in developing countries. background and objective: to implement a simulation model to analyze demand and waiting time (wt) for knee arthroplasty and to compare a waiting list prioritization system (ps) with the usual first-in, first-out (fifo) system. methods: parameters for the conceptual model were estimated using administrative data and specific studies. a discrete-event simulation model was implemented to perform -year simulations. the benefit of applying the ps was calculated as the difference in wt weighted by priority score between disciplines, for all cases who entered the waiting list. results: wt for patients operated under the fifo discipline was homogeneous (standard deviation (sd) between . - . months) with mean . . wt under the ps had higher variability (sd between . - . ) and was positively skewed, with mean . months and % of cases over months. when wt was weighted by priority score, the ps saved . months ( % ci . - . ) on average. the ps was favorable for patients with priority scores over , but penalized those with lower priority scores. conclusions: management of the waiting list for knee arthroplasty through a ps was globally more effective than through fifo, although patients with low priority scores were penalized with higher waiting times. background: we developed a probabilistic linkage procedure for the linking of readmissions of newborns from the dutch paediatrician registry (lnr). % of all newborns ( . ) are admitted to a neonatal ward. the main problems were the unknown number of readmissions per child and the identification of admissions of twins. objective: to validate our linking procedure in a double blinded study. design and methods: a random sample of admissions from children from the linked file has been validated by the caregivers, using the original medical records. results: response was %. for admissions of singletons the linkage contained no errors except for the small uncertain area of the linkage. for admissions of multiple births a high error rate was found. conclusion and discussion: we successfully linked the admissions of singleton newborns with the developed probabilistic linking algorithm. for multiple births we did not succeed in constructing valid admission histories due to too low data quality of twin membership variables. validation showed alternative solutions for linking twin admissions. we strongly recommend that linkage results should always be externally validated. background: salmonella typhimurium definitive phage type (dt) has emerged as an important pathogen in the last two decades. a -fold increase in cases in the netherlands during september-november prompted an outbreak investigation. objective: the objective was to identify the source of infection to enable preventive measures. methods: a subset of outbreak isolates was typed by molecular means. in a case-control study, cases (n = ) and matched population controls (n = ) were invited to complete self-administered questionnaires. results: the molecular typing corroborated the clonality of the isolates. the molecular type was similar to that of a recent s. typhimurium dt outbreak in denmark associated with imported beef. the incriminated shipment was traced after having been distributed sequentially through several eu member states. sampling of the beef identified s. typhimurium dt of the same molecular type as the outbreak isolates. cases were more likely than controls to have eaten a particular raw beef product. conclusions: our preliminary results are consistent with this s. typhimurium dt outbreak being caused by contaminated beef. our findings underline the importance of european collaboration, traceability of consumer products and a need for timely intervention into distribution chains. background: heavy-metals may affect newborns. some of them are presenting tobacco smoke. objectives: to estimate cord-blood levels of mercury, arsenic, lead and cadmium in newborns in areas in madrid, and to assess the relationship with maternal tobacco exposure. design and methods: bio-madrid study obtained cord-blood samples from recruited trios (mother/father/ newborn). cold-vapor atomic absorption spectrophotometry (aas) was used to measure mercury and graphite-furnace aas for the other metals. mothers answered a questionnaire including tobacco exposure. median, means and standard errors were calculated and logistic regression used to estimate or. results: median levels for mercury and lead were . mg/l and . mg/l. arsenic and cadmium were undetectable in % and % of samples. preliminar analysis showed a significant association of maternal tobacco exposure and levels of arsenic (or: . ; % ci: . - . ), cadmium (or: . ; % ci: . - . ), and lead (or: . ; % ci: . - . ). smoking in pregnancy was associated to arsenic (or: . ; % ci: . - . ), while passive exposure was more related to lead (or: . ; % ci: . - . ) and cadmium (or: . ; % ci: . - . ). conclusion: madrid newborns have high cord-blood levels of mercury. tobacco exposure in pregnancy might increase levels of arsenic, cadmium and lead. background: road traffic accidents (rta) are the leading cause of death for young. rta police reports provide no health information other then the number of deaths and injured, while health databases have no information on the accident dynamics. the integration of the two databases would allows to better describe the phenomenon. nevertheless, the absence of common identification variables through the lists makes the deterministic record linkage (rl) impossible. objective: to test feasibility of a probabilistic rl between rta and health information when personal identifiers are lacking. methods: health information came from the rta integrated surveillance for the year . it integrates data from ed visits, hospital discharges and deaths certificates. a deterministic rl was performed with police reports, where the name and age of deceased were present. results of the deterministic rl was then used as gold standard to evaluate the performance of the probabilistic one. results: the deterministic rl resulted in ( . %) linked records. the probabilistic rl, where the name was omitted, was capable to correctly identify ( . %). conclusions: performance of the probabilistic rl was good. further work is needed to develop strategies for the use of this approach in the complete datasets. background: overweight constitutes a major public health problem. the prevalence of overweight is unequally distributed between socioeconomic groups. risk group identification, therefore, may enhance the efficiency of interventions. objectives: to identify which socioeconomic variable best predicts overweight in european populations: education, occupation or income. design: european community household panel data were obtained for countries (n = , ). overweight was defined as a body mass index >= kg/m . uni-and multivariate logistic regression analyses were employed to predict overweight in relationship to socioeconomic indicators. results: major socioeconomic differences in overweight were observed, especially for women. for both sexes, a low educational attainment was the strongest predictor of overweight. after control for confounders and the other socioeconomic predictors, the income gradient was either absent or positive (men) or negative (women) in most countries. similar patterns were found for occupational level. for women, inequalities in overweight were generally greater in southern european countries. conversely, for men, differences were generally greater in other european countries. conclusion: across europe, educational attainment most strongly predicts overweight. therefore, obesity interventions should target adults and children with lower levels of education. background: because incidence and prevalence of most chronic diseases rise with age, their burden will increase in ageing populations. we report the increase in prevalence of myocardial infarction (mi), stroke (cva), diabetes type ii (dm) and copd based on the demographic changes in the netherlands. in addition, for mi and dm the effect of a rise in overweight was calculated. methods: calculations were made for the period - with a dynamic multi-state transition model and demographic projections of the cbs. results: based on ageing alone, between and prevalence of dm will rise from . to . (+ %), prevalence of mi from . to . (+ %), stroke prevalence from . to . (+ %) and copd prevalence from . to . (+ %). a continuation of the dutch (rising) trend in overweight prevalence would in lead to about . diabetics (+ %). a trend resulting in american levels would lead to over million diabetics (+ %), while the impact on mi was much smaller: about . (+ %) in . conclusion: the burden of chronic disease will substantially increase in the near future. a rising prevalence of overweight has an impact especially on the future prevalence of diabetes background: there has been increasing concern about the effects of environmental endocrine disruptors (eeds) on human reproductive health. eeds include various industrial chemicals, as well as dietary phyto-estrogens. intra-uterine exposures to eeds are hypothesized to disturb normal foetal development of male reproductive organs and specifically, to increase the risk of cryptorchidism, hypospadias, testicular cancer, and a reduced sperm quality in offspring. objective: to study the associations between maternal and paternal exposures to eeds and the risks of cryptorchidism, hypospadias, testicular cancer and reduced sperm quality. design and methods: these associations are studied using a case-referent design. in the first phase of the study, we collected questionnaire data of the parents of cases with cryptorchidism, cases with hypospadias and referent children. in the second phase, we will focus on the health effects at adult age: testicular cancer and reduced sperm quality. in both phases, we will attempt to estimate the total eed exposure of parents of cases and referents at time of pregnancy through an exposure-assessment model in which various sources of exposure, e.g. environment, occupation, leisure time activities and nutrition, are combined. in addition, we will measure hormone receptor activity in blood. background: eleven percent of the pharmacotherapeutic budget is spent on acid-suppressive drugs (asd); % of patients are chronic user. most of these indications are not conform to dyspepsia guidelines. objectives: we evaluated the implementation of an asd rationalisation protocol among chronic users, and analysed effects on volume and costs. method: in a cohort study patients from gp's with protocol were compared to a control group of patients from gp's without. prescription data of - were extracted from agis health database. a log-linear regression model compared standardised outcomes of number of patients that stopped or reduced asd (> %) and of prescription volume and costs. results: gp's and patients in both groups were comparable. % in the intervention group had stopped; % in the control group (p< . ). the volume had decreased in another % of patients; % in control group (p< . ). compared to the baseline data in the control group ( %) the adjusted or of volume in the intervention group was . %. the total costs adjusted or was . %. the implementation significantly reduced the number of chronic users, and substantially dropped volume and costs. active intervention from insurance companies can stimulate rationalisation of prescription. background/objective: today, % of lung cancers are resectable (stage i/ii). -year survival is therefore low ( %). spiral computed tomography (ct) screening detects more lung cancers than chest x-ray. it is unknown if this will translate into a lung cancer mortality reduction. the nelson trial investigates whether detector multi-slice ct reduces lung cancer mortality with at least % compared to no screening. we present baseline screening results. methods: a questionnaire was sent to , men and women. of the , respondents, , high-risk current and former smokers were invited. until december , , , of them gave informed consent and were randomised ( : ) in a screen arm (ct in year , and ) and control arm (no screening). data will be linked with the cancer registry and statistics netherlands to determine cancer mortality and incidence. results: of the first , baseline ct examinations % was negative (ct after one year), % indeterminate (ct after months) and % positive (referral pulmonologist). seventy percent of detected tumours were resectable. conclusion/discussion: ct screening detects a high percentage of early stage lung cancers. it is estimated that the nelson trial is sufficiently large to demonstrate a % lung cancer mortality reduction or more. background: due to diagnostic dilemmas in childhood asthma, drug treatment of young children with asthmatic complaints often serves as a trial treatment. objective: to obtain more insight into patterns and continuation of asthma medication in children during the first years of life. design: prospective birth cohort study methods: within the prevention and incidence of asthma and mite allergy (piama) study (n = , children) we identified children using asthma medication in their first year of life. results: about % of children receiving asthma medication before the age of one, discontinued use during follow-up. continuation of therapy was associated with male gender (adjusted odds ratio [aor] . , % confidence interval [ci]: . - . ), a diagnosis of asthma (aor . , % ci: . - . ) and receiving combination or controller therapy (aor , , % ci: . - . ). conclusion: patterns of medication use in preschool children support the notion that both beta -agonist and inhaled corticosteroids are often used as trial medication, since % discontinues. the observed association between continuation of therapy and both an early diagnosis of asthma and a prescription for controller therapy suggests that, despite of diagnostic dilemmas, children in apparent need of prolonged asthma therapy are identified at this very early age. background: this study explored the differences in birthweight between infants of first and second generation immigrants and infants of dutch women, and to what extent maternal, fetal and environmental characteristics could explain these differences. method: during months all pregnant women in amsterdam attending their first prenatel screening were asked to fill out a questionnaire (sociodemographic status, lifestyle) as part of the amsterdam born children and their development (abcd)-study; women ( %) responded. only singleton deliveries with pregnancy duration = weeks were included (n = ). results: infants of all first and second generation immigrant groups (surinam, the antilles, turkey, morocco, ghana, other countries) had lower birthweights (range: - gram) than dutch infants ( gram). linear regression revealed that, adjusted for maternal height, weight, age, parity, smoking, marital status, gestational age and gender, infants of surinamese women ( st and nd generation), antillean and ghanaian women (both st generation) were still lighter than dutch infants ( . , . , . , and . grams respectively; p< . ). conclusion: adjusted for maternal, fetal and environmental characteristics infants of some immigrant groups had substantial lower birthweights than infants of dutch women. other factors (like genetics, culture) can possibly explain these differences. introduction: missing data is frequently seen in cost-effectiveness analyses (ceas). we applied multiple imputation (mi) combined with bootstrapping in a cea. objective: to examine the effect of two new methodological procedures of combining mi and bootstrapping in a cea with missing data. methods: from a trial we used direct health and non-health care costs and indirect costs, kinesiophobia and work absence data assessed over months. mi was applied by multivariate imputation by chained equations (mice) and non-parametric bootstrapping was used. observed case analyses (oca), where analyses were conducted on the data without missings, were compared with complete case analysis (cca) and with analyses when mi and bootstrapping were combined after to % of cost and effect data were omitted. results: by the cca effect and cost estimates shifted from negative to positive and cost-effectiveness planes and acceptability curves were biased compared to the oca. the methods of combining mi and bootstrapping generated good cost and effect estimates and the cost-effectiveness planes and acceptability curves were almost identical to the oca. conclusion: on basis of our study results we recommend to use the combined application of mi and bootstrapping in data sets with missingness in costs and effects. background: since the s, coronary heart disease (chd) mortality rates have halved with approximately % of this decrease being attributable to medical and surgical treatments. objective: this study examined the cost-effectiveness of specific chd treatments. design and methods: the impact chd model was used to calculate the number of life-years-gained (lyg) by specific cardiology interventions given in , and followed over ten years. this previously validated model integrates data on patient numbers, median survival in specific groups, treatment uptake, effectiveness and costs of specific interventions. cost-effectiveness ratios were generated as costs per lyg for each specific intervention. results: in , medical and surgical treatments together prevented or postponed approximately , chd deaths in patients aged - years; this generated approximately , extra life years. aspirin and beta-blockers for secondary prevention of myocardial infarction and heart failure, and spironolactone for heart failure all appeared highly cost-effective (<e , per lyg). less cost effective, however, were revascularisation for chronic angina (cabg surgery e , and angioplasty e , per lyg), or statins for primary prevention (e , per lyg) or for community patients with angina (e , per lyg). conclusions: cost effectiveness ratios generally favoured simple medical treatments for myocardial infarction, secondary prevention, and heart failure. objective: is population-based primary prevention more favourable than secondary prevention (risk factor reduction in coronary heart disease (chd) patients)? methods: the previously validated im-pact model was used to integrate data describing chd patient numbers, specific treatment uptake levels, trends in major risk factors, and the mortality benefits of these risk factor changes in healthy people and in chd patients. results: approximately , fewer deaths were attributable to reductions in the three major risk factors between and . declining smoking prevalence resulted in approximately fewer deaths: in healthy people and in known chd patients. decreasing population total cholesterol concentrations resulted in approximately , fewer deaths attributable to dietary changes ( , in healthy people and in chd patients) plus fewer deaths attributable to statin treatment ( in people without chd and in chd patients). decreasing mean population blood pressure resulted in approximately fewer deaths attributable to secular falls in blood pressure ( in healthy people and in chd patients) plus approximately fewer deaths attributable to antihypertensive treatments in people without chd. conclusions: compared with secondary prevention ( fewer deaths), primary prevention ( , fewer deaths) achieved a three-fold larger reduction in chd deaths. background: carotenoid intake has been positively associated with plasma concentrations in different populations. however, the influence of body mass index (bmi) on this association is mostly unknown. objectives: we explored the relationship between intake of carotenoids and vitamin c, and their plasma concentrations by bmi categories in spanish elderly people. desgin/methods: a dietary interview using a -item food-frequency questionnaire was conducted with men and women, and older, participating in the eureye study in spain. blood samples were collected for measurement of carotenoids and vitamin c. correlation coefficients (r) between energy adjusted nutrient intakes and plasma concentrations were calculated by categories of bmi, adjusting for sex, age and smoking. results: significant correlations between acarotene, ß-carotene, lycopene, ß-cryptoxanthin and vitamin c intakes, and plasma concentrations were observed, r= . ; . ; . ; . , . , respectively (p< . ). correlations for carotenoids changed substantially by bmi categories, with the highest correlations for a-carotene, ß-carotene, and, to a lesser extension, ß-cryptoxanthin and lycopene, in subjects with bmi< ( . ; . ; . ; . respectively), and the lowest, in subjects with bmi= ( . ; . ; . ; . respectively). correlations for vitamin c remained unchanged by bmi. conclusions: our data suggest that carotenoids in plasma may be good markers of dietary intake in elderly subjects with lower bmi. background: reduced heart rate variability (hrv) is associated with an increased mortality, morbidity and worse prognosis of cardiovascular diseases (cvd). there is a lack of population-based data to examine the distribution of hrv, its association with cardiovascular risk factors (cvrf), and its role as a potential mediator of the effect of established risk factors on cvd. objective: to study the distribution of hrv and its association with cvrf in an elderly eastern german population. design and methods: this analysis is based on data of participants ( - years) of an ongoing cross-sectional study. time-and frequency-domain measures of hrv were computed. their association with cvrf was determined by linear regression modelling. results: age-and heart rate-adjusted standard deviation of the durations of nn intervals and high frequency power was significantly higher in women. significant inverse associations of timeand frequency-domain hrv with anthropometric parameters, triglycerides, blood pressure, prevalent diabetes and age were observed in both sexes. there was a graded inverse association of hrv with age except for the oldest age-group. discussion/conclusion: established cvrf are associated with a reduced hrv. therefore reduced hrv could be a mediator between cvrf and cvd, which will be examined in a follow-up study. background: aalen's model and cox's proportional hazard model postulate different relationship between hazard and covariates and the subject matter seldom suggests which of the models are to be preferred. actually the assumption of constant effects, either additive or multiplicative, may be incorrect especially in cancer survival analysis. objectives: the aim of the study is to analyse survival for a breast cancer cohort where typical prognostic factors lack of proportionality. design and methods: the cohort consists of invasive breast cancer cases from turin cancer registry within a high resolution study in co-operation with the breast cancer screening program, diagnosed from through , aged - . an additive-multiplicative cox-aalen model with age at diagnosis, categorized tumour size (t) and nodal status (n) is fitted. results: the prognostic role of age at diagnosis, t and n are confirmed. proportionality assumption holds only for age (hazard ratio = . , p-value< . ). t, particularly the category referring to 'extended to chest wall or skin' versus 'less than cm', has a significant time-varying effect (p-value = . ) after months since diagnosis. discussion: more flexible regression tools than cox model which is routinely used for analyzing cancer survival are needed. cox-aalen proves to be an appealing alternative. background: the major software packages for meta-analysis of causal -notably intervention -research are generally too expensive for developing countries. moreover, interactive educational software for students and teachers is largely unavailable. we recently developed the 'mix' program, an easy-to-maintain, interactive, and educational meta-analysis add-in for excel, available as free download at http://www.mix-for-meta-analysis.info. objectives: to formally establish whether the program's numerical and graphical output is comprehensive and accurate enough for scientific standards. design and methods: the study was designed as a software validation. data sets, primarily intervention studies, with substantially different features were analyzed by three investigators with mix and stata, using the latter program as golden standard. additionally, a feature (software options) comparison was made with modern programs for meta-analysis, such as cma . , metawin . , weasyma . , and revman . . results: the results of the validation project will be fully reported at the congress. preliminary results indicate that the mix program's output is comparable to stata output. it distinguishes itself from other software by the variety of graphical output, the excel-based interface, and free availability. conclusion and discussion: the mix program appears to provide valid output and may be an attractive, feature-rich alternative to existing meta-analysis software. background: an ankle-arm index (aai) < . of systolic blood pressure (sbp) as marker of peripheral arterial disease is a predictor of coronary and carotid artery disease. the present gold standard examination with doppler-ultrasound and sphygmomanometric cuff is subject to observer bias and requires intensive training. for epidemiologic studies, valid and easy-to-use examination methods are needed. objective: to validate the easy-to-use omron hem- cp for sbp measurements in the ankle and for aai determination in epidemiologic studies. design/methods: in a population-based sample of subjects ( - years), arm and ankle sbp measurements by the omron hem- cp and corresponding aai were compared with those using a hand-held doppler and sphygmomanometric cuff. bland-altman plots of sbp and aai differences, validation criteria for use in clinical practice and diagnostic values for the detection of an aai< . were employed. results: omron measured higher sbp than doppler. sbp and aai differences increasing towards higher sbp levels. specificity, sensitivity and negative predictive value for the detection of aai< . were > % (positive predictive value was %). conclusion: omron fails the validation criteria for ankle sbp measurement. however, the ease of use of the device could outweigh the inaccuracy if used as a screening tool for aai< , in epidemiologic studies. background: associations exist between: ) parental birth weight and child birth weight; ) birth weight and adult psychopathology; and ) maternal psychopathology during pregnancy and birth weight of the child. this study is the first to combine those associations. objective: to investigate the different pathways from parental birth weight and parental psychopathology to child birth weight in one model. design and methods: depression and anxiety scores on , mothers and , fathers during weeks pregnancy and birth weights from , children were available. path analyses with standardized regression coefficients were used to evaluate the different effects. results: in the unadjusted path analyses significant effects existed between: maternal (r = . ) and paternal birth weight (r = . ) and child birth weight; maternal birth weight and maternal depression (r=). ) and anxiety (r=). ); and maternal depression (r = . ) and anxiety (r = . ) and child birth weight. after adjustment for confounders, only maternal (r = . ) and paternal (r = . ) birth weight and maternal depression (r=). ) remained significantly related to child birth weight. conclusion after adjustment maternal depression, and not anxiety, remained significantly related to child birth weight. discussion future research should focus on the different mechanisms of maternal anxiety and depression on child birth weight. background: most patients with peripheral arterial disease (pad) die from coronary artery disease (cad). non-invasive cardiac imaging can assess the presence of coronary atherosclerosis and/or cardiac ischemia. screening in combination with more aggressive treatment may improve prognosis. objective: to evaluate whether a non-invasive cardiac imaging algorithm, followed by treatment will reduce the -year-risk of cardiovascular events in pad patients free from cardiac symptoms. design and methods: this is a multicenter randomized controlled clinical trial. patients with intermittent claudication and no history of cad are eligible. one group will undergo computed tomography (ct) calcium scoring. the other group will undergo ct calcium scoring and ct angiography (cta) of the coronary arteries. patients in the latter group will be scheduled for a dobutamine stress magnetic resonance imaging (dsmr) test to assess cardiac ischemia, unless a stenosis of the left main (lm) coronary artery (or its equivalent) was found on cta. patients with cardiac ischemia or a lm/lm-equivalent stenosis will be referred to a cardiologist, who will decide on further (interventional) treatment. patients are followed for years. conclusion: this study assesses the value of non-invasive cardiac imaging to reduce the risk of cardiovascular events in patients with pad free from cardiac symptoms. background: hpv is the main risk factor for cervical cancer and also a necessary cause for it. participation rates in cervical cancer screening are low in some countries and soon hpv vaccination will be available. objectives: aim of this systematic review was to collect and analyze published data on knowledge about hpv. design and methods: a medline search was performed for publications on knowledge about hpv as a risk factor for cervical cancer and other issues of hpv infection. results: of individual studies were stratified by age of study population, country of origin, study size, publication year and response proportion. heterogeneity was described. results: knowledge between included studies varied substantially. thirteen to % (closed question) and . to . % (open question) of the participants knew about hpv as a risk factor for cervical cancer. women had consistently better knowledge on hpv than men. there was confusion of hpv with other sexually transmitted diseases. conclusion and discussion: studies were very heterogeneous, thus making comparison difficult. knowledge about hpv infections depended on the type of question used, gender of the participants and their professional background. education of the general public on hpv infections needs improvement, specially men should also be addressed. background: influenza outbreaks in hospitals and nursing homes are characterized by high attack rates and severe complications. knowledge of the virus' specific transmission dynamics in healthcare institutions is scarce but essential to develop cost-effective strategies. objective: to follow and model the spread of influenza in two hospital departments and to quantify the contributions of the several possible transmission pathways. methods: an observational prospective cohort study is performed on the departments of internal medicine & infectious diseases and pulmonary diseases of the umc utrecht during the influenza season. all patients and regular medical staff are checked daily on the presence of fever and cough, the most accurate symptoms of influenza infection. nose-throat swabs are taken for pcr analysis for both symptomatic individuals and a sample of asymptomatic individuals. to determine transmission, contact patterns are observed between patients and visitors and patients and medical staff. results/discussion: spatial and temporal data of influenza cases will be combined with contact data in a mathematical model to quantify the main transmission pathways. among others the model can be used to predict the effect of vaccination of the medical staff which is not yet common practice in the studied hospital. background: the long term maternal sequelae of stillbirths is unknown. objectives: to assess whether women who experienced stillbirths have an excess risk of long term mortality. study design: cohort study. methods: we traced jewish women from the jerusalem perinatal study, a population-based database of all births to west jerusalem residents ( - who gave birth at least twice during the study period, using unique identity numbers. we compared the survival to - - of women who had at least one stillbirth (n = ) to that of women who had only live births (n = ) using cox proportional hazards models. results: during a median follow up of . years, ( . %) mothers with stillbirths died compared to , ( . %) unexposed women; crude hazard ratio (hr) . ( % ci: . - . ). the mortality risk remained significantly increased after adjustments for sociodemo-graphic variables, maternal diseases, placental abruption and preeclampsia (hr: . , % ci: . - . ). stillbirth was associated with increased risk of death from cardiovascular (adjusted hr: . , . - . ), circulatory ( . , . - . ) and genitourinary ( . , . - . ) causes. conclusions: the finding of increased mortality among mothers of stillbirths joins the growing body of evidence demonstrating long term sequelae of obstetric events. future studies should elucidate the mechanisms underlying these associations. resilience is one of the essential characteristics of successful ageing. however, very little is known about the determinants of resilience in old age. our objectives were to identify resilience in the english longitudinal study of ageing (elsa) and to investigate social and psychological factors determining it. the study design was a crosssectional analysis of wave of elsa. using structural equation modelling, we identified resilience as a latent variable indicated by high quality of life in the face of six adversities: ageing, limiting long-standing illness, disability, depression, perceived poverty and social isolation and we regressed it on social and psychological factors. the latent variable model for resilience showed a highly significant degree of fit (weighted root mean square resid-ual= . ). determinants of resilience included good quality of relationships with spouse (p = . ), family (p = . ), and friends (p< . ), good neighbourhood (p< . ), high level of social participation (p< . ), involvement in leisure activities (p = . ); perception of not being old (p< . ); optimism (p = . ), and high subjective probability of survival to an older age (p< . ). we concluded that resilience in old age was as much a matter of social engagement, networks and context as of individual disposition. implications of this on health policy are discussed. background: there is extensive literature concluding that ses is inversely related to obesity in developed countries. several studies in developing populations however reported curvilinear or positive association between ses and obesity. objectives: to assess the social distribution of obesity in men and women in middle-income countries of eastern and central europe with different level of economic development. methods: random population samples aged - years from poland, russia and czech republic were examined between - as baseline for prospective cohort study. we used body-mass index (bmi) and waist/hip ratio (whr) as obesity measures. we compared age-adjusted bmi and whr for men and women by educational levels in all countries. results: the data collection was concluded in summer . we collected data from about , subjects. lower ses increased obesity risk in women in all countries (the strongest gradient in the czech republic and the lowest in russia), and in czech men. there was no ses gradient in bmi in polish men and positive association between education and bmi in russian men. conclusions: our findings strongly agree with previous literature showing that the association between ses status and obesity is strongly influenced by overall level of country economic development. background: inaccurate measurements of body weight, height and waist circumference will lead to an inaccurate assessment of body composition, and thus of the general health of a population. objectives: to assess the accuracy of self-reported body weight, height and waist-circumference in a dutch overweight working population. design and methods: bland and altman methods were used to examine the individual agreement between self-reported and measured body weight and height in overweight workers ( % male; mean age . +/) . years; mean body mass index [bmi] . +/) . kg/m ). the accuracy of self-reported waistcircumference was assessed in a subgroup of persons ( % male; mean age . +/) . years; mean bmi . +/) . kg/ m ), for whom both measured and self-reported waist circumference was available. results: body weight was underreported by a mean (standard deviation) of . ( . ) kg, body height was on average over-reported by . ( . ) cm. bmi was on average underreported by . ( . ) kg/m . waist-circumference was overreported by . ( . ) cm. the overall degree of error from selfreporting was between . and . %. conclusion and discussion: self-reported anthropometrics seem satisfactorily accurate for the assessment of general health in a middle-aged overweight working population. the incidence of breast cancer and the prevalence of metabolic syndrome are increasing rapidly in chile, but this relationship is still debated. the goal of this study is to assess the association between metabolic syndrome and breast cancer before and after menopause. a hospital based case-control study was conducted in chile during . cases with histologically confirmed breast cancer and age matched controls with normal mammography were identified. metabolic syndrome was defined by atpiii and serum lipids, glucose, blood pressure and waist circumference were measured by trained nurses. data of potential confounders such as, obesity, socioeconomic status, exercise and diet were obtained by anthropometric measures and a questionnaire. odds ratios (ors) and % confidence intervals (cis) were estimated by conditional logistic regression stratified by menopause. in postmenopausal women, a significant increase risk of breast cancer was observed in women with metabolic syndrome (or = . , % ci = . - . ). the elements of metabolic syndrome strongly associated were high levels of glucose and hypertension. in conclusion, postmenopausal women with metabolic syndrome had % of excess risk of breast cancer. these findings support the theory that there is a different risk profile of breast cancer after and before menopause. background: physical exercise during pregnancy has numerous beneficial effects on maternal and foetal health. it may, however, affect early foetal survival negatively. objectives: to examine the association between physical exercise and spontaneous abortion in a cohort study. design and methods: in total, , women recruited to the danish national birth cohort in early pregnancy, provided information on amount and type of exercise during pregnancy and on possible confounding factors. , women experienced foetal death before gestational weeks. hazard ratios for spontaneous abortion in four periods of pregnancy () , - , - , and - weeks) according to amount (min/week) and type of exercise, respectively, were estimated using cox regression. various sensitivity analyses to reveal distortion of the results from selection forces and information bias were made. results: the hazard ratios of spontaneous abortion increased stepwise with amount of physical exercise and were largest in the earlier periods of pregnancy (hrweek - = . (ci . - . ) for min/week, compared to no exercise). weight bearing types of exercise were strongest associated with abortion, while swimming showed no association. these results remained stable, although attenuated, in the sensitivity analyses. discussion: handling of unexpected findings that furthermore challenge official public health messages will be discussed. hemodialysis (hd) patients with a low body mass index (bmi) have an increased mortality risk, but bmi changes over time on dialysis treatment. we studied the association between changes in bmi and all-cause mortality in a cohort of incident hd patients. patients were followed until death or censoring for a maximum follow-up of years. bmi was measured every months and changes in bmi were calculated over each -mo period. with a time-dependent cox regression analysis, hazard ratios (hr) were calculated for these -mo changes on subsequent mortality from all causes, adjusted for the mean bmi of each -mo period, age, sex and comorbidity. men and women were included (age: ? years, bmi: . ? . kg/m , -y mortality: %). a loss of bmi> % was independently associated with an increased mortality risk (hr: . , %-ci: . - . ), while a loss of - % showed no difference (hr: . , . - . ) compared to no change in bmi () % to + % change). a gain in bmi of - % showed beneficial (hr: . , . - . ), while a gain of bmi> % was not associated with a survival advantage (hr: . , . - . ). in conclusion, hd patients with a decreasing bmi have an increased risk of all-cause mortality. background: within the tripss- project, impact of clinical guidelines (gl) on venous thromboembolism (vte) prophylaxis was evaluated in a large italian hospital. gl were effective in increasing the appropriateness of prophylaxis and in reducing vte. objectives: we performed a cost-effectiveness analysis by using a decision-tree model to estimate the impact of the adopted gl on costs and benefits. design and methods: a decision-tree model compared prophylaxis cost and effects before and after gl implementation. four risk profiles were identified (low, medium, high, very high). possible outcomes were: no event, major bleeding, asymptomatic vte, symptomatic vte and fatal pulmonary embolism. vte patients risk and probability of receiving prophylaxis were defined using data from the previous survey. outcome probabilities were assumed from literature. tariffs and hospital figures were used for costing the events. results: gl introduction reduced the average cost per patient from e to () %) with an increase in terms of event free patients (+ %). results are particularly relevant in the very high risk group. conclusion: the implementation of locally adapted gl may lead to a gain in terms of costs and effects, in particular for patients at highest vte risk. background: assisted reproductive techniques are used to overcome infertility. one reason of success is the use of ovarian stimulation. objectives: compare two ovarian stimulation protocols, gonadotropin-releasing hormone agonists/antagonists, assessing laboratorial and clinical outcomes, to provide proper therapy option. identify significant predictors of clinical pregnancy and ovarian response. design and methods: retrospective study (agonist cycles, ; antagonist cycles, ) including ivf/intracytoplasmic sperm injection cycles. multiple logistic and regression models, with fractional polynomial method were used. results: antagonist group exhibited lower length of stimulation and dose of recombinant follicle stimulating hormone (rfsh), higher number of retrieved and fertilized oocytes, and embryos. agonist group presented thicker endometrium, better fertilization, implantation and clinical pregnancy rates. clinical pregnancy has shown positive correlation with endometrial thickness and use of agonist; negative correlation with age and number of previous attempts. retrieved oocytes shown positive correlation with estradiol on day of human chorionic gonadotrophin (hcg) and use of antagonist; negative correlation with rfsh dose. conclusion: patients from antagonist group are more likely to get more oocytes and quality embryos, despite those from agonist group are more likely to get pregnant. background: prevalence studies of the metabolic syndrome require fasting blood samples and are therefore lacking in many countries including germany. objectives: to narrow the incertitude resulting from extrapolation of international prevalence estimates, with a sensitivity analysis of the prevalence of the metabolic syndrome in germany using a nationally representative but partially non-fasting sample. methods: stepwise analysis of the german health examination survey , using the national cholesterol education program (ncep) criteria, hemoglobin a c (hba c), non-fasting triglycerides and fasting time. results: among participants aged - years, the metabolic syndrome prevalence was (i) . % with . % inconclusive cases using the unmodified ncep criteria, (ii) . % with . % inconclusive cases using hba c > . % if fasting glucose was missing, (iii) . % with . % inconclusive cases when additionally using non-fasting triglycerides = th percentile stratified by fasting time, and (iv) . % to . % with < % inconclusive cases using different cutoffs (hba c . %, non-fasting triglycerides and mg/dl). discussion: despite a lower prevalence of obesity in germany compared to the us, the prevalence of the metabolic syndrome is likely to be in the same order of magnitude. this analysis may help promote healthy life styles by stressing the high prevalence of interrelated cardiovascular risk factors. background: epidemiologic studies that directly examine fruits and vegetables (f&v) consumption and other lifestyle factors in relation to weight gain are sparse. objective: we examined the associations between the f&v intake and -y weight gain among spanish adult people. design/methods: the study was conducted with a sub-sample of healthy people aged y and over at baseline in , who participated in a population-based nutrition survey in valencia-spain. data on diet, lifestyle factors and body weight (direct measurement) were obtained in and . information on weight gain was available for participants in . results: during the -y period, participants tended to gain on average . kg (median = . kg). in multivariate analyses, participants with the highest tertile of f&v intake at baseline had a % of lower risk of gaining = . kg compared with those who had the lowest intake tertile after adjustment for sex, age, education, smoking, tv-viewing, bmi, and energy intake (or = . ; % ci: . . ;p-fortrend = . ). for every g/d increase in f&v intake, the or was reduced by % (or = . ; . - . ;p-trend= . ). tvviewing at baseline was positively associated with weight gain, or for- h-increase= . ( . - . ;p-trend= . ). conclusions: our findings suggest that a high f&v intake and low tv-viewing may reduce weight gain among adults. background: diabetic patients develop more readily atherosclerosis thus showing greatly increased risk for cardiovascular disease. objective: the heinz nixdorf recall-study is a prospective cohort-study designed to assess the prognostic value of new risk stratification methods. here we examined the association between diabetes, previously unknown hyperglycemia and the degree of coronary calcification (cac). methods: a population-based sample of , men and women aged - years was recruited from three german cities between - . baseline examination included amongst others a detailed medical history, blood analyses and electron-beam tomography. we calculated adjusted prevalence ratios (pr), adjusting for age, smoking, bmi and %-confidence intervals ( % ci) with log-linear binomial regression. results: the prevalence of diabetes is . % (men: . %, women: . %), for hyperglycemia . % (men: . %, women: . %). prevalence ratio for cac in male diabetics without overt coronary heart disease is . ( % ci: . - . ), for those with hyperglycemia . ( . - . ). in women the association is even stronger: . ( . - . ) with diabetes, . ( . - . ) with hyperglycemia. conclusion: the data support the concept of regarding diabetic patients as being in a high risk category meaning > % hard events in years. furthermore, even persons with elevated blood glucose levels already show higher levels of coronary calcification. background: birth weight is associated with health in infancy and later in life. socioeconomic inequality in birth weight is an important marker of current and future population health inequality. objective: to examine the effect of maternal education on birth weight, low birth weight (lbw, birth weight< , g), and small for gestational age (sga) background: in clinical practice patient data are obtained gradually and health care practitioners tune prognostic information according to available data. prognostic research does not always reproduce this sequential acquisition of data: instead, 'worst', discharge or aggregate data are often used. objective: to estimate prognostic performance of sequentially updated models. methods: cohortstudy of all very-low-birth-weight-babies (< g) admitted to the study neonatal icu < days after birth ( eligible from to ) and followed-up until years old ( . % lost-to-follow-up). main outcomes: disabling cerebral palsy at years ( , . %) or death ( , . % ) % in the first weeks). main prognostic determinants: neonatal cerebral lesions identified with cranial ultrasound (us) exams performed per protocol on days , , and at discharge. logistic regression models were updated with data available at these different moments in time during admission. results: at days , and respectively, main predictor (severe parenchymal lesion) adjusted odds ratio: , and ; us model c-statistic: . , . and . . discussion: prognostic models performance in neonatal patients improved from inception to discharge, particularly for identification of the high risk category. time of data acquisition should be considered when comparing prognostic instruments. in epidemiological longitudinal studies one is often interested in the analysis of time patterns of censored history data. for example, how regular a medication is used or how often someone is depressed. our goal is to develop a method to analyse time patterns of censored data. one of the tools in longitudinal studies is a nonhomogeneous markov chain model with discrete time moments and categorical state space (for example, use of various medications). suppose we are interested only in the time pattern of appearance of a particular state which is in fact a certain epidemiological event under study. for this purpose we construct a new homogeneous markov chain associated with this event. the states of this markov chain are the time moments of the original nonhomogeneous markov chain. using the new transition matrix and standard tools from markov chain theory we can derive the probabilities of occurence of that epidemiological event during various time periods (including ones with gaps). for example, probabilities of cumulative use of medication during any time period. in conclusion, the proposed approach based on markov chain model provides a new way of data representation and analysis which is easy to interpret in practical applications. background: tuberculosis (tb) cases that belong to a cluster of the same mycobacterium tuberculosis dna fingerprint are assumed to be consequence of recent transmission. targeting interventions to fast growing clusters may be an efficient way of interrupting transmission in outbreaks. objective: to assess predictors for large growing clusters compared to clusters that remain small within a years period. design and method out of the culture confirmed tb patients diagnosed between and , ( %) had unique fingerprints while were part of a cluster. of the clustered cases were in a small ( to cases within the first years) and in a large cluster (more than cases within the first years). results independent risk factors for being a case within the first years of a large cluster were non-dutch nationality (or = . % ci [ . - . background: passive smoking causes adverse health outcomes such as lung cancer or coronary heart disease (chd). the burden of passive smoking on a population level is currently unclear and depends on several assumptions. we investigated the public health impact of passive smoking in germany. methods: we computed attributable mortality risks due to environmental tobacco smoke (ets). we considered lung cancer, chd, stroke, copd and sudden infant death. frequency of passive smoking was derived from the national german health survey. sensitivity analyses were performed using different definitions of exposure to passive smoking. results: in total, deaths every year in germany are estimated to be caused by exposure to passive smoking at home (women , men ). most of these deaths are due to chd ( ) and stroke ( ). additional consideration of passive smoking at workplace increased the number of deaths to . considering any exposure to passive smoking and also active smokers who report exposure to passive smoking increased the number of deaths further. conclusions: passive smoking has an important impact on mortality in germany. even the most conservative estimate using exposure to ets at home led to a substantial number of deaths related to passive smoking. des daughters have a strongly increased risk of clear-cell adenocarcinoma of the vagina and cervix (ccac) at a young age. longterm health problems, however, are still unknown. we studied incidence of cancer, other than ccac, in a prospective cohort of des daughters (desnet project). in , questionnaires were sent to des daughters registered at the des center in utrecht. also, informed consent was asked for linkage with disease registries. for this analysis, data of , responders and nonresponders were linked to palga, the dutch nationwide network and registry of histo-and cytopathology. mean age at the end of follow-up was . years. a total of incident cancers occurred. increased standardized incidence rates (sir) were found for vaginal/vulvar cancers (sir = . , % confidence interval ( % ci) . - . ), melanoma (sir = . , % ci . - . )) and breast cancer (sir= . , % ci . - . ) as compared to the general population. no increased risk was found for invasive cervical cancer, possibly due to effective screening. results for breast and cervical cancer are consistent with the sparse literature. the risk of melanoma might be due to surveillance bias. future analyses will include non-invasive cervical cancer, stage specific sirs for melanoma and adjustment for confounding (sister control group) for breast cancer. background: contact tracing plays an important role in the control of emerging infectious diseases in both human and farm animal populations, but little is known yet about its effectiveness. here we investigate in a generic setting for well-mixed populations the dependence of tracing effectiveness on the probability that a contact is traced, the possibility of iteratively tracing yet asymptomatic infectives, and delays in the tracing process. methods and findings: we investigate contact tracing in a mathematical model of an emerging epidemic, incorporating a flexible infectivity function and incubation period distribution. we consider isolation of symptomatic infected as the basic scenario, and determine the critical tracing probability (needed for effective control) in relation to two infectious disease parameters: the reproduction ratio under isolation and the duration of latent period relative to the incubation period. the effect of tracing delays is considered, as is the possibility of single-step tracing vs. iterative tracing of quarantined invectives. finally, the model is used to assess the likely success of tracing for influenza, smallpox, sars, and foot-and-mouth disease epidemics. conclusions: we conclude that single-step contact tracing can be effective for infections with a relatively long latent period or a large variation in incubation period, thus enabling backwards tracing of super spreading individuals. the sensitivity to changes in the tracing delay varies greatly, but small increases may have major consequences for effectiveness. if single-step tracing is on the brink of being effective, iterative tracing can help, but otherwise it will not improve much. we conclude that contact tracing will not be effective for influenza pandemics, only partially for fmd epidemics, and very effective for smallpox and sars epidemics. abstract: infections of highly pathogenic h n avian influenza in humans underline the need for tracking of the ability of these viruses to spread among humans. here we propose a method of analysing outbreak data that allows determination of whether and to what extent transmission in a household has occurred after an introduction from the animal reservoir. in particular, it distinguishes between onward transmission from humans that were infected from the animal reservoir (primary human-to-human transmission) and onward transmission from humans who were themselves infected by humans (secondary human-to-human transmission). the method is applied to data from an epidemiological study of an outbreak of highly pathogenic avian influenza (h n ) in the netherlands in . we contrast a number of models that differ with respect to the assumptions on primary versus secondary human-to-human transmission. session: mathematical modelling of infectious diseases presentation: oral. usually models for the spread of an infection in a population are based on the assumption of a randomly mixing population, where every individual may contact every other individual. however, the assumption of random mixing seems to be unrealistic, therefore one may also want to consider epidemics on (social) networks. connections in the network are possible contacts, e.g. if we consider sexually transmitted diseases and ignore all spread by other than sexual ways, the connections are only between people that may have intercourse with each other. in this talk i will compare the basic reproduction ratio, r and the probability of a major outbreak of network models and for randomly mixing populations. furthermore, i will discuss which properties of the network are important and how they can be incorporated in the model. in this talk a reproductive power model is proposed that incorporates the following points met when an epidemic disease outbreak is modeled statistically: ) the dependence of the data is handled with a non-homogeneous birth process. ) the first stage of the outbreak is described with an epidemic sir model. soon control measures will start to influence the process. these measures are in addition to the natural epidemic removal process. the prevalence is related to the censored infection times in such a way that the distribution function, and therefore the survival function, satisfies approximately the first equation of the sir model. this leads in a natural way to the burr-family of distributions. ) the non-homogeneous birth process handles the fact that in practice, with some delay, it is the infected that are registered and not the susceptibles. ) finally the ending of the epidemic caused by the measures taken is incorporated by modifying the survival function with a finalsize parameter in the same way as is done in long-term survival models. this method is applied to the dutch classical swine individual and area (municipality) measures of income, marital and employment status were obtained. there were , suicides and , controls. after controlling for compositional effects, ecological associations of increased suicide risk with declining area levels of employment and income and increasing levels of people living alone were much attenuated. individual-level associations with these risk factors were little changed when controlling for contextual effects. we found no consistent evidence that associations with individual level risk factors differed depending on the areas characteristics (cross-level interactions). this analysis suggests the ecological associations reported in previous studies are likely to be due in greater part to the characteristics of the residents in those areas than area-influences on risk, rather than to contextual effects. were found to be at higher risk. risk was significantly greater in women whose first full-term pregnancy was at age or more (or . , ). in addition, more than full-term pregnancies would be expected to correlate with an increase in the risk (x . , p< . ). in multivariate analysis, history of breast feeding is a significant factor in decreasing risk (or . , % ci . - . the euregion meuse-rhine (emr) is an area with different regions, regarding language, culture and law. organisations and institutions received frequently signals about an increasing and region-related consumption of addictive drugs and risky behaviour of adolescents. as a reaction institutions from regions of the emr started a cross-border cooperation project 'risky behaviour adolescents in the emr'. the partners intend to improve the efficiency of prevention programmes by investigating the prevalence and pre-conditional aspects related to risky behaviour, and creating conditions for best-practice-public-health. the project included two phases: study. two cross-border (epidemiological) studies where realized: a quantitative study of the prevalence of risky behaviour ( pupils) and a qualitative study mapped preconditional aspects of risky behaviour and possibilities to preventive programmes. implementation. this served bringing about recommendations on policy level as well as on prevention level. during this phase the planning and realisation of cross-border preventionprogrammes and activities started. there is region-related variance of prevalence in risky-behaviour of adolescents in de emr. also there are essential differences in legislation and regulation, (tolerated) policy, prevention structures, political and organizational priorities and social acceptance toward stimulants. cross-border studies and cooperation between institutions have resulted in best-practice-projects in (border) areas of the emr. abstract background: beta-blockers increase bone strenght in mice and may reduce fracture risk in humans. therefore, we hypothesized that inhaled beta- agonists may increase risk of hip fracture. objective: to determine the association between daily dose of beta- agonist use and risk of hip fractures. methods: a case-control study was conducted among adults who were enrolled in the dutch phar-mo database (n = , ). cases (n = , ) were patients with a first hip fracture. the date of the fracture was the index date. four controls were matched by age, gender and region. we adjusted our analyses for indicators of asthma/copd severity, and for disease and drug history. results: low daily doses (dds) (< ug albuterol eq.) of beta -agonists (crude or . , % ci . - . ) did not increase risk of hip fracture, in contrast to high dds (> ug albuterol eq., crude or . , % ci . . - . ). after extensive adjustment for indicators of the severity of the underlying disease, (including corticosteroid intake), fracture risk in the high dd group decreased to . ( % ci . - . ). conclusions: high dds of beta- agonists are linked to increased risk of hip fracture. extensive adjustments for the severity of the underlying disease is important when evaluating this association. abstract salivary nitrate arises from ingested nitrate and is the main source of gastric nitrite, a precursor of carcinogenic n-nitroso compounds. we examinated the nitrate and nitrite levels in saliva of children who used private wells for their drinking water supply. saliva was collected in the morning, from children aged - years. control group (n = ) drank water containing . - . mg/l (milligrams/litre) nitrate. exposure groups consisting of subjects (n = ) who used private wells with nitrate levels in drinking water below mg/l (mean ± standard deviation . ± . mg/l) and above mg/l (n = ) ( . ± . mg/l) respectively. the nitrate and nitrite of saliva samples was determined by high performance liquid chromatographs method. the values of nitrate in saliva samples from exposed groups ranged between . to . mg/l ( . ± . mg/l). for control groups, the levels of . to . mg/l ( . ± . mg/l) were registered. no differences between levels of salivary nitrite from control and exposed groups were found. regression analysis on water nitrate concentrations and salivary nitrate showed significant correlations. in conclusion, we estimate that salivary nitrate may be used as biomarkers of human exposure to nitrate. abstract disinfection of public drinking water supplies produces trihalomethanes. epidemiological studies have associated chlorinated disinfection by-products with cancer, reproductive and developmental effects. we studied the levels of trihalomethanes (chloroform, dibromochloromethane, bromodichloromethane, bromoform) in drinking water delivered to the population living in some urban areas (n= ). the water samples (n= ) were analysed using gas chromatographic method. assessment of exposure to trihalomethanes in tap water has been on monitoring data collected over - months periods and that we averaged over entire water system. analytical data revealed that total trihalomethanes levels were higher in the summer: mean ± standard deviation . ± . lg/l (micrograms/litre). these organic compounds were present in the end of distribution networks ( . ± . lg/l). it is noted that, sometimes, we found high concentrations of chloroform exceeding the sanitary norm ( lg/l) in tap water (maximum value . lg/l). results of sampling programs showed stronger correlations between chlorine and trihalomethanes value (correlation coefficient r = . to . , credible %interval). in conclusion, the population drank water with the law concentration of trihalomethanes, especially chloroform. abstract objective: to determine the validity of a performance-based assessment of knee function, dynaport[rsymbol] kneetest (dpkt), in first-time consulters with non-traumatic knee complaints in general practice. methods: patients consulting for nontraumatic knee pain in general practice aged years and older were enrolled in the study. at baseline and -months follow-up knee function was assessed by questionnaires and the dpkt; a physical examination was also performed at baseline. hypothesis testing assessed cross-sectional and longitudinal validity of the dpkt. results: a total of patients were included for dpkt of which were available for analysis. the studied population included women ( . %), median age was (range - ) years. at follow-up, patients ( . %) were available for dpkt. only out of ( %) predetermined hypotheses concerning cross-sectional and longitudinal validity were confirmed. comparison of the general practice and secondary care population showed a major difference in baseline characteristics, dynaport knee score, internal consistency and hypotheses confirmation concerning the construct validity. conclusion: the validity of the dpkt could not be demonstrated for first-time consulters with non-traumatic knee complaints in general practice. measurement instruments developed and validated in secondary care are not automatically also valid in primary care setting. abstract although animal studies have described the protective effects of dietary factors supplemented before radiation exposure, little is known about the lifestyle effects after radiation exposure on radiation damage and cancer risks in human. the purpose of this study is to clarify whether lifestyle can modify the effects of radiation exposure on cancer risk. a cohort of , japanese atomic-bomb survivors, for whom radiation dose estimates were currently available, had their lifestyle assessed in . they were followed during years for cancer incidence. the combined effect of smoking, drinking, diet and radiation exposure on cancer risk was examined in additive and multiplicative models. combined effects of a diet rich in fruit and vegetables and ionizing radiation exposure resulted in a lower cancer risk as compared to those with a diet poor in fruit and vegetables and exposed to radiation. similarly, those exposed to radiation and who never drink alcohol or never smoke tobacco presented a lower oesophagus cancer risk than those exposed to radiation and who currently drink alcohol or smoke tobacco. there was no evidence to reject either the additive or the multiplicative model. a healthy lifestyle seems beneficial to persons exposed to radiation in reducing their cancer risks. abstract background: clinical trials have shown significant reduction in major adverse cardiac events (mace) following implantation of sirolimus-eluting (ses) vs. bare-metal stents (bms) for coronary artery disease (cad). objective: to evaluate long-term clinical outcomes and economic implications of ses vs. bms in usual care. methods: in this prospective intervention study, cad patients were treated with bms or ses (sequential control design). standardized patient and physician questionnaires , , and months following implantation documented mace, disease-related costs and patient quality of life (qol). results: patients treated with ses (mean age ± , % male), with bms (mean age ± , % male). there were no significant baseline differences in cardiovascular risk factors and severity of cad. after months, % ses vs. % bms patients had suffered mace (p< . ). initial hospital costs were higher with ses than with bms, but respective month follow-up direct and indirect costs were lower ( , ± vs. , ± euro and ± vs. , ± euro, p = ns). overall, disease-related costs were similar in both groups (ses , ± , bms , ± , p = ns) . differences in qol were not significant. conclusions: as in clinical trials, ses patients experienced significantly fewer mace than bms patients during -month follow-up with similar overall costs and qol. abstract background: meta-analyses that use individual patient data (ipd-ma) rather than published data have been proposed as an improvement in subgroup-analyses. objective: to study ) whether and how often ipd-ma are used to perform subgroup-analyses ) whether the methodology used for subgroup-analyses differs between ipd-ma and meta-analyses of published data (map) methods: ipd-ma were identified in pubmed. related article search was used to identify map on the same objective. metaanalyses not performing subgroup-analysis were excluded from further analyses. differences between ipd-ma and map were analysed, reasons for discrepancies were described. we recently developed a simple diagnostic rule (including history and physical findings plus d-dimer assay results) to safely exclude the presence of deep vein thrombosis (dvt) without the need for referral in primary care patients suspected of dvt. when applied to new patients, the performance of any (diagnostic or prognostic) prediction rule tends to be lower than expected based on the original study results. therefore, rules need to be tested on their generalizability. the aim was to determine the generalizability of the rule. in this cross-sectional study, primary care patients with suspicion of dvt were prospectively identified. the rule items were obtained from each patient plus ultrasonography as reference standard. the accuracy of the rule was quantified on its discriminative performance, sensitivity, specificity, negative predictive value, and negative likelihood ratio, with accompanying % confidence interval. dvt could be safely excluded in % ( % in the original study) of the patients, without referral. none of these patients had dvt ( . % in the derivation population). in conclusion, the rule appears to be a safe diagnostic tool for excluding dvt in patients suspected of dvt in primary care. abstract background: long-term exposure to very low concentrations of asbestos in the environment and relation to incidence of mesothelioma contributes to insight into the dose-response relationship and public health policy. aim: to describe regional differences in the occurrence mesothelioma in the netherlands in relation to the occurrence in the asbestos polluted area around goor and to determine whether the increased incidence of pleural mesothelioma among women in this area could be attributed to environmental exposure to asbestos. methods: mesothelioma cases were selected in the period - from the netherlands cancer register (n = ). for the women in the region goor (n = ) exposure to asbestos due to occupation, household or environment was verified from the medical files, the general practitioner and next-of-kin for cases. results: in goor the incidence of pleural mesothelioma among women was -fold increased compared with the netherlands and among men fold. of the additional cases among women, cases were attributed to the environmental asbestos pollution and in cases this was the most likely cause. the average cumulative asbestos exposure was estimated at . fiber-years. . temporal trends and gender differences were investigated by random slope analysis. variance was expressed using median odds ratio (mor). results: ohcs appeared to be more relevant than administrative areas for understanding physicians' propensity to follow prescription guidelines (mor_ohc = . and mor_aa = . ). conclusion and discussion: as expected, the intervention increased prevalence and decreased variance, but at the end of the observation period practice variation remained high. these results may reflect inefficient therapeutic traditions, and suggest that more intensive interventions may be necessary to promote rational statin prescription. abstract background: mortality rates in ska˚ne, sweden have decreased in recent years. if this decline has been similar for different geographical areas have not been examined closely. objectives: we wanted to illustrate trends and geographical inequities in all cause mortality between the municipalities in ska˚ne, sweden from to . we also aimed to explore the application of multilevel regression analysis (mlra) in our study, since it is a relatively new methodology when describing mortality rates. design and methods: we used linear mlra with years at the first level and municipalities at the second to model direct age-standardized rates. temporal trends were examined by random slope analysis. variance across time was expressed using intra-class correlation (icc). results: the municipality level was very relevant for understanding temporal differences in mortality rates (icc = %). in average, mortality decreased by / Ù along the study period but this trend varied considerably between municipalities, geographical inequalities along the years were u-shaped with lowest variance in the s (var = ). conclusion: mortality has decreased in ska˚ne but municipality differences are increasing again. mlra is a useful technique for modelling mortality trends and variation among geographical areas. abstract background: ozone has adverse health effects but it is not clear who is most susceptible. objective: identification of individuals with increased ozone susceptibility. methods: daily visits for lower respiratory symptoms (lrs) in general practitioner (gp) offices in the north of the netherlands ( - , patients) were related to daily ozone levels in summer. ozone effects were estimated for patients with asthma, copd, atopic dermatitis, and cardiovascular diseases (cvd) and compared to effects in patients without these diseases. generalized additive models adjusting for trend, weekday, temperature, and pollen counts were used. results: the mean daily number of lrs-visits in summer in the gp-offices varied from . to . . mean (sd) -hour maximum ozone level was . ( . ) lg/m . rrs ( % ci) for a lg/m increase (from th to th percentile) in the mean of lag to of ozone for patients with/ without disease are: abstract asthma is a costly health condition, its economic effect is greater than that estimated for aids and tuberculosis together. following global initiative for asthma recommendations that require more data about the burden of asthma, we have determined the cost of this illness from - . an epidemiological approach based on population studies was made to estimate global as well as direct and indirect costs. data were obtained mainly from the national health ministry database, the national statistics institute of spain and the national health survey. the costs were averaged and adjusted to e. we have found a global burden (including private medicine) of million e. indirect and direct costs account for , and , %.the largest components within direct costs were pharmaceutical ( . %), primary health care systems ( , %), hospital admissions ( . %) and hospital non-emergency ambulatory visits ( . %). within indirect costs, total cessation of work days ( . %), permanent labour incapacity ( . %) and early mortality ( . %) costs were the main components. pharmaceutical cost is the first component as in most studies from developed countries, followed by primary health care systems unlike some reports that consider hospital admissions in second place. finally, direct costs represent . % of the total health care expenditure. abstract background: it is well known that fair phenotypical characteristics are a risk factor for cutaneous melanoma. the aim of our study was to investigate the analogous associations between phenotypical characteristics and uveal melanoma. design/methods: in our casecontrol study we compared incident uveal melamom patients with population controls to evaluate the role of phenotypical characteristics like iris-, hair-and skin color and other risk factors in the pathogenesis of this tumor. a total of patients and controls matched on sex, age and region were interviewed. conditional logistic regression was used to calculate odds ratio (or) and % confidence intervals ( % ci). results: risk of uveal melanoma was increased among people with light iris color (or = . % ci . - . ) and light skin color was slightly associated with an increased risk of uveal melanoma (or . % . - . ). hair color, tanning ability, burning tendency and freckles as a child showed no increased risk. results of the combined analysis of eye-and hair color, burning tendency and freckles showed that only light iris color was clearly associated with uveal melanoma risk. conclusion: among potential phenotypical risk factors only light iris-and skin color were identified as risk factor for uveal melanoma. abstract background: between-study variation in estimates of the risk of hcv mother-to-child transmission (mtct) and associated risk factors may be due to methodological differences or changes in population characteristics over time. objective: to investigate the effect of sample size and time on risk factors for mtct of hcv. design and methods: heterogeneity was assessed before pooling data. logistic regression estimated odds ratios for risk factors. results: the three studies included mother-child pairs born between and , born between and , and between and . there was no evidence of heterogeneity of the estimates for maternal hcv/hiv co-infection and mode of delivery (q = . , p = . and q = . , p = . , respectively). in pooled analysis the proportion of hcv/hiv co-infected mothers significantly decreased from % before to % since (p< . ). the pooled adjusted odds ratios for maternal hcv/hiv co-infection and elective caesarean section delivery were . ( %ci . - . ), p< . and . ( %ci . - . ), p = . respectively. there was no evidence that the effect of risk factors for mtct changed over time. conclusion: although certain risk factors have become less common, their effect on mtct of hcv has not changed substantially over time. abstract background: the need to gain insight into prevailing eating pattrerns and their health effects is evident. objective: to identify dietary patterns and their relationship with total mortality in dutch older women. methods: principal components analysis on food groups was used to identify dietary patterns among , women ( - y) included in the dutch epic-elderly cohort (follow-up $ . y). mortality ratios for three major principle components were assessed using cox proportional hazard analysis. results: the most relevant principal components were a 'mediterranean-like' pattern (high in vegetable oils, pasta/rice, sauces, fish, and wine), a 'traditional dutch dinner' pattern (high in meat, potatoes, vegetables, and alcoholic beverages) and a 'healthy traditional' pattern (high in vegetables, fruit, non-alcoholic drinks, dairy products, and potatoes). in , person years deaths occurred. independent of age, education, and other life style factors only the 'healthy traditional' pattern score was associated with a lower mortality rate, women in the highest tertile experienced a percent reduced mortality risk. conclusion: from this study a healthy traditional dutch diet, rather than a mediterranean diet, appears beneficial for longevity and feasible for health promotion. this diet is comparable to other reported 'healthy' or 'prudent' diets that have been shown to be protective. parents of (aged - ) and (aged - ) children were sent a questionnaire, as were adolescents (aged - ). to assess validity, generic outcome instruments were included (infant toddler quality of life questionnaire (itqol) or the child health questionnaire (chq) and the euroqol- d). response rate was - %. internal consistency of hobq and boq-scales was good (cronbach's alpha's > . in all but two scales). test-retest results showed no differences in - % of scales. high correlations between hobq-and boq-scales and conceptually equivalent generic outcome instruments were found. the majority of hobq ( / ) and boq scales ( / ) showed significant differences between children with a long versus short length of stay. the dutch hobq and boq can be used to evaluate functional outcome after burns in children. the study estimated caesarean section rates and odds ratios for caesarean section in association with maternal characteristics in both public and private sectors; and maternal mortality associated with mode of delivery in the public sector, adjusted for hypertension, other disorders, problems and complications, as well as maternal age. results: the caesarean section rate was . % in the public sector, and . % in the private sector. the odd ratio for caesarean section was . ( %ci: . - . ) for women with or more years of education. the odd ratio for maternal mortality associated with caesarean section in the public sector was . ( %ci: . - . ). conclusion and discussion: sao paulo presented high caesarean section rates. caesarean section compared to vaginal delivery in the public sector presented higher risk for mortality even when adjusted for hypertension, other disorders, problems and complications, as well as maternal age. we show that serious bias in questionnaires can be revealed by bland-altman methods but may remain undetected by correlation coefficients. we refute the argument that correlation coefficients properly investigate whether questionnaires rank subjects sufficiently well. conclusions: the commonly used correlation approach can yield misleading conclusions in validation studies. a more frequent and proper use of the bland-altman methods would be desirable to improve epidemiological data quality. abstract screening performance relies on quality and efficiency of protocols and guidelines for screening and follow-up. evidence of low attendance rates, over-screening of young women and low smear specificity gathered by the early 's in the dutch cervical cancer screening program called for an improvement. several protocols and guidelines were redefined in , with emphasis on assuring that these would be adhered to. we assessed improvement since by changes in various indicators: coverage rates, follow-up compliance and number of smears. information on all cervix uteri tests in the netherlands registered until st march was retrieved from the nationwide registry of histo-and cytopathology (palga). five-year coverage rate in the age group - years rose to %. the percentage of screened women in follow-up decreased from % to %. fourteen percent more women with abnormal smears were followed-up, and the time spent in follow-up decreased. a % decrease in the annual number of smears made was observed, especially among young women. in conclusion, the changes in protocols and guidelines, and their implementation have increased coverage and efficiency of screening, and decreased the screening-induced negative side effects. similar measures can be used to improve other mass screening programmes. abstract background: it is common knowledge that in low endemic countries the main transmission route of hepatitis b infection is sexual contact, while in high endemic regions it is perinatal transmission and horizontal household transmission in early childhood. objectives: to get insight into what determines the main transmission route in different regions. design and methods: we used a formula for the basic reproduction number r for hepatitis b in a population stratified by age and sexual activity to investigate under which conditions r > . using data extracted from the literature we investigated how r depends on fertility rates, rates of horizontal childhood transmission and sexual partner change rates. results: we identified conditions on the mean offspring number and the transmission probabilities for which perinatal and horizontal childhood transmission alone ensures that r > . those transmission routes are then dominant, because of the high probability for children to become chronic carriers. sexual transmission dominates if fertility is too low to be the driving force of transmission. conclusion: in regions with high fertility rates hepatitis b can establish itself on a high level of prevalence driven by perinatal and horizontal childhood transmission. therefore, demographic changes can influence hepatitis b transmission routes. abstract background: the artificial oestrogen diethylstilboestrol is known to be fetotoxic. thus, intrauterine exposure to other artificial sex hormones may increase the risk of fetal death. objective: to study if use of oral contraceptive months prior to or during pregnancy is associated to an increased risk of fetal death. design and methods: a cohort study of pregnant women who were recruited into the danish national birth cohort during the years - and interviewed about exposures during pregnancy, either during the first part of their pregnancy (n = ) or following a fetal loss (n = ). cox regression analyses with delayed entry were used to estimate the risk of fetal death. results: in total ( . %) women took oral contraceptives during pregnancy. use of combined oestrogen and progesterone oral contraceptives (coc) or progesterone only oral contraceptives (poc) during pregnancy were not associated with increased hazard ratios of fetal death compared to non-users, hr . ( % ci . - . ) and hr . ( % ci . - . ) respectively. neither use of coc nor poc prior to pregnancy was associated with fetal death. conclusion: use of oral contraceptive months prior to conception or during pregnancy is not related to an increased risk of fetal death. abstract background: few studies have been performed to assess if water fluoridation reduces social inequalities among groups of different socioeconomic status, and none of them was conducted in developing countries. objectives: to assess socioeconomic differences between brazilians towns with and without water fluoridation, and to compare dental caries indices among socioeconomic strata in fluoridated and non-fluoridated areas. design and methods: a countrywide survey of oral health performed in - and comprising , children aged years provided information about dental caries indices in brazilian towns. socioeconomic indices, the coverage and the fluoride status of the water supply network of participating towns were also appraised. multivariate regression models were performed. inequalities in dental outcomes were compared in towns with and without fluoridated tap water. results: better-off towns tended to present a higher coverage by the water supply network, and were more inclined to add fluoride. fluoridated tap water was associated with an overall improved profile of caries, concurrent with an expressively larger inequality in the distribution of dental disease. conclusion: suppressing inequalities in the distribution of dental caries requires an expanded access to fluoridated tap water; a strategy that can be effective to foster further reductions in caries indices. objective: to investigate the role of family socioeconomic trajectories from childhood to adolescence on dental caries and associated behavioural factors. design and methods: a population-based birth cohort was carried out in pelotas, brazil. a sample (n= ) of the population of subjects born in were dentally examined and interviewed at aged . dental caries index, care index, toothbrushing, flossing, and pattern of utilization of dental services were the outcomes. these measures were compared among four different family income trajectories. results: adolescents who were always poor showed, in general, a worse dental caries profile, whilst adolescents who never were poor had a better dental caries profile. adolescents who had moved from poverty in childhood to nonpoverty in adolescence and those who had moved from non-poverty in childhood to poverty in adolescence had similar dental profiles to those who were always poor except for pattern of utilization of dental services which was higher in the first group. conclusion: poverty in at least one stage of the lifespan has a harmful effect on dental caries, oral behaviours and utilization of dental services. we assessed contextual and individual determinants of dental caries in the brazilian context. a country-wide survey of oral health informed the dental status of , twelve-year-old schoolchildren living in towns in . a multilevel model fitted the adjustment of untreated caries prevalence to individual (socio-demographic characteristics of examined children) and contextual (geographic characteristics of participating towns) covariates. being black (or = . ; % ci: . - . ), living in rural areas (or = . ; . - . ) and studying in public schools (or = . ; . - . ) increased the odds of having untreated decayed teeth. the multilevel model identified the fluoride status of water supplies (ß=) . ), the proportion of households linked to the water network (ß=) . ) and the human development index (ß=) . ) as town-level covariates of caries experience. better-off brazilian regions presented an improved profile of dental health, besides having a less unequal distribution of dental treatment needs between blacks and whites, rural and urban areas, and public and private schools. dental caries experience is prone to socio-demographic and geographic inequalities. monitoring contrasts in dental health outcomes is relevant for programming socially appropriate interventions aimed both at overall improvements and at the targeting of resources for groups of population presenting higher levels of needs. abstract background: ultraviolet radiation (uvr) is the main cause of nonmelanoma skin cancer but has been hypothesised to protect against development of prostate cancer (pc). if this is true, skin cancer patients should have lower pc incidence than the general population. objectives: to study the incidence of pc after a diagnosis of skin cancer. design methods: using the eindhoven cancer registry, a cohort of male skin cancer patients diagnosed since ( squamous cell carcinoma (scc), basal cell carcinoma (bcc) and melanoma (cm)) was followed up for incidence of invasive pc. observed incidence rates of pc amongst skin cancer patients were compared to those in the reference population, resulting in standardised incidence ratios (sir). results: scc (sir . ( %ci: . ; . )) and bcc (sir . ( %ci: . ; . )) showed a decreased incidence of pc, cm did not. patients with bccs occurring in the chronically sun-exposed head and neck area (sir . ( %ci: . ; . ) had significantly lower pc incidence rates. conclusion discussion: although numbers of scc and cm were too small to obtain unequivocal results, this study partly supports the hypothesis that uvr protects against pc and also illustrate that cm patients are different from nmsc patients in several aspects. abstract introduction: hypo-and hyperthyroidism have been associated to various symptoms and metabolic dysfunctions in men and women. incidences of these diseases have been estimated in a cohort of middle-aged adults in france. methods: the su.vi.max (sup-ple´mentation en vitamines et mine´raux antioxydants) cohort study included volunteers followed-up for eight years since - . the incidence of hypo-and hyperthyroidism was estimated retrospectively from scheduled questionnaires and the data transmitted by the subjects during their follow-up. factors associated to incident cases have been identified by cox proportional hazards models. results: among the subjects free of thyroid dysfunction at inclusion, incident cases were identified. after an average follow-up of . years, the incidence of hyper-and hypothyroidism was . % in men, . % in - year old women, and . % in - year old women. no associated factor was identified in men. in women, age and alcohol consumption (> grams/day) increased the risk of hypo-or hyperthyroidism, while a high urinary thiocyanate level in - would be a protective factor. conclusion: the incidences of hypo and hyperthyroidism observed in our study as well as the associated risk factors found are in agreement with the data of studies performed in other countries. abstract background: lung cancer is the most frequent malignant neoplasm world-wide. in , the number of new lung cancer cases was estimated at . million, which makes over % of all new cases of neoplasm registered all round the globe. it is also the leading cause of cancer deaths. objective: the objective of this paper is to provide a systematic review of life-related factors for lung cancer risk. methods: data sources were medline from january to december , title in the field. search terms included: lung cancer, tobacco smoke, education, diet, alcohol consumption or physical activity terms. book chapters, monographs, relevant news reports, and web material were also reviewed to find articles. results: the results of the literature review suggest that smoking is a major, unquestionable factor of lung cancer risk. exposure to environmental tobacco smoke (ets) and education could also play a role in the occurrence of the disease. diet, alcohol consumption and physical activity level are other important but less extended determinants of lung cancer. conclusions: effective prevention programs against some of the life style-related factors for lung cancer, especially against smoking must be developed to minimize potential health risks and prevent the future cost of health. stedendriehoek twente and south (n = ), additional data (co-morbidity, complications after surgery and follow-up) were gathered. cox-regression analyses were used. results: the proportion resections declined from % of patients < to % of patients aged > = years, whereas primary radiotherapy increased from % to %. in the two regions patients ( %) underwent resection. co-morbid conditions did not influence the choice of the therapy. % had complications. postoperative mortality was %. in multivariate analysis, only treatment had an independent effect.two year survival was % for patients undergoing surgical resection and % for those receiving radiotherapy (p< . ). conclusion: number of co-morbid conditions did not influence choice of treatment, postoperative complications, and survival in patients with nsclc > = years. the epidemiology of oesophageal cancer has changed in recent decades. the incidence has increased sharply, mainly comprising men, adenocarcinoma and tumours of the lower third of the oesophagus. the eurocare study suggested large variation in survival between european countries, primarily related to early mortality. to study potential explanations, we compared data from the rotterdam and thames cancer registry. computer records from , patients diagnosed with oesophageal cancer in the period - were analysed by age, gender, histological type, tumour subsite, period and region. there was a large variation in resection rates between the two regions, % for rotterdam versus % for thames (p< . ). resection rates were higher for men, younger patients, adenocarcinoma and distal tumours. postoperative mortality (pom) was defined as death within days of surgery and was . % on average. pom increased with age from . % for patients younger than years to . % for patients older than years. pom was significantly lower in high-volume hospitals (> operations per year), . % versus . % (p< . ). this study shows a large variation in treatment practice between the netherlands and the united kingdom. potential explanations will need to be studied in detail. abstract russia has experienced tremendous decline in life expectancy after break up of the ussr. surprisingly, im has also been decreasing. less is known on the structure of im in different regions of russia. the official im data may be underestimated partly due to misreporting early neonatal deaths (end) as stillbirths (sb). end/sb ratio considerably exceeding : indicates misreporting. we present the trends and structure of im in arkhangelsk oblast (ao), north-west russia from to as obtained from the regional statistical committee. im decreased from . to . per live births. cause-specific death rates (per , ) decreased from to . for infectious diseases, from to for respiratory causes, from to for traumas, from to for inborn abnormalities but did not change for conditions of the perinatal period ( in both and ) . the end/sb ratio increased from to . . in , im from infections and respiratory causes in the ao are much lower than in russia in general. the degree of misreporting end as sb in the ao is lower than in russia in general. other potential sources of underestimation of im in russia will be discussed. abstract background: epidemiological studies that investigated malocclusion and different physical aspects in adolescents are rare in the literature. objective: we studied the impact of malocclusion on adolescents' self-image regardless of other physical aspects. design and methods: a cross-sectional study nested in a cohort study was carried out, in pelotas, brazil. a random sample of yearsold adolescents was selected. the world health organization ( ) criteria were used to define malocclusion. interviews about self-reported skin colour and appearance satisfaction were administered. the body mass index was calculated. gender, birth weight and socioeconomic characteristics were obtained from the birth phase of the cohort study. poisson regression models were performed. results: the prevalence of moderate or severe malocclusion was . % [ %ci . ; . ] in the whole sample without significant difference between boys and girls. a higher statistically significant difference of appearance dissatisfaction was identified in girls ( . %) than in boys ( . %). a positive association between malocclusion and appearance dissatisfaction was observed only in girls, after adjusting for other physical and socioeconomic characteristics. conclusions: malocclusion influenced appearance dissatisfaction only in young women. abstract background: factors for healthy aging with good functional capacity and those which increase the risk of death and disability need to be identified. objectives: we studied the prevalence of low functional capacity and its associations in a small city in southern brazil. design and methods: a population based cross sectional study was carried out with a random sample size of elderly people. a home-applied questionnaire including socioeconomic, demographic, house conditions, socioeconomic self-perception characteristics was applied. the low functional capacity was defined as the difficulty in the performance of or more activities or inability to carry out of those activities according to scale proposed by rikli and jones. descriptive statistics, association using chi-square test as well as the multiple logistic regression analysis were performed. abstract introduction: assessment of trichiuriasis spatial distribution is important to evaluate sanitation conditions. our objective was to identify risk areas for the trichuris trichiura infection. methods: cross sectional study was held in census tracts of duque de caxias county, rio de janeiro, brazil. collection and analysis of fecal specimens and a standardize questionnaire were carry out in order to evaluate socio-economic and sanitation conditions in a sample of , children between and years old. geoestatistics techniques were used to identify risk areas for trichiuriasis. results: the mean age of the studied population was . years old, which % were females and % were males. the prevalence of trichuris trichiura in the sample was %. children whose mothers studied for years or less had odds ratio (or) = . than children whose mothers studied for more than years old. children who were living in houses without water supply had or = . comparing to children living in houses with water supply. the spatial analysis identified risk areas for infection. conclusion: the results show association between socio-economic conditions and the proliferation of trichuris trichiura infection. the identification of risk areas can guide efficient actions to combat the disease. abstract background: refuge life and diabetes mellitus can affect the healthrelated quality of life (hrqol). objective: to assess how both aspects influence hrqol of the diabetic refugees in gaza strip. methods: overall subjects filled a self-administered questionnaire including world health organization quality of life questionnaire (whoqol-bref) and some socio-demographic information. the sample consisted of three frequency matched groups for gender and sex, each. first group were refugees with diabetes mellitus, second refugees without diabetes and third diabetes patients with no refugee history. the response rate was % on average. global score consisting of all four domains of who-qol-bref was dichotomized by the value of and logistic regression was used for the analysis. results: crude odds ratios (or) for lower quality of life were . ( % ci . - . ) for diabetes refugees compared to diabetes non-refugees and . ( . - . ) compared to non-diabetes refugees. after adjusting for age, gender, education, employment, income status and number of persons depending on the respondents or was . ( . - . ) and . ( . - . ), respectively. additionally, adjusting for length of diabetes and complications reduced the or to . ( . - . ) for diabetes refugees compared to diabetes non-refugees. conclusion: quality of life is highly reduced in refugees with diabetes. abstract background: pesticides have a significant public health benefit by increasing food production productivity and decreasing diseases. on the other hand, public concern has been raised about the potential health effects of the exposure to pesticides on the developing fetus and child. objectives: to review the available literature to find an epidemiological studies dealing with the exposure to pesticides and children health. design and methods: epidemiological studies were identified during search of the literature basis. following health effects were taking into account: adverse reproductive and developmental disorders, childhood cancer, neurodevelopmental effects and the role of pesticides as endocrine disrupters. results: pesticides were associated with wide range of reproductive disorders. the association between exposure to pesticides and the risk of childhood cancer and neurodevelopmental effects was found in several studies. epidemiological studies have been limited by luck of specific pesticide exposure, exposure based on job title, small size of examined groups. conclusions: in the light of existing although still limited evidence of adverse effects of pesticide exposure it is necessary to reduce the exposure. the literature review suggests a great need to increase awareness of people who are occupationally or environmentally exposed to pesticides about its potential negative influence on their children. in order to match local health policy more with the needs of citizens, the municipal health service utrecht started the project 'demand-orientated prevention policy'. one of the aims was to explore the needs of the utrecht citizens. the local health survey from contained questions about needs of information and support with regard to disorders and lifestyle. do these questions about needs give other results compared to questions about prevalence of health problems? in total utrecht citizens aged to years returned the health questionnaire (response rate %). most needs were observed on subjects concerning overweight and mental problems, and were higher among women, moroccans, turks, low educated people and citizens of deprived areas. the prevalence of disorders and unhealthy lifestyles did not correlate well to the needs (majority correlation coefficients: < , ). most striking, of the utrecht population % were smokers and % excessive alcohol drinkers, while needs related to these topics were low. furthermore, higher needs among specific groups did not always correspond to higher prevalences of related health problems in these groups. these results show the importance of including questions about needs in a health survey, because they add additional information to questions about prevalences. abstract background: recent studies associated statin therapy with better outcome in patients with pneumonia. because of an increased risk of pneumonia in patients with diabetes we aimed to assess the effects of statin use on pneumonia occurrence in diabetic patients managed in primary care. methods: we performed a case-control study nested in , patients with diabetes. cases were defined as patients with a diagnosis of pneumonia. for each case, up to controls were matched by age, gender, practice, and index date. patients were classified as current statin user when the index date was between the start and end date of statin therapy. results: statins were currently used in . % of , cases and in . % of , controls (crude or: . , % ci . - . ). after adjusting for potential confounders, statin therapy was associated with a % reduction in pneumonia risk (adjusted or: . , % ci . - . ). the association was consistent among relevant subgroups (stroke, heart failure, and pulmonary diseases) and independent of age or use of other prescription drugs. conclusions: use of statins was significantly associated with reduced pneumonia risk in diabetic patients and may apart from lipid lowering properties be useful in prevention of respiratory infections. abstract introduction: cigarette smoking is the most important risk factor for copd development. therefore, smoking cessation is the best preventive measure. aim: to determine the beneficial effect of smoking cessation on copd development. methods: incidence of copd (gold stage > = ) was studied in smokers without copd who quitted or continued smoking during yr of followup. we performed logistic regression analyses on pairs of observations. correlations within a subject and time, and time between successive surveys were taken into account. abstract objectives: to describe the prevalence and severity of dental caries in adolescents of the city of porto, portugal, and to assess socioeconomic and behavioral covariates of dental caries experience. methods: a sample of thirteen-year-old schoolchildren underwent dental examination. results from the dental examination were linked to anthropometric information and to data supplied by two structured questionnaires assessing nutritional factors, sociodemographic characteristics and behaviors related to health promotion. dental caries was appraised in terms of the dmft index, and two dichotomous outcomes, one assessing the prevalence of dental caries (dmft = ); the other assessing the prevalence of a high level of dental caries (dmft = ). results: consuming soft drinks derived from cola two or more times per week, attending a public school, being girl and having parents with low educational attainment were identified as risk factors both for having dental caries and for having a high level of dental caries. conclusion: the improvement of oral health status in the portuguese context demands the implementation of polices to reduce the frequency of sugar intake, and could benefit from an overall and longstanding expansion of education in society. abstract background: migrant mortality does not conform to a single pattern of convergence towards rates in the host population. to better understand how migrant mortality will develop, there is a need to further investigate how the underlying behavioural determinants change following migration. objective: we studied whether behavioural risk factors among two generations of migrants converge towards the behaviour in the host population. design and methods: cross-sectional interview-data were used including moroccan and turkish migrants, aged - . questions were asked about smoking, alcohol consumption, physical inactivity and weight/height. age-adjusted prevalence rates among first and second generation migrants were compared with prevalence rates in the host population. results: converging trends were found for smoking, physical inactivity and overweight. for example, we found a higher prevalence of physical inactivity in first generation turkish women as compared to ethnic dutch (or = . ( . - . )), whereas among second generation no differences were found (or = . ( . - . )). however, this trend was not found in all subgroups. additionally, alcohol consumption remained low in all subgroups and did not converge. conclusion and discussion: behavioural risk factors in two generations of migrants seem to converge towards the prevalence rates in the host population. although, some groups and risk factors showed a deviant pattern. abstract background/relevance: arm-neck-shoulder complaints are common in general practice. for referral in these complaints, only guidelines exist for shoulder complaints and epicondylitis. besides, other factors can be important. objective: what factors are associated with referral to physiotherapy or specialist in non-traumatic armneck-shoulder complaints in general practice, during the first consultation? design/methods: general practitioners (gps) recruited consulters with new arm, neck or shoulder complaints. data on complaint-, patient-, gp-characteristics and management were collected. the diagnosis was categorised into: shoulder specific, epicondylitis, other specific or non-specific. multilevel analyses (adjustment for treating gp) were executed in procgenmod to assess associated variables (p< . ). results: during the first consultation, % was referred for physiotherapy and % for specialist care. indicators of reference to physiotherapy were: long duration of complaint, recurrent complaint and gp located in a little/not urbanised area. while having shoulder specific or other specific diagnoses was negatively associated. indicators of reference to specialist care were: having other specific diagnosis, long duration of complaint, musculoskeletal co-morbidity, functional limitations and consulting a less experienced gp. conclusion/discussion: most referrals were to physiotherapy and only a minority to specialist care. mainly diagnosis and other complaint variables indicate on 'who goes where'. besides gp-characteristics can play a role. abstract background: the ruhr area has for years been a synonym for a me´gapolis of heavy industry with a high population density. presently, % of the population of the state of north rhine-westphalia live there, i.e. more than five million people. objectives: for the first time, social and health indicators of nrw's health indicator set were brought together for this me´gapolis area. design and methods: new standard tables were constructed for the central area of 'ruhr-city' including seven cities with more than inhabitants/km and the peripheral zone with eight districts and cities. for the pilot phase, four socio-demographic and four health indicators were recalculated. comparability of the figures was achieved by age standardization. the results obtained were submitted to a significance test by identifying % confidence intervals. results: the centre of 'ruhr-city' is characterised by elderly, unemployed, foreign, low-income citizens living closely together. infant mortality lies above nrw's average, male life expectancy is . years lower and female life expectancy . years lower than life expectancy in nrw (without 'ruhr-city'). several avoidable deaths' rate in the ruhr area are significantly higher than the average in nrw. specific intervention strategies are required to improve the health status in 'ruhr-city'. abstract background: general practitioners (gps) have a fundamental role to play in tobacco control, since they reach a high percentage of the target population. objectives: to evaluate specific strategies to enhance promotion of smoking cessation in general practice. design and methods in a cluster-randomized trial, medical practices were randomized following a · factorial design. patients aged - years who smoked at least cigarettes per day (irrespective of their intention to stop smoking) were recruited. the intervention included (ti) the provision of a two-hour physician group training in smoking cessation methods plus direct physician payments for every participant not smoking months after recruitment; and (tm) provision of the same training plus direct participant reimbursements for pharmacy costs associated with nicotine replacement therapy or bupropion treatment. results: in the mixed logistic regression model, no effect was identified for intervention ti (odds ratio (or) = . , % confidence interval (ci) . - . ), but intervention tm strongly increased the odds of cessation (or = . , % ci . - . ). conclusion and discussion: the cost-free provision of effective medication along with improved training opportunities for gps may be an effective measure to enhance smoking cessation promotion in general practice. in europe, little research on international comparison of health surveys has been accomplished, despite a growing interest in this field. smoking prevalence is chosen to explore data comparability. we aim to illustrate methodological problems encountered when comparing data from health surveys and investigate international variations in smoking behaviour. we examined a sample . individuals aged and more, from six european health surveys performed in - . problems met during the comparisons are described. we took the example of current smoking as an indicator allowing a valid comparison of the prevalences. the differences in age and sex distribution between countries were adjusted through direct standardisation. additionally, multivariate analysis will assess variations in current smoking between countries, when controlling for sex, age, and educational level. methodological problems concern comparability of socioeconomic variables. the percentage of current smokers varies from % to %. smoking patterns observed by age groups, sexes and educational level are similar, although rates per country differ. further results will determine if the variations in smoking related to socioeconomic status are alike. this international comparison of health surveys highlights methodological problems encountered when comparing data of several countries. furthermore, variations in smoking may call for adaptations in public health programs. from research it appears that adolescent alcohol use in the achterhoek is much higher than in the rest of the netherlands and rapidly increasing. excessive alcohol use has consequences for health and society. parents play an important role in preventing excessive adolescent alcohol use, but are not aware of the problem and consequences. for this reasons the municipalities in the achterhoek launch an alcohol moderation programme, starting with a regional media campaign to increase problem awareness among parents. the objective of this study is to assess the impact of this media campaign in the achterhoek. three successive independent cross-sectional telephone surveys, interviewing approximately respondents each, will be conducted before, during and after the campaign. respondents will be questioned on knowledge and awareness of excessive adolescent alcohol use, its consequences and the role child raising can play. also the reach and appreciation of the different activities of the campaign will be investigated. results: of the surveys before and during the implementation will be known by may . with these first findings the unawareness of the problem among parents and partly the reach and appreciation of the campaign can be assessed. abstract background: obesity is a growing problem, increasingly so in children and adolescents. overweight is partly 'programmed' during pregnancy, but few comprehensive studies looked prospectively into the changes of body composition and metabolic factors from birth. objectives: the aim of the population-based birth-cohort study within gecko is to study the etiology and prognosis of overweight and the metabolic syndrome during childhood. design and methods: the gecko drenthe will be a population based observational birth-cohort study, which includes all children born from april to april in drenthe, one of the northern provinces of the netherlands. during the first year of life, the study includes repeated questionnaires, extensive anthropometric measurements and blood measurements at birth (cord blood) and at the age of eleven months. results: the number of babies born in the drenthe province is about . per year. the results from a feasibility study conducted in february will be presented. conclusion: gecko drenthe is a unique project that will contribute to the understanding of the development of obesity in childhood and its tracking into adulthood. this will enable early identification of children at risk and opens the way for timely and tailored preventive interventions. abstract background: tunisia is facing an epidemiologic transition with the extension of chronic diseases that share common risk factors. obesity is a leading risk factor and happens to occur frequently in early life. objective: to study the prevalence and the risk factors of obesity and overweight among urban schoolchildren in sousse, tunisia. methods: cross sectional study of a tunisian sample of schoolchildren aged between and years living in the urban area of sousse, tunisia. a representative sample of school children selected by multistage cluster sampling procedure. measurements: weight and height, blood pressure measured by electronic system, fasting blood lipids. questionnaire assessment was used for family history of cardiovascular disease, smoking habits, physical activity and diet. abstract background: quality of life (qol) measurements are acknowledged as very important in the evaluation of health care. objectives: we studied the validity and the reliability of the hungarian version of the whoqol-bref among people living in small settlements. method: a questionnaire-based cross-sectional study was conducted in a representative sample (n = ) of persons aged years and over in south-east-hungary, in . data were analysed by the spss . . the internal consistency was evaluated using cronbach's alpha; for comparison of the qol scores amongst the various groups the two-tailed t-tests were used; convergent validity was assessed by spearman coefficients. results: the male:female ratio was . to . %, and the average age . (sd: . ) years. the domain scores were . (sd: . ) for the physical, . (sd: . ) for psychological, . (sd: . ) for the social, and . (sd: . ) for the environment domains. the cronbach's alpha values ranged from . to . across domains. the whoqol-bref seemed to be suitable to distinguish healthy and unhealthy people. the scores for all domains correlated with the self-evaluated health, and overall quality of life (p< . ). conclusion: our study supported that the whoqol-bref provided a valid, reasonable and useful determination of the qol of people living in hungarian villages. abstract background: further than a cardiovascular disease, arterial hypertension (aht) is the main cardiovascular risk factor. in spain, the aht prevalence reaches %, placed in the third position after germany and finland in affecters percentage. although its high morbi-mortality, the aht is a forecast factor. the treatment's objective (pharmacological and life style modifications) of hypertensive patients is not only to reduce blood pressure levels to optimum levels but also to treat all modifiable vascular risk factors. objective: economic impact evaluation of direct costs due to aht pathology (cie -mc - ) in spain in , according to autonomous region. design and methods: descriptive and transversal study of costs estimation in the period between january to december in spain according to autonomous region. the study is based on data available from the national health ministry database and the national statistics institute of spain. results: the national health service assigned million e to aht treatment. , % of the total cost is owe to pharmaceutical service expenses, , % to primary health care and a , % to hospital admissions. conclusion and discussion: the costs generated by aht are mainly due to the pharmaceutical service. the costs distribution is modified according to the geographical region. abstract background: over the last decades, for low-stage cervical cancer less surgical treatment and for high-stage cervical cancer chemoradiotherapy was recommended in the national guidelines. objectives: to describe changes and variation in treatment and survival in cervical cancer in the regions of the comprehensive cancer centre stedendriehoek twente (cccst) and south (cccs) in the netherlands. design and methods. newly diagnosed cervical cancer cases were selected from both cancer registries in the period - . patient characteristics, tumour characteristics, treatment and follow-up data were collected from the medical records. results: in figo stages ia -ib the percentage hysterectomy decreased from % in - to % in - (p<. ) and survival improved comparing - with - . figo stages iii-ivb had mostly received radiotherapy only ( %). no differences in survival between years of diagnosis were found. in the cccs-region more chemoradiotherapy was given in these stages ( % versus % in the cccst-region in the whole period). conclusion and discussion:. abstract background: the reason for the increased prevalence of depression in type diabetes (dm ) is unknown. objective: we investigated whether depression is associated with metabolic dysregulation or that depression is rather a consequence of having dm . methods: baseline data of the utrecht health project were used. subjects with cardiovascular disease were excluded. , subjects (age: . +/) ) were classified into four mutually exclusive categories: normal fasting plasma glucose (fpg < . mmol/l), impaired fpg (> = . and < . mmol/l), undiagnosed dm (fpg > = . mmol/l), and diagnosed dm . depression was defined as either a score of or more on the depression subscale of the symptom check list- or use of antidepressants. results: subjects with impaired fasting glucose and undiagnosed dm had no increased prevalence of depression. diagnosed dm patients had an increased prevalence of depression (or = . ( . - . )) after adjustment for gender, age, body mass index, smoking, alcohol consumption, physical activity, education level and number of chronic diseases. conclusions: our findings suggest that depression is not related to disturbed glucose homeostasis. the increased risk of depression in diagnosed dm only, suggests that depression is rather a consequence of the psychosocial burden of diabetes. abstract background: breast-conserving surgery (bcs) followed by radiotherapy (bcs-rt) is a safe treatment option for the large majority of patients with tumours less than cm. aim: the use of bcs and bcs-rt in pt (? cm) and pt -tumours ( - cm) was investigated in the netherlands in the period and . methods: from the netherlands cancer registry patients were selected with invasive pt (? . cm) or pt ( . - . cm) tumours, without metastasis at time of diagnosis. trends in the use of bcs and rt after bcs were determined for different age groups and regions. results: in the period - , pt -tumours and , pt -tumours were diagnosed. the %bcs in pt -tumours increased in all age groups. it remained lowest in patients years and older ( % in ). in pt -tumours a decrease was observed in patients years and older (from % to % in ). in both pt and pt tumours the %bcs-rt increased in patients years and older to respectively % and %. between regions and hospitals large differences were seen in %bcs and %bcs-rt. conclusion: multidisciplinary treatment planning, based on specific guidelines, and patient education could increase the use of bcs combined with rt in all age groups. abstract this is a follow-up study on the adverse health effects associated with pesticide exposure among cut-flower farmers. survey questionnaires and detailed physical and laboratory examinations were administered to and respondents, respectively, to determine pesticide exposure, work and safety practices, and cholinesterase levels. results showed that pesticide application was the most frequent activity associated with pesticide exposure, and entry was mostly ocular and dermal. majority of those exposed were symptomatic. on physical examination, or . % of those examined were found to have abnormal peak expiratory flow rate (pefr). eighty-two percent had abnormal temperature, followed by abnormal general survey findings (e.g. cardiorespiratory distress). % had cholinesterase levels below the mean value of . ? ph/hour, and . % exhibited a more than % depression in the level of rbc cholinesterase. certain hematological parameters were also abnormal, namely hemoglobin, hematocrit, and eosinophil count. using pearson's r, factors strongly associated with illness due to pesticides include using a contaminated piece of fabric to wipe sweat off (p.= . ) and reusing pesticide containers to store water (p.= . ). the greatest adverse effect of those exposed is an abnormal cholinesterase level which confirms earlier studies on the effect of pesticides on the body. objectives: this pair-study was performed to find out the rate of spontaneous abortions in female workers exposed to organic solvents from the wood-processing industry. methods: the level of organic solvents was assessed within the workplaces during a year period. exposed female workers from the wood-processing industry were examined. the occupational and non-occupational data associated with their fertility were obtained by applying a standard epidemiological computed questionnaire. the reference group consisted of female workers non-exposed to hypo-fertilizing agents, residing in the same locality. the rate of spontaneous abortions was evaluated in both groups as an epidemiological fertility indicator. results: within the studied period, the organic solvents levels exceeded several time the maximal admissible concentrations in all workplaces. the long-term exposure to organic solvents caused a significant increase in rate of spontaneous abortions compared to the reference group (p< . ). the majority of abortions ( %) have happened in the first trimester of pregnancy. conclusions: the long-term exposure to organic solvents may cause low fertility on female workers because of the spontaneous abortions. it is advised to reduce the organic solvents level in the air of all workplaces, as well as to stop working the pregnant women in exposure to organic solvents. abstract introduction: rio de janeiro city (rj) presents a fast aging of the population with changes in morbi-mortality. cardiovascular diseases are the first cause of death in elderly population. more than a half of ischemic heart diseases (ihd) cases occur in aged people (> years old). objective: describe the spatial distribution of ihd mortality in the elderly population in rj and associations with socio-demographics variables. methods: data were gathered from information on mortality system of the ministry of health and the demographic census of the foundation of the brazilian institute for geography and statistics. the geographic distributions of the standardized coefficient of mortality due to ihd and socio-demographics variables, by districts, in were analyzed in arcgis . . spatial autocorrelation of ihd was assessed by the moran and geary indices. a conditional autoregressive model was used to evaluate the association between idh and socio-demographics variables. results: association between idh mortality and income, educational level, family type and to possess computer, videocassette and microwave was found. conclusion: spatial analysis of the idh mortality and socio-demographics factors influence are fundamental to subsidize more efficient public policies in sense to prevention and control of this important injury of health. abstract purpose: to evaluate the prognostic impact of isolated loco-regional recurrences on metastatic progression among women treated for invasive stage i or ii breast cancer (within phase iii trials concerning the optimal management of breast cancer). patients and methods: the study population consisted of , women primary surgically treated for early stage breast cancer, enrolled in eortc trials , , or , by breast conservation ( %) and mastectomy ( %) with long time of follow-up (median: . range: . - . ). data were analysed in a multi-state model by using multivariate cox regression models, including a state-dependent covariate. results: after the incidence of the loco-regional recurrence, a positive nodal status at baseline is a significant prognostic risk factor for distant metastases. the effects of the young ages at diagnosis and larger tumour size, become less significant after the incidence of loco-regional recurrences. the presence of a locoregional recurrence in itself is a significant prognostic risk factor for distant metastases after loco-regional recurrences. the effect of the time to the loco-regional recurrence is not a significant prognostic factor. conclusion: the presence of local recurrence is an important risk factor for outcome in patients with early breast cancer. abstract background: the relationship between the antral follicles and ovarian reserve tests (ort) to determine ovarian response in ivf is extensively studied. we studied the role of follicle size distribution in the response on the various orts in a large group of subfertile patients. methods: in a prospective cohort study, female patients were included if they had regular ovulatory cycles, subfertility for > months, > = ovary and > = patent ovarian tube. antral follicles were counted by ultrasound and blood was collected for fsh, including a clomiphene challenge test (ccct), inhibin b, and estradiol before and after administration of puregon [rsymbol] . (efort test). statistical analysis was performed using spss . for windows. results: of eligible patients, participated. mean age was . years and mean duration of subfertility was . months. age, baseline fsh, ccct and efort correlated with the number of small follicles ( - mm) but not with large follicles ( - mm). regression analysis confirmed that the number of small follicles and average follicle size contributed to ovarian response after correction for age, while large follicles did not. conclusion: small antral follicles are responsible for the hormonal response in ort and may be suitable to predict ovarian response in ivf. abstract background: dengue epidemics account annually for several million cases and deaths worldwide. the high endemic level of dengue fever and its hemorrhagic form (dhf) correlates to extensive house infestation by aedes aegypti and multiple viral serotype human infection. objective: to describe dengue incidence evolutionary patterns and spatial distribution in brazil. methods: it is a review study that analyzed serial case reports registered since until . results: it was shown that defined epidemic waves followed the introduction of every serotype (den to ) , and reduction in susceptible people possibly responded for downward case frequency. conclusions and discussion: an incremental expansion of affected areas and increasing occurrence of dhf with high lethality were noted in recent years. in contrast, efforts based solely on chemical vectorial combat have been insufficient. moreover, some evidence demonstrated that educational action do not permanently modify population habits. in this regard it was stated that while vaccine is not available, further dengue control would depend on potential results gathered from basic interdisciplinary research and intervention evaluation studies, integrating environmental changes, community participation and education, epidemiological and virological surveillance, and strategic technological innovations aimed to stop transmission. abstract background: patient participation in treatment decisions can have positive effects on patient satisfaction, compliance and health outcomes. objectives: study objectives were to examine attitudes regarding participation in decision-making among psoriasis patients and to evaluate the effect of a decision-aid for discussing treatment options. methods: a 'quasi experiment' was conducted in a large dermatological hospital in italy: a questionnaire evaluating the decision-making process and knowledge on treatments was selfcompleted by a consecutive sample of psoriasis patients after routine clinical practice and by a second sample of patients exposed to a decision-board. results: in routine clinical practice . % of patients wanted to be involved in treatment decisions, . % wanted to leave decisions entirely to the doctor and . % preferred making decisions alone. . % and . % of the control and decision-board group had good knowledge level. at multivariate analysis good knowledge on treatments increased the likelihood of preferring an active role (or = . ; %ci . - . ; p = . ). the decision-board only marginally improved patient knowledge and doctor-patient communication. conclusion and discussion: in conclusion, large proportions of patients want to participate in decision-making, but insufficient knowledge can represent a barrier. further research is needed for developing effective instruments for improving patient knowledge and participation. abstract background: the only available means of controlling infections caused by the dengue virus is the elimination of its principal urban vector (aedes aegypti). brazil has been implementing programs to fight the mosquito; however, since the 's the geographic range of infestation has been expanding steadily, resulting in increased circulation of the virus. objective: to evaluate the effectiveness of the dengue control actions that have been implemented in the city of salvador. methods: prospective design, serologic inquiries were made in a sample population of residents of urban 'sentinel areas'.the seroprevalence and one year seroincidence of dengue are estimated and the relationship between intensity of viral circulation and the standards of living and vector density is analysed. results: there were high overall seroprevalence ( . %) and seroincidence ( . %) for the circulating serotypes (denv- and denv- ). the effectiveness of control measures appears to be low, and although a preventable fraction of . % was found, the incidence of infections in these areas was still very high ( . %). conclusions and discussions: it is necessary to revise the technical and operational strategies of the infection control program in order to attain infestation levels that are low enough to interrupt the circulation of the dengue virus. this study investigates the difference in cancer mortality risks between migrant groups and the native dutch population, and determines the extent of convergence of cancer mortality risks according to migrants' generation, age at migration and duration of residence. data were obtained from the national population & mortality registries in the period - ( person years, cancer deaths). we used poisson regression to compare the cancer mortality rates of migrants originating from turkey, morocco, surinam, netherlands antilles/aruba to the rates for the native dutch. results: all-cancer mortality among all migrant groups combined was significantly lower compared to the native dutch population (rr= . ci: . - . ). mortality rates for all cancers combined were higher among nd generation migrants, among those with younger age at migration, and those with longer duration of residence. this effect was particularly pronounced in lung cancer and colorectal cancer. for most cancers, mortality among nd generation migrants remained lower compared to the native dutch population. surinamese migrants showed the most consistent pattern of convergence of cancer mortality. conclusions: the generally low risk of cancer mortality for migrants showed some degree of convergence but the cancer mortality rates did not yet reach the levels of the native dutch population. abstract background: legionnaires' disease (ld) is a notifiable disease in the netherlands. ld cases are reported to authorities for national surveillance. supplementary, a national ld outbreak detection program (odp) is installed in the netherlands. these two registration systems have their own information exchange process and databases. objectives: surveillance systems are known to suffer from incompleteness of reported data. co-existence of two databases creates the opportunity to investigate accuracy and reliability in a national surveillance system. design and methods: comparison was made between the outcome 'diagnosis by culture' in both databases and physical presence of legionella strains in laboratories for patients. accuracy is described using the parameters sensitivity and correctness. for reliability, cohen's kappa coefficient (?) was applied. results: accuracy and reliability were significantly higher in the odp database, but not optimal in both databases when compared to data in laboratories. the odp database was moderately reliable (? = . ; %ci . - . ), the surveillance database slightly (? = . ; %ci . - . ). conclusion: our findings suggest that diagnostic notification data concerning ld patients are most accurate and reliable when derived directly from diagnostic laboratories. discussion: involvement of data-entry persons in outbreak detection results in higher reliability. unreliable data may have considerable consequences during outbreaks of ld. the aim of the study was to investigate the medical students' plans to emigrate, quantify the scale of migration in the near future and to build a profile of the possible emigrants. data were collected based on anonymous questionnaire delivered to random group of medical students (katowice). we used the binary logistic regression and multivariate analysis to identify the differences between groups preferring to go abroad or stay in poland. % respondents confirmed that considerate the emigration; . % of them declared they are very likely to move and further . % is certain. , % of those considering emigration confirmed having taken practical steps towards moving. binary logistic regression showed no difference between people who were certain or almost certain to go and those who were not considering going for most baseline characteristics: hometown size, socio economic background and having family tradition of the medical profession (p = . ). only marks' mean differentiates between the two groups: . for those who will definitely stay vs . for students who will definitely move (p = . ). the multivariate analysis gave similar results. conclusions: most of the students consider the emigration, but the declarations of will to departure are more frequent among those with the worse marks. abstract background: falls incidence in home resident elderly people varies from % to %. falls induce loss of self-sufficiency and increase mortality and morbidity. objectives: to evaluate falls incidence and risk factors in a group of general practice elderly patients. design : prospective cohort study with year follow-up methods: eight hundreds elderly (> years) were visited by practitioners for a baseline assessment. information on current pathologies and previous falls in the last six months was collected. functional status was evaluated using: short portable mental state questionnaire, geriatric depression scale, activities of daily living (adl), instrumental activities of daily living, total mobility tinetti score. falls were monitored through phone-interviews at and months. data were analyzed through logistic regression. results: twenty-eight percent of the elderly fell in the whole period. sixty percent of falls were not reported to the practitioner. independent predictors for falls were adl score (adl< : or = . ; % ci . - . ) and previous falls (or = . ; % ci . - . ). tinetti score was significantly associated to falls only in univariate analysis. conclusions: practitioners can play a key-role in identifying at-risk subjects and managing prevention interventions. falls monitoring and a continuous practice of comprehensive geriatric assessment should be encouraged. abstract background: oral health represents an important indicator of health status. socio-economic barriers to oral care among elderly are considerable. in the lazio region, a public health program for oral rehabilitation was implemented to offer dentures to elderly people with social security. objectives: to compare hospitalisation between elderly enrolled in the program and a control group. design and methods: for each elderly enrolled in the program living in rome, three controls, matched for sex and age, were selected from rome municipality register. hospital admissions in the two-year period before enrollment were traced by record-linkage with the hospital discharge register. results: totally, , admissions occurred. the annual admissions rate was per elderly among controls and in the program group (incidence rate ratio: irr = . ; % ci . - . ). when comparing diagnosis-specific rates, significant excesses were observed in the program group for respiratory diseases ( abstract background: herpes simplex virus (hsv) type and are important viral sexually transmitted diseases (sti) and can cause significant morbidity. in the netherlands, data about prevalences in the general population are hampered. objective: description of the seroprevalences of hsv- and hsv- and associated factors in the netherlands. design and methods: a population based serum bank survey in the netherlands with an age-stratified sample was used ( ) ( ) . antibodies against hsv- and hsv- were determined using elisa. a questionnaire was used to get information on demographics and risk factors. a logistic regression adjusting for age and full multiple regression were done to establish risk factors. results: questionnaires and sera were available for persons. both hsv- and hsv- seroprevalence increased with age. seroprevalence of hsv- was . % and was amongst others associated with female sex and being divorced. seroprevalence of hsv- was . % and was amongst others associated with being divorced and a history of sti. conclusion: seroprevalence is higher in certain groups like teenagers, women, divorced people and those with a history of sti. prevention should be focused on those groups. more research is needed on prevention methods, which can be used in the netherlands, like screening or vaccination. abstract background: frequently, statistically significant prognostic factors are reported with suggestions that patient management should be modified. however, the clinical relevance of such factors is rarely quantified. objectives: we evaluated the accuracy of predicting the need for invasive treatment among bph patients managed conservatively with alpha -blockers. methods: information on eight prognostic factors was collected from patients treated with alpha -blockers. with phm regression coefficients a risk score for retreatment was calculated for each patient. the analyses were repeated on groups of patients sampled from the original case series. these bootstrap results were compared to the original results. results: three significant predictors of retreatment were identified. the % of patients with the highest risk score had an -month risk of retreatment of only %. analyses of less than half of all the bootstrap samples resulted in the same three significant prognostic factors. the % of patients with the highest risk score in each of the samples experienced a highly variable risk of retreatment of % to %. conclusions: four of the five high risk patients would be overtreated with a modified policy. internal validation procedures may warn against the invalid translation of statistical significance into clinical relevance. background: e-cadherin expression is frequently lost in human epithelium derived cancers, including bladder cancer. for two genetic polymorphisms in the region of the e-cadherin gene (cdh ) promoter, a reduced transcription has been reported: a c/a single nucleotide polymorphism (snp) and a g/ga snp at bp and bp, respectively, upstream of the transcriptional start site. objective: we studied the association between both polymorphisms and the risk of bladder cancer. methods: patients with bladder cancer and population controls were genotyped for the ) c/ a and the ) g/ga promoter polymorphisms using pcr-rflp. results: a significantly increased risk for bladder cancer was found for a allele carriers compared to the homozygous c allele carriers (or . ; % ci: . - . ). the risk for the heterozygous and homozygous a allele carriers, was increased approximately . and -fold, respectively. the association was stronger for more aggressive tumors. we did not find any association between the ) g/ga snp and bladder cancer. conclusion: this study indicates that the ) c/a snp in the e-cadherin gene promoter is a low-penetrance susceptibility factor for bladder cancer. background: health problems, whether somatic, psychiatric or accident-related cluster within persons. the study of allostatic load as a unifying theme (salut) aims to identify risk factors that are shared by different pathologies and that could explain this clustering. studying patients with repetitive injuries might be helpful to identify risk factors that are shared by accident-related and other health problems. objectives: to study injury characteristics in repetitive injury (ri) patients as compared to single injury (si) patients. methods: the presented study included ri patients and si patients. medical records provided information about injury characteristics and patients were asked for possible causes and context. results: ri patients suffered significantly more from contusions than si patients ( % vs %). regarding the context, si patients were significantly more injured in traffic ( % vs %). in both groups most injuries were attributed to 'mere bad luck' (ri %, si %), closely followed by 'clumsiness or inattention' (ri %, si %). ri patients pointed out aggression or substance misuse significantly more often than si patients ( % vs %). conclusion: ri patients seem to have more 'at risk' behavior (i.e. aggression, impulsivity), which will increase their risk for psychiatric health problems. abstract background: breastfeeding may have a protective effect on infant eczema. bias as a result of methodological problems may explain the controversial scientific evidence. objective: we studied the association between duration of breastfeeding and eczema when taking into account the possible influence of reverse causation. design and methods: information on breastfeeding, determinants and outcomes at age one year was collected by repeated questionnaires in mother infant-pairs participating in the koala study ( cases of eczema). to avoid reverse causation, a periodspecific-analysis was performed in which only 'at risk' infants were considered. results: no statistically significant association between the duration of breastfeeding (> weeks versus formula feeding) and the risk of eczema in the first year was found (or . %ci . - . ). after excluding from the analysis all breastfed infants with symptoms of eczema reported in the same period as breastfeeding, also no statistical significant association was found for the duration of breastfeeding and eczema between and months (or . %ci . - . ). conclusion and discussion: in conclusion, no evidence was found for a protective effect of breastfeeding duration on eczema. this conclusion was strengthened by risk period-specific-analysis which made the influence of reverse causation unlikely. abstract background: the internet can be used to meet health information needs, provide social support, and deliver health services. the anonymity of the internet offers benefits for people with mental health problems, who often feel stigmatized when seeking help from traditional sources. objectives: to identify the prevalence of internet use for physical and mental health information among the uk population. to investigate the relationship of internet use with current psychological status. to identify the relative importance of the internet as a source of mental health information. design and methods: self-completion questionnaire survey of a random sample of the uk population (n = ). questions included demographic characteristics, health status (general health questionnaire), and use of the internet and other information sources. results: % of internet users had sought health information online, and % had sought mental health information. use was higher among those with current psychological problems. only % of respondents identified the internet as one of the most accurate sources of mental health information, compared with % who identified it as one of the sources they would use. conclusions: health service providers must recognise the increasing use of the internet in healthcare, even though it is not always regarded as being accurate. abstract old age is a significant risk factor for falls. approximately % of people older than are falling at least once a year, mostly in the own homes. resulting hip fractures cause at least partial immobility in - % of the affected persons. almost % are sent to nursing homes afterwards. in mecklenburg-west pomerania, ageing of the population proceeds particularly fast. to prevent falls and the loss of independent living a falls prevention module was integrated in a community-bbased study conducted in cooperation with a general practitioner (gp). in the patients homes' a trained nurse performed a test to estimate the falls risk of each patient and a consultation how to reduce risk, e.g. eye sight check, gymnastic exercise. in the feasibility study ( %) out of patients (average age years), agreed to a visiting of each room of their homes in search for tripping dangers. the evaluation was assisted by a standardized, computer-based documentation. the prevention module received a considerable acceptance despite the extensive home visiting. within one month the patients started to transfer advice into practice. during the first follow up visits of the nurse three patients reported e.g. to have started gymnastics and/or wear stable shoes. abstract background: the emergence of drug resistant m. tuberculosis (mtb) is an increasing problem in both developed and developing countries. objectives: investigation of isoniazid (inh) and rifampin (rif) susceptibility patterns among mtb isolates from patients. design and methods: in total sputum samples were collected. smears were prepared for acid fast staining and all the isolates were identified as m. tuberculosis by preliminary cultural and biochemical tests. the isolates were examined for inh and rifampin resistance using conventional mic method and pcr technique by using specific inh (kat g) and rifampin (rpo b) resistant primers. results: seven isolates were resistant to both inh and rifampin by mic method. in pcr technique, and out of above mentioned strains showed resistant to inh and rifampin respectively. coclusion: the epidemiology of drug resistance is . % in region of study which is significant. discussion: conventional mic method despite being time consuming is more sensitive for evaluation of drug resistance, however, pcr as a rapid and sensitive technique is recommended additionally to conventional method for having quicker results to start treatment and disease control management. abstract background and objectives: we studied in literature which design characteristics of food frequency questionnaires (ffqs) influence their validity to assess both absolute and relative levels of energy intake in adults with western food habits, and to rank them according to these intakes. this information is required in harmonizing ffqs for multi centre studies. design and methods: we performed a review of studies investigating the validity or reproducibility of ffqs, published since . the included studies validated ffqs against doubly labeled water (for energy expenditure) as a gold standard, or against food records or hour recalls for assessing relative validity (for energy intake). the design characteristics we studied were the number of food items, the reference period, the administration mode, and inclusion of portion size questions. results: and conclusion: for this review we included articles representing the validation of questionnaires. three questionnaires were validated against dlw, ten against urinary n and against -hour recalls or food records. in conclusion a positive linear relationship (r = . , p< . ) was observed between the number of items on the ffq and the reported mean energy intake. details about the influence of other design characteristics on validity will be discussed at the conference. abstract background: high ethanol intake may increase the risk of lung cancer. objectives: to examine the association of ethanol intake with lung cancer in epic. design and methods: information on baseline and past alcohol consumption, lifetime tobacco smoking, diet, and anthropometrics of , participants was collected between and . cox proportional hazard regression was used to examine the association of ethanol intake at recruitment ( cases) and mean lifelong ethanol intake ( cases) with lung cancer. results: non-consumers at recruitment had a higher lung cancer risk than low consumers ( . - . g/day) [hr = . , % ci . - . ]. lung cancer risk was lower for moderate ethanol intake at recruitment ( . - . g/day) compared with low intake (hr = . , % ci . - . ); no association was seen for higher intake. compared with lifelong low consumers, lifelong non-consumers did not have a higher lung cancer risk (hr = . , % ci . , . ) but lifelong moderate consumers had a lower risk (hr = . , % ci: . - . ). lung cancer risk tended to increase with increasing lifelong ethanol intake (= vs. . - . g/ day hr = . , % ci: . - . ). conclusion: while lung cancer risk was lower for moderate compared with low ethanol intake in this study, high lifelong ethanol intake might increase the risk. abstract background: one of the hypotheses to explain the increasing prevalence of atopic diseases (eczema, allergy and asthma) is imbalance between dietary intake of omega- and omega- fatty acids. objectives: we evaluated the role of perinatal fatty acid (fa) supply from mother to child in the early development of atopy. design and methods: fa composition of breast milk was used as a marker of maternal fa intake and placental and lactational fa supply. breast milk was sampled months postpartum from mother-infant pairs in the koala birth cohort study, the netherlands. the infants were followed for atopic symptoms (repeated questionnaires on eczema and wheezing) and sensitisation at age (specific serum ige against major allergens). multivariate logistic regression analysis was used to adjust for confounding factors. results: high levels of omega- long chain polyunsaturated fas were associated with lower incidence of eczema in the first year (odds ratio for the highest vs lowest quintile . , % confidence interval . - . ; trend over quintiles p = . ). wheeze and sensitisation were not associated with breast milk fa composition. conclusion and discussion: the results support the omega- / hypothesis. we suggest that anti-inflammatory activity of omega- eicosanoid mediators is involved but not allergic sensitisation. abstract background: acute myocardial infarction (ami) is among the main causes of death in italy and is characterized by high fatality associated with a fast course of the disease. consequently timeliness and appropriateness of the first treatment are paramount for a positive recovery. objectives: investigate the differences among italian regions of ami first treatment and in-hospital deaths. design and methods: following the theoretical care pathway (from onset of ami to hospitalization and recovery or death), regional in-hospital deaths are decomposed into the contributions of attack rate, hospitalization and in-hospital fatality. hospital discharges, death and population data are provided by the official statistics. results: generally in northern and central regions there is an excess of observed in-hospital deaths, while the opposite occurs in southern regions. conclusion: in northern and central regions the decomposition method suggests a more frequent and severe illness, generally accompanied by a higher availability of hospitals; exceptions are lombardia and lazio, where some inefficiencies in the hospital system are highlighted. in most southern regions the decomposition confirms a less frequent and less severe illness; exceptions are campania and sicilia, where only the less severe patients reach the hospital and then recover, while the others die before reaching the hospital. abstract background: atherosclerotic lesions have typical histological and histochemical compositions at different stages of their natural history. the more advanced atherosclerotic lesions contain calcification. objective: we examined the prevalence of and associations between calcification in the coronary arteries, aortic arch and carotid arteries assessed by multislice computed tomography (msct). methods: this study was part of the rotterdam study,a population-based study of subjects aged years and over. calcification was measured and quantified in subjects. correlation coefficients were computed using spearman's correlation coefficients. results: the prevalence of calcification increased with age throughout the vascular bed. in subjects aged and over, up to % of men had calcification in the coronary arteries and up to % of women had calcification in the aortic arch. in men, the strongest correlation was found between calcification in the aortic arch and the carotid arteries (r= . , p< . ). in women, the relations were somewhat lower, the strongest correlation was found between calcification in the coronary arteries and the carotid arteries (r = . , p< . ). conclusion and discussion: in conclusion, the prevalence of calcification was generally high and increased with increasing age. the study confirms the presence of strong correlations between atherosclerosis in different vessel beds. abstract background: health status deteriorates with age and can be affected by transition from active work to retirement. objective: to assess the effect of retirement on age related deterioration of health. methods: secondary analysis of the german health survey (bundesgesundheitssurvey ) and california health interview survey (chis ) . subjective health was assessed by a single question regarding respondent's health status from = excellent to = poor. locally weighted regression was used for exploratory analysis and b-splines for the effect estimation in regression models. results: subjective health decreased in an obviously non-linear manner with age. in both cases the decrease could be reasonably approximated by two linear segments, however the pattern was different in the german and california sample. in germany, the change point of the slope describing deterioration of health was located at . abstract objective: to assess the effectiveness of physiotherapy compared to general practitioners' care alone, in patients with acute sciatica. design, setting and patients: a randomised clinical trial in primary care with a -months follow-up period. patients with acute sciatica (recruited - ) were randomised in two groups: ) intervention group received physiotherapy (active exercises), and ) control group received general practitioners' care only. main outcome measures the primary outcome was patients' global perceived effect. secondary outcomes were severity of leg and back pain, severity of disability, general health and absence from work. the outcomes were measured at , , and weeks after randomisation. results: at months follow-up, % of the intervention group and % of the control group reported improvement (rr . ; % ci . to . ). at months follow-up, % of the intervention group and % of the control group reported improvement (rr . ; % ci . ; . ). no significant differences in secondary outcomes were found at short-term or long-term follow-up. conclusion: at months follow-up, evidence was found that physiotherapy added to general practitioners' care is more effective in the treatment of patients with acute sciatica than general practitioners' care alone. abstract background: only little is known about the epidemiology of skin melanoma in the baltic states. objectives: the aim of this study was to provide insights into the epidemiology of skin melanoma in lithuania by analyzing population-based incidence and mortality ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) time trends and relative survival based on skin melanoma. methods: we calculated age-standardized incidence and mortality rates (cases per , ) using the european standard population and calculated period estimates of relative survival. for the period - , % of all registered cases were checked by reviews of the medical charts. results: about % of the cases of the period - were reported to the cancer registry indicating a high quality of cancer registration of skin melanoma in lithuania. the incidence rates increased from (men: . , women: . ) to (men: . , women: . ). mortality rates increased from (men: . , women: . ) to (men: . , women: . ). relative -year relative survival rates among men were % lower than among women. the overall difference in survival is mainly due to a more favorable survival among women aged - years. conclusions: overall prognosis is less favorable among men most likely due to diagnoses at later stages. abstract background: only little is known about the epidemiology of skin melanoma in the baltic states. objectives: the aim of this study was to provide insights into the epidemiology of skin melanoma in lithuania by analyzing population-based incidence and mortality ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) time trends and relative survival based on skin melanoma. methods: we calculated age-standardized incidence and mortality rates (cases per , ) using the european standard population and calculated period estimates of relative survival. for the period - , % of all registered cases were checked by reviews of the medical charts. results: about % of the cases of the period - were reported to the cancer registry indicating a high quality of cancer registration of skin melanoma in lithuania. the incidence rates increased from (men: . , women: . ) to (men: . , women: . ). mortality rates increased from (men: . , women: . ) to (men: . , women: . ). relative -year relative survival rates among men were % lower than among women. the overall difference in survival is mainly due to a more favorable survival among women aged - years. conclusions: overall prognosis is less favorable among men most likely due to diagnoses at later stages. abstract background: multifactorial diseases share many risk factors, genetic as well as environmental. to investigate the unresolved issues on etiology of and individual susceptibility to multifactorial diseases, the research focus must move from single determinantoutcome relations to modification of universal risk factors. objectives: the aim of the lifelines project is to study universal risk factors and their modifiers for multifactorial diseases. modifiers can be categorized into factors that determine the effect of the studied risk factor (eg gen-expression), those that determine the expression of the studied outcome (eg previous disease), and generic factors that determine the baseline risk for multifactorial diseases (eg age). design and methods: lifelines is carried out in a representative sample of . participants from the northern provinces of the netherlands. apart from questionnaires and clinical measurements, a biobank is constructed (blood, urine, dna). lifelines will employ a three-generation family design (proband design with relatives), which has statistical advantages, enables unique possibilities to study social characteristics, and offers practical benefits. conclusion: lifelines will contribute to the understanding of how disease-overriding risk factors are modified to influence the individual susceptibility to multifactorial diseases, not only at one stage of life but cumulatively over time: the lifeline. abstract background: obesity-related mortality is a major public health problem, but few studies have been conducted on severely obese individuals. objectives: we assessed long-term mortality in treatment-seeking, severely obese persons. design and methods: we enrolled persons in six centres for obesity treatment in four italian regions, with body mass index (bmi) at first visit => kg/m and age => . after exclusion of duplicate registrations and persons with missing personal or clinical data, persons were followed up; as ( . %) could not be traced, persons ( men, women) were retained for analysis. results: there were ( men, women) deaths; the standardized mortality ratios (smrs) and % confidence intervals were ( - ) among men and ( - ) among women. mortality increased with increasing bmi, but the trend was not monotonic in men. lower smrs were observed among persons recruited more recently. excess mortality was inversely related to age attained at follow-up. conclusions and discussion: the harmful, long-term potential of severe obesity we documented confirms observations from studies carried out in different nutritional contexts. the decrease in mortality among most recently recruited persons may reflect better treatment of obesity and of its complications. abstract background: in finland, every cancer patient should have equal access to high quality care provided by the public sector. therefore no regional differences in survival should be observed. objectives: the aim of the study was to find any regional differences in survival, and to elaborate whether possible differences could be explained, e.g., by differences in distributions of prognostic factors. design and methods: the study material consisted of , patients diagnosed in to with cancer at one of the major primary sites. the common closing date was dec. . finland was divided into five university hospital regions. stage, age at diagnosis and sex were used as prognostic factors. the relative survival rates for calendar period window, - , were tabulated using period method and modelled. results: survival differences between the regions were not significant for most primary sites. for some sites, the differences disappeared in the modelling phase after adjusting for the prognostic factors. for a few of the primary sites (e.g., carcinoma of the ovary), regional differences remained after modelling. conclusion: we were able to highlight certain regional survival differences. ways to improve the equity of cancer care will be considered in collaboration with the oncological community. abstract background: the prevalence of cardiovascular disease (cvd) is extremely high in dialysis patients. disordered mineral metabolism, including hyperphosphatemia and hypercalcaemia, contributes to the development of cvd in these patients. objectives: to assess associations between plasma calcium, phosphorus and calciumphosphorus product levels and risk of cvd-related hospitalization in incident dialysis patients. design and methods: in necosad, a prospective multi-centre cohort study in the netherlands, we included consecutive patients new on haemodialysis or peritoneal dialysis between and . risks were estimated using adjusted time-dependent cox regression modeling. results: mean age was ± years, % was male, and % was treated with haemodialysis. cvd was the cause of hospitalization in haemodialysis patients ( % of hospitalizations) and in peritoneal dialysis patients ( %). most common cardiovascular morbidities were peripheral vascular disease and coronary artery disease in both patient groups. in haemodialysis patients risk of cvd-related hospitalization increased with elevated plasma calcium (hazard ratio: . ; % ci: . to . ) and calcium-phosphorus product levels ( . ; % ci: . to . ). in peritoneal dialysis patients, we observed similar effects that were not statistically significant. conclusion: tight control of plasma calcium and calcium-phosphorus product levels might prevent cvd-related hospitalizations in dialysis patients. abstract background: nurses are at health risk due to the nature of their work. analysis of morbidity among nurses was conducted to provide insight concerning the relationship between their occupational exposure and health response. methods: self reported medical history, was collected from an israeli female-nurses cohort (n = , + years old) and their siblings (n = , age matched +/) years) using a structured questionnaire. to compare disease occurrence between the two groups we used chi-square tests and hazard ratio (hr) was calculated by cox-regression to account for age of onset. results: cardiovascular diseases were more frequent among the nurses compared to the controls: heart diseases . % vs. . , p = . (hr= . , p = . ); hypertension . % vs. . %, p<. (hr= . , p = . ). the frequency of hyperlipidemia was . % among the nurses, and only . % among the controls. (hr= . ,p = . ). for the following chronic diseases the occurrence were significantly higher among the nurses and the hrs were significantly higher than : thyroid, hr= . ; liver, hr= . . total cancer and diabetes rates were similar in the groups (hr$ ). conclusions: the results suggest an association between working as a nurse and the existence of risk factors for cardiovascular diseases. the specific related determinants of their work should be further evaluated. abstract background: early referral (er) to a nephrologist and arteriovenous fistulae as first vascular access (va) reduce negative outcomes in chronic dialysis patients (cdp). objectives: to evaluate the effect of nephrologist referral timing and type of the first va on mortality. design and methods: prospective cohort study of incident cdp notified to lazio dialysis registry (italy) in - . late referral (lr) was a patient not referred to nephrologists within months before starting dialysis. we dichotomized va as fistulae versus catheters. to estimate mortality hazard ratios (hr) a multivariate cox model was performed. results: we observed . % lr subjects and . % catheters as first va; proportion of catheters was . % vs. . % (p< . ) for lr and er, respectively. we found a higher mortality hr for patient with a catheter as first va both for er (hr = . ; %c.i. = . - . ) and lr (hr = . ; %c.i. = . - . ); the interaction between referral and va was slight significant (p = . ). conclusions: the originality of our study was to investigate the influence of nephrologist referral timing and va on cdp mortality using a population registry, area-based: we found that a catheter as first va has an independent effect for mortality and modifies the effect of referral timing on this outcome. abstract patients with idiopathic venous thrombosis (vt) without known genetic risk factors but with a positive family history might carry yet unknown genetic defects. to determine the role of unknown hereditary factors in unexplained vt we calculated the risk associated with family history. in the multiple environmental and genetic assessment of risk factors for vt (mega) study, a large population-based case-control study, we collected blood samples and questionnaires on acquired risk factors (surgery, immobilisation, malignancy, pregnancy and hormone use) and family history of patients and control subjects. overall, positive family history was associated with an increased risk of vt (or ( % ci): . ( . - . )), especially in the absence of acquired risk factors (or ( % ci): . ( . - . ) ). among participants without acquired factors but with a positive family history, prothrombotic defects (factor v leiden, prothrombin a, protein c or protein s deficiency) were identified in out of ( %) patients compared to out of ( %) control subjects. after excluding participants with acquired or prothrombotic defects, family history persisted as a risk factor (or ( % ci): . ( . - . )). in conclusion, a substantial fraction of thrombotic events is unexplained. family history remains an important predictor of vt. abstract background: alcohol may have a beneficial effect on coronary heart disease (chd) through elevation of high-density lipoprotein cholesterol (hdlc) or other alterations in blood lipids. data on alcohol consumption and blood lipids in coronary patients are scarce. objectives: to assess whether alcohol consumption and intake of specific types of beverages are associated with blood lipids in older subjects with chd. design and methods: blood lipids (total cholesterol, hdlc, ldl cholesterol, triglycerides) were measured in myocardial infarction patients aged - years ( % male), as part of the alpha omega trial. intake of alcoholic beverages, total ethanol and macro and micronutrients were assessed by food-frequency questionnaire. results: seventy percent of the subjects used lipidlowering medication. mean total cholesterol was . mmol/l and hdlc was . mmol/l. in men but not in women, ethanol intake was positively associated with hdlc (difference of . mmol/l for = g/d vs. g/d, p = . ) after adjustment for diet, lifestyle, and chd risk factors. also, liquor consumption was weakly positively associated with hdlc in men (p = . ). conclusion and discussion: moderate alcohol consumption may elevate hdlc in (treated) myocardial infarction patients. this is probably due to ethanol and not to other beneficial substances in alcoholic beverages. session: posters session : july presentation: poster. abstract objective: early detection and diagnosis of silicosis among dust exposed workers is based mainly on the presence of rounded opacities on radiographs. it is thus important to examine how reliable the radiographic findings are in comparison to pathological findings. methods: a systematic literature search via medline was conducted. validity of silicosis detection and its influence on risk estimation in epidemiology was evaluated in a sensitivity analysis. results: studies on comparison between radiographic and pathological findings of silicosis were identified. the sensitivity of radiographic diagnosis of silicosis (ilo / ) varied between % and %, and specifity between % and %. under the realistic assumption of silicosis prevalence between % and % in dust exposed workers, % to % of silicosis identified may be falsely diagnosed. the sensitivity analysis indicates that invalid diagnostics alone may lead to the finding of an increased risk of lung cancer among patients with silicosis. it may also lead to findings of % to % of radiographic silicosis even when there is no case of silicosis. however, the risk of silicosis could also be underestimated if the prevalence of silicosis exceeds %. conclusion: epidemiological studies based on patients with silicosis should be interpreted with caution. abstract introduction: epidemics of dengue occurring in various countries have stimulated investigators to seek innovative ways of improving current knowledge on the issue. objective: to identify the characteristics of spatial-temporal diffusion of the first dengue epidemic in a major brazilian city (salvador, bahia). methods: notified cases of dengue in salvador in were georeferenced according to census sector (cs) and by epidemiological week. kernel density estimation was used to identify the spatial diffusion pattern. results: of the cs in the city, cases of dengue were registered in ( %). spatial distribution showed that in practically the entire city had been affected by the virus, with a greater concentration of cases in the western region, comprising cs of high population density and predominantly horizontal dwellings. conclusion and discussion: the pattern found showed characteristics of a contagious diffusion process. it was possible to identify the epicenter of the epidemic from which propagation initiated. the speed of progression suggested that even if a rapid intervention was initiated to reduce the vector population, it would probably have little effect in reducing the incidence of the disease. this finding confirms the need for new studies to develop novel technology for prevention of this disease. abstract background: knowing the size of drug user hidden populations in a community is important to plan and evaluate public health interventions. objectives: the aim of this study was to estimate the prevalence of opiate and cocaine users in liguria region by using the covariate capture-recapture method applied to four data sources. methods: we performed a cross-sectional study in the resident population aged - years ( . people at census). during individual cases identified as opiate or cocaine primary users were flagged by four sources (drug dependence services, social service at prefectures, therapeutic communities, hospital discharges). poisson regression models were fitted, adjusting for dependence among sources and for heterogeneity in catchability among categories of the two examined covariates: age ( - and - years) and gender. results: the prevalence of opiate or cocaine users was , % ( % c.i., , - , %) and , % ( % c.i.= , - , %) respectively. conclusions: the estimated prevalence of opiate and cocaine users is consistent with that found in inner london: . % and . % respectively (hickman m., ; hope v.d., ) . the covariate capture-recapture method applied to four data sources allowed identifying a large cocaine-using population and resulted appropriate to determine drug user hidden populations. abstract background: in - we performed a population based diabetes screening programme. objectives: to investigate whether the yield of screening is influenced by gp and practice characteristics. methods: a questionnaire containing items on the gp (age, gender, employment, specialty in diabetes, applying insulin therapy) and the practice (setting, location, number of patients from ethnic minority groups, specific diabetes clinic, involvement of practice assistant and practice nurse in diabetes care, cooperation with a diabetes nurse) was sent to general practitioners (gps) in practices in the southwestern region of the netherlands. multiple linear regression analysis was performed. outcome measure was the ratio screen detected diabetic patients/known diabetic patients per practice (sdm/kdm). results: sdm/kdm was independently associated with higher age of the gp (regression coefficient . ; % confidence interval . to . ), urban location () . ; ) . to ) . ) and involvement of the practice assistant in diabetes care ( . ; . to . ) . conclusion: a lower yield of screening, assumably reflecting a lower prevalence of undiagnosed diabetes, was found in practices of younger gps and in urban practices. a lower yield was not associated with an appropriate practice organization nor with a specialty of the gp in diabetes. session: posters session : july presentation: poster. background: since few years increased incidence rates for childhood cancer were reported from industrialized countries. these findings were discussed controversial, because increases could be caused by changing of potential risk factors. objectives: the question is: are observed increasing rates due to actual changes in incidence rates or mainly caused by changes in registration practice or artefacts? methods: for europe, data from the accis project (pooled data from european population-based cancer registries, performed at iarc, lyon; responsible: e. steliarova-foucher) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) , and for germany, data of the german childhood cancer registry available from onwards were used. results: accis data (based on , cases) show significantly increased data with an overall average annual percentage change of about % and it is seen for mainly all diagnostic subgroups. for germany, increases are seen for neuroblastoma (due to screening programmes) and brain tumours (due to improved registration). for acute leukaemia the observed increase is explained by changes in classification. conclusion and discussion: the increased incidence for europe can only partly be explained by registration artefacts or improved diagnostic methods. the observed patterns suggest that an actual change exists. in germany, from till now the observed increased rates could be explained by artefacts. abstract suicide is the fourth most common cause of death among working age finns. among men socioeconomic status is strongly and inversely associated with suicide mortality, but little is known about socioeconomic differences in female suicide. we studied the direct and indirect effects of different socioeconomic indicators -education, occupation-based social class and income -on suicide among finnish women aged - . also the effect of main economic activity was studied. we used individual level data from the census linked to the death register for the years - . altogether over million person-years were included and suicides were committed. age-adjusted rii conducted using poissonregression model was . ( % ci . - . ) for education, . ( . - . ) for social class and . ( . - . ) for income. however, almost all of the effect of education was mediated by social class. fifteen per cent of social class was explained by education and per cent was mediated by income. the effect of income on suicide was mainly explained by economic activity. in conclusion, net of other indicators occupation-based social class is a strong determinant of socioeconomic differences in female suicide mortality, and actions aimed at preventing female suicide should target this group. abstract c-reactive protein levels (crp) in the range between and mg/ l independently predict the risk of future cardiovascular events. besides being a marker of atherosclerotic processes, high-normal crp levels may also be a sign of a more pronounced response to everyday inflammatory stimuli. the aim of our study is to assess the association between response to everyday stimuli and the risk of myocardial infarction. we will perform a population based case-control study including a total of persons. cases (n = ) are first myocardial infarction (mi) patients. controls (n = ) are partners of the patients. offspring of the mi patients (n = ) are included because disease activity and the use of medication by the mi patients may influence the inflammatory response. in order to assess the inflammatory response in mi patients the mean genetically determined inflammatory response in the offspring will be assessed and used as a measure for the inflammatory response in the mi patients. the offspring is free of disease and medication use. partners of the offspring (n = ) are the controls for the offspring. influvac vaccine will be given to assess crp concentration, i.e. inflammatory response, before and after the vaccination. abstract background:. ischemic heart disease risk may be influenced by long-term exposure to electromagnetic fields (emf) in vulnerable subjects, but epidemiological data is inconsistent. objectives: we studied whether the long-term occupational exposure to emf is related to an increased myocardial infarction (mi) risk. design and methods: we conducted a prospective case-control study, which involved mi cases and controls. emf exposure in cases and controls was assessed subjectively. the effect of emf exposure on mi risk was estimated using multivariate logistic regression. results: after adjustment for age, smoking, blood pressure, body mass index and psychological stress the odds ratios for emf exposure < years was . ; % ci . - . , for emf exposure - years - . ; % ci . - . and for emf exposure > years - . ; % ci . - . . conclusion: longterm occupational exposure to emf may increase the risk of mi. our crude estimates of emf exposure might have impact on excess risk because of nondifferential misclassification in assigning exposure. abstract background: it has been suggested that noise exposure is associated with ischemic heart disease risk, but epidemiological evidence is still limited. objectives: we studied whether road traffic noise exposure increases the risk of myocardial infarction (mi). design and methods: we conducted population-based prospective casecontrol study, which involved mi cases and controls. we measured traffic-related noise levels at the electoral districts and linked these levels to residential addresses. we used multiple logistic regression to assess effect of noise exposure on mi risk. results: after adjustment for age, smoking, blood pressure, body mass index, and psychological stress the risk of mi was higher for the men exposed to - dba ( background: some studies have suggested that patients, depressed following acute myocardial infarction (mi), experience poorer survival. however, i) other studies show no significant association, when adjusted for recognized prognostic indicators and ii) some 'natural' responses to mi may be recorded in questionnaires as indicators of depression. method: depression was assessed in mi patients, by interview on two measures (gwb and sf ) - weeks after discharge, clinical data were abstracted from patients' medical records and vital status was assessed at - years. survivals of depressed, marginally depressed and normal patients were calculated by kaplan meier method and comparisons made by log rank and cox proportional hazard modelling. results: crude survival at years in patients was higher for depressed and marginally depressed ( %) than for normals ( %), although not significantly. in multivariate analysis, four patient characteristics contributed significantly to survival: age (p< . ), previous mi (< . ), diabetes (< . ) and sex (< . ): other potential explanatory variables, including hypertension, infarct severity and depression were excluded by the model. abstract background: the low coronary heart disease (chd) incidence in southern europe could result in lower low density lipoprotein cholesterol oxidation (oxldl). objective. the aim of this study was to compare oxldl levels in chd patients from several european countries. methods: a cross-sectional multicenter study included stable chd male subjects aged to years from northen (finland and sweden), central (germany), and southern europe (greece and spain). lipid peroxidation was determined by plasma oxldl. results: the score of adherence to mediterranean diet, antioxidant intake, alcohol intake, and lipid profile were significantly associated with oxldl. oxldl levels were higher in northern ( . u/l) than in centre ( . u/l) and southern populations ( . u/l), p = . , in the adjusted models. the probability of northern europe to have the highest oxldl levels was . %, and . , % in logarithm of triglyceride-adjusted and fully adjusted models, respectively. the probability of this order to hold after adjustment for country was . %. conclusion: a gradient on lipoperoxidation from north to central and southern europe is very likely to exist, and parallels that observed in the chd mortality, and incidence rates. southern populations may have more favourable environmental factors against oxidation than northern europe. abstract background: whereas socioeconomic status (ses) has been established as a risk factor for a range of adverse health outcomes, little literature exists examining socio-economic inequalities in the prevalence of congenital anomalies. objectives: to investigate the relationship between the ses and the risk of specific congenital anomalies, such as neural tube defects (ntd), oral clefts (oc) and down's syndrome (ds). design and methods: a total of cases of congenital anomaly and non-malformed control births were collected between and from the italian archive of the certificates of delivery care. as a measure of ses, cases and controls were given a value for a level deprivation index. data were analysed using a logistic regression model. results: we found cases of dtn, cases of sof and cases of ds. the risk of having a baby with ntd was significantly higher for women of low ses (or = . ;c.i.: . - . ), as well as for oc (or = . ; c.i.: . - . ). no significant evidence for ses variation was found for ds. conclusion and discussion: our data suggest risk factors linked to ses, such as nutritional factors, lifestyle, and access to health services, may play a role in the occurrence of some malformations. abstract background: general practitioners (gp) are well-regarded by their patients and have the opportunity to play an active role in providing cessation advice. objectives: this study was run to examine whether a public health programme based on a carefully adapted programme of continuing education can increase gps' use of cessation advice and increase the success rates of such advice. methods: the particular context due to a randomization of gp leads us to consider a cluster randomization trial. marginal models, estimated by gee and mixed generalized linear models are used for this type of design. results: the cessation rate is relatively high for all smokers enrolled in the trial (n = ): a total of smokers were ex-smokers at one year ( . %). patients who were seen by trained gps were more likely to successfully stop smoking than those seen by the control gps ( . % vs . %). motivated subjects, aged over , lower had anxiety scores, and confidence in their ability to stop smoking, were predictive of successful cessation at one year follow-up. conclusions: cluster analysis indicated that factors important to successful cessation in this population of smokers are factors commonly found to influence cessation. abstract background: and purpose conventional meta-analysis showed no difference in primary outcome for coronary bypass surgery without (offpump) or with (onpump) the heart-lung machine. secondary outcome parameters such as transfusion requirements or hospitalization days favored offpump surgery. combined individual data analysis improves precision of effect estimates and allows accurate subgroup analyses. objective: our objective is to obtain accurate effect estimates for stroke, myocardial infarction, or death, after offpump versus onpump surgery, by meta-analysis on pooled individual patient data. method and results: bibliographic database search identified eleven large trials (> patients). the obtained data for trials data included patients. primary endpoint was composite (n = ), secondary endpoints were death (n = ), stroke (n = ) and myocardial infarction (n = ). hazard ratio for event-free survival after offpump vs onpump ( % ci) was: composite endpoint . ( . ; . ), death . ( . ; . ) myocardial infarction . ( . ; . ), stroke . ( . ; . ). after stratification for diabetes, gender and age the results slightly favored offpump for high-risk groups. hazard ratios remained not statistically significant. conclusion: no clinical or statistical significant differences were found for any endpoint or subgroup. offpump coronary bypass surgery is at least equal to conventional coronary bypass surgery. offpump surgery therefore is a justifiable option for cardiac surgeons for cardiac bypass surgery. in - an outbreak of pertussis occurred, mostly among vaccinated children. since then the incidence has remained high. therefore, a fifth dose with acellular booster vaccine for -yearolds was introduced in october . the impact of this vaccination on the age-specific pertussis incidence was assessed. mandatory notifications and hospitalisations were analysed for - and compared with previous years. surveillance data show 'epidemic' increases of pertussis in , , and . the total incidence/ , in ( . ) was higher than in the previous epidemic year ( . ). nevertheless, the incidence of notifications and hospitalisations in the age-groups targeted for the booster-vaccination had decreased with respectively % and % compared to . in contrast, the incidence in adolescents and adults almost doubled. unlike other countries that introduced a pre-school booster, the incidence of hospitalised infants < months also decreased ( % compared with ). as expected, the booster-vaccination for -year-olds has decreased the incidence among the target-population itself. more importantly, the decreased incidence among infants < months suggests that transmission from siblings to infants has also decreased. in further exploration of the impact of additional vaccination strategies (such as boostering of adolescents and/or adults) this effect should not be ignored. abstract acute respiratory infections (ari) are responsible for considerable morbidity in the community, but little is known about the presence of respiratory pathogens in asymptomatic individuals. we hypothesized that asymptomatic persons could have a sub clinical infection and so act as a source of transmission. between and all patients with ari who visited their sentinel general practitioner were reported to estimate the incidence of ari in dutch general practices. a random selection of them (cases) and an equal number of asymptomatic persons visiting for other complaints (controls) were included in a case-control study. nose/ throat swabs of participants were tested for a broad range of pathogens. the overall incidence of ari was per , person years, suggesting that in the dutch population an estimated , persons annually consult their general practitioner for respiratory complaints. viruses were detected in % of the cases, ?-haemolytic streptococci group a in % and mixed infections in %. besides, pathogens were detected in approximately % of controls, particularly in the youngest age groups. this study confirms that most ari are viral and supports the reserved policy of prescribing antibiotics. furthermore, we demonstrated that asymptomatic persons might be a neglected source of transmission. abstract background: the baking and flour producing industries in the netherlands agreed on developing a health surveillance system to reduce the burden of and improve prognosis of occupational allergic diseases. objectives: to develop and validate a diagnostic model for sensitization to wheat and fungal amylase allergens, as triage instrument to detect occupational allergic diseases. design and methods: a diagnostic regression model was developed in bakers from a cross-sectional study with ige serology to wheat and or amylase allergens as the reference standard. model calibration was assessed with hoshmer-lemeshow goodness of fit test; discriminative ability using area under receiver operating characteristic curve (auc); and internal validity using bootstrapping procedure. external validation was conducted in other bakers. results: the diagnostic model consisted of four questionnaire items (history of asthma, rhinitis, conjunctivitis, and work-related allergic symptom) showed good calibration (p = . ) and discriminative ability (auc . ; % ci . to . ). internal validity was reasonable (correction factor of . and optimism corrected auc of . ). external validation showed good calibration (p = . ) and discriminative ability (auc . ; % ci . to . ). conclusions and discussion: this easily applicable diagnostic model for sensitization to flour and enzymes shows reasonable diagnostic accuracy and external validation. abstract background: in the netherlands the baking and flour producing industries ( , small bakeries, industrial bakeries, and flour manufactures) agreed to reduce the high rate (up to %) of occupational related allergic diseases. objectives: to conduct health surveillance for early detection of occupational allergic diseases by implementing a diagnostic model as triage instrument. design and methods: in the preparation phase, a validated diagnostic regression model for sensitization to wheat and or a-amylase allergens was converted into score chart for use in occupational health practice. two cut off points of the sum scores were selected based on diagnostic accuracy properties. in the first phase, a questionnaire including the diagnostic predictors from the model was applied in . bakers. surveillance simulation was done in bakers recently enrolled in the surveillance. workers with high questionnaire scores were referred for advanced medical examination. results: implementing the diagnostic questionnaire model yielded %, %, and % bakers in the low, intermediate, and high score groups. workers with high scores showed the highest percentage of occupational allergic diseases. conclusions and discussion: with proper cut off points for referral, the diagnostic model could serve as triage instrument in health surveillance to early detect occupational allergic diseases. abstract background: the prevalence of cardiovascular risk factors in spain is high but myocardial infarction incidence is lower than in other countries. objective: to determine the role of basic lipid profile on coronary heart disease (chd) incidence in spain. methods: a cohort of , healthy spanish individuals aged to years was followed for years. the end-points were fatal and non-fatal myocardial infarction, and angina. results: the participants who developed a coronary end-point were significantly older ( vs ), more often diabetic ( % vs %), smoker ( % vs %) and hypertensive ( % vs %) than the rest, and their average total and hdl-cholesterols (mg/dl) were: vs (ns) and vs , (p< . ), respectively. chd incidence among individuals with low hdl levels (< in men/< in women) was higher than in the rest: . &aeyear- vs . &aeyear- (p< . ) in men, and . &aeyear- vs . &aeyear- (p< . ) in women. hdl-cholesterol was the only lipid related variable significantly associated with chd: hazard ratio for mg/dl increase was . ( % ci: . - . ) in men, and . ( % ci: . - . ) in women, after adjusting for classical risk factors. conclusion: hdl-cholesterol is the only classical lipid variable associated with chd incidence in spain. abstract background: it is widely recognized that health service interventions may reduce infant mortality/imr rate which usually occurs alongside with economic growth. however, there are reports showing that imr decrease under adverse economic and social conditions, indicating the presence of other unknown determinants. objective: this study aims to analyze temporal tendency of infant mortality in brazil during a recent period ( to ) of economic crisis. methods: temporal series study using data from the mortality information system, censuses (ibge) and epidemiological information (funasa). applying arima -autoregressive integrated moving average, it was described series parameters and, spearman correlation coefficients were used to evaluate the association between infant mortality coefficient and some determinants. results: the infant mortality showed a declining tendency () . %) and strong correlation to the majority of the indicators analyzed. however, only correlation between infant mortality coefficient and total fecundity and birth rates differed significantly within decades. conclusions/discussion: fecundity variation was responsible to the persistence of mortality decline during the eighties. in the next period those indicators of life conditions, mostly health care, could be more important. abstract background: across european union (eu) member states, considerable variation exists in the structure and performance of surveillance systems for communicable disease prevention and control. objectives: the study aims to support the improvement of surveillance systems of communicable diseases in europe while using benchmarking for the comparison of national surveillance systems. design and methods: surveillance systems from england & wales, finland, france, germany, hungary and the netherlands were described and analysed. benchmarking processes were performed with selected criteria (e.g. case definitions, early warning systems). after the description of benchmarks, best practices were identified and described. results: the six countries have in general wellfunctioning communicable disease control and prevention systems. nevertheless, different strengths and weaknesses in could be identified. practical examples for best practice from various surveillance systems demonstrated fields for improvement. conclusion and discussion: benchmarking national surveillance systems is applicable as a new tool for the comparison of communicable disease control in europe. a gold standard of surveillance systems in various countries is very difficult to achieve because of heterogeneity (e.g. in disease burden, personal and financial resources). however, to improve the quality of surveillance systems across europe, it will be useful to benchmark surveillance systems of all eu member states. abstract background: therapeutic decisions in osteoarthritis (oa) often involve trade-offs between accepting risks of side effects and gaining pain relief. data about the risk levels patients are willing to accept are limited. objectives: to determine patients' maximum acceptable risk levels (marls) for different adverse effects from typical oa medications and to identify the predictors of these risk attitudes. design and methods: marls were measured with a probabilistic threshold technique for different levels of pain relief. baseline pain and risk levels were controlled for in a x factorial design. clinical and sociodemographic characteristics were assessed using a selfadministered questionnaire. results: for subjects, marls distributions were skewed, and varied by level of pain relief, type of adverse effect, and baseline risk level. given a % baseline risk, for a -point ( - scale) pain benefit the mean (median) marls were . % ( %) for heart attack/stroke; . % ( %) for stomach bleed; . % ( . %) for hypertension; and . % ( . %) for dyspepsia. most clinical and sociodemographic factors were not associated with marls. conclusion: subjects were willing to trade substantial risks of side effects for pain benefits. this study provides new data on risk acceptability in oa patients that could be incorporated into practice guidelines for physicians. background: several independent studies have shown that single genetic determinants of platelet aggregation are associated with increased ihd risk. objectives: to study the effects of clustering prothombotic (genetic) determinants on the prediction of ihd risk. design and methods: the study is based on a cohort of , women, aged to years, who were followed from to . during this period, there were women with registered ihd (icd- - ) . a nested case cohort analysis was performed to study the relation of plasma levels vwf and fibrinogen, blood group genotype and prothrombotic mutations in the gene of a b , gpvi, gpib and aiibb to ihd. results: blood group ab, high vwf concentrations and high fibrinogen concentrations were associated with increased incidence of acute ihd. when the effects of blood group ab/o genotype, plasma levels fibrinogen and vwf were clustered with a score, there was a convincing relationship between a high prothrombotic score and increased incidence of acute ihd: the full-adjusted hr ( % confidence interval) was . ( . - . ) for women with the highest score when the lowest score was taken as reference. conclusions: clustering of prothrombotic markers is a major determinant of increased incidence of acute ihd. abstract background: studies have revealed heart rate variability (hrv) was a predictor of hypertension; however its h-recording has not been analysed with the -hour ambulatory blood pressure. objective: we studied the relationship between hrv and blood pressure. methods: hrv and blood pressure were measured by -hour ambulatory recordings, in randomly selected population without evidence of heart disease. cross-sectional analyses were conducted in women and men (mean age: . ± . ). hrv values, measured by the standard deviation of rr intervals (sdnn), were compared after logarithmic transformation between the blood pressure levels ( / mmhg), using analysis of variance. stepwise multiple-regression was performed to assess on sdnn the cumulative effects of systolic and diastolic blood pressure, clinical obesity, fasting glycaemia, c-reative protein, treatments, smoking and alcohol consumption. results: sdnn was lower in hypertensive men and women (p< . ), independently of drug treatments. after adjustment for factors associated with hypertension, sdnn was no more associated with hypertension, but with obesity, glycaemia and c-reative protein in both genders. sdnn was negatively associated with diastolic blood pressure in men (p = . ) in the multivariate approach. conclusion: whereas blood pressure levels were not related to the sdnn in the multivariate analysis, diastolic blood pressure contributed to sdnn in men. it has been proposed that n- fatty acids may protect against the development of allergic disease, while n- fatty acids may promote its development. in the piama (prevention and incidence of asthma and mite allergy) study we investigated associations between breast milk fatty acid composition of allergic and non allergic mothers and allergic disease (doctor diagnosed asthma, eczema or hay fever) in their children at the age of year and at the age of years. in children of allergic mothers prevalences of allergic disease at age and at age were relatively high if the breast milk they consumed had a low content (wt%) of n- fatty acids and particularly of n- long chain polyunsaturates (lcps), a low content of trans fatty acids, or a low ratio of n- lcps/n- lcps. the strongest predictor of allergic disease was a low breast milk n- lcps/n- lcps ratio (odds ratios ( % ci) of lowest vs highest tertile, adjusted for maternal age, parity and education: . ( . to . ) for allergic disease at age and . ( . to . ) for allergic disease at age ). in children of non allergic mothers no statistically significant associations were observed. abstract background/relevance: to find out about the appropriateness of using two vision related quality of life instruments to measure outcome of visually impaired elderly in a mono-and multidisciplinary rehabilitation centre. objective/design: to evaluate sensitivity of the vision quality of life core measure (vcm ) and the low vision quality of life questionnaire (lvqol) to measure changes in vision related quality of life in a non-randomised followup study. methods: visually impaired patients (n= ) recruited from ophthalmology departments administered questionnaires at baseline ( ) ( ) ( ) ( ) , months and year after rehabilitation. person measures were analysed using rasch analyses for polytomous rating scales. results: paired sample t-tests for the vcm showed improvement at months (p = . ; effect size = . and p = . ; effect size= . ) for the monodisciplinary and the multidisciplinary groups respectively. at year only the multidisciplinary group showed improvement on the vcm (p = . ; effect size = . ). on the lvqol, no significant improvement or deterioration was found for both groups. discussion: although, vcm showed improvement in vision related quality of life over time, the effect sizes appeared to be quite small. we conclude that both instruments lack sensitivity to measure changes. another explanation is that rehabilitation did not contribute to quality of life improvements. abstract background: the natural history of asthma severity is poorly known. objective: to investigate prognostic factors of asthma severity. methods: all current asthmatics identified in / in the european community respiratory health survey were followed up and their severity was assessed in by using the global initiative for asthma categorization (n = ). asthma severity was related to baseline/follow-up potential determinants by a multinomial logistic model, using intermittent asthmatics as reference category for relative risk ratios (rrr). results: patients in the lowest/highest levels of severity at baseline had an % likelihood of remaining in a similar level. severe persistent had a poorer fev %predicted at baseline, higher ige levels (rrr= . ; % ci: . - . ), higher prevalence of chronic cough/phlegm ( . ; . - . ) than intermittent asthmatics. moderate persistent showed similar associations. mild persistent were similar to intermittent asthmatics, although the former showed a poorer control of symptoms than the latter. subjects in remission had a lower probability of an increase in bmi than current asthmatics ( . ; . - . ). allergic rhinitis, smoking, respiratory infections in childhood were not associated with severity. conclusion: asthma severity is a relatively stable condition, at least for patients at the two extremes of the severity spectrum. high ige levels and persistent cough/phlegm are strong markers of moderate/severe asthma. abstract background: thyroid cancer (tc) has a low, yet growing, incidence in spain. ionizing radiation is the only well established risk factor. objectives: this study sought to depict the municipal distribution of tc mortality in spain and to argue about possible risk factors. design and methods: posterior distribution of relative risk for tc was computed using a single bayesian spatial model covering all municipal areas of spain ( , ) . maps were plotted depicting standardised mortality ratios, smoothed municipal relative risk (rr) using the besag, york and mollie`model, and the distribution of the posterior probability that rr> . results: from to a total of , tc deaths were registered in , municipalities. there was a higher risk of death in some areas of canary islands, galicia and asturias. abstract igf-i is an important growth factor associated with increased breast cancer risk in epidemiological and experimental studies. lycopene intake has been associated with decreased cancer risk. although some data indicate that lycopene can influence the igfsystem, this has never been extensively tested in humans. the purpose of this study is to evaluate the effects of a lycopene intervention on the circulating igf-system in women with an increased risk of breast cancer. we conducted a randomized, placebo-controlled cross-over intervention study on the effects of lycopene supplementation ( mg/day, months) in pre-menopausal women with ) history of breast cancer (n = ) and ) high familial breast cancer risk (n = ). drop-out rate was %. mean igf-i and igfbp- concentrations after placebo were . ± . ng/ml and . ± . mg/ml respectively. lycopene supplementation did not significantly alter serum total igf-i (mean lycopene effect: ) . ng/ml; % ci: ) . - . ) and igfbp- () . mg/ml; ) . - . ) concentrations. dietary energy and macronutrient intake, physical activity, body weight, and serum lycopene concentrations were assessed, and are currently under evaluation. in conclusion, this study shows that months lycopene supplementation has no effect on serum igf-system components in a high risk population for breast cancer. abstract introduction: patients who experience burden during diagnostic tests may disrupt these tests. the aim was to describe the perception of melanoma patients with lymph node metastases of the diagnostic tests. methods: patients were requested to complete a self-administrated questionnaire. experienced levels of embarrassment, discomfort and anxiety were calculated, as well as (total) scores for each burden. the non-parametric friedman test for related samples was used to see if there was a difference in burden. results: the questionnaire was completed by patients; response rate was %. overall satisfaction was high. in total % felt embarrassment, % discomfort and % anxiety. overall, % felt some kind of burden. there was no difference in anxiety between the three tests. however, patients experienced more embarrassment and discomfort during the pet (positron emission tomography) scan (p = . and p = . ). conclusion: overall levels of burden were low. however, patients experienced more embarrassment and discomfort during the pet scan, possibly as a result of lying immobile for a long time. the accuracy, costs and patients upstaged will probably be the most important to determine the additional value of fdg-pet and ct, but it is reassuring to know that only few patients experience severe or extreme burden. abstract gastric cancer (gc) is the second most frequent cause of cancer death in lithuania. some intercultural aspects of diet that is related to the outcome could be the risk factors of the disease. the objective of the study was to assess an associations between risk of gc and dietary factors. a case-control study included cases with diagnose of gc and controls that were cancer and gastric diseases free. a questionnaire used to collect information on possible risk factors. the odds ratios (or) and % confidence intervals (ci) estimated by the conditional logistic regression model. after controlling for possible confounders that were associated with gc, use of salted meat (or = . ; % ci = . - . ; > - times/week vs. almost never) smoked meat (or = . ; % ci = . - . ; > - times/week vs. less), smoked fish (or = . ; % ci = . - . ; > - times/week vs. less) was associated with increased risk of gc. higher risk of gc was associated with frequent use of butter, eggs and noodles. while frequent consumption of carrots, cabbages, broccolis, tomatoes, garlic, beans decreased the risk significantly. the data support a role of salt processed food and some animal foods in increasing the risk of gc and plant foods in reducing the risk of the disease. abstract background: standards for the evaluation of measurement properties of health status measurement instruments (hsmi), including explicit criteria for what constitutes good measurement properties, are lacking. nevertheless, many systematic reviews have been performed to compare and select hsmi, using different criteria to judge the measurement properties. objectives: ( ) to determine which measurement properties are reported in systematic reviews of hsmi and how these properties are defined, ( ) which standards are used to define how measurement properties should be assessed, and ( ) which criteria are defined for good measurement properties. methods: a systematic literature search was performed in pubmed, embase and psychlit. articles were included if they met the following inclusion criteria: ( ) systematic review, ( ) hsmi were reviewed, and ( ) the purpose of the review is to identify all measurement instruments assessing (an aspect of) health status and to report on the clinimetric properties of these hsmi. two independent reviewers selected the articles. a standardised data-extraction form was used. preliminary results: systematic reviews were included. conclusions: large variability in standards and criteria used for evaluating measurement properties was found. this review can serve as basis for reaching consensus on standards and criteria for evaluating measurement properties of hsmi. abstract residential exposure to nitrogen dioxide is an air quality indicator and could be very useful to assess the effects of air pollution on respiratory diseases. the present study aims at developing a model to predict residential exposure to no , combining data from questionnaires and from local monitoring stations (ms). in the italian centres of verona, torino and pavia, participating in ecrhs-ii, no concentrations were measured using passive samplers (ps-no ) placed outside the kitchen of subjects for days. simultaneously, average no concentrations were collected from all the mss of the three centres (ms-no ). a multiple regression model was set up with ps-no concentrations as response variable and questionnaire information and ms-no concentrations as predictors. the model minimizing the root mean square error (rmse), obtained from a ten fold cross validation, was selected. the model with the best predictive ability (rmse= . ), had as predictors: ms-no concentrations, season of the survey, centre, type of building, self-reported intensity of heavy vehicle traffic. the correlation coefficient between predictive and observed values was . ( % ci: . - . ). in conclusion, this preliminary analysis suggests that the combination of questionnaire information and routine data from the mss could be useful to predict the residential exposure to no . abstract background: currently only % of dutch mothers comply with the who recommendation to give exclusive breastfeeding for at least six months. therefore, the dutch authorities consider policies on breastfeeding. objectives: quantification of the health effect of several breastfeeding policies. methods: a systematic literature search of published epidemiological studies conducted in the general 'western' population. based on this overview a model is developed. the model simulates incidences of diseases of mother and child depending on the duration that mothers breastfeed. each policy corresponds to a distribution in the duration of breastfeeding. the health effects of each policy are compared to the present situation. results: breastfeeding has beneficial health effects on both the short and the long term for mother and child. the longer the duration of breastfeeding, the larger is the effect. most public health gain is achieved by introducing breastfeeding to all newborns rather than through a policy focusing just on extending the lactation period of women already breastfeeding. conclusion: breastfeeding has positive health effects. policy should focus preferentially on encouraging all mothers to start with breastfeeding. abstract background: constant increase of international trade and travel activities has risen the significance of pandemic infectious diseases worldwide. the / sars outbreak rapidly spread from china to countries, from which were located in europe. in order to control and prevent pandemic infections in europe, systematic and effective public health preparation by every member state is essential. method: supported by the european commission, surveys focusing on national sars (september ) and influenza (october ) preparedness were accomplished. a descriptive analysis was undertaken to identify differences in european infectious disease policies. results: guidelines and guidance for disease management were well established in most european countries. however, the application of control measures, like e.g. measures for mass gatherings or public information policies, had varied among member states. discussion: the results show that european countries are aware of preparing for pandemic infections. yet, the effectiveness of certain control measures is analysed insufficiently. further research and detailed knowledge about factors influencing international spread of diseases is required. 'hazard analysis of critical control points' (haccp) will be applied to evaluate national health response in order to provide comprehensive data for recommendations to european pandemic preparedness. abstract background: influenza is still an important problem for public health. knowing its space-time evolution is of special interest in order to carry out prevention plans. objectives: to analyze the geographical diffusion of the epidemic wave in extremadura. methods: the influenza incidence absolute rates in every town have been calculated, according to the registered cases per week in the compulsory disease declaration system. continuous maps have been represented using a geographical interpolation method (inverse distance weighting (idw) was applied with weighting exponents of ). results: the / season began in the th week of , with a small influenza incidence. there have been concrete cases in those towns until the th week. punctual areas diffusion in the north and southwest of the region between the th and the st weeks. the highest incidence appeared between the nd week of and the rd of . influenza cases started to decrease in the northwest and north of the region from the rd week of , till the th week, when most of the cases were found in the southwest. conclusion: there is a space-time diffusion of influenza, due to the higher population density. we propose to analyze these data combining temperature information. abstract background: acute lower respiratory tract infection (lrti) can cause various complications leading to morbidity and mortality notably among elderly patients. antibiotics are often given to prevent complications. to minimise costs and bacterial resistance, antibiotics are only recommended in case of pneumonia or in patients at serious risk for serious complications. objective: we assessed the course of illness of lrtis among dutch elderly primary care patients and assessed whether gps were inclined to prescribe antibiotics more readily to patients at risk for complications. methods: we retrospectively analysed medical data from , episodes of lrti among patients? years of age presenting in primary care to describe the course of illness. the relation between prescriptions of antibiotics and patients with risk factors for a complicated course was assessed by means of multivariate logistic regression. results: in episodes of acute bronchitis antibiotics were more readily prescribed to patients aged years or older. in exacerbations of copd or asthma gps favoured antibiotics in male patients and when diabetes, neurological disease or dementia was present. conclusion: gp's do not take all high risk conditions into account when prescribing antibiotics to patients with lrti despite recommendations of national guidelines. abstract background: the putative association between antidepressant treatment and increased suicidal behaviour has been under debate. objectives: to estimate the risk of suicide, attempted suicide, and overall mortality during antidepressant treatments. design and methods: study cohort consisted all subjects without non-affective psychosis, hospitalized due to a suicide attempt during the years - , followed up by using nationwide registers. main outcome were completed suicides, attempted suicides, and mortality. main explanatory variable was antidepressant usage. results: suicides, suicide attempts and deaths were observed. when the effect of background variables was taken into account, the risk of suicide attempts was increased markedly during antidepressant treatment (rr for selective serotonin reuptake inhibitors or ssri . , . - . ) compared with no antidepressants. however, the risk of completed suicides was not increased. a lower mortality was observed during ssri use (rr . , . - . ), which was mainly attributable to decrease in cardiovascular deaths. conclusion and discussion: in this suicidal high risk cohort the use of any antidepressant is associated with an increased risk of suicide attempts, but not with the increased risk of completed suicide. antidepressants and, especially, ssri use is associated with a substantial decrease in cardiovascular deaths and overall mortality. abstract background: the quattro study is a rct on the effectiveness of intensified preventive care in primary health care centres in deprived neighbourhoods. additional qualitative research on the execution of interventions in primary care was considered necessary for the explanation of differences in effectiveness. objectives: our question was: can we understand rct outcomes better with qualitative research? design and methods: an ethnographic design was used. in their daily work we observed researchers for months days a week, and practice nurses for days each. two other practice nurses were interviewed. all transcribed observations were analysed thematically. results: from the rct showed differences in effectiveness among the centres and that intensified preventive care provided no additional effect compared to structural physical measurements. ethnographic results show that these differences are due to variations in execution of the intervention among the centres. conclusion: in conclusion ethnographic analysis showed that differences in execution of intervention lead to differences in rct outcomes. the rct conclusion 'no additional effect' is problematic. discussion as variations in primary care influence a rcts' execution they create methodological problems for research. to what extent can additional qualitative research improve rct research. abstract background: acute myocardial infarction is the most important cause of morbidity from ischemic heart disease (ihd) and is the leading cause of death in the western world. objectives: to assess the benefits and harms of 'dan shen' compound for acute myocardial infarction. methods: we searched the cochrane controlled trials register on the cochrane library, medline, embase, chinese biomedical database and the chinese cochrane centre controlled trials register. we included randomized controlled studies lasting at least days. main results: eleven studies with participants in total were included. seven studies compared the mortality in routine treatment plus 'dan shen' compound and single routine treatment. one trial compared the arrhythmia in routine treatment plus 'dan shen' compound injection and single routine treatment. two trials compared the revascularization in routine treatment plus 'dan shen' compound injection and single routine treatment. conclusions: evidence is insufficient to recommend the routine use of 'dan shen' compound because of the small number of included studies and their low quality. no well designed randomized controlled trials with adequate power to provide a more definitive answer have been conducted. in addition, the safety of 'dan shen' compound is unproven, though adverse events were rarely reported. abstract antimicrobial resistance is emerging. to identify the scope of this threat and to be able to take proper actions and evaluate these, monitoring is essential. the remit of earss is to maintain a comprehensive surveillance system that provides comparable and validated data on the prevalence and spread of major invasive bacteria with clinically and epidemiologically relevant antimicrobial resistance in europe. since , earss collects routine antimicrobial susceptibility test (ast) results of invasive isolates of five indicator bacteria, tested according to standard protocols. in , ast results for , isolates were provided by laboratories, serving hospitals, covering million inhabitants in countries. through a biannual questionnaire denominator information was collected. the quality of ast results of laboratories was evaluated by the yearly external quality assessment. currently, earss includes all member and candidate states ( ) of the european union, plus israel, bosnia, bulgaria and turkey. participating hospitals treat a wide range of patients and laboratory results are of sufficient validity. earss identified antimicrobial resistance time trends and found a steady increase for most pathogen-compound combinations. in conclusion, earss is a comprehensive system with sufficient quality to show that antimicrobial resistance is increasing in europe and threatens health-care outcomes. abstract introduction: since chloroform has been detected in drinking waters, the number of studies has increased to identify the presence of trihalomethanes (thms) in drinking waters, as well as to establish the possible effects they may have population health. objectives: to determine thms levels in the water distributing network in the city of valencia. design and methods: over a one-year period, points of the drinking water distributing netowrk have undergone sampling at week intervals. the concentration of these pollutants was determined by gas chromatography. results: our results showed greater concentrations of the species substituted by chlorine and bromine atoms (dichlorobromomethane and dibromochloromethane) in the range of - z lg/l for both, - lg/l for trichloromethane and between - lg/l for tribromomethane. an increase in thms concentration was observed in those points near the sea, although they did not exceed the legal limit of lg/l. conclusion: we established two areas of concentration of these species in water: high and average, according to their proximity to the sea. abstract background: childhood cancer survivors are known to be at increased risk for second malignancies. objectives: we studied longterm risk of second malignancy in -year survivors, according to therapy and follow-up interval. methods: the risk of second malignancies was assessed in -year survivors of childhood cancer treated in the emma children's hospital amc in amsterdam and compared with incidence in the general population of the netherlands. complete follow-up till at least january was obtained for . % of the patients. the median follow-up time was . . results: sixty second malignancies were observed against . expected, yielding a standardized incidence ratio (sir) of ). the absolute excess risk (aer) was . per , persons per year. the sir appeared to stabilize after years of follow-up, but the absolute excess risk increased with longer follow-up (aer follow-up > = years of . ). patients who were treated with radiotherapy experienced the greatest increase of risk. conclusions: in view of the quickly increasing background rate of cancer with ageing of the cohort, it is concerning that even after more than years of follow-up the sir is still increased, as is the absolute excess risk. the chek * delc germline variant has been shown to increase susceptibility for breast cancer and could have an impact on breast cancer survival. this study aimed to determine the proportion of chek * delc germline mutation carriers, and breast cancer survival and tumor characteristics, compared to non-carriers in an unselected (for family history) breast cancer cohort. women with invasive mammary carcinoma, aged < years and diagnosed in several dutch hospitals between and , were included. for all patients, paraffin embedded tissue blocks were collected for dna isolation (normal tissue), and subsequent mutation analyses, and tumor revision. in breast cancer patients, ( . %) chek * delc carriers were detected. chek * delc tumors characteristics, treatment and patient stage did not differ from those of non-carriers. chek * delc carriers had times increased risk of developing a second breast cancer compared to non-carriers. with a mean follow up of years, chek * delc carriers had worse recurrence free and breast cancer specific survival than non-carriers. in conclusion, this study indicates a worse breast cancer outcome in chek * -delc carriers compared to non-carriers. the extension of the presence of the chek * delc germline mutation warrants research into therapy interaction and possibly into screening of premenopausal breast cancer patients. abstract background: for primary or secondary prevention (e.g. myocardial infarction) hormone therapy (ht) is no longer recommended in postmenopausal women. however, physicians commonly prescribe ht to climacteric women as a treatment of hot flashes/night sweats. objective: to assess efficacy and adverse reactions of ht in climacteric women with hot flashes (including night sweats). methods: for our systematic review (sr), we searched databases (medline, embase, cochrane) for randomized controlled trials, other srs and meta-analyses, published to . the quality of the studies was assessed using checklists corresponding to the study type. results: we identified studies of good/excellent quality. they included predominantly caucasian women and lasted - months. in all studies, ht showed a reduction of - % in the number of hot flashes, which was significantly better than placebo. most common adverse events of ht were uterine bleeding and breast pain/tenderness. cardiovascular diseases and neoplasms were reported only sporadically. conclusions: ht is highly effective in treating hot flashes in climacteric women. however, to assess serious adverse events longer studies (including also non-caucasian women) are needed, as there are only sparse data available. abstract igf-i is an important growth factor, and has been associated with increased colorectal cancer risk in both prospective epidemiological and experimental studies. however, it is largely unknown which lifestyle factors are related to circulating levels of the igf-system. studies investigating the effect of isoflavones on the igf-system have thus far been conflicting. the purpose of this study was to evaluate the effects of isoflavones on the circulating igf-system in men with high colorectal cancer risk. we conducted a randomized, placebo-controlled, cross-over study on the effect of a -month isoflavone supplementation ( mg/day) on igf levels in men with a family history of colorectal cancer or a personal history of colorectal adenomas. dropout rate was %, and all but men were more than % compliant. isoflavones supplementation did not significantly alter serum total igf-i () . %; %ci: ) . - . ) and igf binding protein (+ . %, %ci: ) . - . ) concentrations. other covariables, e.g. dietary energy and macronutrient intake, physical activity, and body weight, are currently under evaluation. in conclusion, this study shows that a -month isoflavone supplementation has no effect on serum igf-system components in men with high colorectal cancer risk. abstract background/objective: eurociss-european cardiovascular indicators surveillance set project, funded under the health monitoring programme of european commission, aims developing health indicators and recommendations for monitoring cardiovascular diseases (cvd). methods: prioritise cvd according to their importance in public health; identify morbidity and mortality indicators; develop data collection and harmonizing recommendations; describe data collection, validation procedures and discuss their comparability. population (geographical area, age, gender), methods (case definition, icd codes), procedures (record linkage, validation), morbidity indicators (attack rate, incidence, case fatality) collected by questionnaire. results: the main outcome was the inventory of acute myocardial infarction (ami) populationbased registers in the european partner countries: countries have no register, regional, of which also national. registers differ for: icd codes (only ami or also acute and subacute ischemic forms), ages ( - , - , all) , record linkage (probabilistic, personal identification number), calendar years, validation (monica, esc/acc diagnostic criteria). differences make morbidity indicators difficult to compare. conclusion: new diagnostic criteria led to a more exhaustive definition of myocardial necrosis as acute coronary syndrome (acs). given the high burden of ami/acs, efforts are needed to implement population-based registers in all countries. application of recommended indicators, validated through standardized methodology, will provide reliable, valid and comparable data. abstract objective: the objective of this paper was to compare and discuss the use of odd ratios and prevalence ratios using real data with complex sampling design. method: we carried out a cross-sectional study using data obtained from a two-stage stratified cluster sample from a study conducted in - (n = , ) . odds ratios and prevalence ratios were obtained by unconditional logistic regression and poisson regression, respectively, for later comparison using the stata statistical package (v. . ). confidence intervals and design effects were considered in the evaluation of the precision of estimates. two outcomes of a cross-sectional study with different prevalence were evaluates: vaccination against influenza ( . %) and self-referred lung disease ( . %). results: in the high-prevalence scenario, using prevalence ratios the estimates were more conservative and we found narrower confidence intervals. in the low-prevalence scenario, we found no important differences between the estimates and standard errors obtained using the two techniques. discussion: however, it is the researcher's task to choose which technique and measure to use for each set of data, since this choice must remain within the scope of epidemiology. abstract background: in italy coronary heart disease chd mortality has been falling since the s. objective: to examine how much of the fall between and could be attributed to trends in risk factors, medical and surgical treatments. methods: a validated model was used to combine and analyse data on uptake and effectiveness of cardiological treatments and risk factor trends. published trials, meta-analyses, official statistics, longitudinal studies, surveys are main data sources. results: chd mortality fell by % in men and % in women aged - ; , fewer deaths in . approximately half mortality fall was attributed to treatments in patients and half to population changes in risk factors: in men, mainly improvements in cholesterol ( %) and smoking ( %) rather than blood pressure ( %). in women / mortality fall attributable to improvements in cholesterol ( %) and blood pressure ( %); adverse trends in smoking () %). adverse trends also in bmi () % in both genders) and diabetes () % in men; ) . % in women). conclusion: half chd mortality fall was attributable to risk factors reductions, principally cholesterol in men and women and smoking in men; in women rising smoking rates generated substantial additional deaths. a comprehensive strategy promoting primary prevention is needed. objective: to investigate the efficacy of ni in the post exposure prophylaxis (pep), i.e. in persons who had contact with an influenza case. design and methods: we conducted a systematic electronic data base review for the period between and . studies were selected and graded by two independent reviewers. the proportion of influenza-positive patients was chosen as primary outcome. for all analyses fixed effect models were used. weighted relative risks (rr) and % confidence intervals (ci) were calculated on an intention-to-treat basis. results: randomized controlled trials (n= , ) were included in the analysis. zanamivir and oseltamivir were effective against an infection with influenza (rr= . , % ci . - . and rr= . , % ci . - . , respectively). prophylactic efficacy was comparable in the subgroup of persons who had contact with an index case with lab-confirmed influenza ( studies, all ni, rr= . , % ci . - . ). conclusions: the available evidence suggests that ni are effective in the pep of influenza. discussion: results have to be interpreted with caution when transferred into general medical practice because study populations mainly included young and healthy adults without chronic diseases. abstract an important risk factor of breast cancer, mammographic breast density (mbd) is inversely associated with reproductive factors (age at first childbirth, and lactating). as pregnancy and lactating are highly correlated, whether this decline is induced by pregnancy or lactating is still unclear. we hypothesize that lactation reduces mbd independent of age at first pregnancy and parity. a study was done on women in the third sub-cohort of the dom project who had complete data regarding lactating, dy, had a child but varied by duration of lactating. multiple logistic regression analysis was done using dy (yes/ no) as outcome variable. explanatory variables added into the model were age, bmi, parity and age at first childbirth. a significant univariate relation was seen between lactating of the first child and dy. or . (ci % . ; . ). adjusted for explanatory variables, the or changed to . (ci % . ; . ). lactating seems to contribute independently to the reduction of mbd over and above pregnancy itself. given the limitations of the dichotomous dy ratio scores, additional studies will address which part; either glandular mass or fat tissue is responsible for the observed relation which will be measured from mammograms to be digitized. abstract background: alcohol consumption is common, but little is known about whether drinking patterns vary across geographic regions. objectives: to examine potential disparities in alcohol consumption across census regions and urban, suburban, and rural areas of the united states. design and methods: the data source was the national epidemiologic survey on alcohol and related conditions, an in-person interview of approximately , adults. the prevalence of abstinence and, among drinkers, the prevalences of heavy and daily drinking were calculated by census region and metropolitan status. multivariate logistic regression analyses were conducted to test for differences in abstinence and per drinker consumption after controlling for confounders. results: the odds of abstinence, heavy, and daily drinking varied widely across geographic areas. additional analyses stratified by census region revealed that rural residents in the south and northeast as well as urban residents in the northeast had higher odds of abstinence. rural residents in the midwest had higher odds of heavy drinking. conclusion and discussion: heavy alcohol consumption is of particular concern among drinkers living in the rural areas of the united states, particularly the rural midwest. other nations should consider testing for similar differences as they implement policies to promote safe alcohol consumption. abstract background: long-term exposure to particulate air pollution (pm) has been suggested to accelerate atherogenesis. objective: we examined the relationship between long-term exposure to traffic emissions and the degree of coronary artery calcification (cac), a measure of atherosclerosis. methods: in a population based, crosssectional study, distances between participants' home addresses and major roads were calculated with a geographic information system. annual mean pm . -exposure at the residence was derived from a small scale geostatistical model. cac, assessed by electronbeam computer tomography, was modelled with linear regression by proximity to major roads, controlling for background pm . air pollution and individual level risk factors. results: of participants lived within m of a major road. background-pm . ranged from . to . lg/m (mean . ). mean cacvalues were strongly dependent on age, sex and smoking status. reducing the distance to major roads by % leads to increases in cac by . % ( %ci . - . %) in the unadjusted model and , % ( %ci ) . - . ) in the adjusted model. stronger effects (adjusted model) were seen in men ( , %, %ci ) . - . ) and male non-smokers ( , %, %ci ) . - . ). conclusions: this study provides epidemiologic evidence that long-term exposure to traffic emissions is an important risk factor for coronary calcification. abstract background: this polymorphism has been associated risk factor levels and in one study with a reduced risk of acute myocardial infarction (ami). yet, the risk relation has not been confirmed. objectives: we investigated the role of this polymorphism on occurrence of ami, coronary heart disease (chd) and stroke in healthy dutch women. design and methods: a case-cohort study in a prospective cohort of initially healthy dutch from until january st . results: we applied a cox proportional hazards model with an estimation procedure adapted for case-cohort designs. a lower ami (n= ) risk was found among carriers of the ala allele (n= ) compared with those with the more common pro pro genotype (hazard ratio= . ; % ci, . to . ). no relation was found for chd (n= ;hr . ; % ci, . - . ) and for stroke (n= ;hr . ; % ci, . - . ). in our data little evidence was found for a relation between pparg and risk factors. conclusion and discussion: this study shows the pro ala polymorphism in pparg gene is modestly related to a reduced risk of ami in our study. no statistically significant relation was found for chd and stroke. abstract background: pseudo cluster randomization was used in a services evaluation trial because individual randomization risked contamination and cluster randomization risked selection bias due to expected treatment arm preferences of recruiting general practitioners (gps). gps were randomized in two groups. depending on this randomization, participants were randomized in majority to one study arm: intervention:control/ : or intervention:control/ : . objectives: to evaluate internal validity of pseudo cluster randomization. have gps treatment arm preferences? what is the effect on allocation concealment and selection bias? design and methods: we compared the baseline characteristics of participants to study selection bias. using a questionnaire, gps indicated their treatment arm preferences on a visual analogue scale (vas) and the allocation proportions they believed were used to allocate their patients over treatment arms. results: gps preferred allocation to the intervention (vas . (sd . ); - : indicates strongly favoring the intervention arm). after recruitment % of gps estimated a randomization ratio of : was used. the participants showed no relevant differences at baseline. conclusion and discussion: gps profoundly preferred allocation to the intervention group. few indications of allocation disclosure or selection bias were found in the dutch easycare trial. pseudo cluster randomization proofs to be a valid randomization method. abstract background: epidemiological studies rely on self-reporting to acquire data on participants, although such data are often limited in reliability. the aim here is to assess nuclear magnetic resonance (nmr) based metabonomics for evaluation of self-reported data on paracetamol use. method: four in-depth -hour dietary recalls and two timed -hour urine collections were obtained for each participant in the intermap study. a mhz h nmr spectrum was acquired for each urine specimen (n = , ). training and test sets involved two strata, i.e., paracetamol metabolites yes or no in the urine spectra, selected from all population samples by a principal component analysis model. the partial least squares-discriminant analysis (pls-da) model based on a training set of samples was validated by test set (n = ). the model correctly predicted stratum for of samples ( %) after removal of outliers not fitting the model, sensitivity . %, specificity %. this model was used to predict paracetamol status in all intermap specimens. it identified participants ( . %) who underreported analgesic use, of whom underreported analgesic use in both clinical visits. conclusion: nmr-based metabonomics can be used as a tool to enhance reliability of self-reported data. abstract background: in patients with asthma, the decline in forced expiratory volume in one second (fev ) is accelerated compared with non-asthmatics. objective: to investigate long-term prognostic factors of fev change in asthmatics from the general population. methods: a cohort of asthmatics ( - years-old) was identified in the frame of the european community respiratory health survey ( / ), and followed up in / . spirometry was performed on both occasions. the annual fev decrease (?fev ) was analysed by multi-level regression models, according to age, sex, height, bmi, occupation, familiarity of asthma, hospitalization for asthma (baseline factors); cumulative time of inhaled corticosteroid (ics) use and annual weight gain during the follow-up; lifetime pack-years smoked. results: when adjusting for all covariates, ics use for > years significantly reduced ?fev , with respect to non-users, of . ( %ci: . - . ) ml/year. ?fev was . ( . - . ) ml/year lower in women than in men. it increased by . ( . - . ) ml/year for every additional year in patient age and by . ( . - . ) ml/year for every additional kg/year in the rate of weight gain. conclusion: long-term ics use (> years) seems to be associated with a reduced ?fev over a -year followup. body weight gain seems a crucial factor in determining lung function decrease in asthmatics. abstract background: effectiveness of screening can be predicted by episode sensitivity, which is estimated by interval cancers following a screen. full-field digital or cr plate mammography are increasingly introduced into mammography screening. objectives: to develop a design to compare performance and validity between screen-film and digital mammography in a breast cancer screening program. methods: interval cancer incidence was estimated by linking screening visits from - at an individual level to the files of the cancer registry in finland. these data were used to estimate the study size requirements for analyzing differences in episode sensitivity between screen-film and digital mammography in a randomized setting. results: the two-year cumulative incidence of interval cancers per screening visits was estimated to be . to allow the maximum acceptable difference in the episode sensitivity between screen-film and digital arm to be % ( % power, . significance level, : randomization ratio, % attendance rate), approximately women need to be invited. conclusion: only fairly large differences in the episode sensitivity can be explored within a single randomized study. in order to reduce the degree of non-inferiority between the screen-film and digital mammography, meta-analyses or pooled analyses with other randomized data are needed. according to the literature up to % of colorectal cancers worldwide is preventable by dietary change. however the results of the epidemiologic studies are not consistent across the countries. the objective of the study is to evaluate the role of dietary nutrients on colorectal cancer risk in poland. the hospital-based case-control study was carried out in - . in total, histologically confirmed cancer cases and controls were recruited. adjustment for age, sex, education, marital status, multivitamin use, alcohol consumption, cigarette smoking, family history and energy consumption was done by logistic regression model. low tertile of daily intake in the control group was defined as a reference level. the lower colorectal cancer risk was found in cases with high daily intake of dietary fiber (or = , ; %ci: , - , ) and vitamin e (or = , ; %ci: , - , ). on the other hand, an increased risk for high monosaccharides consumption was observed. the risk pattern wasn't changed after additional adjustment for physical activity and body mass index. the results of the present study support the protective role of dietary fiber and some antyoxidative vitamins in the etiology of colorectal cancer. additionally they suggest that high consumption of monosaccharides may lead to elevated risk of investigated cancers. abstract assessment of nutrition is very difficult in every population, but in children there's additional question if child can properly recognize and recall foods that have been eaten. the aim of this study was to assess if dietary recall administered to adolescents can be used in epidemiological studies on nutrition. subjects were children, - years old, and they caretakers. -h recall was used to evaluate children's nutrition. both, child and caretaker were asked to recall all products, drinks and dishes eaten by child during the day before recall. the statistical analyses were done separately for each meal. we have noticed statistically significant differenced for intake of energy and almost all nutrients from the lunch. the observed spearman rank correlation coefficients between child and his caretaker ranged from . for vitamin c up to . for intake of carbohydrates. only calcium intake ( . vs. . mg/day) differentiated groups for the breakfast and b-carotene for the supper. the study showed that the recall with adolescents could be helpful source of data for the research in the population aspect. however, one shouldn't use such data for the examination of the individual nutritional habit of children, especially information about dinner can be biased. abstract background: acute bronchitis is one of the most common diagnoses made by primary care physicians. in addition to antibiotics, chinese medicinal herbs may be a potential medicine of choice. objectives: this review aims to summarize the existing evidence of comparative effectiveness and safety of chinese medicinal herbs for treating uncomplicated acute bronchitis. methods: we searched the cochrane central register of controlled trials, medline, embase, chinese biomedical database and etc. we included randomised controlled trials comparing chinese medicinal herbs with placebo, antibiotics or other western medicine for treating uncomplicated acute bronchitis. at least two authors extracted data and assessed trial quality. main results: four trials reported the time to improvement of cough, fever, and rales; two trials reported the proportion of patients with improved signs and symptoms; thirteen trials analyzed the data of global assessments of improvement. one trial reported the adverse effect during treatment. conclusions: there is insufficient quality data to recommend the routine use of chinese herbs for acute bronchitis. the benefit found in individual studies and this systematic review could be due to publication bias and study design limitations. in addition, the safety of chinese herbs is unknown, though adverse events are rarely reported. design and methods: patients with a definite ms and classified as dead or alive at st january were included in this retrospective observational study. influence of demographic and clinical variables was assessed with kaplan meier and cox methods. standardised mortality ratios were computed to compare patients' mortality with the french general population. results: a total of patients were included ( men, women). the mean age at ms onset was +/) years and the mean follow-up duration was +/) years ( patients-years). by , deaths occurred ( per patients-years). male gender, progressive course, polysymptomatic onset and high relapse rate were related to a worse prognosis. ms did not increase the number of deaths in our cohort compared to the general french population ( expected), except for highly disabled patients ( observed, expected). conclusion: this study gave precise insights on mortality in multiple sclerosis in west france. mattress dust. methods: we performed nested case-control studies within ongoing birth cohort studies in germany, the netherlands, and sweden and selected approximately sensitised and non-sensitised children per country. we measured levels of bacterial endotoxin, ß( -> )-glucans, and fungal extracellular polysaccharides (eps) in dust samples collected on the children's mattresses. results: combined across countries, higher amounts of dust and higher endotoxin, ß( -> )-glucans, and eps loads of mattress dust were associated with a significantly decreased risk of sensitization to inhalant allergens, but not food allergens. after mutual adjustment, only the protective effect of the amount of mattress dust remained significant [odds ratio ( % confidence interval) . ( . - . )]. conclusion: higher amounts of mattress dust might decrease the risk of allergic sensitization to inhalant allergens. the effect might be partly attributable to endotoxin, ß( -> )-glucans, and eps. it is not possible to distinguish with certainty, which component relates to the effect, since microbial agents loads are highly correlated with amount of dust and with each other. abstract background: postmenopausal hormone therapy (ht) increases mammographic density, a strong breast cancer risk factor, but effects vary across women. objective: to investigate whether the effect of ht use on changes in mammographic density is modified by polymorphisms in the estrogen (esr ) and progesterone receptor (pgr) genes. design and methods: information on ht use, dna and two consecutive mammograms were obtained from ht users and never ht users of the dutch prospect-epic and the english epic-norfolk cohorts. mammographic density was assessed using a computer-assisted method. changes in density between mammograms before and during ht use were analyzed using linear regression. results: a difference in percent density change between ht users and never users was seen in women with the esr pvuii pp or pp genotype ( . %; p< . ), but not in those with the pp genotype ( . %; p = . ). similar effects were observed for the esr xbai and the pgr + g/a polymorphisms. the pgr progins polymorphism did not appear to make women more susceptible to the effects of ht use. discussion and conclusion: our results suggest that specific polymorphisms in the esr and pgr genes may make women more susceptible to the effects of ht use on mammographic density. abstract background: there is a paucity of data on the cancer risk of turkish migrant children in germany. objectives: to identify cancer cases of turkish origin in the german childhood cancer registry (gccr) and to compare the relative incidence of individual cancers among turkish and non-turkish children. design and methods: we used a name algorithm to identify children of turkish origin among the , cancer cases below years of age registered since . we calculated proportional cancer incidence ratios (pcir) stratified for sex and time period. results: the name algorithm performed well (high sensitivity and specificity), and turkish childhood cancers were identified. overall, the relative frequency of tumours among turkish and non-turkish children is similar. there are specific sites and cancers for which pcirs are different; these will be reported during the conference. conclusion: our study is the first to show differences in the relative frequency of cancers among turkish and non-turkish children in germany. discussion: case control studies could help to explain whether observed differences in the relative frequency of cancers are due to differences in genetic disposition, lifestyle or socio-economic status. mutations in the netherlands cohort study on diet and cancer. data from , participants, cases and , subcohort members were analysed from a follow-up period between . to . years after baseline. adjusted gender-specific incidence rate ratios (rr) and % confidence intervals (ci) were calculated over tertiles of folate intake in case-cohort analyses. high folate intake did not reduce overall colon cancer risk. however, in men only, it was inversely associated with apc[csymbol] colon tumours (rr . , % ci . - . for the highest versus the lowest tertile of folate intake), but positively associated with apc+ colon tumours (highest vs. lowest tertile: rr . , ci . - . ). folate intake was neither associated with overall rectum cancer risk, nor with rectum cancer when apc mutation status was accounted for. we observed opposite associations between folate intake and colon cancer risk with or without apc mutations in men, which may implicate a distinct mutated apc pathway mediated by folate intake in men. abstract background and objectives: ten years after completion of the first serum bank of the general population to evaluate the long-term effects of the national immunisation programme (nip) a new serum collection is desirable. the objective is to provide insight into age-specific estimates of the immunity to childhood diseases and estimates of the incidence of infectious diseases with a frequent sub clinical course. design and methods: a two-stage cluster sampling technique was used to draw a nationwide sample. in each of five geographic regions, eight municipalities were randomly selected proportionally to their size. within each municipality, an age-stratified sample of individuals ( - yr) will be drawn from the population register. in addition eight municipalities will be selected with lower immunization coverage to obtain insight into the immune status of persons who often refuse vaccination on religious grounds. furthermore over sampling of migrants will be performed to study whether their immune status is satisfactory. participants will be asked to fill in a questionnaire and to allow blood to be taken. extra blood will be taken for a genetic study. results and conclusion: the design of a population-based serum collection aimed at the establishment of a representative serum bank will be presented. abstract background: during the last decade, the standard of diabetes care evolved to require more intensive management focussing on multiple cardiovascular risk factors. treatment decisions for lipidlowering drugs should be based on cholesterol and blood pressure levels. objectives: to investigate the influence of hba c, blood pressure and cholesterol levels on subsequent intensification of lipid-lowering therapy between - . design and methods: we conducted a prospective cohort study including , type diabetes patients who had at least two consecutive annual visits to a diabetes nurse. treatment intensification was measured by comparing drug regimes per year, and defined as initiation of a new drug class or dose increase of an existing drug. results: between - , the prevalence of lipid-lowering drug use increased from % to %. rates of intensification of lipid-lowering therapy remained low in poorly controlled patients ( % to %;tc/hdl ratio> ). intensification of lipid-lowering therapy was only associated with tc/hdl ratio (age-adjusted or = . ; %ci . - . ) and this association became slightly stronger over time. blood pressure was not found to be a predictor of the intensification of lipid-lowering therapy (or = . ). conclusion: hypercholesterolemia management intensified between - , but therapy intensification was only triggered by elevated cholesterol levels. more attention for multifactorial risk assessment is needed. abstract background: there are no standard severity measures that can classify the range of illness and disease seen in general practice. objectives: to validate new scales of morbidity severity against age, gender, deprivation and poor physical function. design and methods: in a cross-sectional design, morbidity data for consulters in a -month period was linked to their physical function status . there were english older consulters ( years +) and dutch consulters ( years +). consulters for morbidities classified on four gp-defined ordinal scales of severity ('chronicity', 'time course', 'health care use' and 'patient impact on activities of daily living') were compared to consulters for morbidity other than the , by age-groups, gender, and dichotomised deprivation and physical function scores. results: for both countries, on all scales, there was an increasing association between morbidity severity and older ages, female gender, more deprivation (minimum p< . ) and poor physical function (all trends p< . ). the estimates for categories, for example, within the 'chronicity' scale was ordered as follows: 'acute' (unadjusted odds ratio . ), 'acute-on-chronic' ( . ), 'chronic' ( . ) and 'life-threatening' ( . ). conclusions: new validated measures of morbidity severity indicate physical health status and offer the potential to optimise general practice care. hospitalization or death. calibration and discriminative capacity were estimated. results: among episodes of lrti in elderly patients with dm, endpoints occurred (attack rate %). reliability of the model was good (goodness-of-fit test p = . ). the discriminative properties of the original rule was acceptable (area under the receiver-operating curve (auc): . , % ci: . to . ). conclusion: the prediction rule for the probability of hospitalization or death derived from an unselected elderly population with lrti appeared to have acceptable discriminative properties in diabetes patients and can be used to target management of these common diseases. confounding by indication is a major threat to the validity of nonrandomized studies on treatment effects. we quantified such confounding in a cohort study on the effect of statin therapy on acute respiratory disease (ard) during influenza epidemics in the umc utrecht general practitioner research database among persons aged > = years. the primary endpoint was a composite of pneumonia or prednisolone-treated ard during epidemic, non-epidemic and summer seasons. to quantify confounding, we obtained unadjusted and adjusted estimates of associations for outcome and control events. in all, , persons provided , persons-periods, statin therapy was used in . % and in , person-periods an outcome event occurred. without adjustments, statin therapy was not associated with the primary endpoint during influenza epidemics (relative risk [rr] . ; % confidence interval [ %ci]: . - . ). after applying multivariable generalized estimating equations (gee) and propensity score analysis the rrs were . ( % ci: . - . ) and . ( % ci: . - . ). the findings were consistent across relevant strata. in non-epidemic influenza and summer seasons the rr approached . while statin therapy was not associated with control event rates. observed confounding in the association between statin therapy and acute respiratory outcomes during influenza epidemics masked a potential benefit of more than %. abstract background: despite several advances in the treatment of schizophrenia, the currently available pharmacotherapy does not change the course of illness or prevent functional deterioration in a substantial number of patients. therefore, research efforts into alternative or adjuvant treatment options are needed. in this project, called the 'aspirine trial', we investigate the effect of the antiinflammatory drug acetylsalicylic acid as an add-on to regular antipsychotic therapy on the symptoms of schizophrenia. objectives: to objective is to study the efficacy of acetylsalicylic acid in schizophrenia on positive and negative psychotic symptoms, immune parameters and cognitive functions. design and methods: a randomized placebo controlled double-blind add-on trial of inpatients and outpatients with schizophrenia, schizophreniform or schizoaffective disorder is performed. patients are : randomized to either months mg acetylsalicylic acid per day or months placebo, in addition to their regular antipsychotic treatment. all patients receive pantoprazole treatment for gastroprotection. participants are recruited from various major psychiatric hospitals in the netherlands. the outcomes of this study are -month change in psychotic and negative symptom severity, cognitive function, and several immunological parameters. status around participants have been randomized. no interim analysis was planned. abstract background: congenital cmv infection is the most prevalent congenital infection worldwide. epidemiology and outcome are known to vary with socio-economic background, but few data are available on epidemiology and outcome in a developing country, where the overall burden of infectious diseases is high. objective: to determine prevalence, riskfactors and outcome of congenital cmv infection in an environment with high infectious disease burden methods: as part of an ongoing birth cohort study, baby and maternal samples were collected at birth, and tested with an inhouse pcr for the presence of cmv. standardised clinical assessment were performed by a paediatrician. placental malaria was also assessed. follow-up is ongoing till the age of years. preliminary results: the prevalence of congenital cmv infection was / ( . %). the infected children were more often first born babies ( . % vs . %, p< . ). while no seasonality was observed, placental malaria was more prevalent among congenitally infected children ( . % vs . %,p = . ). there were no symptomatic babies detected. conclusion: this prevalence of congenital cmv is much higher than reported in industrialised countries, in the absence of obvious clinical pathology. further follow up is needed to assess impact on response to vaccinations, growth, and morbidities. of wheeze or cough at night in the first years. data on respiratory symptoms and dda were collected by yearly questionnaires. in total, symptomatic children with and without an early dda were included in the study population. results: fifty-one percent of the children with and % of the children without an early dda had persistent respiratory symptoms at age . persistence of symptoms was associated with parental atopy, eczema, nose symptoms without a cold, or a combination of wheeze and cough in the first years. conclusions: monitoring the course of symptoms in children with risk factors for persistent symptoms, irrespective of a diagnosis of asthma, may contribute to early recognition and treatment of asthma. little is known about the response mechanisms of survivors of disasters. objective: to examine selective non-response and to investigate whether attrition has biased the prevalence estimates among survivors of a disaster. design and methods: a longitudinal study was performed after the explosion of a fireworks depot in enschede, the netherlands. survivors completed a questionnaire weeks (t ), months (t ) and years post-disaster (t ). prevalence estimates resulting from multiple imputation were compared with estimates resulting from complete case analysis. results: non-response differed between native dutch and nonwestern immigrant survivors. for example, native dutch survivors who participate at t only were more likely to have health problems at t such as depression than native dutch who participated at all three waves (or = . , % ci: . - . ) . in contrast, immigrants who participated at t only were less likely to have depression at t (or = . , % ci: . - . ). conclusion and discussion: among native dutch survivors, the imputed estimates of t health problems tended to underestimated than the complete case estimates. the imputed t estimates among immigrants were unaffected or somewhat overestimated than the complete case estimates. multiple imputation is a useful statistical technique to examine whether selective non-response has biased the prevalence estimates. session: posters session : july presentation: poster. background: several epidemiologic studies have shown decreased colon cancer risk in physically active individuals. objectives: this review provides an update of the epidemiologic evidence for the association between physical activity and colon cancer risk. we also explored whether study quality explains discrepancies in results between different studies. methods: we included cohort (male n = ; female n = ) and case-control studies (male n = ; female n = ) that assessed total or leisure time activities in relation to colon cancer risk. we developed a specific methodological quality scoring system for this review. due to the large heterogeneity between studies, we refrained from statistical pooling. results: in males, the cohort and case-control studies lead to different conclusions: the case-control studies provide strong evidence for a decreased colon cancer risk in the physically active while the evidence in the cohort studies is inconclusive. these discrepant findings can be attributed to either misclassification bias in cohort or selection bias in case-control studies. in females, the small number of high quality cohort studies precludes a conclusion and the case-control studies indicate an inverse association. conclusion: this review indicates a possible association of physical activity and reduction of colon cancer risk in both sexes but the evidence is not yet convincing. abstract background/objectives: radiotherapy after lumpectomy is commonly applied to reduce recurrence of breast cancer but may cause acute and late side effects. we determined predictive factors for the development of late toxicity in a prospective study of breast cancer patients. methods: late toxicity was assessed using the rtog/ eortc classification among women receiving radiotherapy following lumpectomy after a mean follow-up time of months. predictors of late toxicity were modelled using cox regression in relation to observation time, adjusting for age, bmi and biologically effective dose in the maximum at the skin. results: ( . %) patients presented with telangiectasia and ( . %) patients with fibrosis. we observed a strong association between development of telangiectasia and fibrosis (p< . ). increasing patient age was a risk factor for telangiectasia and fibrosis (p for trend . and . , respectively). boost therapy (hazard ratio (hr) . , % ci . - . ) and acute skin toxicity (hr . , % ci . - . ) significantly increased risk of telangiectasia. risk of fibrosis was elevated among patients with atopic diseases (hr . , % ci . - . ). discussion: our study revealed several risk factors for late complications of radiotherapy. further understanding of differences in response to irradiation may enable individualized treatment and improve cosmetic outcome. doctor-diagnosed asthma and respiratory symptoms (age ) were available for (rint) and (no) children. results: the discriminative capacities of rint and exhaled no were statistically significant for the prediction of doctor-diagnosed asthma, wheeze (rint only) and shortness of breath (rint only). due to the low prevalence of disease in this general population sample, the positive predictive values of both individual tests were low. however, the positive predictive value of the combination of increased rint (cutoff . kpa.l- .second) and exhaled no (cut-off ppb) was % for the prediction of doctor-diagnosed asthma, with a negative predictive value of %. combinations of rint or exhaled no with atopy of the child showed similar results. conclusions: the combination of rint, exhaled no and atopy may be useful to identify high-risk children, for monitoring the course of their symptoms and to facilitate early detection of asthma. abstract background: in a cargo aircraft crashed into apartment buildings in amsterdam, killing people, and destroying apartments. an extensive, troublesome aftermath followed with rumours on toxic exposures and health consequences. objectives: we studied the long-term physical health effects of occupational exposure to this disaster among professional assistance workers. design and methods: in this historical cohort study we compared the firefighters and police officers who were occupationally exposed to this disaster (i.e. who reported one or more disasterrelated tasks) with their nonexposed colleagues (n = , and n = , respectively), using regression models adjusted for background characteristics. data collection took place from january to march , and included various clinical blood and urine parameters (including blood count and kidney function), and questionnaire data on occupational exposure, physical symptoms, and background characteristics. the overall response rate was %. results: exposed workers reported various physical symptoms (including fatigue, skin and musculoskeletal symptoms) significantly more often than their nonexposed colleagues. in contrast, no consistent significant differences between exposed and nonexposed workers were found regarding clinical blood and urine parameters. discussion and conclusion: this epidemiological study demonstrates that professional assistance workers involved in a disaster are at risk for long-term unexplained physical symptoms. abstract background and objectives: recent studies indicate that women with cosmetic breast implants have significantly increased risk of suicide. reasons for elevated risk are not known. it is suggested that women with cosmetic breast implants differ in their characteristics and have more mental problems than women of general population. aim of this study was to find out possible associations between physical or mental health and postoperative quality of life among finnish women with cosmetic breast implants. design and methods: information was collected from patient records of women and structured questionnaires mailed to women of the same cohort. data was analysed by using pearson chi square testing and logistic regression modelling. results: although effects of implantation on postoperative quality of life in different areas were mainly reported as positive or neutral, % of the women reported decreased state of health. postoperative dissatisfaction and decreased quality of life were significantly associated with diagnoses of depression (p = . ) and local complication called capsular contracture (p< . ). conclusion: our results are consistent with previous results finding most of the cosmetic breast surgery patients satisfied after implantation. however, this study brings new information on associations between depression, capsular contracture and decreased quality of life. abstract cancer and its treatments often produce significant persistent morbidities that reduce quality of life (qol) in cancer survivors. research indicates that both, physical exercise and psycho-education might enhance qol. therefore, we developed a -week multidisciplinary rehabilitation program that combines physical training with psycho-education. the aim of the present multicenter study is to determine the effect of multidisciplinary rehabilitation on qol as compared to no treatment and, additionally, to physical training alone. furthermore, we will explore which variables are related to successful outcome (socio-demographic, disease related, physiological, psychological and environmental characteristics). participants are needed to detect a medium effect. at present, cancer survivors are randomised to either the multidisciplinary or physical rehabilitation program or a -month waiting list control group. outcome assessment will take place before, halfway, directly after, and months following the intervention by means of questionnaires. physical activity will be measured before, halfway and directly after rehabilitation using maximal and submaximal cycle ergometer testing and muscle strength measurement. effectiveness of multidisciplinary rehabilitation will be determined by analysing changes between groups from baseline to post-intervention using multiple linear and logistic regression. positive evaluation of multidisciplinary rehabilitation may lead to implementation in usual care. continuous event recorders (cer) have proven to be successful in diagnosing causes of palpitations but may affect patient qol and increase anxiety. objectives: determine qol and anxiety in patients presenting with palpitations, and to evaluate the burden of the cer on qol and anxiety in patients presenting to the general practitioner. methods: randomized clinical trial in general practice. the short form- (sf- ) and state-trait anxiety inventory (stai) were administered at study inclusion, -weeks and months. results: at baseline, patients with palpitations (n = ) reported lower qol and more anxiety than a healthy population for both males and females. there were no differences between the cer arm and usual gp care at -weeks. at -months the usual care group (n = ) showed minimal qol improvement and less anxiety compared to the cer group (n = ). type of diagnosis did not account for any of these reported differences. conclusion: anxiety decreases and qol increases in both groups at -weeks and -month follow-up. hence it is a safe and effective diagnostic tool, which is applicable for all patients with palpitations in the general practice. abstract background: clinical benefits of statin therapy are accepted, but their safety profiles have been under scrutiny, particularly for the most recently introduced statin, rosuvastatin, relating to serious adverse events involving muscle, kidney and liver. objective: to study the association between statin use and the incidence of hospitalizations for rhabdomyolysis, myopathy, acute renal failure and hepatic impairment (outcome events) in real life. methods: in and , , incident rosuvastatin users, , incident other statin users and , patients without statin prescriptions from the pharmo database of > million dutch residents were included in a retrospective cohort study. potential cases of hospitalization for myopathy, rhabdomyolysis, acute renal failure or hepatic impairment were identified using icd- -cm codes and validated using hospital records. results: there were validated outcome events in the three cohorts including one case each of myopathy (other statin group) and rhabdomyolysis (non-treated group). there were no significant differences in the incidence of outcome events between rosuvastatin and other statin users. discussion: this study indicated that the number of outcome events is less than per person years. rosuvastatin does not lead to an increased incidence of rhabdomyolysis, myopathy, acute renal failure and hepatic impairment compared to other statins. the aim: the aim of the study was to assess the influence of insulin resistance (ir) on the coronary artery disease (cad) occurrence in middle aged women with normal glucose tolerance (ngt) material and methods: in - year women aged - , participants of the polish multicenter study on diabetes epidemiology were examined. anthropometric, biochemical (fasting lipids, fasting and after glucose load plasma glucose and insulin) and blood pressure determinations were performed . ir was defined as the matsuda index (irmatsuda) below the lower quartile of the irmatsuda distribution in ngt population the questionnaire examination of the lifestyle, present and past diseases was performed. results: ir was observed in % of all examined women and in . % with ngt. cad was diagnosed in , % of all examined women and in , % of those with ngt. the relative risk of cad related to ir in ngt and normotensive women was , ( % ci: , - , ) (p< . ). regular menstruation was observed in , % of cad women. irmatsuda was not different for cad menstruating and non menstruating women (respectively , ± , and , ± , ). conclusion: in middle aged, normotensive and normal glucose tolerant women ir seems to be an important risk factor of cad abstract background: in germany, primary prevention at population level is provided by general practitioners (gp). little is known about gps' strategies to identify patients at high risk for vascular diseases using standardised risk scores. objectives: we studied gp attitudes and current practice in using risk scores. methods-a cross-sectional survey was conducted among gps in north rhine-westphalia, germany, using mailed self-administered questionnaires on attitudes and current practice in identification of patients at high risk for vascular diseases. results: in , gps participated in the study. . % of gps stated to know the framingham-score, . % the procam-score and . % the score-score. . % of gps reported regular use of standardised risk scores to identify patients at high risk for vascular diseases, most frequently procam-score ( . %), followed by score-score ( . %) and framingham-score ( . %). main reasons for not using standardised risk scores were assumed rigid assessment of individual patients' risk profile ( . %), time-consuming appliance ( . %) and higher confidence in own work experience ( . %). conclusion: use of standardised risk scores to identify patients at high risk for vascular diseases is common among gps in germany. however, more educational work might be useful to strengthen gps' belief in the flexible appliance of standardised risk scores in medical practice. among epilepsy patients than in general population, but effects of specific antiepileptic drugs on birth rate are not well known. objectives: to estimate birth rate in epilepsy patients on aed treatment or without aeds and in a population-based reference cohort without epilepsy. design and methods: patients (n = , ) with reimbursement for aeds for the first time between and and information on their aed use, were identified from the databases of social insurance institution of finland. reference cohort without epilepsy (n = , ) and information on live births were identified from the finnish population register centre. the analyses were performed using poisson regression modelling. results:birth rate was decreased in epilepsy patients in relation to reference cohort without epilepsy in both genders regardless of aed use. in relation to untreated patients, women on any of the aeds had non-significantly lower birth rates. among men, birth rate was decreased in men on oxcarbazepine (rr = . , % ci = . , . ), but was not clearly lower among those on carbamazepine (rr = . , % ci = . , . ) or valproate (rr = . , % ci = . , . ) when compared to untreated patients. conclusion: our results suggest that birth rate is decreased among epilepsy patients on aeds, more so in men. abstract background: hereditary hemochromatosis (hh), characterised by excessive iron absorption, subsequent iron storage and progressive clinical features, can when diagnosed at an early stage be successfully treated. high prevalence of the c y-mutation on the hfe-gene in the hh patient population may motivate genetic screening. objectives: in first-degree relatives of c y-homozygotes we studied the gender and age -related biochemical penetrance of hfe-genotype to define a high-risk population eligible for screening. design and methods: one-thousand-six first-degree family members of probands with clinically overt hfe-related hh from five medical centres in the netherlands were approached. data on levels of serum iron parameters and hfe-genotype were collected. elevation of serum ferritin was defined using the centre-specific normal-values by age and gender. results: among the participating relatives, highest serum iron parameters were found in male c y-homozygous siblings aged > years: % had elevated levels of serum ferritin. generally, male gender and increased age are related with higher iron values. discussion and conclusion: genetic screening for hh is most relevant in male and elderly first-degree relatives of patients with clinically overt hfe-related hh, enabling regular investigations of iron parameters in homozygous individuals. abstract background: nosocomial infection causes increased hospital morbidity and mortality rates. although handwashing is known to be the most important action in its prevention, adherence of health care workers to recommended hand hygiene procedures is extremely poor. objective: evaluation of compliance of hand hygiene recommendations in health care workers of a tertiary hospital in barcelona after a course on hand hygiene was given to all nurses in the hospital during the previous year. methods: by means of nondeclared observation, compliance (handwashing or disinfecting, not solely glove exchange) of recommendations given by the center for disease control related to opportunities for hand hygiene was registered, in procedures of diverse risk level for infection, both in physicians and nurses. results: in opportunities for hand hygiene carried out by health care workers compliance of recommendations was . %. adherence differed between wards ( . % in intensive care units, . % in medical wards and . % in surgical wards) and slightly between health care workers ( . % in physicians, . % in nurses). discussion: in conclusion, after one year of an intervention on education, adherence to hand hygiene recommendations is very low. these results enhance the need of reconsidering the type of interventions implemented. type of comorbidity affects qol most. objectives: we studied whether qol differed in subjects with dm with and without comorbidities. in addition, we determined differences in type of comorbidity. design and methods: cross-sectional data of dm patients, participants of a population-based dutch monitoring project on risk factors for chronic disease (morgen) were analyzed. qol was measured by the short form . we compared the means of subdimensions for dm patients with one comorbidity (cardiovascular diseases (cvd), musculoskeletal diseases (msd) and asthma/copd) to dm patients without this comorbidity, by regression analyses adjusted for age and sex. results: the prevalences of cvd, msd and asthma/copd were . %, . %, and . %. all comorbidities were associated with lower qol, especially for physical functioning. the mean difference ( % ci) was . abstract background: the extent or increase of ueds is suggested repeatedly, but never before the scientific literature was systematic studied. objectives: a systematic appraisal of the worldwide incidence and prevalence rates of upper extremity disorders (ueds) available in scientific literature was executed to gauge the range of these estimates in various countries and to determine whether the rates are increasing in time. design and methods: studies that recruited at least people, collected data by using questionnaires, interviews and/or physical examinations, and reported incidence or prevalence rates of the whole upper-extremity including neck, were included. results: no studies were found with regard to the incidence of ueds and studies that reported prevalence rates of ueds were included. the point prevalence ranged from . - %; the months prevalence ranged from . - %. one study reported on the lifetime prevalence ( %). we did not find evidence of a clear increasing or decreasing pattern over time. it was not possible to pool the date, because the definitions used for ueds differed enormously. conclusions: there are substantial differences in reported prevalence rates on ueds. main reason for this is the absence of a universally accepted way of labelling or defining ueds. abstract background: the absolute number of women diagnosed with breast cancer increased from , in to , in in the netherlands. likewise, the age standardized rate increased from . to . per , women. besides the current screening programme, changes in risk profile could be a reason for the increased incidence. objective: we studied the changes in breast cancer risk factors for women in nijmegen. methods: in the regional screening programme in nijmegen, almost , women aged - years filled in a questionnaire about risk factors in [ ] [ ] . similar questions were applied in the nijmegen biomedical study in , where women of - year participated. the median age in both studies was years. results: the frequency of a first-degree relative with breast cancer was . % and . % in and , respectively . none of the other risk factors, as the age of women at st birth ( . % respectively . %), nulliparity ( . % resp. . %), age at menarche ( . % resp. . %), age at menopause ( . % resp. . %) and obesity ( . % resp. . %), changed in time. conclusion: the distribution of risk factors hardly changed, and is unlikely to explain the rise in breast cancer incidence from onwards. abstract background: a single electronic clinical history system has been developed in the bac (basque autonomous community) for general use for all health centres, thus making it possible to collect information online on acute health problems as well as chronic ailments. method: the prevalence of diabetes, high blood pressure and copd (chronic obstructive pulmonary disease) was estimated using icd- -cm diagnosis performed by primary care physicians. an estimate was also made of the prevalence of cholesterolemia based on the results of analyses requested by physicians. results: in , , patients (out of a total population of , , ) were assessed for serum cholesterol levels. based on this highly representative sample, it was estimated that . % had serum cholesterol levels above mg/dl. the prevalence of diabetes mellitus in people over the age of was . %. the prevalence of high blood pressure in people over was %. discussion: the primary care database makes it possible to access information on problems related to chronic illnesses. knowing the prevalence of diabetes patients enables doctors to analyse all aspects related to services used by the diabetic population. it also makes it possible to monitor analytical data in real time and evaluate health service outcomes. examinations were used to asses risk factors for diabetes. cases (n = ) were matched on age and sex to controls (n = ) who were not treated with antidiabetic drugs. logistic regression was used to calculate odds ratios (or). results: the or of incident diabetes for acei-use versus non-acei use was . ( %ci : . - . ). for ace dd homozygotes the or was . ( %ci: . - . ) and for ace-i allele carriers . ( %ci: . - . ). the interaction or was . ( %ci: . - . ). the agt and at r genotypes did not modify the association between acei use and diabetes. abstract background: lignans have antioxidant and estrogen-like activity, and may therefore lower cardiovascular and cancer risk. objective: we have investigated whether intake of four plant lignans (lariciresinol, pinoresinol, secoisolariciresinol, matairesinol) was inversely associated with coronary heart disease (chd), cardiovascular diseases (cvd), cancer, and all-cause mortality. design: the zutphen elderly study is a prospective cohort study in which men aged - y were followed for years. lignan intake was estimated using a recently developed database, and related to mortality using cox proportional hazards analysis. results: median total lignan intake in was lg/d. beverages such as tea and wine, vegetables, bread, and fruits were the major lignan sources. total lignan intake was not related to mortality. however, matairesinol was inversely associated with chd, cvd, cancer, and all-cause mortality. multivariate adjusted rrs ( % ci) per sd increase in intake were . ( . - . ) for chd, . ( . - . ) for cvd, . ( . - . ) for cancer, and . ( . - . ) for allcause mortality. conclusions: total lignan intake was not associated with mortality. the intake of matairesinol was inversely associated with mortality from chd, cvd, cancer, and all-causes. we can not rule out that this is due to an associated factor, such as wine consumption. abstract despite the drastic increase in the amount of research into neighbourhood-level contextual effects on health, studies contrasting these effects between different domains of health within one contextual setting are strikingly sparse. in this study we use multilevel logistic regression models to estimate the existence of neighbourhood-level variations of physical health functioning (pcs) and mental well-being (ghq) in the helsinki metropolitan area and assess the causes of these differences. the individual-level data are based on a health-survey of - year old employees of the city of helsinki (n = , response rate %). the metropolitan area is divided into neighbourhoods, which are characterised using a number of area-level indicators (e.g. unemployment rate). our results show moderate but systematic negative effect of indicators of neighbourhood deprivation on physical functioning, whereas for mental health the effect is absent. these effects were strongest for proportion of manual workers; odds ratio for poor physical functioning was . for respondents living in areas with low proportion of manual workers. part of this effect was mediated by differences in health behaviour. analyses on cross-level interactions show that individual-level socioeconomic differences in physical health are smallest in most deprived areas, somewhat contradicting the results of earlier studies. abstract background: the second-eye cataract surgery is beneficial, nevertheless, there is a considerable proportion of unmet needs. objective: to estimate the proportion of second-eye cataract surgery in the public health system of catalonia, and explore differences in utilisation by patients' gender, age, and region of residence. methods: a total of , senile cataract surgeries performed between and were included. proportions observed were adjusted through independent logarithmic regression models for each study factor. results: the proportion of second-eye surgery showed an increasing trend (r . %) from . % ( % ci . ; . ) in november to . % ( % ci . ; . ) in december , and its projection to years was , % ( % ci . ; . ). the proportion of second-eye surgery was % ( % ci . ; . ) greater in women than in men. patients years or older had a lowest proportion ( . %; % ic . ; . ), which nevertheless increased during the period, unlike that of patients aged less than years. differences among regions were moderate and decreased throughout the period. conclusions: if the observed trends persist, there will be a substantial proportion of unmet need for bilateral surgery. we predict greater use of second-eye surgery by older patients. abstract background: persistence with bisphosphonates is suboptimal which could limit prevention of fractures in daily practice. objectives: to investigate the effect of long term persistent bisphosphonate usage on the risk of osteoporotic fractures. methods: the pharmo database, including drug-dispensing and hospital discharge records for > two million subjects in the netherlands, was used to identify new female bisphosphonate users > years from jan ' -jun ' . persistence with bisphosphonates was determined using the method of catalan. a nested matched case-control study was performed. cases had a first hospitalization for an osteoporotic fracture (index-date). controls were matched : to cases on year of inclusion and received a random index-date. the association with fracturerisk was assessed for one and two year persistent bisphosphonate use prior to the index-date. analyses were adjusted for di fferences in patient characteristics. results: , bisphosphonate users were identified and had a hospitalization for osteoporotic fracture during follow-up. one year persistent bisphosphonate use resulted in a % lower fracture rate (or . ; % ci . - . ) whereas two year persistent use resulted in a % lower rate (or . ; % ci . - . ). conclusion and discussion: these results emphasize the importance of persistent bisphosphonate usage to obtain maximal protective effect of treatment. abstract background: in the who recommended all countries to add hepatitis b (hbv) vaccination to their national immunization programs. the netherlands is a low hbv endemic country and therefore adopted a vaccination policy targeted towards high-risk groups. methods: during , epidemiological data and blood samples were collected from all reported patients with an acute hbv infection. a fragment of the s-gene was sequenced and phylogenetically analysed to clarify transmission patterns between risk groups. results: of hbv cases reported, % was infected through sexual contact ( % homo-/bisexual, % heterosexual). for patients samples were available for genotyping. phylogenetic analysis identified genotypes: a( %), b( %), c( %), d( %), e( %) and f( %). of men who have sex with men (msm), % were infected with genotype a. among heterosexuals, all genotypes were found. in many cases, genotypes b-f were direct or indirect related to countries abroad. only injecting drug user was found (genotype a). conclusion: genotype a is predominant in the netherlands, including most of the msm. migrant hbv carriers play an important role in the dutch hbv epidemic. genotyping provides insight into the spread of hbv among highrisk groups. this information will be used to evaluate the vaccination policy in the netherlands. abstract background: excess weight might affect the perception of both physical and mental health in women. objective: to examine the relationship between body mass index (bmi) and hrqol in women aged -to -year-old in a rural zone of galicia. design and methods: population-based cross-sectional study covering women, personally interviewed, from villages. hrqol was assessed with sf- questionnaire, through personal interviews. each scale of sf- was dichotomised in suboptimal or optimal hrqol using previously defined cut-offs. odds ratios (or) obtained from logistic regression summarize the relationship of bmi with each scale, adjusting for sociodemographic variables, sedentary leisuretime, number of chronic diseases and sleeping hours. results: a . % of women were obese (bmi = kg/m ) and . % overweight kg/m ) . frequency of suboptimal physical function was higher among overweight women (adjusted or: . ; % ci: . - . ) and obesity (adjusted or: . ; % ci: . - . ). furthermore, obese women had higher frequency of suboptimal scores on the general health scale (adjusted or: . ; % ci: . - . ). no differences were observed regarding mental health scores among women with different bmi categories. conclusion: in women from rural villages, overweight is associated with worse hrqol in physical function and general health. abstract background: pneumococcal vaccination among elderly is recommended in several western countries. objectives: we estimate the cost-effectiveness of a hypothetical vaccination campaign among the + general population in lazio region (italy). methods: a cohort was followed during a years timeframe. we estimated the incidence of invasive pneumococcal disease, in absence of vaccine, based on actual surveillance and hospital data. the avoided deaths and cases have been estimated from literature according to trial results. health expenditures included: costs of vaccine program, inpatient and some outpatient costs. cost-effectiveness was expressed as net healthcare costs per episode averted and life-year gained (lyg) and was estimated at baseline and in deterministic and stochastic sensitivity analyses. all parameters were age-specific and varied according to literature data. results: at baseline net costs per event averted and lyg at prices were, respectively, e , ( % ci: e , -e , ) and , ( % ci: e , -e , ). in the sensitivity analysis, bacteraemic pneumonia incidence and vaccine effectiveness increased the net cost per lyg by % and % in the worst-case scenario, and decreased it to e , in the best-case. conclusions: the intervention was not cost saving. the uncertainties concerning invasive pneumococcal disease incidence and vaccine effectiveness make the cost-effectiveness estimates instable. spain - abstract background: spatial data analysis can detect possible sources of heterogeneity in spatial distribution of incidence and mortality of diseases. moreover small area studies have greater capacity to detect local effects linked to environmental exposures. objective: to estimate the patterns of cancer mortality at municipal level in spain using smoothing techniques in a single spatial model. design and methods: cases were deaths due to cancer, registered at a municipal level nation-wide for the period - . expected cases for each town were calculated using overall spanish mortality rates and standard mortality ratios were computed. to plot the maps, smoothed municipal relative risks were calculated using besag york and mollie`model and markov chain monte carlo simulation methods. as an example maps for stomach and lung cancer neoplasms are shown. results: it was possible to obtain the posterior distribution of relative risk by a single spatial model including towns and the adjacencies. maps showed the singular patterns for both cancer locations. conclusion: the municipal atlas allows to avoid edge local effects, improving the detection of spatial patterns. discussion: bayesian modelling is a very efficient way to detect spatial heterogeneity by cancer and other causes of death. abstract background: little is known about the impact of socioeconomic status (ses) on outcomes of surgical care. objectives: we estimated the association between ses and outcomes of selected complex elective surgical procedures. methods: using hospital discharge registries (icd-ix-cm codes) of milan, bologna, turin and rome we identified patients undergoing cardiovascular operations (coronary artery bypass grafting, valve replacement, carotid endarterectomy, repair of unruptured thoracic aorta aneurysm) (n = , ) and cancer resections (pancreatectomy, oesophagectomy, liver resection, pneumonectomy, pelvic evisceration) (n = , ) in four italian cities, - . an area-based income index was calculated. post-operative mortality (in-hospital or within days) was the outcome. logistic regression adjusted for gender, age, residence, comorbidities, concurrent and previous surgeries. results: high income patients were older and had fewer comorbidities. mortality varied by surgery type (cabg , %, valve , %, endartectomy , %, aorta aneurysm , %, cancer . %). low income patients were more likely to die after cabg (or = . abstract background: an important medical problem of renal transplant patients who receive immunosuppression therapy, is the development of a malignancy during the long term follow-up. however, existing studies are not in agreement over whether patients who undergo renal transplantation have an increased risk of melanoma. objective: the aim of this study was to determine the incidence of melanoma in renal transplantation patients in the northern part of the netherlands. methods: we linked a cohort of patients who received a renal transplantation in the university medical centre groningen between and with the cancer registry of the comprehensive cancer centre north-netherlands, to identify all melanoma patients in this cohort. results: only patient developed a melanoma following the renal transplantation; no significant increase in the risk of melanoma was found. conclusion: although several epidemiologic studies have shown that the risk of melanoma is increased in renal transplantation patients who receive immunosuppression therapy to prevent allograft rejection, this increased risk was not found in the present study. the lower level of immunosuppressive agents given in the netherlands might be responsible for this low incidence. abstract background: socio-economic health inequalities are usually studied for self-reported income, although the validity of self-reports is uncertain. objectives: to compare self-reports of income by respondents to health surveys with their income according to tax registries, and determine to what extent choice of income measure influences the health-income relation. methods: around . respondents from the dutch permanent survey on living conditions were linked to data from dutch tax and housing registries of . both self-reported and registry-based measures of household equivalent income were calculated and divided into deciles. the association with less than good self-assessed health was studied using prevalence rates and odds ratios. results: around % of the respondents did not report their income. around % reported an income deciles lower or higher than the actual income value. the relation between income and health was influenced by choice of income measure. larger health inequalities were observed with selfreports compared to registry-based measures. while a linear healthincome relation was found using self-reported income, a curvilinear relation (with the worst health in the second lowest deciles) was observed for registry-based income. conclusion: choice of the income source has a major influence on the health-income relation that is found in inequality research. abstract background: while many health problems are known to affect immigrant groups more than the native dutch population, little is known about health differences within immigrant groups. objectives: to determine the association between self assessed health and socioeconomic status (ses) among people of turkish, moroccan, surinamese and antillean origin. methods: data were obtained from a social survey held among immigrants - years in the netherlands, with almost respondents per immigrant group. ses differences in the prevalence of 'poor' self-assessed health were measured using prevalence rate ratios estimated with regression techniques. results: within each immigrant group, poor health was much more common among those with low ses. the health of women was related to their educational level, occupational position, household income, financial situation and (to a lesser extent) their parents' education. similar relationships were observed for men, except that income was the strongest predictor of poor health. the health differences were about as large as those known for the native dutch population. conclusion and discussion: migrant groups are not homogenous. also within these groups, low ses is related to poor general health. in order to identify subgroups where most health problems occur, different socioeconomic indictors should be used. abstract background: genetic damage quantification can be considered as biomarker of exposure to genotoxic agents and as early-effect biomarker regarding cancer risk. objectives: to assess genetic damage in newborns and its relationship with anthropometrical, sociodemographic variables, maternal tobacco consumption and pollution perception. design and methods: the bio-madrid study recruited trios (mother/father/newborn) from areas in madrid to assess the impact of pollutants in humans. parents answered a questionnaire about socio-economic characteristics, pregnancy, life-style and perception of pollution. genetic damage in newborns were measured with the micronucleus(mn) test in peripheral lymphocytes poisson regression models were fitted using mn frequency per binucleated cells as dependent variable. explanatory variables included sex, parents age, tobacco, area and reported pollution level. results: the mean frequency of mn was . per (range: - ). no differences were found regarding area, sex and maternal tobacco consumption. mn frequency was higher in underweighted newborns and in those residing near heavy traffic roads. in recent years minimally invasive surgery procedures underwent rapid diffusion and laparoscopic cholecystectomy has been among the first to be introduced. after its advent, increasing rates of overall and laparoscopic cholecystectomy have been observed in many countries. we evaluated the effect of the introduction of laparoscopic procedure on the rates of cholecystectomy in friuli venezia giulia region, performing a retrospective study. from regional hospitals discharge data we selected all records with procedure code of laparoscopic (icd cm: ) or open ( ) cholecystectomy and diagnosis of uncomplicated cholelithiasis (icd cm: . ; . ; , ) or cholecystitis ( , ; , ), in any field, from to . in the year study period, the number of overall cholecystectomies increased from to (+ , %), mainly for the relevant increase of laparoscopic interventions from procedures, ( , % of overall cholecystectomies), to ( , %). rates of laparoscopic cholecystectomies increased from , to , per admitted patients with diagnosis of cholelithiasis or cholecystitis. the introduction of laparoscopic cholecystectomies was followed not only by a shift towards laparoscopically performed interventions but also by an increase in overall cholecystectomies in friuli venezia giulia region. abstract background: although a diminished doses scheme of -valent pneumococcal conjugate vaccination (pcv ) may offer protection against invasive pneumococcal disease, it might affect pneumococcal carriage and herd immunity. long term memory has to be evaluated. objective: to compare the influence of a and -doses pcv -vaccination scheme on pneumococcal carriage, transmission, herd immunity and anti-pneumococcal antibody levels. methods: in a prospective, randomized, controlled trial infants are randomly allocated to receive pcv at ages and months; ages , and months and the age of months only. nasopharyngeal (np) swabs are regularly obtained from infants and family members. the np swabs are cultured by conventional methods and pneumococcal serotypes are determined by quellung reaction. antibody levels are obtained at and months from infants in group i and ii and from infants in group iii. one thousand infants are needed to detect a % difference in pneumococcal carriage (a = . , ß = . ) between the three groups. results: so far, infants have been included. preliminary results show that prior to vaccination pneumococcal carriage was %. conclusion: this trial will provide insight into the effects of a diminished dose scheme on herd immunity and long-term antipneumococcal antibody development. abstract background: oil-spills cause important environmental damages and acute health problems on affected populations. objectives: to assess the impact of the prestige oil-spill in the hrqol of the exposed population. design and methods: we selected residents in coastal areas heavily affected by the oil-spill and residents in unaffected inland villages through random sampling, stratified by age and sex. hrqol was measured with the sf- questionnaire in personal interviews. individual exposure was also explored. mean differences in sf- scores > points were considered 'clinically relevant'. odds ratios (or) summarized the association between area of residence (coast vs inland) and suboptimum hrqol (lower than percentile th), adjusting for possible confounders. results: neither clinically relevant nor statistically significant differences were observed in most of the sf- scales regarding place of residence or individual exposure. worse scores (inland = , ; coast = , ; p< , ) abstract background: patient comorbidities are usually measured and controlled in health care outcome research. hypertension is one of the most commonly used comorbidity measures. objectives: this study aims to assess underreporting of hypertension in ami patients, and to analyze the impact of coding practices among italian regions or hospitals' type. methods: a cohort of ami hospitalisations in italy from november to october was selected. patients with a previous hospital admission reporting a diagnosis of complicated hypertension within the preceding months were studied. a logistic model was constructed. both crude and adjusted probability of reporting a hypertension in ami admissions, depending from the number of diagnosis fields compiled in discharge abstracts, and presence of other diseases were estimated. results: in . % of patients hypertension was not reported. probability of reporting hypertension increased with the number of compiled diagnosis fields (adjusted ors range: . - . ). there were no significant differences among italian regions, while private hospitals' reporting was less accurate. disorders of lipoid metabolism were more probably coded with hypertension (adjusted or: . ). conclusions: information from both ami and previous hospitali-sations would be needed to include hypertension in a comorbidity measure. abstract background: the angiotensin converting enzyme inhibitors (acei) should be considered the standard initial treatment of the systolic heart failure. this treatment is not recommended in patients with hypotension, although figures of systolic blood pressure around - mmhg during the treatment are allowed if the patient remains asymptomatic. objectives: to know the proportion of patients with systolic heart failure receiving treatment with acei, and the proportion of these patients with signs oh hypotension. design and methods: the electronic clinical records of all the patients diagnosed of systolic heart failure were reviewed. the electronic information system covers a % of the population of the basque country, approximately. diagnosis of heart failure was defined as the presence of any of the following cie- codes: or . or . . to evaluate the blood pressure, the last available determination was considered. results: out of patients with left heart failure, ( . %) have been prescribed acei. among the patients with blood pressure lower than mmhg (systolic) or than mmhg (diastolic), ( . %) were also receiving this treatment. conclusions: acei are clearly underprescribed in the basque country for the treatment of heart failure. attention should be given to the group at risk of hypotension. abstract background: epidemiologic studies have shown an association between c-reactive protein (crp) and cardiovascular endpoints in population samples. methods: in a longitudinal study of myocardial infarction (mi) survivors, crp was measured repeatedly (up to times) within a period of months. data on disease history and life style were collected at baseline. we examined the association between different variables and the level of crp using a random effects model. results: in total crp samples were collected in athens, augsburg, barcelona, helsinki, rome and stockholm. mean levels of crp were . , . , . , . , . , . [mg/l] respectively. body mass index (bmi) and chronic bronchitis (ever diagnosed) had the largest effect on crp ( % (for kg/m ) and % change from the mean level, respectively, p< . ). age classes showed a cubic function with a minimum at ages to . glycosylated hemoglobin (hba c) < . % as a measure of long-term blood glucose control and being male were found to be protective () % and ) % respectively, p< . ). conclusion: it was shown that bmi and history of bronchitis are important in predicting the level of crp. other variables, like alcohol intake, play a minor role in this large sample of mi patients. abstract background: during the last decades a remarkable increase in incidence rates of malignant lymphoma was seen. although some reasons are known or suspect underlying risk factors are not well understood. objectives: we studied the influence of medical radiation (x-ray, radiotherapy and szintigraphy) on the risk of malignant lymphoma. methods: we analysed data from a population-based case-control study with incident lymphoma cases in germany from - . after informed consent cases were pair-matched with controls recruited from registration office by age, gender and study region. data was collected in a personal interview. we analysed data using conditional logistic regression. results: the linear model shows an or = . /msv due to x-ray exposure and or = . ( %-ci = . - . ) comparing higher with lower exposure. radiotherapy shows an or = . (n = cases). there is no association between all lymphomas and szintigraphies but in the subgroup containing multiple myeloma, cll, malt-and marginalcell lymphoma we found an or = . ( %-ci = . - . ) in the multivariate model. discussion: no excess risk was observed for x-ray examinations. ionising radiation may increase risk for specific lymphoma subgroups. however, it should be noted that numbers in the subgroups are small and that radiation dose may be somehow inaccurate as no measures were available. abstract background: varus-alignment (bow-leggedness) is assumed to correlate with knee osteoarthritis (oa), but it is unknown whether varus-alignment precedes the oa or whether varus-alignment is a result of oa. objective: to assess the relationship between varusalignment and the development, as well as progression, of knee oa. methods: , participants in the rotterdam study were selected. knee oa at baseline and at follow-up (mean follow-up . years) was defined as kellgren & lawrence (k&l) grade , and progression of oa as an increase of k&l degree. alignment was measured by the femoro-tibial angle on baseline radiographs. multivariable logistic regression for repeated measurements was used. results: of , knees, . % showed normal alignment, . % varus-alignment, and . % valgus-alignment. comparison of high varus-alignment versus normal, low and mediate varus-alignment together, showed a two-fold increase in the development of knee oa. (or = . ; %ci = . - . ). the risk of progression was higher in the high varus group compared to the normal, low and mediate varus group (or = . ; %ci = . - . ). stratification for overweight gave similar odds ratio's in the overweight group, but weaker odds ratio's in the non-overweight group. conclusion: a higher value of varus-alignment is associated with the onset and progression of knee oa. abstract background: echocardiographic image quality in copd patients can be hampered by hyperinflated lungs. cardiovascular magnetic resonance imaging (cmr) may overcome this problem and provides accurate and reproducible information about the heart without geometric assumptions. objective: to determine the value of easily assessable cmr parameters compared to other diagnostic tests in identifying heart failure (hf) in copd patients. design and methods: participants were recruited from a cohort of copd patients = years. a panel established the diagnosis of hf during consensus meetings using all diagnostic information, including echocardiography. in a nested case-control study design, copd patients with hf (cases) and a random sample of copd patients without hf (controls) received cmr. the diagnostic value of cmr for diagnosing hf was quantified using univariate and multivariate logistic modelling and roc-area analyses. results: four easily assessable cmr measurements had significantly more added diagnostic value beyond clinical items (roc-area . ) than amino-terminal pro b-type natriuretic peptide (roc-area . ) or electrocardiography (roc-area . ). a 'cmr model' without clinical items had an roc-area of . . conclusion: cmr has excellent capacities to establish a diagnosis of heart failure in copd patients and could be an alternative for echocardiography in this group of patients. abstract background: the prevalence of overweight (i.e, body mass index [bmi] > = kg/m ) is increasing. new approaches to address this problem are needed. objectives: ) to assess the effectiveness of distance counseling (i.e., by phone and e-mail/internet) on body weight and bmi, in an overweight working population. ) to assess differences in effectiveness of the two communication methods. design and methods: overweight employees ( % male; mean age . ± . years; mean bmi . ± . kg/m ) were randomized to a control group receiving general information on overweight and lifestyle (n = ), a phone based intervention group (n = ) and an internet based intervention group (n = ). the intervention took months and used a cognitive behavioral approach, addressing physical activity and diet. the primary outcome measures, body weight and bmi, were measured at baseline and at six months. statistical analyses were performed with multiple linear regression. results: the intervention groups (i.e., phone and e-mail combined) lost . kg (bmi reduced by . kg/m ) over the control group (p = . ). the phone group lost . kg more than the internet group (p = . ). abstract objective: although an inverse gradient education-mortality has been shown in the general population, little is known about this trend in groups with higher risks of death.we examine differences in mortality by education and hiv-status among injecting drug users (idus) before and after introduction of highly active antiretroviral therapy (haart) in . methods: communitybased cohort study of idus recruited in aids prevention centres ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) abstract background: pancreatic cancer is an aggressive cancer with low survival time, with health-related quality of life (hrqol) being of major importance. objectives: the aim of our study was to assess both generic and disease-specific hrqol in patients with pancreatic cancer. methods: patients with suspected pancreatic cancer were consecutively included at admission to the hospital. hrqol was determined with the disease-specific european organization for research and treatment of cancer (eortc) health status instrument and generic euroqol (eq- d). results: a total of patients (mean age years ± , % men) were admitted with suspected pancreatic cancer. of these patients, ( %) had pancreatic cancer confirmed as final diagnosis. hrqol was significantly impaired in patients with pancreatic cancer for most eortc and eq- d scales in comparison to norm populations. the ed- d visual analogue scale (vas) and utility values were significantly correlated to the five functional scales, to the global health scale and to some but not all of the eortc symptom scales/items. conclusions: hrqol was severely impaired in patients with pancreatic cancer. there was a significant correlation between most eortc and eq- d scales. our results may facilitate further economic evaluations and aid health policy makers in resource allocation. abstract background: organised violence has health impact both on those who experience the violence directly and indirectly. the numbers of people affected by mass violence is alarming. substantial knowledge on the long-term health impact of organized violence is of importance for public health and for epidemiology. objectives: to investigate research results of long term mental health impact of organised violence. design and methods: a search of papers for the keywords genocide, organised violence, transgenerational effects, mental health was carried out in pubmed, science citation index and psychinfo. results: the systematic review on the long-term health impact of genocide showed that exposure to organised violence has an impact on mental health. methodological strenghts and weaknesses varied between studies. the found mental health consequences were associated with the country of research and the time of study. overall data showed organised violence has transgenerational impact on mental health of individuals and societies. conclusion: longitudinal studies have to be carried out to get further insight into the long-term health effects of organised violence. discussion: research results on mental health effects of organised violence have to be analysed in the context of changing concepts of illness. overweight is increasing and associated with various health problems. there are no well-structured primary care programs for overweight available in the netherlands. therefore, we developed a -month multidisciplinary treatment program in a primary care setting. the aim of the present study is to determine the feasibility and efficacy of a multidisciplinary treatment program on weight loss and risk profile in an adult overweight population. hundred participants of the utrecht health project are randomised to either a dietetic group or a dietetic plus physiotherapy group. the control group consist of another participants recruited from the utrecht health project and receives routine health care. body weight, waist circumference, blood pressure, serum levels, energy-intake and physical activity are measured at baseline, halfway and at the end of the treatment program. feasibility of the treatment program is assessed by response, compliance and program-associated costs and workload. efficacy is determined by analysing changes in outcome measures between groups over time using t-tests and anova repeated measurements. the treatment program is considered effective with at least a % difference in mean weight change over time between groups. positive evaluation of the multidisciplinary treatment program for overweight may lead to implementation in routine primary health care. abstract background: examining patient's quality of life (qol) before icu admission will permit to compare and analyze its relation with other variables. objectives: analyze qol of patients admitted to a surgical icu before admission and study its relation with baseline characteristics and outcome. design and methods: the study was observational and prospective in a surgical icu, enrolling all patients admitted between november and april . baseline characteristics of patients, history of co morbidities and quality of life survey score (qolss) were recorded. assessment of the relation between each variable or outcome and the total score of qolss was performed by multiple linear regression. results: total qolss demonstrated worse qol in patients with hypertension, cerebrovascular disease, renal insufficiency, severely ill (as measured by saps and asa physical status), and in older patients. there was no relation between qol and longer icu los. conclusions: preadmission qol correlates with age, severity of illness, comorbidities and mortality rates but is an able to predict longer icu stay. discussion: qolss appears to be a good indicator of outcome and severity of illness. abstract background: transient loss of consciousness (tloc) has a cumulative lifetime incidence of %, and can be caused by various disorders. objectives: to assess the yield and accuracy of initial evaluation (standardized history, physical examination and ecg), performed by attending physicians in patients with tloc, using follow-up as a gold standard. design and methods: adult patients presenting with tloc to the academic medical centre between february and may were included. after initial evaluation physicians made a certain, likely or no initial diagnosis. when no diagnosis was made additional cardiological testing, expert history taking and autonomic function testing were performed. the final diagnosis, after years follow-up, was determined by an expert committee. results: patients were included. after initial evaluation, % of the patients were diagnosed with a certain and % with a likely cause for their episodes. overall diagnostic accuracy was % ( %ci - %); % ( %ci - %) for the certain diagnoses and % ( %ci - %) for the likely diagnoses. conclusion and discussion: attending physicians make a diagnosis in % of patients with tloc after initial evaluation, with high accuracy. the use of abundant additional testing can be avoided in many patients. abstract background: the possibility of an influenza pandemic is one of the major public health challenges of today. risk perceptions among the general public may be important for successful public health measures to better control an outbreak situation. objectives: we investigated risk perception and efficacy beliefs related to an influenza pandemic in the general population in countries in europe and asia. design and methods: telephone interviews were conducted in . risk perception of an influenza pandemic was measured on a -point scale and outcome-and self-efficacy on a point scale (low-high). the differences in risk perception by country, sex and age were assessed with a general linear model including interaction effects. results: , persons were interviewed. the mean risk perception of flu was . and was significantly higher in europe ( . ) compared to asia ( . ) (p< . ) and higher in women ( . ) than men ( . ) (p< . ). outcome-and self-efficacy were lower in europe than asia. conclusion: in europe higher risk perceptions and lower efficacy beliefs were found as compared to asia. in developing preparedness plans for an influenza pandemic specific attention should therefore be paid to risk communication and how perceived self-efficacy can be increased. abstract background: increased survival of patients with cf has prompted interest towards their hrqol. objectives: .to measure hrqol and its predictors in cf patients cared for at the bambino gesuc hildren's hospital in rome; . to assess the psychometric properties of the italian version of the cf specific hrqol instrument (cystic fibrosis questionnaire, cfq). design and methods: crosssectional survey. all cf patients aged years or more were asked to complete the cfq (age-specific format). psychological distress was assessed through standardized questionnaires in patients (achenbach and general health questionnaire, ghq) and their parents (ghq and sf- ). results: one-hundred-eighteen patients ( males, females, age range to years) participated in the study (response rate %). internal consistency of cfq was satisfactory (cronbach alpha from . to . ); all item-test correlation were greater than . . average cfq standardized scores were very good in all domains (> on a - scale), except perceived burden of treatments ( ) and degree of socialization ( ). multiple regression analysis was performed to identify factors associated with different hrqol dimensions. conclusion: support interventions for these patients should concentrate on finding a balance between need to prevent infections and promotions of adequate, age-appropriate social interactions. abstract background: the metabolic syndrome (metsyn) -a clustering of metabolic risk factors with diabetes and cardiovascular diseases as the primary clinical outcomes -is thought to be highly prevalent with an enormous impact on public health. to date, consistent data in germany are missing. objective: the study was conducted to examine the prevalence of the metsyn (according to ncap atp iii-definition) among german patients in primary care. methods: the german-wide cross-sectional study run two weeks in october with randomly selected general practitioners included. blood glucose and serum lipids were analyzed, waist circumference and blood pressure assessed, data on smoking, dietary and exercise habits, regional and sociodemographic characteristics collected. abstract background: excessive infant crying is a common and often stress inducing problem than can ultimately result in child abuse. from previous research is known that maternal depression during pregnancy is related to excessive crying, but so far little attention is paid to paternal depression. objective: we studied whether paternal depression is independently associated to excessive infant crying. design and methods: in a prospective multiethnic population-based study we obtained depression scores of , mothers and , fathers at weeks pregnancy using the brief symptom inventory, and information on crying behaviour of , infants at months. we used logistic regression analyses in which we adjusted for depression of the mother, level of education, smoking and alcohol use. results: paternal depressive symptomatology was related to the widely used wessel's criteria for excessive crying (adusted odds ratio . , . - . ). conclusion: our findings indicate that paternal depressive symptomatology might be a risk factor for excessive infant crying. discussion genetic as well as other direct (e.g. interaction between father and child) or indirect (e.g. marital distress or poor circumstances) mechanisms could explain the found association. abstract background: in studying genetic background of congenital anomalies the comparison of affected cases to non-affected controls is popular method. investigation of case-parent triads uses observation of cases and their parents exclusively. methods: both casecontrol approach and log-linear case-parent triads model were implemented to spina bifida (sb) cases and their parents ( triads) and controls in analysis of impact of the c t and a c mthfr polymorphisms on occurrence of sb. results: observed frequencies for tt genotype were , % in sb children, , % in mothers, , % in fathers, , % in controls and for cc genotype were , % of sb children, , % of mothers, , % of fathers and , % of controls. both genotype frequencies in sb triads did not differ significantly from controls. case-control approach showed nonsignificant increase in risk of having sb for t allele carriers either in homozygous (or = , ) or heterozygous form (or = , ) and for c allele carriers in heterozygous form (or = , ). log-linear model revealed significant relative risk of sb in children with both tt and ct genotype (rr = , and rr = , respectively). child's genotype at a c and mother's genotypes did not contribute to the risk. conclusions: caseparent triads approach adds new information regarding impact of parental imprinting on congenital anomalies. abstract background: previous studies showed an association of autonomic dysfunction with coronary heart disease (chd) and with depression as well as an association of depression with chd. however, there is limited information on autonomic dysfunction as potential mediator of the adverse effect of depression on chd. objectives: to examine the role of autonomic dysfunction as a potential mediator of the association of depression with chd. design/ methods: we used data of participants aged - years of the ongoing population-based cross-sectional carla study ( % male). time-and frequency-domain parameters of heart rate variability (hrv) as a marker of autonomic dysfunction were calculated. prevalent myocardial infarction (mi) was defined as selfreported physician-diagnosed mi or diagnostic minnesota code in the electrocardiogram. depression was defined based on the cesd-depression scale. logistic regression was used to assess associations between depression, hrv and mi. results: in ageadjusted logistic regression models, there was no statistically significant association of hrv with depression, of depression with mi, or of hrv with mi in men and women. discussion/conclusion: the present analyses do not support the hypothesis of an intermediate role of autonomic dysfunction on the causal path from depression to chd. abstract background: hypertension is an established risk factor for cardiovascular disease. however, prevalence of untreated or uncontrolled hypertension is often high (even in populations at high risk). objectives: to assess the prevalence of untreated and of uncontrolled hypertension in an elderly east german population. design and methods preliminary data of a cross-sectional, populationbased examination of men and women aged - years were analysed. systolic (sbp) and diastolic blood pressure (dbp) were measured and physician-diagnosed hypertension and use of antihypertensive drugs were recorded. prevalence of hypertension was calculated according to age and sex. results: of all participants, . % were hypertensive ( . % of men, . % of women). of these, . % were untreated, . % treated but uncontrolled, and . % controlled. women were more often properly treated than men. the prevalence of untreated hypertension was highest in men aged - years ( . %) and lowest in men and women aged > = years ( . %). uncontrolled hypertension increases with age in both sexes. conclusion and discussion: in this elderly population, there is a high prevalence of untreated and uncontrolled hypertension. higher awareness in the population and among physicians is needed to prevent sequelae such as cardiovascular disease. abstract background: exposure to pesticides is a potential risk factor for subfertility, which can be measured by time-to-pregnancy (ttp). as female greenhouse workers constitute a major group of workers exposed to pesticides at childbearing age, a study was performed among these and a non-exposed group of female workers. objectives: to measure the effects of pesticide exposure on time-topregnancy. design and methods: data were collected through postal questionnaires with detailed questions on ttp, lifestyle factors, and work tasks (e.g. application of pesticides, re-entry activities, and work hours) during six months prior to conception of the most recent pregnancy. associations between ttp and exposure to pesticides were studied in cox's proportional hazards models among female greenhouse workers and referents. results: the initial fecundability ratio (fradjusted) for greenhouse workers versus referents was . ( %ci: . - . ). this fr proved to be biased by the reproductively unhealthy worker effect. restricting the analyses to fulltime workers only gave an fradjusted of . ( %ci: . - . ). among primigravidous greenhouse workers, an association was observed between prolonged ttp and gathering flowers (fr = . , %ci: . - . ). conclusion and discussion: this study adds some evidence to the hypothesis of adverse effects of pesticide exposure on time-topregnancy, but more research is needed. abstract background: hfe-related hereditary hemochromatosis (hh) is an iron overload disease for which screening is recommended to prevent morbidity and mortality. however, discussion has risen on the clinical penetrance of the hfe-gene mutations. objective: in the present study the morbidity and mortality of families with hferelated hh is compared to a normal population. methods: c yhomozygous probands with clinically overt hfe-related hh and their first-degree relatives filled in a questionnaire on health, diseases and mortality among relatives. laboratory results on serum iron parameters and hfe-genotype were collected. the self-reported morbidity, family mortality and laboratory results were compared with an age and gender matched subpopulation of the nijmegen biomedical study (nbs), a population-based survey conducted in the eastern part of the netherlands. results: twohundred-twenty-eight probands and first-degree relatives participated in the hefas. serum iron parameters were significantly elevated in the hefas population compared to the nbs controls. also, the morbidity within hefas families was significantly increased for fatigue, hypertension, liver disease, myocardial infarction, osteoporosis and rheumatism. mortality among siblings, children and parents of hefas probands and nbs participants was similar. discussion: the substantially elevated morbidity within hefas families justifies further exploration for a family cascade screening program for hh in the netherlands. abstract objectives: to evaluate awareness levels and effectiveness of warning labels in cigarette packs, among portuguese students enrolled in the th to the th grades. design and methods: a cross sectional-study was carried out in may ( ) in a high school population ( th- th grades) in the north of portugal (n = ). a confidential self-reported questionnaire was administered. warning labels effectiveness was evaluated by changes in smoking behaviour and cigarette consuption, during the period between june/ (before the implementation of the tobacco warnings labels in portugal) and may/ . continuous variables were compared by the t-test for paired samples and kruskal-wallis test. crude and adjusted odds ratios and confidential intervals were calculated by logistic regression analysis. results: the majority of students ( . %) have a high level of awareness about warning labels content. this knowledge was significantly associated with school grade and current smoking status. none of these variables was significantly associated with changes in smoking behaviour. although not reaching statistic significance, the majority of teenagers ( . %) increased or kept their smoking pattern. awareness level was not associated with smoking prevalence or consumption decreases. conclusions: current warning labels are ineffective in changing smoking behaviour among portuguese adolescents. abstract background: injuries are an important cause of morbidity. the presence of pre-existing chronic conditions (pecs) have been shown to be associated with higher mortality. objectives: aim of this study is to evaluate the association between pecs and risk of death in elder trauma patients. methods: an injury surveillance, based on the integration between emergency, hospital, and mortality databases of lazio region, year , was used. patients were the elder people visited at the emergency departments, and hospitalised. pecs were evaluated on the basis of the charlson comorbidity index (cci). to measure the effect of pecs on the probability of death, we used logistic regression. results: patients were admitted to the hospital. the . % of the injured subjects were affected by one or more chronic conditions. risk of death for non urgent and urgent patients increased at increasing cci score abstract background: c-reactive protein (crp) was shown to predict prognosis in heart failure (hf). objective: to assess variability of crp over time in patients with stable hf. methods: we measured high-sensitivity crp (hscrp) times ( -week intervals) in patients with stable hf. patients whose hscrp was > mg/dl or whose clinical status deteriorated were excluded. two consecutive hscrp measurements were available for patients: men, mean(sd) age . ( . ) years, % depressed left ventricular systolic function. forty-four patients had a third measurement. using the cutoff point of . mg/dl for prediction of adverse cardiac events we assessed the proportion of patients who changed risk category. results: median(p -p ) baseline hscrp was . mg/dl( . - . ). hscrp varied largely particularly for higher levels. the th and th percentiles of differences between first two measurements were ) . mg/dl and + . mg/dl. correlation coefficient between these measurements: . , p< . . eleven ( %) patients changed risk category, kappa = . , p< . . among patients whose first two measurements were concordant, . % changed category in third measurement, kappa = . , p< . . conclusion: large variability in hscrp in stable hf may decrease the validity of risk stratification based on single measurements. it remains to be demonstrated whether the pattern of change over time adds predictive value in hf patients. abstract background: instrumental variables can be used to adjust for confounding in observational studies. this method has not yet been applied with censored survival outcomes. objectives: to show how instrumental variables can be combined with survival analysis. design and methods: in a sample of patients with type- diabetes who started renal-replacement therapy in the netherlands between and , the effect of pancreas-kidney transplantation versus kidney transplantation on mortality was analyzed using region as the instrumental variable. because the hospital could not be chosen with this type of transplantation, patients can be assumed to be naturally randomized across hospitals. we calculated an adjusted difference in survival probabilities for every time point including the appropriate confidence interval (ci %). results: the -year difference in survival probabilities between the two transplantation methods, adjusted for measured and unmeasured confounders, was . (ci %: . - . ) favoring the pancreas-kidney transplantation. this is substantially larger than the intention-to-treat estimate of . (ci %: . - . ) where policies are compared. conclusion and discussion: instrumental variables are not restricted to uncensored data, but can also be used with a censored survival outcome. hazard ratios with this method have not yet been developed. the strong assumptions of this technique apply similarly with survival outcomes. . ] . sir of coronary heart disease was . [ %ci: . - . ] and remained significantly increased up to years of follow-up. cox regression analysis showed a . -fold ( % ci, . - . ) increased risk of congestive heart failure after anthracyclines and a . -fold ( % ci, . - . ) increased risk of coronary heart disease after radiotherapy to the mediastinum. conclusion: the incidence of several cardiac diseases was strongly increased after treatment for hl, even after prolonged follow-up. anthracyclines increased the risk of congestive heart failure and radiotherapy to the mediastinum increased the risk of coronary heart disease. abstract background: the concept of reproductive health is emerging as an essential need for health development. objectives: to know the opinions of parents, teachers and students about education of reproductive health issues to students of mid and high schools. design and methods: focus group discussions (fgd) as a qualitative research was chosen. a series of group discussions with participation of persons ( students, teachers, and parents) was held. each group had included to persons. results: all the participants noted to a true need in education of puberty health in order to provide essentials for pre-adolescent students to adopt the psycho-and somatic changes of puberty. however, a few fathers and a group of mothers believed that education of family planning is not suitable for students. a need for education of aids and marital problems for students was the major concern in all groups. the female students emphasized a need for programming counseling in pre-marital period. conclusion: essentials in puberty health, family planning, aids and marital problems should be provided in mid-and high schools in order to narrow the knowledge gap of the students. abstract background: the association between social support and hypertension in pregnancy remains controversial. objective: the objective of this study was to investigate whether level of social support is a protective factor against preeclampsia and eclampsia. design and methods: a case-control study was carried out in a public high-risk maternity hospital in rio de janeiro, brazil. between july -may , all cases, identified at diagnosis, and controls, matched on gestational age, were included in the study. participants were interviewed about clinical history, socio-demographic and psychosocial characteristics. the principal exposure was the level of social support available during the pregnancy, using the medical outcomes study scale. adjusted odds ratios were estimated using multivariate conditional logistic regression. results: multiparous women with a higher level of social support had a lower risk of presenting with preeclampsia and eclampsia (or = . ), although this association was not statistically significant ( % ci . - . ). in primiparous women, a higher level of social support was seen amongst cases (or = . ; % ci . - . ). an interaction between level of social support and stressful life events was not identified. these results contribute to increased knowledge of the relationship between preeclampsia and psychosocial factors in low-income pregnant and puerperal women. abstract background: current case-definitions for cfs/me are designed for clinical-use and not appropriate for health needs assessment. a robust epidemiological case-definition is crucial in order to achieve rational allocation of resources to improve service provision for people with cfs/me. objectives: to identify the clinical features that distinguish people with cfs/me from those with other forms of chronic fatigue and to develop a reliable epidemiological case-definition. methods-primary care patient data for unexplained chronic fatigue was assessed for symptoms, exclusionary and comorbid conditions and demographic characteristics. cases were assigned to disease and non-disease groups by three members of the chief medical officer's working group on cfs/me (reliability-cronbach's alpha . ). results: preliminary multivariate analyses were conducted and classification and regression tree analysis included a -fold cross-validation approach to prevent over fitting. the results suggested that there were at least four strong discriminating variables for cfs/ me with 'post-exertional malaise' being the strongest predictor. risk and classification tables showed an overall correct classification rate of . %. conclusion: the analyses demonstrated that the application of the combination of the four discriminating variables (the defacto epidemiological case-definition) and predefined comorbid conditions had the ability to differentiate between cfs/me and non-cfs/me cases. abstract background: infection with high-risk human papillomavirus (hpv) is a necessary cause for cervical cancer. vaccines against the most common types (hpv , hpv ) are being developed. relatively little is known about factors associated with hpv or hpv infection. we investigated associations between lifestyle factors and hpv and hpv infection. methods: uk women aged - years with a recent abnormal cervical smear underwent hpv testing and completed a lifestyle questionnaire. hpv and hpv status was determined using type-specific pcrs. associations between lifestyle factors and hpv status were assessed by multivariate logistic regression models. results: . % ( %ci . %- . %) of women were hpv -positive. . % ( % ci . %- . %) were hpv -positive. for both types, the proportion testing positive decreased with increasing age, and increased with increasing grade of cytological abnormality. after adjusting for these factors, significant associations remained between (i) hpv and marital, employment, and smoking status and (ii) hpv and marital status and contraceptive pill use. gravidity, ethnicity, barrier contraceptive use and socio-economic status were not related to either type. conclusions we identified modest associations between several lifestyle factors and hpv and hpv . studies of this type help elucidate hpv natural history in different populations and will inform development of future vaccine delivery programmes. in ageing men testosterone levels decline, while cognitive function, muscle and bone mass, sexual hair growth, libido and sexual activity decline and the risk of cardiovascular diseases increase. we set up a double-blind, randomized placebo-controlled trial to investigate the effects of testosterone supplementation on functional mobility, quality of life, body composition, cognitive function, vascular function and risk factors, and bone mineral density in older hypogonadal men. we recruited men with serum testosterone levels below nmol/l and ages - years. they were randomized to either four capsules of mg testosterone undecanoate (tu) or placebo daily for weeks. primary endpoints are functional mobility and quality of life. secondary endpoints are body composition, cognitive function, aortic stiffness and cardiovascular risk factors and bone mineral density. effects on prostate, liver and hematological parameters will be studied with respect to safety. measure of effect will be the difference in change from baseline visit to final visit between tu and placebo. we will study whether the effect of tu differs across subgroups of baseline waist girth, testosterone level, age, and level of outcome under study. at baseline, mean age, bmi and testosterone levels were (yrs), (kg/m ) and .x (nmol/l), respectively. abstract at a student population, the carie's prevalence was , %. objectives: to evaluate the efficiency between two types of oral health education programmes and the adherence towards tooth brushing. study design: case control study: youngsters took part, in the case group. an health education programme was carried out in schools and included two types of strategies: a participative strategy (learning based on the colouring of the dental plaque) towards a case group; and a traditional strategy (oral expository method) towards a control group. during the outcome of the programmes, the oral health condition evaluation was done through cpo index, the adherence towards tooth brushing and the (iho's) oral hygiene index. results: in the initial dental exam the (iho) average was of , . three months after the application of the oral health programme, there was a general decrease in the average of iho's to , . discussion and conclusion: in the case group the decrease was higher: , to , . the students submitted to a session of oral health education based on the colouring of the dental plaque showed an lower iho's average and higher knowledge. this can be due to the teaching session being more active, participative and demonstrative. abstract background: violence perpetuated by adolescents is a major problem in many societies. objectives: the aim of this study is to examine high school students' violent behaviour and to identify predictors. design and methods: a cross-sectional study was conducted in timis county, romania between may-june . the sample consisted of randomly selected classes, stratified proportionally according to grades - , high school profile, urban and rural environment. the students completed a self administered questionnaire in their classroom. a weighting factor was applied to each student record to adjust for non-response and for the varying probabilities of selection. results: a total of students were included in the survey. during the last months, . % of adolescents got mixed into a physical fight outside school and . % on school property. abstract background: drug use by adolescents has become an increasing public health problem in many countries. objectives: the aim of this study is to identify prevalence of drug use and to examine high school students' perceived risks of substance use. design and methods: a cross-sectional study was conducted in timis county, romania between may-june . the sample consisted of randomly selected classes, stratified proportionally according to grades - , high school profile, urban and rural environment. the students completed a self administered questionnaire in their classroom. eighteen items regarding illicit drug use suggesting different intensity of use were listed. the response categories were 'no risk', 'slight risk', 'moderate risk', 'great risk' and 'don't know'. results: a total of students were included in the survey. the lifetime prevalence of any illicit drug was . %. significant beliefs associated with drug use are: trying marijuana once or twice (p< . ), smoking marijuana occasionally (p = . ), trying lsd once or twice (p = . ), trying cocaine once or twice (p = . ), trying heroine once or twice (p = . ). conclusion: the overall drug use prevalence is small. however, use of some drugs once or twice is not seen as a very risky behaviour. abstract background: the health ombudsman service was created in ceara´, brazil, in , with the objective of receiving user opinions about public services. objectives: to describe user profiles, evaluating their satisfaction with health services and the ombudsman service itself. design and methods: a transversal and exploratory study with a random sample of users who had used the service in the last three months. the data were analyzed with the epi info program. results: women were those who used the service most ( . %). the users sought the service for complaints ( . %), guidance ( . %) and commendation ( . %). users made the following complaints about health services: lack of care ( . %), poor assistance ( . %) lack of medication ( . %). in relation to the ombudsman service, the following failures were mentioned: lack of autonomy ( . %), delay in solving problems ( . %) and few ombudsmen ( . %). conclusion: participation of the population in use of the serviced is small. the service does not satisfy the expectations of users, it is necessary to publicize the service and try to establish an effective partnership between users and ombudsmen so that the population finds in the ombudsman service an instrument to put into effect social control and improve the quality of health services. in chile, the rates of breast cancer and diabetes have dramatically increased in the last decade. the role of insulin resistance in the development of breast cancer, however, remains unexplored. we conducted a hospital-based case-control to assess the relationship of insulin resistance (ir) and breast cancer in chilean pre and postmenopausal women. we compared women, - y, with incident breast cancer diagnosed by biopsy and controls with normal mammography. insulin and glucose were measured in blood and ir was calculated by homeostasis model assessment method. anthropometric measurements and socio-demographic and behavioural data were also collected. odds ratios (ors) and % confidence intervals (cis) were estimated by multivariate logistic regression. the risk of breast cancer increased with age. ir was significantly associated to breast cancer in postmenopausal women (or = . , %ci = . - . ), but not in premenopausal (p> . ). socioeconomic status and smoking appeared as important risk factors for breast cancer. obesity was not associated with breast cancer at any age (p> . ). in these women, ir increased the risk of breast cancer only after menopause. overall, these results suggest a different risk pattern for breast cancer before and after menopause. keywords: insulin resistance; breast cancer; chile. abstract background: previous european community health indicators (echi) projects have proposed a shortlist of indicators as a common conceptual structure for health information. the european community health indicators and monitoring (echim) is a -year project to develop and implement health indicators and to develop health monitoring. objectives: our aim is to assess the availability and comparability of the echi-shortlist indicators in european countries. methods: four widely used health indicators i) perceived general health ii-iii) prevalence of any and certain chronic diseases or conditions iv) limitations in activities of daily living (adl) were evaluated. our evaluation of available sources for these indicators is based on the european health interview & health examination surveys database ( surveys in chile, breast cancer, obesity and sedentary behaviour rates are increasing. the role of specific nutrients and exercise in the risk of breast cancer remains unclear. the aim of the present study was to evaluate the role of fruits and vegetables intake and physical activity in the prevention of breast cancer. we undertook an age matched case-control study. cases were women with breast cancer histologically confirmed and controls were women with normal mammography, admitted to the same hospital. a structured questionnaire was used to obtain dietary information and measurement of physical activity was obtained from the international physical activity questionnaire. odds ratios (ors) and % confidence intervals (cis) were estimated by conditional logistic regression adjusted by obesity, socioeconomic status and smoking habit. a significant association was found with fruit intake (or = . , %ci = . - . ). the consumption of vegetables (or = . , %ci = . - . ), moderate (or = . , %ci = . - . ) and high physical activity (or = . , %ci = . - . ) were not observed as protective factors. in conclusion, the consumption of fruit is protective in breast cancer. these findings need to be replicated at chile to support the role of diet and physical activity in breast cancer and subsequence contribution in public health policy. keywords: diet; physical activity; breast cancer; chile. the role of trace elements in pathogenesis of liver cirrhosis and its complications is still not clearly understood. serum concentrations of zinc, copper, manganese and magnesium were determinated in patients with alcoholic liver cirrhosis and healthy subjects by means of plasma sequential spectrophotometer. serum levels of zinc were significantly lower (median . vs . lmol/l, p = . ) in patients with liver cirrhosis in comparison to controls. serum levels of copper were significantly higher in patients with liver cirrhosis ( . vs . lmol/l, p< . ) as well as manganese ( . vs . lmol/l, p = . ). concentration of magnesium was not significantly different between patients with liver cirrhosis and controls ( . vs . mmol/l, p = . ). there was no difference in trace elements concentrations between child-pugh groups. zinc level was significantly lower in patients with hepatic encephalopathy in comparison to cirrhotic patients without encephalopathy ( . vs . lmol/l, p = . ). manganese was significantly higher in cirrhotic patients with ascites in comparison to those without ascites ( . vs . lmol/l, p = . ). correction of trace elements concentrations might have beneficial effect on complications and maybe progression of liver cirrhosis. it would be recommendable to provide analyzis of trace elements as a routine. abstract background: respiratory tract infections (rti) are very common in childhood and knowledge of pathogenesis and risk factors is required for effective prevention. objective: to investigate the association between early atopic symptoms and occurrence of recurrent rti during first years of life. design and methods: in the prospective prevention and incidence of asthma and mite allergy birth cohort study, children were followed from birth to the age of years. information on atopic symptoms, potential confounders, and effect modifiers like passive smoking, daycare attendance and presence of siblings was collected at ages months and year by parental questionnaires. information on rti was collected at ages , , , and years. results: children with early atopic symptoms, i.e. itchy skin rash and/or eczema or doctordiagnosed cow's milk allergy at year of age had a slightly higher risk to develop recurrent rti (aor . ( . - . ); and . ( . - . ), respectively). the association between atopic symptoms and recurrent rti was stronger in children whose mother smoked during pregnancy and who had siblings (aor . ( . - . ) the aim : the aim of the study was to assess the relative risk (rr) of obesity and abdominal fat distribution on the insulin resistance (ir), diabetes, hyperlipidemia and hypertension in polish population. materials and methods: subjects at age - , were randomized and invited to the study. in participants anthropometric and blood pressure examination was performed. fasting lipids, fasting and after glucose load glucose and insulin were determined. ir was defined as the upper quartile of the homa-ir distribution for the normal glucose tolerant population. results: overweight and obesity was observed in , % and , % of subjects. visceral obesity was found in subjects ( , %-men and , %-women). rr of ir in obesity was , ( % ci: , - , ), for obese subjects at age below was , ( % ci: , - , ). in men with visceral obesity rr of ir was the highest for men aged below . rr of diabetes was increasing with the increase of body weight, in obese subjects with abdominal fat distribution was , ( %ci: , - , ). the same was observed for the hypertension and hyperlipidemia. conclusions: obesity and the abdominal fat distribution seems to be an important risk factor of ir, diabetes, hypertension, hiperlipidemia, especially in the younger age groups. abstract background: age as an effect modifier in cardiovascular risk remains unclear. objective: to evaluate age-related differences in the effect of risk factors for acute myocardial infarction (ami). methods: in a population-based case-control study, with data collected by trained interviewers, consecutive male cases of first myocardial infarction (participation rate %) and randomly selected male control dwellers (participation rate %) were compared. effect-measure modification was evaluated by the statistical significance of a product term of each independent variable with age. unconditional logistic regression was used to estimate ors in each age stratum (< years/> years). results: there was a statistically significant interaction between education (> vs. < years), sports practice, diabetes and age: the adjusted (education, ami family history, dyslipidemia, hypertension, diabetes, angina, waist circumference, sports practice, alcohol and caffeine consumption, and energy intake) ors ( %ci) were respectively . ( . - . ), . ( . - . ) and . ( . - . ) in younger, and . ( . - . ), . ( . - . ) and . ( . - . ) in older participants. conclusions: in males, age has a significant interaction with education, sports practice and diabetes in the occurrence of ami. the effect is evident in the magnitude but not in the direction of the association. abstract there are few studies on the role of diet in lung cancer etiology. thus, we calculated both, squamous cell and small cell carcinoma risks in relation to the frequency of consumption of vegetables, cooked meat, fish and butter in silesian male in industrial area of poland. in the case-control study, the studied population comprised men with squamous cell carcinoma and men with small cell carcinoma, and healthy controls. multivariate logistic regression was employed to calculate lung cancer risk in the relation to simultaneous influence of dietary factors. the relative risk was adjusted for age and smoking. we observed a significant decrease in lung cancer risk related to more frequent consumption of raw vegetables, cooked meat and fish. however, stronger protective effect was reported for squamous cell carcinoma. frequent fish consumption significantly decreases the risk especially in cigarette smokers. the frequent consumption of pickles lowers squamous cell carcinoma risk in all cases but small cell carcinoma risk only in smokers. the presence of butter, cooked meat, fish and vegetables in diet significantly decreases the lung cancer risk especially in smokers. the association between diet and lung cancer risk is more pronounced for squamous cell carcinoma. abstract background: in functional disease research selection mechanisms need to be studied to assure external validity of trial results. objective: we compared demographic and disease-specific characteristics, history, co-morbidity and psychosocial factors of patients diagnosed, approached and randomised for a clinical trial analysing the efficacy of fibre therapy in irritable bowel syndrome (ibs). design and methods: in primary care patients were diagnosed with ibs by their gp in the past two years. characteristics were compared between ( ) randomised patients (n = ); ( ) patients who did not give their informed consent (n = ); ( ) patients who decided not to participate (n = ); and ( ) those not responding to the mailing (n = ). results: the groups showed no significance differences in age and gender ( % females, mean age years, s.d. ). patients consulting their gp for the trial compared to patients not attending their gp showed significant more severe ibs symptoms, more abdominal pain during the previous three months, and a longer history of ibs (p< , ). patients randomised have more comorbidity (p = , ). conclusion and discussion: patients included in this ibs trial differ from no participating and excluded patients mainly in ibs symptomatology, history and comorbidity. this may affect the external validity of the trial results. abstract objectives: to evaluate smoking prevalence among teenagers and identify associated social-behavioral factors. study design and methods: a cross sectional-study was carried out in may ( ) in high school population ( th- th grades) in the north of portugal (n = ). a confidential self-reported questionnaire was administered. crude and adjusted odds ratios and confidence intervals were calculated by logistic regression analysis. results: overall smoking prevalence was . % (boys = . %; girls = . %) (or = . ; ci % = . - . ; p< . ). smoking prevalence was significantly and positively associated with gender, smoking parents, school failure and school grade; in the group of students with smoking relatives, smoking was significantly associated with parents who smoke near the student (or = . ; ci % = . - . ; p< . ); in the group of the secondary grade ( th- th grades) smoking was significantly associated with belonging to 'non science-courses' (or = . ; ci % = . - . ; p = . ). conclusions: smoking is a growing problem among portuguese adolescents, increasing with age, prevailing among males, although major increases have been documented in the female population. parents' behaviours and habits have an important impact in their children's smoking behaviour. school failure is also an important factor associated with smoking. there is a need for further prevention programmes that should include families and consider students' social environment. abstract background: social environment of school can contribute to etiology of health behaviors. objective: to evaluate the role of school context for substance abuse in youth. design: a cross-sectional study was carried out in , using self-completed classroomadministered questionnaire. subjects: from a representative sample of students, a sub-sample of students was selected (including / classes with at least persons without missing data)*. methods: substance abuse was measured by: tobacco smoking at present, episodes of drunkenness and marijuana use in the lifetime. overall index was created as main independent variable, ranging - (cronbach's alpha = . ). class membership, type of school, gender, place of domicile, and school climate were included as contextual variables, measured on individual or group level. results: on individual level, the mean index was equal to . (sd = . ), and ranged from . in general comprehensive schools to . in basic vocational schools and from . to . for separated classes. about . % of total variance in this index may be attributed to differences between classes. conclusion: individual differences in substance abuse in youth could be partly explained by factors at school level. * project no po d . abstract background: rates of c-section in brazil are very high, . % in . brazil illustrates an extreme example of medicalization of birth. c-section, as any major surgery, increases the risk of morbidity, which can persist long after discharge from hospital. objectives: to investigate how social, reproductive, prenatal care and delivery factors interact after hospital discharge, influencing post partum complications. design and methods: a cross-sectional study of women gathered information through home interviews and clinical examination during post-partum. a hierarchical logistic regression model of factors associated with post-partum complications was applied. results: physical and emotional post partum complications were almost twice as high among women having c-section. most of this effect were associated with lower socioeconomic conditions which influences, were mainly explained by longer duration of delivery (even in the presence of medical indications), and less social support when returning home. conclusion: risk of c-section complications is higher among women from the lower socioeconomic strata. social inequalities mediate the association between type of delivery and postpartum complications. discussion: c-section complications should be taken into account when decisions concerning type of delivery are made. social support after birth, from the public health sector, has to be provided for women in socioeconomic deprivation. the relationship between unemployment and increased mortality was previously reported in western countries. the aim of this study was to assess the influence of the changes in unemployment rate on survival in general population in northern poland at the time of economic transition. to analyze the association between the unemployment and risk of death we collected survival data from death certificates and data on rates of unemployment from regions of gdansk county from period - . kaplan-meier method and cox proportional hazard model were used in univariate and multivariate analysis. a change of unemployment (percentage) in the year of death in the area of residence, sex and educational level ( categories) were included into multivariate analysis. the change of unemployment rate was associated with significantly worse overall survival: hazard ratio . % confidence interval . to . . the highest risk associated with the change of unemployment in the area of residence was for death from congenital defects (hazard ratio . % confidence interval . to . ) and for death from cardiovascular diseases (hazard ratio . % confidence interval . to . abstract background: there is no evidence from randomized trials as to whether or not educational interventions improve voluntary reporting systems in terms of quantity or quality. objectives: evaluation of the effectiveness of educational outreach visits aimed at improving adverse drug reaction (adr) reporting by physicians design and methods: cluster-randomized controlled trial covering all health system physicians in northern portugal. four spatialclusters assigned to intervention group (n = ) received outreach visits tailored to training needs detected in previous study and clusters were assigned to the control (n = ). the main was the total number of reported adr; the second was the number of serious, unexpected, high-causality and new-drug-related adr. a follow-up was conducted for a period of months. results: the intervention increased reports as follows: total adr, . -fold (p< . ); serious adr, . -fold (p = . ); high-causality adr, . -fold (p< . ); unexpected adr, . -fold (p< . ); and newdrug-related adr, . -fold (p = . ). the intervention had its maximum effect during the first four months ( . -fold increase, p< . ), yet the effect was nonetheless maintained over the four month periods post-intervention (p = . ). discussion and conclusion: physician training based on academic detailing visits improves reporting quality and quantity. this type of intervention could result in sizeable improvements in voluntary reporting in many countries. there were no evidence of differences in absolute indications between the years. conclusion: most of the increase in rates in the period may be attributable to relative and non-medical indications. discussion policies to promote rational use of c-sections should take into account the role played by obstetrician's convenience and the increased medicalization of birth on cesarean rates. abstract background: the changing environment has led to unhealthy dietary habits and low physical activity of children resulting in overweight/obesity and related comorbid conditions. objective: idefics is a five-year multilevel epidemiological approach proposed under the sixth eu framework to counteract the threatening epidemic of diet-and lifestyle-induced morbidity by evidence-based interventions. design and methods: a population-based cohort of . children to years old will be established in nine european countries to investigate the aetiology of these diseases. culturally adapted multi-component intervention strategies will be developed, implemented and evaluated prospectively. results: idefics compares regional, ethnic and sex-specific distributions of the above disorders and their key risk factors in children within europe. the impact of sensory perception, genetic polymorphisms and the role of internal/external triggers of food choice and children's consumer behaviour are elucidated. risk profile inventories for children susceptible to obesity and its co-morbid conditions are identified. based on controlled intervention studies an evidencebased set of guidelines for health promotion and disease prevention is developed. conclusions: provision of effective intervention modules, easy to implement in larger populations, may reduce future obesity related disease incidence. discussion: transfer of feasible guidelines into practice requires involvement of health professionals, stakeholders and consumers. abstract background: non-medically indicated cesarean deliveries increase morbidity and health care costs. brazil has one of the highest rates of caesarean sections in the world. variations in rates are positively associated with socioeconomic status. objectives: to investigate factors associated with cesarean sections in public and private sector wards in south brazil. design and methods: cross sectional data from post partum interviews and clinical records of consecutive deliveries ( in the main public and in a private maternity) was analyzed using logistic regression. results: multiple regression showed privately insured women having much higher cesarean rates than those delivering in public sector wards (or = . ; ci %: . - . ). obstetricians individual rates varied from %- %. doctors working in both, public and private sectors had a higher rates of cesarean in private wards (p< . ). wanting and having a cesarean was significantly more common among privately insured women. conclusion: women from wealthier families are at higher risk of cesarean, particularly those willing this type of delivery and whose obstetrician works in the private sector. discussion: women potentially at lower clinical risk are more like to have a caesarean. the obstetricians' role and women's preferences must be further investigated to tackle this problem. abstract background: in the netherlands, bcg-vaccination is offered to immigrant children and children of immigrant parents in order to prevent severe tuberculosis. the effectiveness of this policy has never been studied. objectives: assessing the effectiveness of the bcg-vaccination policy in the netherlands. design and methods: we used data on the size of the risk population per year (from statistics netherlands), number of children with meningitis or miliary tuberculosis in the risk population per year, and vaccination status of those cases (from the netherlands tuberculosis register) over the period - . we estimated the vaccine efficacy and annual risk of acquiring meningitis or miliary tuberculosis by log-linear modelling and treating the vaccination coverage as missing data. results: in the period - cases of meningitis or miliary tuberculosis were registered. the risk for unvaccinated to children to acquire such a serious tuberculosis infection was . ( %ci . - . ) per per year; the reduction in risk for vaccinated children was % ( %ci - %). conclusion and discussion: this means that, discounting future effects with %, a ( %ci: - ) extra children should be vaccinated to prevent one extra case of meningitis or miliary tuberculosis. given that bcg-vaccination is relatively inexpensive, the current policy could even be cost-saving. abstract background: psychotic symptom experiences in the general population are frequent and often longlasting. objectives: the zurich cohort study offered the opportunity of differentiating the patterns of psychotic experiences over a span of years. design and methods: the zurich study is based on a stratified community sample of persons born in (women) and (men). the data were collected at six time points since . we examined variables from two subscales of the scl- -r -'paranoid ideation' and 'psychoticism' -using factor analysis, cluster analysis and polytomous logistic regression. results: two new subscales were derived representing 'thought disorders' and 'schizotypal signs'. continously high symptom load on one of these subscales (both subscales) was found in % ( . %) of the population. cannabis use was the best predictor of continuously high symptom load in the 'thought disorders' subscale, whereas several variables representing adversity in childhood / youth were associated with continuously high symptom load in the 'schizotypal signs' subscale. conclusion and discussion: psychotic experiences can be divided at least in two different syndromes -thought disorders and schizotypal signs. despite similar longitudinal course patterns and also similar outcomes these syndromes rely on different risk factors, thus possibly defining separate pathways to psychosis. abstract background: the reasons for the rise in asthma and allergies remain unclear. to identify influential factors several european birth cohort studies on asthma and allergic diseases have been initiated since . objective: the aim of one work package within the global allergy and asthma european network (ga len), sponsored by the european commission, was to identify and compare european birth cohorts specifically designed to examine asthma and allergic diseases. methods: for each study, we collected detailed information (mostly by personal visits) regarding recruitment process, study setting, follow-up rates, subjective/objective outcomes and exposure parameters. results: by june , we assessed european birth cohort studies on asthma and allergic diseases. the largest recruited over children each. most studies determined specific immunoglobulin e levels to various allergens or used the isaac questionnaire for evaluation of asthma or allergic rhinitis symptoms. however, the assessment of other objective and subjective outcomes (e.g. lung function or definitions of eczema) were rather heterogeneous across the studies. conclusions due to the unique cooperation within the ga len project a common database was established containing study characteristics of european birth cohorts on asthma and allergic diseases. the possibility to pool data and perform meta-analyses is currently being evaluated. abstract background: birth weight is an important marker of health in infancy and health trajectories later in life. social inequality in birth weight is a key component in population health inequalities. objective: to comparatively study social inequality in birth weight in denmark, finland, norway, and sweden from to . design and methods as part of the nordic collaborative project on health and social inequality in early life (norchase), register-based data covering all births in all involved countries - was linked with national registries on parental socioeconomic position, covering a host of different markers including income, education and occupation. also, nested cohort studies provide opportunity to test hypotheses of mediation. results: preliminary results show that the social inequality in birth weight, small for gestational age, and low birth weight has increased in denmark through out the period. also, preliminary results from finland, norway and sweden will be presented. discussion: crosscountry comparisons pose several methodological challenges. these challenges include characterizing the societal context of each country so as to correctly interpret inter-country differences in social gradients, along with dealing with differences in the data collection methods and classification schemes used by different national registries. also, strategies for influencing policy will be discussed. abstract background: modifying the availability of suicide methods is a major issue in suicide prevention. objectives: we investigated changes in the proportion of firearm suicides in western countries since the 's, and their relation to the change of legislation and regulatory measures. design and methods: data from previous publications, from the who mortality database, and from the international crime victims survey (icvs) were used in a multilevel analysis. results: multilevel modeling of longitudinal data confirmed the effect of the proportion of households owning firearms on firearm suicide rates. several countries stand out with an obvious decline in firearm suicides since the s: norway, united kingdom, canada, australia, and new zealand. in all of these countries legislative measures have been introduced which led to a decrease in the proportion of households owning firearms. conclusion and discussion: the spread of firearms is a main determinant of the proportion of firearm suicides. legislative measures restricting the availability of firearms are a promising option in suicide prevention. abstract background: fatigue is a non-specific but frequent symptom in a number of conditions, for which correlates are unclear. objectives: to estimate socio-demographic and clinical factors determining the magnitude of fatigue. methods: as part of a follow-up evaluation of a cohort of urban portuguese adults, socio-demographic and clinical variables for consecutive participants were collected through personal interview. lifetime history of chronic disease diagnosis was inquired (depression, cancer, cardiovascular, rheumatic, and respiratory conditions), anthropometry was measured, and haemoglobin determined. krupp's -item fatigue severity scale was applied and severe fatigue defined as mean score over . mean age (sd) was . ( . ) and . % of participants were females. logistic regression was used to compute adjusted odds ratios, and attributable fractions were estimated using the formula ar = -s(?j/orj). results: adjusted for age and clinical conditions, female gender (or = . , %ci: . - . ) and education (under -years schooling: or = . , %ci: . - . ) were associated with severe fatigue. obesity (or = . , %ci: . - . ) and diagnosed cardiovascular disease (or = . , %ci: . - . ) also increased fatigue. attributable fractions were . % for gender, . % for education, . % for obesity, and . % for cardiovascular disease. conclusion: gender and education have large impact on severe fatigue, and, to a lesser extent, obesity and cardiovascular disease. abstract introduction: analysis of infant mortality allows identification of death contributing factors and assessment of child health care quality. objective: to study characteristics of infant and fetal mortality using data from a committee for prevention of maternal and infant mortality, in sobral, brazil. methods: all cases of infant deaths between and were analyzed. medical records were reviewed and mothers, interviewed. using a tool to identify preventable deaths (seade classification -brazil) the committee characterized causes of death. meetings with governmental groups involved in family health care took place to identify death contributing factors. results: in , infant mortality decreased from . to . . in the next years there was an increase from . to . . the increase in was due to respiratory illnesses. in , was due to diarrhea. analysis of preventable deaths indicated a reduction from to deaths that could have been prevented by adequate gestational care, and an increase in preventable deaths by early diagnosis and treatment. conclusion: pre-natal and delivery care improved whereas care for children less than yr old worsened. analysis of death causes allowed a reduction of infant mortality rate to . abstract objective: to identify dietary patterns and its association with metabolic syndrome. design and methods: we evaluated noninstitutionalised adults. diet was assessed using a semi-quantitative dietary frequency questionnaire, and dietary patterns were identified using principal components analysis followed by cluster analysis (k-means method) with bootstrapping (choosing the clusters presenting the lowest intra-cluster variance). metabolic syndrome (mets) was defined according to the ncep-atp-iii. results: the overall prevalence of metabolic syndrome was . %. in the population sample clusters were identified in females - .healthy, .milk/soup; .fast food; .wine/low calories; and in males - .milk/carbohydrates; . codfish/soup; .fast food; .low calories. in males, using milk/carbohydrates as the reference and adjusting for age and education, high blood pressure (or = . ; %ci: . - . ) and high triglycerides (or = . ; %ci: . - . ) were associated with the fast food pattern, and low calories pattern presented higher frequency of high blood pressure (or = . ; %ci: . - . ). in females, after age and education adjustment, no significant association was found either with metabolic syndrome or its individual features and the dietary patterns identified. conclusion: we found no specific dietary pattern associated with an increased prevalence of metabolic syndrome. however, a fast food diet was significantly more frequent in males with dyslipidemia and high blood pressure. abstract aim: to determine the prevalence of stress urinary incontinence (sui) before, during pregnancy and following childbirth, and also to analyse the impact of a health education campaign about sui prevention, following childbirth in viana district, portugal. methods: participants (n = ), interviewed during hospitalization, after birth and two months later at health centres, were divided into two groups: a first group of non-exposed and a second exposed to a health education campaign. this second group was encouraged to perform an exercise programme and given a 'suiprevention-treatment' brochure, approved by the regional health authority. results: sui prevalence was . %( %ci: . - . ) before pregnancy, . %( %ci: . - . ) during pregnancy and . ( %ci: . - . ) four weeks after birth. less than half of the women with sui sought help from healthcare professionals. statistical significant differences were found between groups: sui knowledge level and practice of pelvic floor muscles re-education exercises were higher in the exposed group ( . and . times, respectively). conclusions: sui affects a great number of women but only a small percentage reveals it. this campaign improved women knowledge and modified their else behaviors. healthcare professionals must be aware of this reality, providing an early and continuous intervention that would optimise the verified benefits of this campaign. abstract background: social inequalities have been associated with poorer developmental outcomes, but little is known about the role of the area of residence. objectives: examine whether the housing infrastructure of the area modifies the effect of socio-economic conditions of the families on child development. design and methods: community-based survey of under-fives in southern brazil applied hierarchical multi-level linear regression to investigate determinants of child development, measured by a score from the denver developmental screening test. results: in multivariable models, the mean score of child development increased with maternal and paternal education and work qualification, family income and better housing and was higher when the mother was in paid work (all p< . ). paternal education had an effect in areas of lower housing quality only; the effect of occupational status and income in these areas were twice as large as in better-provided areas (likelihood test for all interactions p< . ). this model explained % of the variation in developmental score between the areas of residence. conclusion: the housing quality and sanitation of the area modified the effects of socioeconomic conditions on child development. discussion: housing and sanitation programs are potentially beneficial to decrease the negative effect of social disadvantage on child development. abstract background: it is known that both genetic and environmental factors are involved in the early development of type diabetes (t d), and that incidence varies geographically. however we still need to explain why there is variation in incidence. objectives: in order to better understand the role of non-genetic factors, we decided to examine whether prevalence of newborns with high risk genotypes or islet autoantibodies varies geographically. design and methods: the analysis was performed on a cohort of newborns born to non-diabetic mothers, between september and august , who were included in diabetes prediction in ska˚ne study (dipis) in sweden. neighbourhoods were defined by administrative boundaries and variation in prevalence was investigated using multi-level regression analysis. results: we observed that prevalence of newborns with islet autoantibodies differed across the municipalities of ska˚ne (s = . , p < . ), with highest prevalence found in wealthy urban areas. however there was no observed difference in the prevalence of newborns with high risk genes. conclusion and discussion: newborns born with autoantibodies to islet antigens appear to cluster by region. we suggest that non-genetic factors during pregnancy may explain some of the geographical variation in the incidence of t d. abstract background: risk assessment is a science-based discipline used for decision making and regulatory purposes, such as setting acceptable exposure limits. estimation of risks attributed to exposure to chemical substances are traditionally mainly the domain of toxicology. it is recognized, however, that human, epidemiologic data, if available, are to be preferred to data from laboratory animal experiments. objectives: how can epidemiologic data be used for (quantitative) risk assessment? results: we described a framework to conduct quantitative risk assessment based on epidemiological studies. important features of the process include a weight-of-theevidence approach, estimation of the optimal exposure-risk function by fitting a regression model to the epidemiological data, estimation of uncertainty introduced by potential biases and missing information in the epidemiological studies, and calculation of excess lifetime risk through a life table to take into account competing risks. sensitivity analyses are a useful tool to evaluate the impact of assumptions and the variability of the underlying data. conclusion and discussion: many types of epidemiologic data, ranging from published, sometimes incomplete data to detailed individual data, can be used for risk assessment. epidemiologists should better facilitate such use of their data, however. abstract background: high-virulence h. pylori (hp) strains and smoking increase the risk of gastric precancerous lesions. its association with specific types of intestinal metaplasia (im) in infected subjects may clarify gastric carcinogenesis pathways. objectives: to quantify the association between types of im and infection with highvirulence hp strains (simultaneously caga+, vacas and va-cam ) and current smoking. design and methods: male volunteers (n = ) underwent gastroscopy and completed a self-administered questionnaire. participants were classified based on mucin expression patterns in biopsy specimens (antrum, body and incisura). hp vaca and caga were directly genotyped by pcr/reverse hybridization. data were analysed using multinomial logistic regression (reference: normal/superficial gastritis), models including hp virulence, smoking and age. results: high-virulence strains increased the risk of all im types (complete: or = . , %ci: . - . ; incomplete: or = . , %ci: . - . ; mixed: or = . , %ci: . - . ) but smoking was only associated with an increased risk of complete im (or = . %ci: . - . ). compared to non-smokers infected with lowvirulent strains, infection with the high-virulence hp increased the risk of im similarly for smokers (or = . , %ci: . - . ) and non-smokers (or = . %ci: . - . ). conclusion: gastric precancerous lesions, with different potential for progression, are differentially modulated by hp virulence and smoking. the risk of im associated with high-virulence hp is not further increased by smoking. abstract background: in may , the portuguese government created the basic urgency units (buu). these buu must attend at least . persons, be open hours per day, and be at maximum minutes of distance to all the users. objectives: determine the optimal location of buu, considering the existing health centers, in the viseu district, north portugal. methods: from a matrix of distances between population and health centers an accessibility index was created (sum of distances traveled by population to reach a buu). the location-allocation models were used to create simulations based on p-median model, maximal covering location problem (mclp) and set covering location problem (sclp). the solutions were ranked by weighting the variables of accessibility ( %), number of doctors in the health centers ( %), equipments ( %), distance/time ( %) and total number of buu ( %). results: the best solution has buu, doctors, attends users and the accessibility index is . km. conclusions: it was proved that it is impossible to attend all the criterion for creation of a buu. in some areas with low population density, to sum at least persons in a buu, the travel time is necessarily more than hour. background: a prospective observational study of fatigue in staff working a day/ off/ night/ off roster of hour shifts was conducted at a fly-in/fly-out fertilizer mine in remote northern australia. objectives: to determine whether fatigue in staff increased: from the start compared to the finish of shift; with the number of consecutive shifts; and from day-compared to nightshift. methods: data of sleep diaries, the mackworth clock test and the swedish occupational fatigue inventory were obtained at the start and finish of each shift from august to november . results: a total of staff participated in the study. reaction times, sleepiness and lack of energy scores were highest at the finish of nights to . the reaction times increased significantly at both the start and finish of day onwards, and at the finish of night . reaction times and lack of motivation were highest during nightshift. conclusions: from the above results, a disturbed diurnal rhythm and decreased motivation during night-shift; and a roster of more than eight consecutive shifts can be inferred as the primary contributors to staff fatigue. discussion: the implications for changes to workplace practices and environment will be discussed. the aim of this survey was to assess the impact of a meta-analysis comparing resurfacing with nonresurfacing of the patella on the daily practice of experts. participants in this study were experts which had participated in a previous survey on personal preferences regarding patella resurfacing. these experts in the field of patella resurfacing were identified by a thorough search of medline, an internet search (with googletm search engine), and personal references from the identified experts. participants of the 'knee arthroplasty trial' (kat) in the united kingdom were also included. two surveys were sent to the participants, one before and one after the publication of the meta-analysis. the response rate is questionnaires or %. the vast majority of responders are not persuaded to change change their practice after reading the metaanalysis. this is only in part due to the fact that best evidence and practice coincide. other reasons given are methodology related, an observation which is shared by the authors of the review, which force the orthopedic community to improve its research methodology. reasons such as 'i do not believe in meta-analysis' either demands a fundamental discussion or demands the reader to take evidence based medicine more seriously. abstract background: patients with type diabetes (dm ) have a - fold increased risk of cardiovascular disease. delegating routine tasks and computerized decision support systems (cdss) such as diabetes care protocol (dcp) may improve treatment of cardiovascular risk factors hba c, blood pressure and cholesterol. dcp includes consultation-hours exclusively scheduled for dm patients, rigorous delegation of routine tasks in diabetes care to trained paramedics, and software to support medical management. objective: to investigate the effects of dcp, used by practice assistants, on the risk of coronary heart disease for patients with dm . design and methods: in an open-label pragmatic trial in general practices with patients, hba , blood pressure and cholesterol were examined before and prospectively one year after implementation of dcp. the primary outcome was the change in the year ukpds coronary heart disease (chd) risk estimate. results: the median year ukpds chd risk estimate improved significantly from . % to . %. hba decreased from . % to . %, systolic blood pressure from . to . mmhg and total cholesterol from . to . mmol/l. (all p< . ). conclusion: delegating routine task in diabetes care to trained paramedics and using cdss improves the cardiovascular risk of dm patients. tuberculosis in exposed immigrants by tuberculin skin test, ifn-g tests and epidemiologic abstract background: currently immigrants in western countries are only investigated for active tuberculosis (tb) by use of a chest x-ray. recent latent tuberculosis infection (ltbi) is hard to diagnose in this specific population because the only available test method, the tuberculin skin test (tst), has a low positive predictive value (ppv). recently interferon-gamma (ifn-g) tests have become available that measure cellular responses to specific m. tuberculosis antigens and might have a better ppv. objective: to determine the predictive value of tst and two different ifn-g tests combined with epidemiological characteristics for developing active tb in immigrants who are close contacts of smear positive tb patients. methods in this prospective cohort study close contacts will be included. demographic characteristics and exposure data are investigated. beside their normal examination they will all have a tst. two different ifn-g tests will be done in those with a tst induration of ? mm. these contacts will be followed for years to determine the occurrence of tb. results since april , municipal health services have started with the inclusion. preliminary results on the predictive value of tst, both ifn-g tests and epidemiological characteristics will be presented. abstract background: different factors contribute to the quality of ed (emergency department) care of an injured patient. objective: determine factors influencing the disagreement between er diagnoses and those assigned at hospital admission in injuried patients, and evaluate if disagreement between the diagnoses could have worsened the outcome. methods: all the er visits of the emergency departments of lazio region for unintentional injuries followed by hospitalisation in . concordant diagnoses were established on the basis of the barell matrix cells. logistic regression was used to assess the role of individual and er care factors on the probability of concordance. a logistic regression where death within days was the outcome and concordance the determinant was uses. results: , injury er visits were considered. in . % cases, the er and discharge diagnoses were concordant. higher concordance was found with increasing age and less urgent cases. factors influencing concordance were: the hour of the visit, er level, initial outcome, length of stay in hospital. patients who had non concordant diagnoses had a % higher probability of death. conclusions: a correct diagnosis at first contact with the emergency room is associated with lower mortality. methods: a cohort of consecutive patients treated for secondary peritonitis were sent the posttraumatic stress syndrome inventory (ptss- ) and impact of events scale-revised (ies-r) - years following their surgery for secondary peritonitis. results: from the patients operated upon between and , questionnaires were sent to the long-term survivors of which % responded (n = ). ptsd-related symptoms were found in % of patients by both questionnaires. patients admitted to icu (n = ) were significantly older, with higher apache-ii scores, but reported similar ptsd symptomology scores compared to non-icu patients (n = ). traumatic memories during icu and hospital-stay were most predictive for higher scores. adverse memories did not occur more often in the icu group than in the hospital-ward group conclusions: longterm ptsdrelated symptoms in patients with secondary peritonitis were very barthé lé my c cabanas ruiz-carrillo de la cruz den boon jimé nez-moleó n mü ller-nordhorn national evaluation team rich-edwards in the netherlands. design and methods we used the populationbased databases of the netherlands cancer registry, the eindhoven cancer registry (ecr) and the central bureau of statistics. patients from the ecr were followed until - - for vital status and relative survival was calculated. results: the number of breast cancer cases increased from in to . in , an annual increase of . % (p< . ). the death rate decreased , % annually (p< . ), which resulted in deaths in . the relative -yr survival was less than % for patients diagnosed in the seventies, this increased to over % for patient diagnosed since , patients with stage i disease even have a % -yr relative survival. conclusion: the alarming increase in breast cancer incidence is accompanied with a serious improvement in survival rates. this results in a large number of women (ever) diagnosed with breast cancer, about , in of whom % demand some kind of medical care. abstract background: nine % of the population in the netherlands belongs to non-western ethnic minorities. perceived health is worse and health care use different from dutch natives. objectives. which factors are associated with ethnic differences in self-rated health? which factors are associated with differences in utilisation of gp care? methods: during one year all contacts with gps were registered. adult surinam, antillean, turkish, moroccan and dutch responders were included (total n: . ). we performed multivariate analyses of determinants of self-rated health and on the number of contacts with gps. results: self-rated health differ from native dutch: surinam/antillean (or . ) and turkish/moroccan patients (or . / . ) , especially in turkish/moroccan females. more turks visit the gps at least once a year (or . ). less surinamese (or . ) and antillean patients (or . ) visit their gps than the dutch do. people from ethnic minorities in good health visit their gps more often ( . - . consults per year vs. . ). incidence rates of acute respiratory infections and chest complaints were significantly higher than in the dutch. conclusions: ethnicity is independently associated with self-rated health. higher use of gp-care by ethnic minorities in good health, points towards possible inappropriate use of resources. the future: do they fulfil it? first results of the limburg youth monitoring project abstract background: incidence of coronary heart disease (chd) and stroke can be estimated from local, population-based registers. it is unclear, to what extent local register data are applicable on a nationwide level. therefore, we compared german register data with estimates derived with who global burden of disease (gbd) method. methods: incidence of chd and stroke was computed with the gbd method using official german mortality statistics and prevalences from the german national health survey. results were compared to estimates from the kora/monica augsburg register (chd) and the erlangen stroke project in southern germany. results: gbd estimates and register data showed good agreement: chd (age group - years) , (gbd) versus , (register) and stroke (all ages) , versus , incident cases per year. chd incidence among all age groups was estimated with the gbd method to be , per year (no register data available). chd incidence in men and stroke incidence in women were underestimated with the gbd method as compared to register data. conclusions: gbd method is a useful tool to estimate incidence of chd and stroke. the computed estimates may be seen as lower limit for incidence data. differences between gbd estimates and register data are discussed. abstract background: children with mental retardation (mr) are a vulnerable not much studied population. objectives: to investigate psychopharmacotherapy in children with mr and to examine possible factors associated with psychopharmacotherapy. methods: participants were recruited through all facilities for children with mental retardation in friesland, the netherlands, resulting in participants, - years old, including all levels of mental retardation. the dbc and the pdd-mrs were used to assess general behavior problems and pervasive developmental disorders (pdd). information on medication was collected through a parent-interview. logistic regression was used to investigate the relationship between the psychotropic drug use and the factors dbc, pdd, housing, age, gender and level of mr. results: % of the participants used psychotropic medication. main factors associated with receiving psychopharmacotherapy were pdd (or . ) and dbc score (or . ). living away from home and mr-level also played a role whereas gender and age did not. dbc score was associated with clonidine, stimulants and anti-psychotics. pdd was the main factor associated with anti-psychotics use (or . ). discussion: psychopharmacotherapy is especially prevalent among children with mr and comorbid pdd and general behavior problems. although many psychotropic drugs are used off-label, specific drugs were associated with specific psychiatric or behavior problems. abstract background: increased survival in children with cancer has raised interest on the quality-of-life of long-term survivors. objective: to compare educational outcomes of adult survivors of childhood cancer and healthy controls. methods: retrospective cohort study including a sample of adult survivors ( ) treated for childhood cancer in the three existing italian paediatric research hospitals. controls ( ) were selected among siblings, relatives or friends of survivors. when these controls were not available, a search was carried out in the same area of residence of the survivors though random digit dialling. data collection was carried out through a telephone-administered structured questionnaire. results: significantly more survivors than controls needed school support (adjusted odds ratio -oradj- . , % ci . - . ); failed at least a grade after disease onset (oradj . , % ci . - . ); achieved a lower educational level (oradj . , % ci . - . ) and did not reach an educational level higher than their parents' (oradj . , % ci . - . ). subject's age, sex, parents' education and area of residence were taken into account as possible confounders. conclusions: these findings suggest the need to provide appropriate school support to children treated for childhood cancer. abstract background: in italy supplementation with folic acid (fa) in the periconceptional period to prevent congenital malformations (cms) is quite low. the national health institute has recently launched ( ) a programme to improve awareness about the role of fa in reducing the risk of serious defects also by providing . mg fa tablets free of charge to women planning a pregnancy. objectives: we analysed cms that are or may be sensitive to fa supplementation in order to establish an adequate baseline to allow a fa impact assessment in the next years and to investigate spatial differences among cms registries, time trends and time-space interactions. design and methods data collected over - by the italian registries members of eurocat and icbdsr on births and induced abortions with neural tube defects, ano-rectal atresia, omphalocele, oral clefts, cardiovascular, limb reduction and urinary system defects. results: all the cms showed statistically significant differences among registries with the exception of ano-rectal atresia. the majority of cms by registry showed stable or increasing trends over time. conclusions results show the importance of fa intake during the periconceptional period. differences among registries indicate also the need of having a baseline for each registry to follow trends over time. abstract country-specific resistance proportions are more biased by variable sampling and ascertainment procedures than incidence rates. within the european antimicrobial resistance surveillance system (earss) resistance incidence rates and proportions can be calculated. in this study, the association between antimicrobial resistance incidence rates and proportions and the possible effect of differential sampling of blood cultures was investigated. in , earss collected routine antimicrobial susceptibility test data from invasive s. aureus isolates, tested according to standard protocols. via a questionnaire denominator information was collected. the spearman correlation coefficient and linear regression were used for statistical analysis. this year, of hospitals and of laboratories from of earss countries responded to the questionnaire. they reported of, overall, , s. aureus isolates. in the different countries, mrsa proportions ranged from < % to % and incidence rates per , patient days from . ae - to . ae - . overall, the proportions and rates highly correlated. blood culturing rates only influenced the relationship between mrsa resistance proportions and incidence rates for eastern european countries. in conclusion, resistance proportions seem to be very similar to resistance incidence rates, in the case of mrsa. nevertheless, this relationship appears to be dependent of some level of blood culturing. . key: cord- -gvezk vp authors: ahonen, pasi; alahuhta, petteri; daskala, barbara; delaitre, sabine; hert, paul de; lindner, ralf; maghiros, ioannis; moscibroda, anna; schreurs, wim; verlinden, michiel title: safeguards date: journal: safeguards in a world of ambient intelligence doi: . / - - - - _ sha: doc_id: cord_uid: gvezk vp the multiplicity of threats and vulnerabilities associated with ami will require a multiplicity of safeguards to respond to the risks and problems posed by the emerging technological systems and their applications. in some instances, a single safeguard might be sufficient to address a specified threat or vulnerability. more typically, however, a combination of safeguards will be necessary to address each threat and vulnerability. in still other instances, one safeguard might apply to numerous treats and vulnerabilities. one could depict these combinations in a matrix or on a spreadsheet, but the spreadsheet would quickly become rather large and, perhaps, would be slightly misleading. just as the ami world will be dynamic, constantly changing, the applicability of safeguards should also be regarded as subject to a dynamic, i.e., different and new safeguards may need to be introduced in order to cope with changes in the threats and vulnerabilities. the multiplicity of threats and vulnerabilities associated with ami will require a multiplicity of safeguards to respond to the risks and problems posed by the emerging technological systems and their applications. in some instances, a single safeguard might be sufficient to address a specified threat or vulnerability. more typically, however, a combination of safeguards will be necessary to address each threat and vulnerability. in still other instances, one safeguard might apply to numerous treats and vulnerabilities. one could depict these combinations in a matrix or on a spreadsheet, but the spreadsheet would quickly become rather large and, perhaps, would be slightly misleading. just as the ami world will be dynamic, constantly changing, the applicability of safeguards should also be regarded as subject to a dynamic, i.e., different and new safeguards may need to be introduced in order to cope with changes in the threats and vulnerabilities. for the purpose of this chapter, we have grouped safeguards into three main categories: the main privacy-protecting principles in network applications are: • anonymity (which is the possibility to use a resource or service without disclosure of user identity) • pseudonymity (the possibility to use a resource or service without disclosure of user identity, but to be still accountable for that use) • unlinkability (the possibility to use multiple resources or services without others being able to discover that these resources are being used by the same user) prime project, which studied the state of the art in privacy protection in network applications in , pointed out many performance problems and security weaknesses. the challenges in privacy-enhancing technologies for networked applications include developing methods for users to express their wishes regarding the processing of their data in machine-readable form ("privacy policies") and developing methods to ensure that the data are indeed processed according to users' wishes and legal regulations ("licensing languages" and "privacy audits": the former check the correctness of data processing during processing, while the latter check afterwards and should allow checking even after the data are deleted). privacy protection research is still new, and research on privacy protection in such emerging domains as personal devices, smart environments and smart cars is especially still in its infancy. privacy protection for personal mobile devices is particularly challenging due to the devices' limited capabilities and battery life. for these domains, only generic guidelines have been developed (see lahlou et al. ). langheinrich et al. show how difficult it might be to apply fair information practices (as contained in current data protection laws) to ami applications. most of the research on privacy protection is concerned with dangers of information disclosure. other privacy aspects have not received much attention from researchers. for example, the privacy aspect known as "the right to be let alone" is rarely discussed by technology researchers, despite its importance. research is needed with regard to overcoming the digital divide in the context of ami. the european commission has already been sponsoring some research projects which form a foundation for needed future initiatives. projects dealing with accessibility for all and e-inclusion (such as cost : "accessibility for all to services and terminals for next generation mobile networks", ask-it: "ambient intelligence system of agents for knowledge-based and integrated services for camenisch, j. (ed.), first annual research report, prime deliverable d . , . http:// www.prime-project.eu.org/public/prime_products/deliverables/rsch/pub_del_d . .a_ec_ wp . _v _final.pdf such research is, however, going on. an example is the ec-supported connect project, which aims to implement a privacy management platform within pervasive mobile services, coupling research on semantic technologies and intelligent agents with wireless communications (including umts, wifi and wimax) and context-sensitive paradigms and multimodal (voice/graphics) interfaces to provide a strong and secure framework to ensure that privacy is a feasible and desirable component of future ambient intelligence applications. the two-year project started in june . http://cordis.europa.eu/search/index.cfm?fuseaction = proj.simpledocument&pj_rcn = lahlou, s., and f. jegou, "european disappearing computer privacy design guidelines v ", ambient agora deliverable d . , electricité de france, clamart, . mobility impaired users") are concerned with standardisation, intuitive user interfaces, personalisation, interfaces to all everyday tools (e.g., domotics, home health care, computer accessibility for people with disabilities and elderly people), adaptation of contents to the channel capacity and the user terminal and so on. standardisation in the field of information technology (including, e.g., biometrics) is important in order to achieve interoperability between different products. however, interoperability even in fairly old technologies (such as fingerprint-based identification) has not yet been achieved. minimising personal data should be factored into all stages of collection, transmission and storage. the goal of the minimal data transmission principle is that data should reveal little about the user even in the event of successful eavesdropping and decryption of transmitted data. similarly, the principle of minimal data storage requires that thieves do not benefit from stolen databases and decryption of their data. implementation of anonymity, pseudonymity and unobservability methods helps to minimise system knowledge about users at the stages of data transmission and storage in remote databases, but not in cases involving data collection by and storage in personal devices (which collect and store mainly the device owner's data) or storage of videos. the main goals of privacy protection during data collection are, first, to prevent linkability between diverse types of data collected about the same user and, second, to prevent surveillance by means of spyware or plugging in additional pieces of hardware transmitting raw data (as occurs in wiretapping). these goals can be achieved by: • careful selection of hardware (so that data are collected and transmitted only in the minimally required quality and quantity to satisfy an application's goals, and there are no easy ways to spy on raw and processed data) • an increase of software capabilities and intelligence (so that data can be processed in real time) • deleting data as soon as the application allows. in practice, it is difficult to determine what "minimally needed application data" means. moreover, those data can be acquired by different means. thus, we suggest that data collection technologies less capable of violating personal privacy expectations be chosen over those more privacy-threatening technologies even if the accuracy of collected data decreases. software capabilities need to be maximised in order to minimise storage of raw data and avoid storage of data with absolute time and location stamps. we suggest this safeguard in order to prevent accidental logging of sensitive data, because correlation of different kinds of data by time stamps is fairly straightforward. these safeguards are presented below in more detail: • in our opinion, the most privacy-threatening technologies are physiological sensors and video cameras. physiological sensors are privacy-threatening because they reveal what's going on in the human body and, accordingly, reveal health data and even feelings. video cameras, especially those storing raw video data, are privacy-threatening because they violate people's expectations that "nobody can see me if i am hidden behind the wall" and because playback of video data can reveal more details than most people pay attention to in normal life. we suggest that usage of these two groups of devices should be restricted to safety applications until proper artificial intelligence safeguards (see below) are implemented. • instead of logging raw data, only data features (i.e., a limited set of pre-selected characteristics of data, e.g., frequency and amplitude of oscillations) should be logged. this can be achieved by using either hardware filters or real-time preprocessing of data or a combination of both. • time and location stamping of logged data should be limited by making it relative to other application-related information or by averaging and generalising the logged data. • data should be deleted after an application-dependent time, e.g., when a user buys clothes, all information about the textile, price, designer, etc., should be deleted from the clothes' rfid tag. for applications that require active rfid tags (such as for finding lost objects ), the rfid identifier tag should be changed, so that links between the shop database and the clothes are severed. • applications that do not require constant monitoring should switch off automatically after a certain period of user inactivity (e.g., video cameras should automatically switch off at the end of a game). • anonymous identities, partial identities and pseudonyms should be used wherever possible. using different identities with the absolute minimum of personal data for each application helps to prevent discovery of links between user identity and personal data and between different actions by the same user. orr, r.j., r. raymond, j. berman and f. seay, "a system for finding frequently lost objects in the home", technical report - , graphics, visualization, and usability center, georgia tech, . data and software protection from malicious actions should be implemented by intrusion prevention and by recovery from its consequences. intrusion prevention can be active (such as antivirus software, which removes viruses) or passive (such as encryption, which makes it more difficult to understand the contents of stolen data). at all stages of data collection, storage and transmission, malicious actions should be hindered by countermeasures such as the following: privacy protection in networking includes providing anonymity, pseudonymity and unobservability whenever possible. when data are transferred over long distances, anonymity, pseudonymity and unobservability can be provided by the following methods: first, methods to prove user authorisation locally and to transmit over the network only a confirmation of authorisation; second, methods of hiding relations between user identity and actions by, for example, distributing this knowledge over many network nodes. for providing anonymity, it is also necessary to use special communication protocols which do not use device ids or which hide them. it is also necessary to implement authorisation for accessing the device id: currently, most rfid tags and bluetooth devices provide their ids upon any request, no matter who actually asked for the id. another problem to solve is that devices can be distinguished by their analogue radio signals, and this can hinder achieving anonymity. additionally, by analysing radio signals and communication protocols of a personal object, one can estimate the capabilities of embedded hardware and guess whether this is a new and expensive thing or old and inexpensive, which is an undesirable feature. unobservability can be implemented to some extent in smart spaces and personal area networks (pans) by limiting the communication range so that signals do not penetrate the walls of a smart space or a car, unlike the current situation when two owners of bluetooth-enabled phones are aware of each other's presence in neighbouring apartments. methods of privacy protection in network applications (mainly long-distance applications) include the following: • anonymous credentials (methods to hide user identity while proving the user's authorisation). • a trusted third party: to preserve the relationships between the user's true identity and his or her pseudonym. • zero-knowledge techniques that allow one to prove the knowledge of something without actually providing the secret. • secret-sharing schemes: that allow any subset of participants to reconstruct the message provided that the subset size is larger than a predefined threshold. • special communication protocols and networks such as: -onion routing: messages are sent from one node to another so that each node removes one encryption layer, gets the address of the next node and sends the message there. the next node does the same, and so on until some node decrypts the real user address. -mix networks and crowds that hide the relationship between senders and receivers by having many intermediate nodes between them. • communication protocols that do not use permanent ids of a personal device or object; instead, ids are assigned only for the current communication session. communication protocols that provide anonymity at the network layer, as stated in the prime deliverable, are not suitable for large-scale applications: there is no evaluation on the desired security level, and performance is a hard problem. strong access control methods are needed in ami applications. physical access control is required in applications such as border control, airport check-ins and office access. reliable user authentication is required for logging on to computers and personal devices as well as network applications such as mobile commerce, mobile voting and so on. reliable authentication should have low error rates and strong anti-spoofing protection. work on anti-spoofing protection of iris and fingerprint recognition is going on, but spoofing is still possible. we suggest that really reliable authentication should be unobtrusive, continuous (i.e., several times during an application-dependent time period) and multimodal. so far, there has been limited research on continuous multimodal access control systems. authentication methods include the following: camenish, . some experts don't believe that biometrics should be the focus of the security approach in an ami world, since the identification and authentication of individuals by biometrics will always be approximate, is like publishing passwords, can be spoofed and cannot be revoked after an incident. tokens are portable physical devices given to users who keep them in their possession. implants are small physical devices, embedded into a human body (nowadays they are inserted with a syringe under the skin). implants are used for identification by unique id number, and some research aims to add a gps positioning module in order to detect the user's location at any time. with multimodal fusion, identification or authentication is performed by information from several sources, which usually helps to improve recognition rates and anti-spoofing capabilities. multimodal identification and/or authentication can also be performed by combining biometric and non-biometric data. methods for reliable, unobtrusive authentication (especially for privacy-safe, unobtrusive authentication) should be developed. unobtrusive authentication should enable greater security because it is more user-friendly. people are not willing to use explicit authentication frequently, which reduces the overall security level, while unobtrusive authentication can be used frequently. methods for context-dependent user authentication, which would allow user control over the strength and method of authentication, should be developed, unlike the current annoying situation when users have to go through the same authentication procedure for viewing weather forecasts and for viewing personal calendar data. recently, the meaning of the term "access control" has broadened to include checking which software is accessing personal data and how the personal data are processed. access control to software (data processing methods) is needed for enforcing legal privacy requirements and personal privacy preferences. user-friendly interfaces are needed for providing awareness and configuring privacy policies. maintaining privacy is not at the user's focus, so privacy information should not be a burden for a user. however, the user should easily be able to know and configure the following important settings: • purpose of the application (e.g., recording a meeting and storing the record for several years) • how much autonomy the application has • information flow from the user • information flow to the user (e.g., when and how the application initiates interactions with the user). additionally, user-friendly methods are needed for fast and easy control over the environment, which would allow a person (e.g., a home owner but not a thief) to override previous settings, and especially those settings learned by ami technologies. standard concise methods of initial warnings should be used to indicate whether privacy-violating technologies (such as those that record video and audio data, log personal identity data and physiological and health data) are used by ambient applications. licensing languages or ways to express legal requirements and user-defined privacy policies should be attached to personal data for the lifetime of their transmission, storage and processing. these would describe what can be done with the data in different contexts (e.g., in cases involving the merging of databases), and ensure that the data are really treated according to the attached licence. these methods should also facilitate privacy audits (checking that data processing has been carried out correctly and according to prescribed policies), including instances when the data are already deleted. these methods should be tamper-resistant, similar to watermarking. high-level application design to provide an appropriate level of safeguards for the estimated level of threats can be achieved by data protection methods such as encryption and by avoiding usage of inexpensive rfid tags that do not have access control to their id and by minimising the need for active data protection on the part of the user. high-level application design should also consider what level of technology control is acceptable and should provide easy ways to override automatic actions. when communication capabilities move closer to the human body (e.g., embedded in clothes, jewellery or watches), and battery life is longer, it will be much more difficult to avoid being captured by ubiquitous sensors. it is an open question how society will adapt to such increasing transparency, but it would be beneficial if the individual were able to make a graceful exit from ami technologies at his or her discretion. to summarise, the main points to consider in system design are: • data filtering on personal devices is preferred to data filtering in an untrustworthy environment. services (e.g., location-based services) should be designed so that personal devices do not have to send queries; instead, services could simply broadcast all available information to devices within a certain range. such an implementation can require more bandwidth and computing resources, but is safer because it is unknown how many devices are present in a given location. thus, it is more difficult for terrorists to plan an attack in a location where people have gathered. • authorisation should be required for accessing not only personal data stored in the device, but also for accessing device id and other characteristics. • good design should enable detection of problems with hardware (e.g., checking whether the replacement of certain components was made by an authorised person or not). currently, mobile devices and smart dust nodes do not check anything if the battery is removed, and do not check whether hardware changes were made by an authorised person, which makes copying data from external memory and replacement of external memory or sensors relatively easy, which is certainly inappropriate in some applications, such as those involved in health monitoring. • personal data should be stored not only encrypted, but also split according to application requirements in such a way that different data parts are not accessible at the same time. • an increase in the capabilities of personal devices is needed to allow some redundancy (consequently, higher reliability) in implementation and to allow powerful multitasking: simultaneous encryption of new data and detection of unusual patterns of device behaviour (e.g., delays due to virus activity). an increase in processing power should also allow more real-time processing of data and reduce the need to store data in raw form. • software should be tested by trusted third parties. currently, there are many kinds of platforms for mobile devices, and business requires rapid software development, which inhibits thorough testing of security and the privacy-protecting capabilities of personal devices. moreover, privacy protection requires extra resources and costs. • good design should provide the user with easy ways to override any automatic action, and to return to a stable initial state. for example, if a personalisation application has learned (by coincidence) that the user buys beer every week, and includes beer on every shopping list, it should be easy to return to a previous state in which system did not know that the user likes beer. another way to solve this problem might be to wait until the system learns that the user does not like beer. however, this would take longer and be more annoying. • good design should avoid implementations with high control levels in applications such as recording audio and images as well as physiological data unless it is strictly necessary for security reasons. • means of disconnecting should be provided in such a way that it is not taken as a desire by the user to hide. to some extent, all software algorithms are examples of artificial intelligence (ai) methods. machine-learning and data-mining are traditionally considered to belong to this field. however, safeguarding against aml threats requires al methods with very advanced reasoning capabilities. currently, ai safeguards are not mature, but the results of current research may change that assessment. many privacy threats arise because the reasoning capabilities and intelligence of software have not been growing as fast as hardware capabilities (storage and transmission capabilities). consequently, the development of ai safeguards should be supported as much as possible, especially because they are expected to help protect people from accidental, unintentional privacy violation, such as disturbing a person when he does not want to be, or from recording some private activity. for example, a memory aid application could automatically record some background scene revealing personal secrets or a health monitor could accidentally send data to "data hunters" if there are no advanced reasoning and anti-spyware algorithms running on the user's device. advanced ai safeguards could also serve as access control and antivirus protection by catching unusual patterns of data copying or delays in program execution. we recommend that ami applications, especially if they have a high control level, should be intelligent enough to: • detect sensitive data in order to avoid recording or publishing such data provide an automatic privacy audit, checking traces of data processing, data-or code-altering, etc. these requirements are not easy to fulfil in full scale in the near future; however, we suggest that it is important to fulfil these requirements as far as possible and as soon as possible. data losses and identity theft will continue into the future. however, losses of personal data will be more noticeable in the future because of the growing dependence on ami applications. thus, methods must be developed to inform all concerned people and organisations about data losses and to advise and/or help them to replace compromised data quickly (e.g., if somebody's fingerprint data are compromised, a switch should be made to another authentication method in all places where the compromised fingerprint was used). another problem, which should be solved by technology means, is recovery from loss of or damage to a personal device. if a device is lost, personal data contained in it can be protected from strangers by diverse security measures, such as data encryption and strict access control. however, it is important that the user does not need to spend time customising and training a new device (so that denial of service does not happen). instead, the new device should itself load user preferences, contacts, favourite music, etc, from some back-up service, like a home server. we suggest that ways be developed to synchronise data in personal devices with a back-up server in a way that is secure and requires minimal effort by the user. we suggest that the most important, but not yet mature technological safeguards are the following: • communication protocols that either do not require a unique device identifier at all or that require authorisation for accessing the device identifier • network configurations that can hide the links between senders and receivers of data • improving access control methods by multimodal fusion, context-aware authentication and unobtrusive biometric modalities (especially behavioural biometrics, because they pose a smaller risk of identity theft) and by aliveness detection in biometric sensors • enforcing legal requirements and personal privacy policies by representing them in machine-readable form and attaching these special expressions to personal data, so that they specify how data processing should be performed, allow a privacy audit and prevent any other way of processing • developing fast and intuitive means of detecting privacy threats, informing the user and configuring privacy policies • increasing hardware and software capabilities for real-time data processing in order to minimise the lifetime and amount of raw data in a system • developing user-friendly means to override any automatic settings in a fast and intuitive way • providing ways of disconnecting in such a way that nobody can be sure why a user is not connected • increasing security by making software updates easier (automatically or semiautomatically, and at a convenient time for the user), detection of unusual patterns, improved encryption • increasing software intelligence by developing methods to detect and to hide sensitive data; to understand the ethics and etiquette of different cultures; to speak different languages and to understand and translate human speech in many languages, including a capability to communicate with the blind and deaf • developing user-friendly means for recovery when security or privacy has been compromised. the technological safeguards require actions by industry. we recommend that industry undertake such technological safeguards. industry may resist doing so because it will increase development costs, but safer, more secure technology should be seen as a good investment in future market growth and protection against possible liabilities. it is obvious that consumers will be more inclined to use technology if they believe it is secure and will shield, not erode their privacy. we recommend that industry undertake such safeguards voluntarily. it is better to do so than to be forced by bad publicity that might arise in the media or from action by policy-makers and regulators. security guru bruce schneier got it right when he said that "the problem is … bad design, poorly implemented features, inadequate testing and security vulnerabilities from software bugs. … the only way to fix this problem is for vendors to fix their software, and they won't do it until it's in their financial best interests to do so. … liability law is a way to make it in those organizations' best interests." if development costs go up, industry will, of course, pass on those costs to consumers, but since consumers already pay, in one way or another, the only difference is who they pay. admittedly, this is not a simple problem because hardware manufacturers, software vendors and network operators all face competition and raising the cost of development and lengthening the duration of the design phase could have competitive implications, but if all industry players face the same exacting liability standards, then the competitive implications may not be so severe as some might fear. co-operation between producers and users of ami technology in all phases from r&d to deployment is essential to address some of the threats and vulnerabilities posed by ami. the integration of or at least striking a fair balance between the interests of the public and private sectors will ensure more equity, interoperability and efficiency. governments, industry associations, civil rights groups and other civil society organisations can play an important role in balancing these interests for the benefit of all affected groups. standards form an important safeguard in many domains, not least of which are those relating to privacy and information security. organisations should be expected to comply with standards, and standards-setting initiatives are generally worthy of support. while there have been many definitions and analyses of the dimensions of privacy, few of them have become officially accepted at the international level, especially by the international organization for standardization. the iso has at least achieved consensus on four components of privacy, i.e., anonymity, pseudonymity, unlinkability and unobservability. (see section . , p. , above for the definitions.) among the iso standards relevant to privacy and, in particular, information security are iso/iec on evaluation criteria for it security and iso , the code of practice for information security management. the iso has also established a privacy technology study group (ptsg) under joint technical committee (jtc ) to examine the need for developing a privacy technology standard. this is an important initiative and merits support. its work and progress should be tracked closely by the ec, member states, industry and so on. the iso published its standard iso in , which was updated in july . since then, an increasing number of organisations worldwide formulate their security management systems according to this standard. it provides a set of recommendations for information security management, focusing on the protection of information as an asset. it adopts a broad perspective that covers most aspects of information systems security. among its recommendations for organisational security, iso states that "the use of personal or privately owned information processing facilities … for processing business information, may introduce new vulnerabilities and necessary controls should be identified and implemented." by implementing such controls, iso/iec , information technology -security techniques -evaluation criteria for it security, first edition, international organization for standardization, geneva, . the standard is also known as the common criteria. similar standards and guidelines have also been published by other eu member states: the british standard bs was the basis for the iso standard. another prominent example is the german it security handbook (bsi, organisations can, at the same time, achieve a measure of both organisational security and personal data protection. iso acknowledges the importance of legislative requirements, such as legislation on data protection and privacy of personal information and on intellectual property rights, for providing a "good starting point for implementing information security". iso is an important standard, but it could be described more as a framework than a standard addressing specificities of appropriate technologies or how those technologies should function or be used. also, iso was constructed against the backdrop of today's technologies, rather than with ami in mind. hence, the adequacy of this standard in an ami world needs to be considered. nevertheless, organisations should state to what extent they are compliant with iso and/or how they have implemented the standard. audit logs may not protect privacy since they are aimed at determining whether a security breach has occurred and, if so, who might have been responsible or, at least, what went wrong. audit logs could have a deterrent value in protecting privacy and certainly they could be useful in prosecuting those who break into systems without authorisation. in the highly networked environment of our ami future, maintaining audit logs will be a much bigger task than now where discrete systems can be audited. nevertheless, those designing ami networks should ensure that the networks have features that enable effective audits. the oecd has been working on privacy and security issues for many years. it produced its first guidelines more than years ago. its guidelines on the protection of privacy and transborder flows of personal data were (are) intended to harmonise national privacy legislation. the guidelines were produced in the form of a recommendation by the council of the oecd and became applicable in september . the guidelines are still relevant today and may be relevant in an ami world too, although it has been argued that they may no longer be feasible in an ami world. the oecd's more recent guidelines for the security of information systems and networks are also an important reference in the context of developing privacy and security safeguards. these guidelines were adopted as a recommendation of the oecd council (in july ). in december , the oecd published a report on "the promotion of a culture of security for information systems and networks", which it describes as a major information resource on governments' effective efforts to date to foster a shift in culture as called for in the aforementioned guidelines for the security of information systems and networks. in november , the oecd published a -page volume entitled privacy online: oecd guidance on policy and practice, which contains specific policy and practical guidance to assist governments, businesses and individuals in promoting privacy protection online at national and international levels. in addition to these, the oecd has produced reports on other privacy-related issues including rfids, biometrics, spam and authentication. sensible advice can also be found in a report published by the us national academies press in , which said that to best protect privacy, identifiable information should be collected only when critical to the relationship or transaction that is being authenticated. the individual should consent to the collection, and the minimum amount of identifiable information should be collected and retained. the relevance, accuracy and timeliness of the identifier should be maintained and, when necessary, updated. restrictions on secondary uses of the identifier are important in order to safeguard the privacy of the individual and to preserve the security of the authentication system. the individual should have clear rights to access information about how data are protected and used by the authentication system and the individual should have the right to challenge, correct and amend any information related to the identifier or its uses. among privacy projects, prime has identified a set of privacy principles in the design of identity management architecture: principle : design must start from maximum privacy. principle : explicit privacy rules govern system usage. principle : privacy rules must be enforced, not just stated. principle : privacy enforcement must be trustworthy. principle : users need easy and intuitive abstractions of privacy. principle : privacy needs an integrated approach. principle : privacy must be integrated with applications. trust marks and trust seals can also be useful safeguards because the creation of public credibility is a good way for organisations to alert consumers and other individuals to an organisation's practices and procedures through participation in a programme that has an easy-to-recognise symbol or seal. trust marks and seals are a form of guarantee provided by an independent organisation that maintains a list of trustworthy companies that have been audited and certified for compliance with some industry-wide accepted or standardised best practice in collecting personal or sensitive data. once these conditions are met, they are allowed to display a trust seal logo or label that customers can easily recognise. a trust mark must be supported by mechanisms necessary to maintain objectivity and build legitimacy with consumers. trust seals and trust marks are, however, voluntary efforts that are not legally binding and an effective enforcement needs carefully designed procedures and the backing of an independent and powerful organisation that has the confidence of all affected parties. trust seals and trust marks are often promoted by industry, as opposed to consumer-interest groups. as a result, concerns exist that consumers' desires for stringent privacy protections may be compromised in the interest of industry's desire for the new currency of information. moreover, empirical evidence indicates that even some eight years after the introduction of the first trust marks and trust seals in internet commerce, citizens know little about them and none of the existing seals has reached a high degree of familiarity among customers. this does not necessarily mean that trust marks are not an adequate safeguard for improving security and privacy in the ambient intelligence world, it suggests that voluntary activities like self-regulation have -apart from being well designed -to be complemented by other legally enforceable measures. in addition to the general influence of cultural factors and socialisation, trust results from context-specific interaction experiences. as is well documented, computermediated interactions are different from conventional face-to-face exchanges due to anonymity, lack of social and cultural clues, "thin" information, and the uncertainty about the credibility and reliability of the provided information that commonly characterise mediated relationships. in an attempt to reduce some of the uncertainties associated with online commerce, many websites acting as intermediaries between transaction partners are operating so-called reputation systems. these institutionalised feedback mechanisms are usually based on the disclosure of past transactions rated by the respective partners involved. giving participants the opportunity to rank their counterparts creates an incentive for rule-abiding behaviour. thus, reputation systems seek to imitate some of the real-life trust-building and social constraint mechanisms in the context of mediated interactions. so far, reputation systems have not been developed for ami services. and it seems clear that institutionalised feedback mechanisms will only be applicable to a subset of future ami services and systems. implementing reputation systems only makes sense in those cases in which users have real choices between different suppliers (for instance, with regard to ami-assisted commercial transactions or information brokers). ami infrastructures that normally cannot be avoided if one wants to take advantage of a service need to be safeguarded by other means, such as trust seals, iso guidelines and regulatory action. despite quite encouraging experiences in numerous online arenas, reputation systems are far from perfect. many reputation systems tend to shift the burden of quality control and assessment from professionals to the -not necessarily entirely informed -individual user. in consequence, particularly sensitive services should not exclusively be controlled by voluntary and market-style feedbacks from customers. furthermore, reputation systems are vulnerable to manipulation. pseudonyms can be changed, effectively erasing previous feedback. and the feedback itself need not necessarily be sincere, either due to co-ordinated accumulation of positive feedback, due to negotiations between parties prior to the actual feedback process, because of blackmailing or the fear of retaliation. last but not least, reputation systems can become the target of malicious attacks, just like any netbased system. an alternative to peer-rating systems are credibility-rating systems based on the assessment of trusted and independent institutions, such as library associations, consumer groups or other professional associations with widely acknowledged expertise within their respective domains. ratings would be based on systematic assessments along clearly defined quality standards. in effect, these variants of reputation-and credibility-enhancing systems are quite similar to trust marks and trust seals. the main difference is that professional rating systems enjoy a greater degree of independence from vested interests. and, other than in the case of peer-rating systems which operate literally for free, the independent professional organisations need to be equipped with adequate resources. on balance, reputation systems can contribute to trust-building between strangers in mediated short-term relations or between users and suppliers, but they should not be viewed as a universal remedy for the ubiquitous problem of uncertainty and the lack of trust. a possible safeguard is a contract between the service provider and the user that has provisions about privacy rights and the protection of personal data and notification of the user of any processing or transfer of such data to third parties. while this is a possible safeguard, there must be some serious doubt about the negotiating position of the user. it's quite possible the service provider would simply say here are the terms under which i'm willing to provide the service, take it or leave it. also, from the service provider's point of view, it is unlikely that he would want to conclude separate contracts with every single user. in a world of ambient intelligence, such a prospect becomes even more unlikely in view of the fact that the "user", the consumer-citizen will be moving through resnick, p., r. zeckhauser, e. friedman and k. kuwabara, "reputation systems: facilitating trust in internet interactions", communications of the acm, ( ), , pp. - . http:// www.si.umich.edu/~presnick/papers/cacm /reputations.pdf. different spaces where there is likely to be a multiplicity of different service providers. it may be that the consumer-citizen would have a digital assistant that would inform him of the terms, including the privacy implications, of using a particular service in a particular environment. if the consumer-citizen did not like the terms, he would not have to use the service. consumer associations and other civil society organisations (csos) could, however, play a useful role as a mediator between service providers and individual consumers and, more particularly, in forcing the development of service contracts (whether real or implicit) between the service provider and the individual consumer. consumer organisations could leverage their negotiating position through the use of the media or other means of communication with their members. csos could position themselves closer to the industry vanguard represented in platforms such as artemis by becoming members of such platforms themselves. within these platforms, csos could encourage industry to develop "best practices" in terms of provision of services to consumers. government support for new technologies should be linked more closely to an assessment of technological consequences. on the basis of the far-reaching social effects that ambient intelligence is supposed to have and the high dynamics of the development, there is a clear deficit in this area. research and development (at least publicly supported r&d) must highlight future opportunities and possible risks to society and introduce them into public discourse. every research project should commit itself to explore possible risks in terms of privacy, security and trust, develop a strategy to cover problematic issues and involve users in this process as early as possible. a template for "design guidelines" that are specifically addressing issues of privacy has been developed by the "ambient agora" project which has taken into account the fundamental rules by the oecd, notably its guidelines on the protection of privacy and transborder flows of personal data, adopted on september , and the more recent guidelines for the security of information systems and networks. if the state acts as a buyer of strategically important innovative products and services, it contributes to the creation of the critical demand that enables suppliers to reduce their business risk and realise spillover effects. thus, public procurement programmes can be used to support the demand for and use of improved products and services in terms of security and privacy or identity protection. in the procurement of ict products, emphasis should therefore be given to critical issues such as security and trustworthiness. as in other advanced fields, it will be a major challenge to develop a sustainable procurement policy that can cope with ever-decreasing innovation cycles. the focus should not be on the characteristics of an individual product or component, but on the systems into which components are integrated. moreover, it is important to pay attention to the secondary and tertiary impacts resulting from deployment of large technical systems such as ambient intelligence. an evaluation of the indirect impacts is especially recommended for larger (infrastructure) investments and public services. while public procurement of products and services that are compliant with the eu legal framework and other important guidelines for security, privacy and identity protection is no safeguard on its own, it can be an effective means for the establishment and deployment of standards and improved technological solutions. accessibility is a key concept in helping to promote the social inclusion of all citizens in the information society embedded with ami technologies. accessibility is needed to ensure user control, acceptance, enforceability of policy in a user-friendly manner and the provision of citizens with equal rights and opportunities in a world of ambient intelligence. all citizens should have equal rights to benefit from the new opportunities that ami technologies will offer. this principle promotes the removal of direct and indirect discrimination, fosters access to services and encourages targeted actions in favour of under-represented groups. this principle promotes system design according to a user-centric approach (i.e., the concept of "design for all"). the design-for-all concept enables all to use applications (speech technology for the blind, pictures for the deaf). it means designing in a way to make sure applications are user-friendly and can be used intuitively. in short, industry has to make an effort to simplify the usage of icts, rather than forcing prospective users to learn how to use otherwise complex icts. better usability will then support easy learning (i.e., learning by observation), user control and efficiency, thus increasing satisfaction and, consequently, user acceptance. this principle aims to overcome user dependency and more particularly user isolation and stress due to the complexity of new technology, which leads to loss of control. education programmes on how to use new technologies will increase user awareness about the different possibilities and choices offered by ami technologies and devices. training and education help to overcome user dependency and social disruptions. user awareness is important to reduce the voluntary exclusion caused by a misunderstanding on how the technology works. this safeguard is essential in order to prevent almost all facets of dependency, system dependency as well as user dependency. consumers need to be educated about the privacy ramifications arising from virtually any transaction in which they are engaged. an education campaign should be targeted at different segments of the population. school-age children should be included in any such campaign. any networked device, particularly those used by consumer-citizens, should come with a privacy warning much like the warnings on tobacco products. when the uk department of trade and industry (dti) released its information security review, the uk e-commerce minister emphasised that everyone has a role to play in protecting information: "risks are not well managed. we need to dispel the illusion the information security issues are somebody else's problem. it's time to roll up our sleeves." the oecd shares this point of view. it has said that "all participants in the new information society … need … a greater awareness and understanding of security issues and the need to develop a 'culture of security'." the oecd uses the word "participants", which equates to "stakeholders", and virtually everyone is a participant or stakeholder -governments, businesses, other organisations and individual users. oecd guidelines are aimed at promoting a culture of security, raising awareness and fostering greater confidence (i.e., trust) among all participants. there are various ways of raising awareness, and one of those ways would be to have some contest or competition for the best security or privacy-enhancing product or service of the year. the us government's department of homeland security is sponsoring such competitions, and europe could usefully draw on their experience to hold similar competitions in europe. in the same way as the principle that "not everything that you read in the newspapers is true" has long been part of general education, in the ict age, awareness should generally be raised by organisations that are trustworthy and as close to the citizen as possible (i.e., on the local or regional level. questions of privacy, identity and security are, or should be, an integral part of the professional education of computer scientists. we agree with and support the commission's "invitation" to member states to "stimulate the development of network and information security programmes as part of higher education curricula". perhaps one of the best safeguards is public opinion, stoked by stories in the press and the consequent bad publicity given to perceived invasions of privacy by industry and government. new technologies often raise policy issues, and this is certainly true of ambient intelligence. ami offers great benefits, but the risk of not adequately addressing public concerns could mean delays in implementing the technologies, a lack of public support for taxpayer-funded research and vociferous protests by privacy protection advocates. cultural artefacts, such as films and novels, may serve as safeguards against the threats and vulnerabilities posed by advanced technologies, including ambient intelligence. science fiction in particular often presents a dystopian view of the future where technology is used to manipulate or control people, thus, in so doing, such artefacts raise our awareness and serve as warnings against the abuse of technology. a new york times film critic put it this way: "it has long been axiomatic that speculative science-fiction visions of the future must reflect the anxieties of the present: fears of technology gone awry, of repressive political authority and of the erosion of individuality and human freedom." an example of a cultural artefact is steven spielberg's film, minority report, which depicts a future embedded with ambient intelligence, which serves to convey messages or warnings from the director to his audience. minority report is by no means unique as a cultural artefact warning about how future technologies are like a double-edged knife that cuts both ways. to implement socio-economic safeguards will require action by many different players. unfortunately, the very pervasiveness of ami means that no single action by itself will be sufficient as a safeguard. a wide variety of socio-economic safeguards, probably even wider than those we have highlighted in the preceding sections, will be necessary. as implementation of ami has already begun (with rfids, surveillance systems, biometrics, etc.), it is clearly not too soon to begin implementation of safeguards. we recommend, therefore, that all stakeholders, including the public, contribute to this effort. the fast emergence of information and communication technologies and the growth of online communication, e-commerce and electronic services that go beyond the territorial borders of the member states have led the european union to adopt numerous legal instruments such as directives, regulations and conventions on ecommerce, consumer protection, electronic signature, cyber crime, liability, data protection, privacy and electronic communication … and many others. even the european charter of fundamental rights will play an important role in relation to the networked information society. our analysis of the dark scenarios shows that we may encounter serious legal problems when applying the existing legal framework to address the intricacies of an ami environment. our proposed legal safeguards should be considered as general policy options, aimed at stimulating discussion between stakeholders and, especially, policymakers. law is only one of the available sets of tools for regulating behaviour, next to social norms, market rules, "code" -the architecture of the technology (e.g., of cyberspace, wireless and wired networks, security design, encryption levels, rights management systems, mobile telephony systems, user interfaces, biometric features, handheld devices and accessibility criteria) and many other tools. the regulator of ambient intelligence can, for instance, achieve certain aims directly by imposing laws, but also indirectly by, for example, influencing the rules of the market. regulatory effect can also be achieved by influencing the architecture of a certain environment. the architecture of ami might well make certain legal rules difficult to enforce (for example, the enforcement of data protection obligations on the internet or the enforcement of copyright in peer-to-peer networks), and might cause new problems, particularly related to the new environment (spam, dataveillance ). on the other hand, the "code" has the potential to regulate by enabling or disabling certain behaviour, while law regulates via the threat of sanction. in other words, software and hardware constituting the "code", and architecture of the digital world, causing particular problems, can be at the same time the instrument to solve them. regulating through code may have some specific advantages: law traditionally regulates ex post, by imposing a sanction on those who did not comply with its rules (e.g., in the form of civil damages or criminal prosecution). architecture regulates by putting conditions on one's behaviour, allowing or disallowing something, not allowing the possibility to disobey. it regulates ex ante. ambient intelligence is particularly built on software code. this code influences how ambient intelligence works, e.g., how the data are processed, but this code itself can be influenced and accompanied by regulation. thus, the architecture can be a tool of law. this finding is more than elementary. it shows that there is a choice: should the law change because of the "code"? or should the law change "code" and thus ensure that certain values are protected? the development of technology represents an enormous challenge for privacy, enabling increasing surveillance and invisible collection of data. a technology that threatens privacy may be balanced by the use of a privacy-enhancing technology: the "code", as lessig claims, can be the privacy saviour. other technologies aim to limit the amount of data actually collected to the necessary minimum. however, most of the current technologies simply ignore the privacy implications and collect personal data when there is no such need. a shift of the paradigm to privacy-bydesign is necessary to effectively protect privacy. indeed, technology can facilitate privacy-friendly verification of individuals via, for example, anonymous and pseudonymous credentials. leenes and koops recognise the potential of these privacyenhancing technologies (pets) to enforce data protection law and privacy rules. but they also point at problems regarding the use of such technologies, which are often troublesome in installation and use for most consumers. moreover, industry is not really interested in implementing privacy-enhancing technology. they see no (economic) reason to do it. the analysis of leenes and koops shows that neither useful technology, nor law is sufficient in itself. equally important is raising stakeholder awareness, social norms and market rules. all regulatory means should be used and have to be used to respond to problems of the new environment to tackle it effectively. for the full effectiveness of any regulation, one should always look for the optimal mixture of all accessible means. as the impact and effects of the large-scale introduction of ami in societies spawn a lot of uncertainties, the careful demarche implied by the precautionary principle, with its information, consultation and participation constraints, might be appropriate. the application of this principle might inspire us in devising legal policy options when, as regards ami, fundamental choices between opacity tools and transparency tools must be made. opacity tools proscribe the interference by powerful actors into the individual's autonomy, while transparency tools accept such interfering practices, though under certain conditions which guarantee the control, transparency and accountability of the interfering activity and actors. legal scholars do not discuss law in general terms. their way of thinking always involves an application of the law in concrete or exemplified situations. the legislator will compare concrete examples and situations with the law and will not try to formulate general positions or policies. thus, the proposed legal framework will not deal with the ami problems in a general way, but focus on concrete issues, and apply opacity and transparency solutions accordingly. another particularity of legal regulation in cyberspace is the absence of a central legislator. though our legal analysis is based mostly on european law, we emphasise that not everything is regulated at a european level. regulation of (electronic) identity cards, for instance, concerns a crucial element in the construction of an ami environment, but is within the powers of the individual member states. both at european and national level, some decision-making competences have been delegated to independent advisory organs (children's rights commissioners, data protection authorities). hence, there exist many, what we can call, "little legislators" that adjust in some way the often executive power origin of legislation: the article data protection working party, national children's rights commissioners and international standardisation bodies can and do, for example, draft codes of conduct that constitute often (but not always) the basis for new legislation. we do not suggest the centralisation of the law-making process. on the contrary, we recommend respect for the diversity and plurality of lawmakers. the solutions produced by the different actors should be taken into consideration and be actively involved in policy discussions. development of case law should also be closely observed. consulting concerned citizens and those who represent citizens ( including legislators) at the stage of development would increase the legitimacy of new technologies. privacy aims to ensure no interference in private and individual matters. it offers an instrument to safeguard the opacity of the individual and puts limits to the interference by powerful actors into the individual's autonomy. normative in nature, regulatory opacity tools should be distinct from regulatory transparency tools, of which the goal is to control the exercise of power rather than to restrict power. we observe today that the reasonable expectation of privacy is eroding due to emerging new technologies and possibilities for surveillance: it develops into an expectation of being monitored. should this, however, lead to diminishing the right to privacy? ambient intelligence may seriously threaten this value, but the need for privacy (e.g., the right to be let alone) will probably remain, be it in another form adapted to new infrastructures (e.g., the right to be left offline). the right to privacy in a networked environment could be enforced by any means of protecting the individual against any form of dataveillance. such means are in line with the data minimisation principle of data protection law, which is a complementary tool to privacy. however, in ambient intelligence where collecting and processing personal data is almost a prerequisite, new tools of opacity such as the right to be left offline (in time, e.g., during certain minutes at work, or in space, e.g., in public bathrooms) could be recognised. several instruments of opacity can be identified. we list several examples, and there may be others. additional opacity recommendations are made in subsequent sections, for example, with regard to biometrics. we observe that there is not necessarily an internal coherence between the examples listed below. the list should be understood as a wish list or a list with suggestions to be consulted freely. opacity designates a zone of non-interference which should not be confused with a zone of invisibility: privacy, for instance, does not imply secrecy; it implies the possibility of being oneself openly without interference. another word might have been "impermeability" which is too strong and does not contrast so nicely with "transparency" as "opacity" does. see hildebrandt, m., and s. gutwirth the concept of a digital territory represents a vision that introduces the notions of space and borders in future digitised everyday life. it could be visualised as a bubble, the boundaries and transparency of which depend on the will of its owner. the notion of a digital territory aims for a "better clarification of all kinds of interactions in the future information society. without digital boundaries, the fundamental notion of privacy or the feeling of being at home will not take place in the future information society." the concept of digital territories encompasses the notion of a virtual residence, which can be seen as a virtual representation of the smart home. the concept of digital territories could provide the individual with a possibility to access -and stay in -a private digital territory of his own at (any) chosen time and place. this private, digital space could be considered as an extension of the private home. today, already, people store their personal pictures on distant servers, read their private correspondences online, provide content providers with their viewing and consuming behaviour for the purpose of digital rights management, communicate with friends and relatives through instant messengers and internet telephony services. the "prognosis is that the physical home will evolve to 'node' in the network society, implying that it will become intimately interconnected to the virtual world." the law guarantees neither the establishment nor the protection of an online private space in the same way as the private space in the physical world is protected. currently, adequate protection is lacking. for example, the new data retention law requires that telecommunication service providers keep communication data at the disposal of law enforcement agencies. the retention of communication data relates to mobile and fixed phone data, internet access, e-mail and e-telephony. data to be retained includes the place, time, duration and destination of communications. what are the conditions for accessing such data? is the individual informed when such data are accessed? does he have the right to be present when such data are examined? does the inviolability of the home extend to the data that are stored on a distant server? another example of inadequate protection concerns the increasing access to home activities from a distance, for example, as a result of the communication data generated by domestic applications that are connected to the internet. in both examples, there is no physical entrance in the private place. to ensure that these virtual private territories become a private domain for the individual, a regulatory framework could be established to prevent unwanted and unnoticed interventions similar to that which currently applies to the inviolability of the home. a set of rules needs to be envisaged to guarantee such protection, amongst them, the procedural safeguards similar to those currently applicable to the protection of our homes against state intervention (e.g., requiring a search warrant). technical solutions aimed at defending private digital territories against intrusion should be encouraged and, if possible, legally enforced. the individual should be empowered with the means to freely decide what kinds of information he or she is willing to disclose, and that aspect should be included in the digital territory concept. similarly, vulnerable home networks should be granted privacy protection. such protection could be extended to the digital movement of the person, that is, just as the privacy protection afforded the home has been or can be extended to the individual's car, so the protection could be extended to home networks, which might contact external networks. privacy at the workplace has already been extensively discussed. most of the legal challenges, we believe, that may arise can be answered with legal transparency rules. more drastic, prohibitive measures may be necessary in certain situations involving too far-reaching or unnecessary surveillance, which a society considers as infringing upon the dignity of the employee. in addition, transparency rules are needed to regulate other, less intrusive problems. we recall here the specific role of law-making institutions in the area of labour law. companies must discuss their surveillance system and its usage in collective negotiations with labour organisations and organisations representing employees before its implementation in a company or a sector, taking into account the specific needs and risks involved (e.g., workers in a bank vs. workers in public administration). all employees should always be clearly and a priori informed about the employee surveillance policy of the employer (when and where surveillance is taking place, what is the finality, what information is collected, how long it will be stored, what are the (procedural) rights of the employees when personal data are to be used as evidence, etc.). specific cyber territories for children have to be devised along the same lines. the united nations convention on the rights of the child ( ) contains a specific privacy right for children, and sets up monitoring instruments such as national children's rights commissioners. opinions of such advisory bodies should be carefully taken into account in policy discussion. national children's rights commissioners could take up problems relating to the permanent digital monitoring of children. as concluded in the legal analysis of the dark scenarios above, courts are willing to protect one's privacy but, at the same time, they tend to admit evidence obtained through a violation of privacy or data protection. there is a lack of clarity and uniformity regarding the consequence of privacy violations. in that case, the evidence was secured by the police in a manner incompatible with the requirements of article of the european convention on human rights (echr). the court accepted that the admission of evidence obtained in breach of the privacy right is not necessarily a breach of the required fairness under article of echr (the right to a fair trial), since the process taken as a whole was fair in the sense of article . the evidence against the accused was admitted and led to his conviction. the khan doctrine (followed in cases such as doerga v. the netherlands and p.g. and j.h. v. the united followed by at least some national courts. the fact that there is no general acceptance of an exclusionary rule creates legal uncertainty. its general acceptance is, however, necessary to protect the opacity of the individual in a more effective way. the departure from such position by the courts (namely "no inclusion of evidence obtained through privacy and/or data protection law infringements") could be considered and legislative prohibition of the admissibility (or general acceptance of the exclusionary rule) of such obtained evidence envisaged. in ambient intelligence, the use of implants can no longer be considered as a kind of futuristic or extraordinary exception. whereas it is clear that people may not be forced to use such implants, people may easily become willing to equip themselves with such implants on a (quasi) voluntary basis, be it, for example, to enhance their bodily functions or to obtain a feeling of security through always-on connections to anticipate possible emergencies. such a trend requires a careful assessment of the opacity and transparency principles at a national, european and international level. currently, in europe, the issue of medical implants has already been addressed. in ami, however, implants might be used for non-medical purposes. one of our dark scenarios suggests that organisations could force people to have an implant so they could be located anywhere at any time. cannot be achieved by less body-intrusive means. informed consent is necessary to legitimise the use of implants. we agree with those findings. the european group on ethics in science and new technologies goes further, stating that non-medical (profit-related) applications of implants constitute a potential threat to human dignity. applications of implantable surveillance technologies are only permitted when there is an urgent and justified necessity in a democratic society, and must be specified in legislation. we agree that such applications should be diligently scrutinised. we propose that the appropriate authorities (e.g., the data protection officer) control and authorise applications of implants after the assessment of the particular circumstances in each case. when an implant enables tracking of people, people should have the possibility to disconnect the implant at any given moment and they should have the possibility to be informed when a (distant) communication (e.g., through rfid) is taking place. we agree with the european group on ethics in science and new technologies that irreversible ict implants should not be used, except for medical purposes. further research on the long-term impact of ict implants is also recommended. another safeguard to guarantee the opacity of the individual is the possibility to act under anonymity (or at least under pseudonymity or "revocable anonymity"). the article working party has considered anonymity as an important safeguard for the right to privacy. we repeat here its recommendations: (a) the ability to choose to remain anonymous is essential if individuals are to preserve the same protection for their privacy online as they currently enjoy offline. (b) anonymity is not appropriate in all circumstances. (c) legal restrictions which may be imposed by governments on the right to remain anonymous, or on the technical means of doing so (e.g., availability of encryption products) should always be proportionate and limited to what is necessary to protect a specific public interest in a democratic society. (e) some controls over individuals contributing content to online public fora are needed, but a requirement for individuals to identify themselves is in many cases disproportionate and impractical. other solutions are to be preferred. (f) anonymous means to access the internet (e.g., public internet kiosks, prepaid access cards) and anonymous means of payment are two essential elements for true online anonymity. according to the common criteria for information technology security evaluation document (iso ), anonymity is only one of the requirements for the protection of privacy, next to pseudonymity, unlinkability, unobservability, user control/ information management and security protection. all these criteria should be considered as safeguards for privacy. the e-signature directive promotes the use of pseudonyms and, at the same time, aims to provide security for transactions. currently, the directive on electronic signatures states that only advanced electronic signatures (those based on a qualified certificate and created by a secure signature-creation device) satisfy the legal requirements of a signature in relation to data in electronic form in the same manner as a handwritten signature satisfies those requirements in relation to paper-based data and are admissible as evidence in legal proceedings. member states must ensure that an electronic signature (advanced or not) is not denied legal effectiveness and admissibility as evidence in legal proceedings solely on the grounds that it is: (a) in electronic form, (b) not based upon a qualified certificate, (c) not based upon a qualified certificate issued by an accredited certification service-provider, or (d) not created by a secure signature-creation device. in ambient intelligence, the concept of unlinkability can become as important as the concept of anonymity or pseudonymity. unlinkability "ensures that a user may make multiple uses of resources or services without others being able to link these uses together. … unlinkability requires that users and/or subjects are unable to determine whether the same user caused certain specific operations in the system." when people act pseudonymously or anonymously, their behaviour in different times and places in the ambient intelligence network could still be linked and consequently be subject to control, profiling and automated decision-making: linking data relating to the same non-identifiable person may result in similar privacy threats as linking data that relate to an identified or identifiable person. thus, in addition to and in line with the right to remain anonymous goes the use of anonymous and pseudonymous credentials, accompanied with unlinkability in certain situations (e.g., e-commerce), reconciling thus the privacy requirements with the accountability requirements of, e.g., e-commerce. in fact, such mechanisms should always be foreseen when disclosing someone's identity or when linking the information is not necessary. such necessity should not be easily assumed, and in every circumstance more privacy-friendly technological solutions should be sought. however, the use of anonymity should be well balanced. to avoid its misuse, digital anonymity could be further legally regulated, especially stating when it is not appropriate. by the cybercrime convention, which provides a definition for several criminal offences related to cybercrime and for general principles concerning international co-operation. the cybercrime convention, however, allows for different standards of protection. the convention obliges its signatories to criminalise certain offences under national law, but member states are free to narrow the scope of the definitions. the most important weakness of this convention is the slow progress in its ratification by signatory states. council framework decision / /jha also provides for criminal sanctions against cybercrimes. the framework decision is limited, however, both in scope and territory, since it only defines a limited number of crimes and is only applicable to the member states of the european union. international co-operation in preventing, combating and prosecuting criminals is needed and may be facilitated by a wide range of technological means, but these new technological possibilities should not erode the privacy of innocent citizens who are deemed to be not guilty until proven otherwise. cybercrime prosecution, and more importantly crime prevention, might be facilitated by a wide range of technological means, among them, those that provide for the security of computer systems and data against attacks. almost all human activity in ami can be reduced to personal data processing: opening doors, sleeping, walking, eating, putting lights on, shopping, walking in a street, driving a car, purchasing, watching television and even breathing. in short, all physical actions become digital information that relates to an identified or identifiable individual. often, the ambient intelligence environment will need to adapt to individuals and will therefore use profiles applicable to particular individuals or to individuals within a group profile. ami will change not only the amount, but also the quality of data collected so that we can be increasingly supported in our daily life (a goal of ambient intelligence). ami will collect data not only about what we are doing, when we do it and where we are, but also data on how we have experienced things. one can assume that the accuracy of the profiles, on which the personalisation of services depends, will improve as the amount of data collected grows. but as others hold more of our data, so grow the privacy risks. thus arises the fundamental question: do we want to minimise personal data collection? instead of focusing on reducing the amount of data collected alone, should we admit that they are indispensable for the operation of ami, and focus rather on empowering the user with a means to control such processing of personal data? data protection is a tool for empowering the individual in relation to the collection and processing of his or her personal data. the european data protection directive imposes obligations on the data controller and supports the rights of the data subject with regard to the transparency and control over the collection and processing of data. it does not provide for prohibitive rules on data processing (except for the processing of sensitive data and the transfer of personal data to third countries that do not ensure an adequate level of protection). instead, the eu data protection law focuses on a regulatory approach and on channelling, controlling and organising the processing of personal data. as the title of directive / /ec indicates, the directive concerns both the protection of the individual with regard to the processing of personal data and the free movement of such data. the combination of these two goals in directive / /ec reflects the difficulties we encounter in the relations between ambient intelligence and data protection law. there is no doubt that some checks and balances in using data should be put in place in the overall architecture of the ami environment. civil movements and organisations dealing with human rights, privacy or consumer rights, observing and reacting to the acts of states and undertakings might provide such guarantees. it is also important to provide incentives for all actors to adhere to legal rules. education, media attention, development of good practices and codes of conducts are of crucial importance. liability rules and rules aimed at enforcement of data protection obligations will become increasingly important. data protection law provides for the right to information on data processing, access to or rectification of data, which constitute important guarantees of individual rights. however, its practical application in an ami era could easily lead to an administrative nightmare, as information overload would make it unworkable. we should try to remedy such a situation in a way that does not diminish this right. the individual's right to information is a prerequisite to protect his interests. such a right corresponds to a decentralised system of identity (data) management, but it seems useful to tackle it separately to emphasise the importance of the individual's having access to information about the processing of his data. because of the large amounts of data to be processed in an ami world, the help of or support by intelligent agents to manage such information streams seems indispensable. information about what knowledge has been derived from the data could help the individual in proving causality in case of damage. further research on how to reconcile access to the knowledge in profiles (which might be construed as a trade secret in some circumstances) with intellectual property rights would be desirable. the right to be informed could be facilitated by providing information in a machine-readable language, enabling the data subject to manage the information flow through or with the help of (semi-) autonomous intelligent agents. of course, this will be more difficult in situations of passive authentication, where no active involvement of the user takes place (e.g., through biometrics and rfids). thus, information on the identity of the data controller and the purposes of processing could exist both in human-readable and machine-readable language. the way such information is presented to the user is of crucial importance -i.e., it must be presented in an easily comprehensible, user-friendly way. in that respect, the article working party has provided useful guidelines and proposed multilayer eu information notices essentially consisting of three layers: layer -the short notice contains core information required under article of the data protection directive (identity of the controller, purpose of processing, or any additional information which, in the view of the particular circumstances of the case, must be provided to ensure fair processing). a clear indication must be given as to how the individual can access additional information. layer -the condensed notice contains all relevant information required under the data protection directive. this includes the name of the company, the purpose of the data processing, the recipients or categories of recipients of the data, whether replies to the questions are obligatory or voluntary, as well as the possible consequences of failure to reply, the possibility of transfer to third parties, the right to access, to rectify and oppose choices available to the individual. in addition, a point of contact must be given for questions and information on redress mechanisms either within the company itself or details of the nearest data protection agency. layer -the full notice includes all national legal requirements and specificities. it could contain a full privacy statement with possible additional links to national contact information. we recommend that industry and law enforcement agencies consider an approach for ami environments similar to that recommended by the article working party. electronic versions of such notices should be sufficient in most of circumstances. our dark scenarios indicate a new kind of practice that has emerged in recent years in the sector of personal data trading: while some companies collect personal data in an illegal way (not informing the data subjects, transfer of data to third parties without prior consent, usage for different purposes, installing spyware, etc.), these personal data are shared, sold and otherwise transferred throughout a chain of existing and disappearing companies to the extent that the origin of the data and the original data collector cannot be traced back. this practice has been described as "data laundering", with analogy to money laundering: it refers to a set of activities aiming to cover the illegitimate origin of data. in our ami future, we should assume the value of personal data and therefore the (illegal) trading in these data will only grow. a means to prevent data laundering could be to oblige those who buy or otherwise acquire databases, profiles and vast amounts of personal data to check diligently the legal origin of the data. without checking the origin and/or legality of the databases and profiles, one could consider the buyer equal to a receiver of stolen goods and thus held liable for illegal data processing. they could be obliged to notify the national data protection officers when personal data(bases) are acquired. those involved or assisting in data laundering could be subject to criminal sanctions. ami requires efficient, faultless exchanges of relevant data and information throughout the ami network. the need for efficiency requires interoperable data formats and interoperable hardware and software for data processing. dark scenario (about the bus accident) has shown the need for interoperability in ambient intelligence, but it must be recognised that, at the same time, interoperable data and data processing technologies in all sectors and all applications could threaten trust, privacy, anonymity and security. full interoperability and free flow of personal data are not always desirable, and should not be considered as unquestionable. interoperability can entail an unlimited availability of personal data for any purpose. interoperability may infringe upon the finality and purpose specification principles and erode the rights and guarantees offered by privacy and data protection law. moreover, the purposes for which the data are available are often too broadly described (what is "state security", "terrorism", "a serious crime"?). data can become available afterwards for any purpose. interoperability of data and data processing mechanisms facilitates possible function creep (use of data for purposes other than originally envisaged). interoperability could contribute to the criminal use of ambient intelligence, for example, by sending viruses to objects in the network (interoperability opens the door for fast transmission and reproduction of a virus) or abusing data (interoperable data formats make data practical for any usage). interoperability is thus not only a technological issue. awareness -already today -of the possible negative sides of interoperability should bring about a serious assessment of both law and technology before the market comes up with tools for interoperability. legal initiatives in france (e.g., requiring interoperability of the itunes music platform) and sanctions imposed by the european commission (imposing interoperability of the microsoft work group server operating system) indicate clearly that the need for interoperability is desired on a political and societal level. in the communication from the commission to the council and the european parliament on improved effectiveness, enhanced interoperability and synergies among european databases in the area of justice and home affairs of , interoperability is defined as the "ability of it systems and of the business processes they support to exchange data and to enable the sharing of information and knowledge". this is, however, a more technological definition: it "explicitly disconnects the technical and the legal/political dimensions from interoperability, assuming that the former are neutral and the latter can come into play later or elsewhere. … indeed, technological developments are not inevitable or neutral, which is mutatis mutandis also the case for technical interoperability. the sociology of sciences has shown that any technological artefact has gone through many small and major decisions that have moulded it and given it its actual form. hence, the development of information technology is the result of micro politics in action. technologies are thus interwoven with organisation, cultural values, institutions, legal regulation, social imagination, decisions and controversies, and, of course, also the other way round. any denial of this hybrid nature of technology and society blocks the road toward a serious political, democratic, collective and legal assessment of technology. this means that technologies cannot be considered as faits accomplis or extrapolitical matters of fact." this way of proceeding has also been criticised by the european data protection supervisor, according to whom this leads to justifying the ends by the means. taking into account the need for interoperability, restrictions in the use and implementation of interoperability are required based on the purpose specification and proportionality principles. to this extent, a distinction between the processing of data for public (enforcement) to achieve certain purposes, for which access to data has been granted, access to the medium carrying the information (e.g., a chip) may be sufficient, for example, when verifying one's identity. there should always be clarity as to what authorities are being granted access. in the case of deployment of centralised databases, a list of authorities that have access to the data should be promulgated in an adequate, official, freely and easily accessible publication. such clarity and transparency would contribute to security and trust, and protect against abuses in the use of databases. the proportionality and purpose limitation principles are already binding under existing data protection laws. the collection and exchange of data (including interoperability) should be proportional to the goals for which the data have been collected. it will not be easy to elaborate the principles of proportionality and purpose limitation in ambient intelligence; previously collected data may serve for later developed applications or discovered purposes. creation and utilisation of databases may offer additional benefits (which are thus additional purposes), e.g., in the case of profiling. those other (derived) purposes should, as has been indicated in the opinion of the european data protection supervisor, be treated as independent purposes for which all legal requirements must be fulfilled. technical aspects of system operation can have a great impact on the way a system works, and how the proportionality principles and purpose limitation principles are implemented since they can determine, for example, if access to the central database is necessary, or whether access to the chip or part of the data is possible and sufficient. biometric technology can be a useful tool for authentication and verification, and may even be a privacy-enhancing technology. however, it can also constitute a threat to fundamental rights and freedoms. thus, specific safeguards should be put in place. biometric safeguards have already been subject of reflection by european data protection authorities: the article working party has stated that biometric data are in most cases personal data, so that data protection principles apply to processing of such data. on the principle of proportionality, the article working party points out that it is not necessary (for the sake of authentication or verification) to store biometric data in central databases, but in the medium (e.g., a card) remaining in the control of the user. the creation and use of centralised databases should always be carefully assessed before their deployment, including prior checking by data protection authorities. in any case, all appropriate security measures should be put in place. framing biometrics is more than just deciding between central or local storage. even storage of biometric data on a smart card should be accompanied by other regulatory measures that take the form of rights for the card-holders (to know what data and functions are on the card; to exclude certain data or information from being written onto the card; to reveal at discretion all or some data from the card; to remove specific data or information from the card). biometric data should not be used as unique identifiers, mainly because biometric data still do not have sufficient accuracy. of course, this might be remedied in the progress of science and technological development. there remains, however, a second objection: using biometrics as the primary key will offer the possibility of merging different databases, which can open the doors for abuses (function creep). european advisory bodies have considered biometric data as a unique identifier. generally speaking, since the raw data might contain more information than actually needed for certain finalities (including information not known at the moment of the collection, but revealed afterwards due to progress in science, e.g., health information related to biometric data), it should not be stored. other examples of opacity rules applied to biometrics might be prohibitions on possible use of "strong" multimodal biometrics (unless for high security applications) for everyday activities. codes of conduct can be appropriate tools to further regulate the use of technology in particular sectors. ami will depend on profiling as well as authentication and identification technologies. to enable ubiquitous communication between a person and his or her environment, both things and people will have to be traced and tracked. rfid seems to offer the technological means to implement such tracking. like biometrics, rfid is an enabling technology for real-time monitoring and decision making. like biometrics, rfids can advance the development of ami and provide many advantages for users, companies and consumers. no legislative action seems needed to support this developing technology. market mechanisms are handling this. there is, however, a risk to the privacy interests of the individual and for a violation of the data protection principles, as caspian and other privacy groups have stated. rfid use should be in accordance with privacy and data protection regulations. the article working party has already given some guidelines on the application of the principles of eu data protection legislation to rfids. it stresses that the data protection principles (purpose limitation principle, data quality principle, conservation principle, etc.) must always be complied with when the rfid technology leads to processing of personal data in the sense of the data protection directive. as the article working party points out, the consumer should always be informed about the presence of both rfid tags and readers, as well as of the responsible controller, the purpose of the processing, whether data are stored and the means to access and rectify data. here, techniques of (visual) indication of activation would be necessary. the data subject would have to give his consent for using and gathering information for any specific purpose. the data subject should also be informed about what type of data is gathered and whether the data will be used by the third parties. in ami, such rights may create a great burden, both on the data subject, on the responsible data controller and on all data processors. though adequate, simplified notices about the data processors' policy would be welcome (e.g., using adequate pictograms or similar means). in our opinion, such information should always be provided to consumers when rfid technology is used, even if the tag does not contain personal data in itself. the data subject should also be informed how to discard, disable or remove the tag. the right to disable the tag can relate to the consent principle of data protection, since the individual should always have the possibility to withdraw his consent. disabling the tag should at least be possible when the consent of the data subject is the sole legal basis for processing the data. disabling the tag should not lead to any discrimination of the consumer (e.g., in terms of the guarantee conditions). technological and organisational measures (e.g., the design of rfid systems) are of crucial importance in ensuring that the data protection obligations are respected (privacy by design, e.g., by technologically blocking unauthorised access to the data). thus, availability and compliance with privacy standards are of particular importance. the concept of "personal data" in the context of rfid technology is contested. wp states: in assessing whether the collection of personal data through a specific application of rfid is covered by the data protection directive, we must determine: (a) the extent to which the data processed relates to an individual, and (b) whether such data concerns an individual who is identifiable or identified. data relates to an individual if it refers to the identity, characteristics or behaviour of an individual or if such information is used to determine or influence the way in which that person is treated or evaluated. in assessing whether information concerns an identifiable person, one must apply recital of the data protection directive which establishes that "account should be taken of all the means likely reasonably to be used either by the controller or by any other person to identify the said person." and further: "finally, the use of rfid technology to track individual movements which, given the massive data aggregation and computer memory and processing capacity, are if not identified, identifiable, also triggers the application of the data protection directive." article data protection working party, working document on data protection issues related to rfid further research on the rfid technology and its privacy implications is recommended. this research should also aim at determining whether any legislative action is needed to address the specific privacy concerns of rfid technology. further development of codes of conducts and good practices is also recommended. profiling is as old as life, because it is a kind of knowledge that unconsciously or consciously supports the behaviour of living beings, humans not excluded. it might well be that the insight that humans often "intuitively know" something before they "understand" it can be explained by the role profiling spontaneously plays in our minds. thus, there is no reason to prohibit automated profiling and data mining concerning individuals with opacity rules. profiling activities should in principle be ruled by transparency tools. in other words, the processing of personal datacollection, registration and processing in the strict sense -is not prohibited but submitted to a number of conditions guaranteeing the visibility, controllability and accountability of the data controller and the participation of the data subjects. data protection rules apply to profiling techniques (at least in principle). the collection and processing of traces surrounding the individual must be considered as processing of personal data in the sense of existing data protection legislation. both individual and group profiling are dependent on such collection and on the processing of data generated by the activities of individuals. and that is precisely why, in legal terms, no profiling is thinkable outside data protection. there is an ongoing debate in contemporary legal literature about the applicability of data protection to processing practices with data that are considered anonymous, i.e., they do not allow the identification of a specific individual. some contend that data protection rules do not allow processing practices that bring together data on certain individuals without trying to identify the said individual (in terms of physical location or name). some contend that data protection rules do not apply to profiling practices that process data relating to non-identifiable persons (in the sense of the data protection directive). we hold that it is possible to interpret the european data protection rules in a broad manner covering all profiling practices, but the courts have not spoken on this yet. data protection should apply to all profiling practices. when there is confusion in the application and interpretation of the legal instruments, they should be adapted so that they do apply to all profiling practices. profiling practices and the consequent personalisation of the ambient intelligence environment lead to an accumulation of power in the hands of those who control the profiles and should therefore be made transparent. the principles of data protection are an appropriate starting point to cope with profiling in a democratic constitutional state as they do impose good practices. nevertheless, while the default position of data protection is transparency ("yes, you can process, but …"), it does not exclude opacity rules ("no, you cannot process, unless …"). in relation to profiling, two examples of such rules are relevant. on the one hand, of course, there is the explicit prohibition against taking decisions affecting individuals solely on the basis of the automated application of a profile without human intervention (see article of the data protection directive). this seems obvious because in such a situation, probabilistic knowledge is applied to a real we recall that personal data in the eu data protection directive refers to "any information relating to an identified or identifiable natural person" (article ). the role of data protection law and non-discrimination law in group profiling in the private sector." article on automated individual decisions states: " . member states shall grant the right to every person not to be subject to a decision which produces legal effects concerning him or significantly affects him and which is based solely on automated processing of data intended to evaluate certain personal aspects relating to him, such as his performance at work, creditworthiness, reliability, conduct, etc. . subject to the other articles of this directive, member states shall provide that a person may be subjected to a decision of the kind referred to in paragraph if that decision: (a) is taken in the course of the entering into or performance of a contract, provided the request for the entering into or the performance of the contract, lodged by the data subject, has been satisfied or that there are suitable measures to safeguard his legitimate interests, such as arrangements allowing him to put his point of view; or (b) is authorized by a law which also lays down measures to safeguard the data subject's legitimate interests." person. on the other hand, there is the (quintessential) purpose specification principle, which provides that the processing of personal data must meet specified, explicit and legitimate purposes. as a result, the competence to process is limited to welldefined goals, which implies that the processing of the same data for other incompatible aims is prohibited. this, of course, substantially restricts the possibility to link different processing and databases for profiling or data mining objectives. the purpose specification principle is definitely at odds with the logic of interoperability and availability of personal data: the latter would imply that all databases can be used jointly for profiling purposes. in other words, the fact that the legal regime applicable to profiling and data mining is data protection does not give a carte blanche to mine and compare personal data that were not meant to be connected. the european data protection supervisor indicated in his annual report a number of processing operations that are likely to encompass specific risks to the rights and freedoms of data subjects, even if the processing does not occur upon sensitive data. this list relates to processing operations (a) of data relating to health and to suspected offences, offences, criminal convictions or security measures, (b) intended to evaluate personal aspects relating to the data subject, including his or her ability, efficiency and conduct, (c) allowing linkages, not provided for pursuant to national or community legislation, between data processed for different purposes, and (d) for the purpose of excluding individuals from a right, benefit or contract. software can be the tool for regulating one's behaviour by simply allowing or not allowing certain acts. thus, technology constituting the "software code" can affect the architecture of the internet (and thus potentially of ami) and can provide effective means for enforcing the privacy of the individual. for example, cryptology might give many benefits: it could be used for pseudonymisation (e.g., encrypting ip addresses) and ensuring confidentiality of communication or commerce. privacy-enhancing technologies can have an important role to play, but they need an adequate legal framework. the directive on the legal protection of software obliges member states to provide appropriate remedies against a person committing any act of putting into circulation, or the possession for commercial purposes of, any means the sole intended purpose of which is to facilitate the unauthorised removal or circumvention of any technical devices which may have been applied to protect a computer program. this mechanism aims to protect programmes enforcing the intellectual property rights against circumvention. similar legal protection against circumvention of privacy-enhancing technologies could be legally foreseen. technology might go beyond what the law permits (e.g., drm prevents intellectual property infringements but at the same time might limit the rights of the lawful user). negative side effects of such technologies should be eliminated. more generally, when introducing new technology on the market, manufacturers together with relevant stakeholders should undertake a privacy impact assessment. development of a participatory impact assessment procedure would allow stakeholders to quickly identify and react to any negative features of technology (see also section . . ). the european data protection directive imposes obligations on the data controller and gives rights to the data subject. it aims to give the individual control over the collection and processing of his data. many provisions in the data protection directive have several weaknesses in an ami environment. principles of proportionality and fairness are relative and may lead to different assessments in similar situations; obtaining consent might not be feasible in the constant need for the collection and exchange of data; obtaining consent can be simply imposed by the stronger party. individuals might not be able to exercise the right to consent, right to information, access or rectification of data due to the overflow of information. thus, those rules might simply become unworkable in an ami environment. and even if workable (e.g., thanks to the help of the digital assistants), are they enough? should we not try to look for an approach granting the individual even more control? several european projects are involved in research on identity management. they focus on a decentralised approach, where a user controls how much and what kind of information he or she wants to disclose. identity management systems, while operating on a need-to-know basis, offer the user the possibility of acting under pseudonyms, under unlinkability or anonymously, if possible and desirable. among the other examples of such systems, there are projects that base their logic on the assumption that the individual has property over his data, and then could use licensing schemes when a transfer of data occurs. granting him property over the data is seen as giving him control over the information and its usage in a "distribution chain". however, it is doubtful if granting him property over the data will really empower the individual and give him a higher level of protection and control over his data. the property model also assumes that the data are disseminated under a contract. thus, the question might arise whether the data protection directive should serve as a minimum standard and thus limit the freedom of contracts. but as our dark scenarios show, there exist many cases in which the individual will not be able to freely enter into a contract. another question arises since our data are not always collected and used for commercial purposes. in most situations, the processing of personal data is a necessary condition for entering into a contractual relation (whereas the data protection directive states in article that data processing without the individual's consent to use of his personal data is legitimate when such processing is necessary for the performance of a contract). the most obvious example is the collection of data by police, social insurance and other public institutions. the individual will not always be free to give or not give his data away. the property model will not address these issues. it will also not stop the availability of the data via public means. a weakness of the property model is that it might lead to treating data only as economic assets, subject to the rules of the market. but the model's aim is different: the aim is to protect personal data, without making their processing and transfer impossible. regarding data as property also does not address the issue of the profile knowledge derived from personal data. this knowledge is still the property of the owner or the licenser of the profile. the data-as-property option also ignores the new and increasingly invisible means of data collection, such as rfids, cameras or online data collection methods. discussing the issue of whether personal data should become the individual's property does not solve the core problem. on the one hand, treating data as property may lead to a too high level of protection of personal information, which would conflict with the extensive processing needs of ami. on the other hand, it would, by default, turn personal data into a freely negotiable asset, no longer ruled by data protection, but left to market mechanisms and consent of the data subjects (more often than not to the detriment of the latter). finally, the data-as-property option loses its relevance in the light of a focus upon anonymisation and pseudonymisation of data processed in ami applications. the prime consortium proposes identity management systems controlled by data subjects. it aims to enable individuals to negotiate with service providers the disclosure of personal data according to the conditions defined. such agreement would constitute a contract. an intelligent agent could undertake the management on the user side. this solution is based on the data minimisation principle and on the current state of legislation. it proposes the enforcement of (some) current data protection and privacy laws. it seems to be designed more for the needs of the world today than for a future ami world. the user could still be forced to disclose more information than he or she wishes, because he or she is the weaker party in the negotiation; he or she needs the service. the fidis consortium has also proposed a decentralised identity management, the vision of which seems to go a bit further than the prime proposal. it foresees that the user profiles are stored on the user's device, and preferences relevant for a particular service are (temporarily) communicated to the service provider for the purpose of a single service. the communication of the profile does not have to imply disclosure of one's identity. if there is information extracted from the behaviour of the user, it is transferred by the ambient intelligent device back to the user, thus updating his profile. thus, some level of exchange of knowledge is foreseen in this model, which can be very important for the data subject's right to information. a legal framework for such sharing of knowledge from an ami-generated profile needs to be developed, as well as legal protection of the technical solution enabling such information management. such schemes rely on automated protocols for the policy negotiations. the automated schemes imply that the consent of the data subject is also organised by automatic means. we need a legal framework to deal with the situation wherein the explicit consent of the data subject for each collection of data is replaced by a "consent" given by an intelligent agent. in such automated models, one could envisage privacy policies following the data. such "sticky" policies, attached to personal data, would provide for clear information and indicate to data processors and controllers which privacy policy applies to the data concerned. sticky policies could facilitate the auditing and self-auditing of the lawfulness of the data processing by data controllers. in any event, research in this direction is desirable. since ami is also a mobile environment, there is a need to develop identity management systems addressing the special requirements of mobile networks. the fidis consortium has prepared a technical survey of mobile identity management. it has identified some special challenges and threats to privacy in the case of mobile networks and made certain recommendations: • location information and device characteristics both should be protected. • ease of use of the mobile identity management tools and simplified languages and interfaces for non-experts should be enhanced. • a verifiable link between the user and his digital identity has to be ensured. accordingly, privacy should also be protected in peer-to-peer relationships. the importance of consumer protection will grow in ambient intelligence, because of the likelihood that consumers will become more dependent on online products and services, and because product and service providers will strengthen their bargaining position through an increasing information asymmetry. without the constraints of law, ambient intelligence service providers could easily dictate the conditions of participation in new environments. consumer protection should find the proper balance in ami. consumer protection law defines the obligations of the producers and the rights of consumer and consists of a set of rules limiting the freedom to contract, for the benefit of the consumer. consumer protection law plays a role of its own, but can support the protection of privacy and data protection rights. the basis for the european framework for consumer protection rules can be found in article of the ec treaty: "in order to promote the interests of consumers and to ensure a high level of consumer protection, the community shall contribute to protecting the health, safety and economic interests of consumers, as well as to promoting their right to information, education and to organise themselves in order to safeguard their interests." consumer protection at european level is provided by (amongst others) directive / on unfair terms in consumer contracts, directive / on consumer protection in respect of distance contracts and the directive on liability for defective products (discussed below). directive / and directive / were both already discussed (in chapter , sections . . . and . . . ) . in many respects, their rules are not fitted to ami and they need to be re-adapted. this especially relates to extending the scope of protection of those directives, thereby making sure that all services and electronic means of communications and trading are covered (including those services on the world wide web not currently covered by the distance contract directive). due to the increasing complexity of online services, and due to the possibility of information overflow, it seems necessary to find legal ways to assess and recognise contracts made through the intervention of intelligent agents. is the legal system flexible enough to endorse this? moreover, the same should apply to the privacy policies and to the consent of individuals for the collection of data (because, in identity management systems, intelligent agents will decide what data are to be disclosed to whom). here is a challenge: how to technologically implement negotiability of contracts and the framework of binding law in electronic, machine-readable form? suppliers should not be allowed to set up privacy conditions which are manifestly not in compliance with the generally applicable privacy rules and which disadvantage the customer. data protection legislation and consumer protection law could constitute the minimum (or default) privacy protection level. similar rules as those currently applicable under the consumer protection of directive / on unfair terms in consumer contracts could apply. mandatory rules of consumer protection require, inter alia, that contracts be drafted in plain, intelligible language, that the consumer be given an opportunity to examine all terms, that -in cases of doubt -the interpretation most favourable to the consumer prevail. suppliers should not be allowed to unfairly limit their liability for security problems in the service they provide to the consumer. in this respect, more attention could be given to a judgment of the court of first instance of nanterre (france) in in which the online subscriber contract of aol france was declared illegal in that it contained not less than abusive clauses in its standard contractual terms (many of which infringed consumer protection law). the directive on unfair terms in consumer contracts and the directive on consumer protection in respect of distance contracts provide a broad right to information for the consumer. it should be sufficient to dispense such information in electronic form, in view of the large amount of information directed towards consumers that would have to be managed by intelligent agents. an increasing number of service providers will be involved in ami services and it cannot be feasible to provide the required information about all of them. the solution may be to provide such information only about the service provider whom the consumer directly pays and who is responsible towards the consumer. joint liability would apply (for liability issues, see below). the right to withdrawal, foreseen by the directive / on consumer protection with respect to distance contracts, may not apply (unless otherwise agreed) to contracts in which (a) the provision of services has begun with the consumer's agreement before the end of the seven-working-day period and (b) goods have been made to the consumer's specifications or clearly personalised or which, by their nature, cannot be returned or are liable to deteriorate or expire rapidly. in an ami world, services will be provided instantly and will be increasingly personalised. this implies that the right of withdrawal will become inapplicable in many cases. new solutions should be developed to address this problem. currently, insofar as it is not received on a permanent medium, consumers must also receive written notice in good time of the information necessary for proper performance of the contract. in ami, payments will often occur automatically, at the moment of ordering or even offering the service. temporary accounts, administered by trusted third parties, could temporarily store money paid by a consumer to a product or service provider. this can support consumer protection and enforcement, in particular with respect to fraud and for effectively exercising the right of withdrawal. this would be welcome for services that are offered to consumers in the european union by service providers located in third countries, as enforcement of consumer protection rights is likely to be less effective in such situations. the possibility of group consumer litigation can increase the level of law enforcement and, especially, enforcement of consumer protection law. often an individual claim does not represent an important economic value, thus, individuals are discouraged from making efforts to enforce their rights. bodies or organisations with a legitimate interest in ensuring that the collective interests of consumers are protected can institute proceedings before courts or competent administrative authorities and seek termination of any behaviour adversely affecting consumer protection and defined by law as illegal. however, as far as actions for damages are concerned, issues such as the form and availability of group litigation are regulated by the national laws of the member states as part of procedural law. the possibility to bring such a claim is restricted to a small number of states. group litigation is a broad term which captures collective claims (single claims brought on behalf of a group of identified or identifiable individuals), representative actions (single claims brought on behalf of a group of identified individuals by, e.g., a consumer interest association), class action (one party or group of parties may sue as representatives of a larger class of unidentified individuals), among others. these definitions as well as the procedural shape of such claims vary in different member states. waelbroeck d., d. slater and g. the directive on electronic commerce aims to provide a common framework for information society services in the eu member states. an important feature of the directive is that it also applies to legal persons. similar to the consumer protection legislation, the directive contains an obligation to provide certain information to customers. in view of the increasing number of service providers, it may not be feasible to provide information about all of them. providing information about the service provider whom the customer pays directly and who is responsible towards him could be a solution to the problem of the proliferating number of service providers (joint liability may also apply here). the directive should also be updated to include the possibility of concluding contracts by electronic means (including reference to intelligent agents) and to facilitate the usage of pseudonyms, trusted third parties and credentials in electronic commerce. unsolicited commercial communication is an undesirable phenomenon in cyberspace. it constitutes a large portion of traffic on the internet, using its resources (bandwidth and storage capacity) and forcing internet providers and users to adopt organisational measures to fight it (by filtering and blocking spam). spam can also constitute a security threat. the dark scenarios show that spam may become an even more serious problem than it is today. an increase in the volume of spam can be expected because of the emergence of new means of electronic communication. zero-cost models for e-mail services encourage these practices, and similar problems may be expected when mobile services pick up a zero-cost or flat-fee model. as we become increasingly dependent on electronic communication -ambient intelligence presupposes that we are almost constantly online -we become more vulnerable to spam. in the example from the first dark scenario, spamming may cause irritation and make the individual reluctant to use ambient intelligence. fighting spam may well demand even more resources than it does today as new methods of spamming -such as highly personalised and location-based advertising -emerge. currently, many legal acts throughout the world penalise unsolicited communication, but without much success. the privacy & electronic communication directive provides for an opt-in regime, applicable in the instance of commercial communication, thus inherently prohibiting unsolicited marketing. electronic communications are, however, defined as "any information exchanged or conveyed between a finite number of parties by means of a publicly available electronic communications service. this does not include any information conveyed as part of a broadcasting service to the public over an electronic communications network except to the extent that the information can be related to the identifiable subscriber or user receiving the information." the communications need to have a commercial content in order to fall under the opt-in regulation of the privacy & electronic communication directive. consequently, this directive may not cover unsolicited, location-based advertisements with a commercial content that are broadcast to a group of people ("the public"). the impact of this exception cannot be addressed yet since location-based services are still in their infancy. a broad interpretation of electronic communications is necessary (the directive is technology-neutral). considering any unsolicited electronic communication as spam, regardless of the content and regardless of the technological means, would offer protection that is adequate in ambient intelligence environments in which digital communications between people (and service providers) will exceed physical conversations and communications. civil damages address a harm already done, and compensate for damages sustained. effective civil liability rules might actually form one of the biggest incentives for all actors involved to adhere to the obligations envisaged by law. one could establish liability for breach of contract, or on the basis of general tort rules. to succeed in court, one has to prove the damage, the causal link and the fault. liability can be established for any damages sustained, as far as the conditions of liability are proven and so long as liability is not excluded (as in the case of some situations in which intermediary service providers are involved ). however, in ami, to establish such proof can be extremely difficult. as we have seen in the dark scenarios, each action is very complex, with a multiplicity of actors involved, and intelligent agents acting for service providers often undertake the action or decision causing the damage. who is then to blame? how easy will it be to establish causation in a case where the system itself generates the information and undertakes the actions? how will the individual deal with such problems? the individual who is able to obtain damages addressing his harm in an efficient and quick way will have the incentive to take an action against the infringer, thus raising the level of overall enforcement of the law. such an effect would be desirable, especially since no state or any enforcement agency is actually capable of providing a sufficient level of control and/or enforcement of the legal rules. the liability provisions of the directive on electronic commerce can become problematic. the scope of the liability exceptions under the directive is not clear. the directive requires isps to take down the content if they obtain knowledge on the infringing character of the content (notice-and-take-down procedure). however, the lack of a "put-back" procedure (allowing content providers whose content has been wrongfully alleged as illegal, to re-publish it on the internet) or the verification of take-down notices by third parties is said to possibly infringe freedom of speech. it is recommended that the liability rules be strengthened and that consideration be given to means that can facilitate their effectiveness. the directive provides for exceptions to the liability of intermediary service providers (isps) under certain conditions. in the case of hosting, for example, a service provider is not liable for the information stored at the request of a recipient of the service, on condition that (a) the provider does not have actual knowledge of illegal activity or information and, as regards claims for damages, is not aware of facts or circumstances from which the illegal activity or information is apparent or (b) the provider, upon obtaining such knowledge or awareness, acts expeditiously to remove or to disable access to the information. see also section . . . in addition to the general considerations regarding liability presented in this section, we also draw attention to the specific problems of liability for infringement of privacy, including security infringements. currently, the right to remedy in such circumstances is based on the general liability (tort) rules. the data protection directive refers explicitly to liability issues stating that an immediate compensation mechanism shall be developed in case of liability for an automated decision based on inadequate profiles and refusal of access. however, it is not clear whether it could be understood as a departure from general rules and a strengthening of the liability regime. determining the scope of liability for privacy breach and security infringements might also be problematic. in any case, the proof of the elements of a claim and meeting the general tort law preconditions (damage, causality and fault) can be very difficult. opacity instruments, as discussed above, aiming to prohibit the interference into one's privacy can help to provide some clarity as to the scope of the liability. in addition, guidelines and interpretations on liability would be generally welcome, as would standards for safety measures, to provide for greater clarity and thus greater legal certainty for both users and undertakings. as already mentioned, it can be difficult for a user to identify the party actually responsible for damages, especially if he or she does not know which parties were actually involved in the service and/or software creation and delivery. the user should be able to request compensation from the service provider with whom he or she had direct contact in the process of the service. joint and several liability (with the right to redress) should be the default rule in the case of providers of ami services, software, hardware or other products. the complexity of the actions and multiplicity of actors justify such a position. moreover, this recommendation should be supplemented by the consumer protection recommendation requiring the provision of consumer information by the service or product provider having the closest connection with the consumer, as well as the provision of information about individual privacy rights (see above) in a way that would enable the individual to detect a privacy infringement and have a better chance to prove it in court. there is a need to consider the liability regime with other provisions of law. liability regime. in fact, the answer depends on national laws and how the directive has been implemented. the directive applies to products defined as movables, which might suggest that it refers to tangible goods. software not incorporated into a tangible medium (available online) will not satisfy such a definition. there are a growing number of devices (products) with embedded software (e.g., washing machines, microwaves, possibly rfids), which fall under the directive's regime. this trend will continue; the software will be increasingly crucial for the proper functioning of the products themselves, services and whole environments (smart car, smart home). should the distinction between the two regimes remain? strict liability is limited to death or personal injury, or damage to property intended for private use. the damage relating to the product itself, to the product used in the course of business and the economic loss will not be remedied under the directive. currently, defective software is most likely to cause financial loss only, thus the injured party would not be able to rely on provisions of the directive in seeking redress. however, even now in some life-saving applications, personal injury dangers can emerge. such will also be the case in the ami world (see, e.g., the first and second dark scenarios in which software failures cause accidents, property damage and personal injury) so the importance and applicability of the directive on liability for defective products will grow. the increasing dependence on software applications in everyday life, the increasing danger of sustaining personal injury due to a software failure and, thus, the growing concerns of consumers justify strengthening the software liability regime. however, the directive allows for a state-of-the-art defence. under this defence, a producer is not liable if the state of scientific and technical knowledge at the time the product was put into circulation was not such that the existence of the defect would be discovered. it has been argued that the availability of such a defence (member states have the discretion whether to retain it in national laws ) will always be possible since, due to the complexity of "code", software will never be defect-free. these policy and legal arguments indicate the difficulty in broadening the scope of the directive on liability for defective products to include software. reversal of the burden of proof might be a more adequate alternative solution, one that policymakers should investigate. it is often difficult to distinguish software from hardware because both are necessary and interdependent to provide a certain functionality. similarly, it may be difficult to draw the line between software and services. transfer of information via electronic signals (e.g., downloaded software) could be regarded as a service. some courts might also be willing to distinguish between mass-market software and software produced as an individual product (on demand). ami is a highly personalised environment where software-based services will surround the individual, thus the tendency to regard software as a service could increase. strict liability currently does not apply to services. service liability is regulated by national laws. extending such provision to services may have far-reaching consequences, not only in the ict field. the ami environment will need the innovation and creativity of service providers; therefore, one should refrain from creating a framework discouraging them from taking risks. however, some procedural rules could help consumers without upsetting an equitable balance. the consumer, usually the weaker party in a conflict with the provider, often has difficulty proving damages. reversing the burden of proof might facilitate such proof. most national laws seem to provide a similar solution. since national law regulates the issue of service liability, differences between national regulations might lead to differences in the level of protection. the lack of a coherent legal framework for service liability in europe is regrettable. learning from the differences and similarities between the different national legal regimes, as indicated in the analysis of national liability systems for remedying damage caused by defective consumer services, is the first step in remedying such a situation. reversing the burden of proof is less invasive than the strict liability rules where the issue of fault is simply not taken into consideration. such a solution has been adopted in the field of the non-discrimination and intellectual property laws, as well as in national tort regimes. an exception to the general liability regime is also provided in directive / /ec on the community framework for electronic signatures. in that directive, the certification service provider is liable for damage caused by non-compliance with obligations imposed by the directive unless he proves he did not act negligently. technology could potentially remedy the information asymmetry between users and ami service suppliers or data processors. the latter could have an obligation to inform consumers what data are processed, how and when and what is the aim of such activities (thus actually fulfilling their obligations under the data protection directive). this information could be stored and managed by an intelligent agent on behalf of the user, who is not able to deal with such information flow. however, the user would have the possibility to use such information to enforce his rights (e.g., to prove causation). other technological solutions (e.g., watermarking) could also help the user prove his case in court. in many cases, the damage sustained by the individual will be difficult to assess in terms of the economic value or too small to actually provide an incentive to bring an action to court. however, acts causing such damage can have overall negative effects. spam is a good example. fixed damages, similar to the ones used in the united states, or punitive damages could remedy such problems (some us state laws provide for fixed damages such as us$ for each unsolicited communication without the victim needing to prove such damage). they would also provide clarity as to the sanctions or damages expected and could possibly have a deterrent effect. the national laws of each member state currently regulate availability of punitive damages; a few countries provide for punitive and exemplary damages in their tort systems. the universal service directive provides for a minimum of telecommunication services for all at an affordable price as determined by each member state. prices for universal services may depart from those resulting from market conditions. such provisions aim at overcoming a digital divide and allowing all to enjoy a certain minimum of electronic services. the directive is definitely a good start in shaping the information society and the ami environment. the development of new technologies and services generates costs, both on individuals and society. many high-added-value ami services will be designed for people who will be able to pay for them. thus, ami could reinforce the inequalities between the poor and rich. everyone should be able to enjoy the benefits of ami, at least at a minimum level. the commission should consider whether new emerging ami services should be provided to all. some services (e.g., emergency services) could even be regarded as public and provided free of charge or as part of social security schemes. as shown in scenario , ami might cause major problems for current intellectual property protection, because ami requires interoperability of devices, software, data and information, for example, for crucial information systems such as health monitoring systems used by travelling seniors. there is also a growing need to create means of intellectual property protection that respect privacy and allow for anonymous content viewing. intellectual property rights give exclusive rights over databases consisting of personal data and profiles, while the data subjects do not have a property right over their own information collected. we discuss these issues below. the directive on the legal protection of databases provides for a copyright protection of databases, if they constitute the author's own intellectual creation by virtue of his selection or arrangement of their content. the directive also foresees a sui generis protection if there has been a qualitatively and/or quantitatively substantial investment in either the acquisition, verification or presentation of the content. sui generis protection "prevents the extraction and/or the re-utilization of the whole or of a substantial part, evaluated qualitatively and/or quantitatively, of the contents of that database". this implies that the database maker can obtain a sui generis protection of a database even when its content consists of personal data. although the user does not have a property right over his personal data, the maker of a database can obtain an exclusive right over this type of data. hence, a profile built on the personal data of a data subject might constitute somebody else's intellectual property. the right to information about what knowledge has been derived from one's data could, to some extent, provide a safeguard against profiling. we recommend that further research be undertaken on how to reconcile this with intellectual property rights. the copyright directive provides for the protection of digital rights management (drms) used to manage the licence rights of works that are accessed after identification or authentication of a user. but drms can violate privacy, because they can be used for processing of personal data and constructing (group) profiles, which might conflict with data protection law. less invasive ways of reconciling intellectual property rights with privacy should be considered. this not only relates to technologies but also to an estimation of the factual economic position of the customer. for example, the general terms and conditions for subscribing to an interactive television service -often a service offered by just a few players -should not impose on customers a condition that personal data relating to their viewing behaviour can be processed and used for direct marketing or for transfer to "affiliated" third parties. as the article working party advises, greater attention should be devoted to the use of pets within drm systems. in particular, it advises that tools be used to see also section . . , specific recommendation regarding security. article data protection working party, working document on data protection issues related to intellectual property rights (wp/ ), adopted on january . http://ec.europa.eu/ justice_home/fsj/privacy preserve the anonymity of users and it recommends the limited use of unique identifiers. use of unique identifiers allows profiling and tagging of a document linked to an individual, enabling tracking for copyright abuses. such tagging should not be used unless necessary for performance of the service or unless with the informed consent of individual. all relevant information required under data protection legislation should be provided to users, including categories of collected information, the purpose of collecting and information about the rights of the data subject. the directive on the legal protection of software obliges member states to provide appropriate remedies against a person's committing any act of putting into circulation or the possession for commercial purposes of any means the sole intended purpose of which is to facilitate the unauthorised removal or circumvention of any technical device which may have been applied to protect a computer program. the software directive only protects against the putting into circulation of such devices and not against the act of circumventing as such. it would be advisable to have a uniform solution in that respect. drms can also violate consumer rights, by preventing the lawful enjoyment of the purchased product. the anti-circumvention provisions should be then coupled with better enforcement of consumer protection provisions regarding information disclosure to the consumer. the consumer should always be aware of any technological measures used to protect the content he wishes to purchase, and restrictions in use of such content as a consequence of technological protection (he should also be informed about technological consequences of drms for his devices, if any, e.g., about installing the software on his computer). product warnings and consumer notifications should always be in place and should aim to raise general consumer awareness about the drms. as interoperability is a precondition for ami, ami would have to lead to limitations on exclusive intellectual property rights. one could argue that software packages should be developed so that they are interoperable with each other. those restrictions might, inter alia, prevent the user from making backups or private copies, downloading music to portable devices, playing music on certain devices, or constitute the geographical restrictions such as regional coding of dvds. privacy protection. a broader scope of the decompilation right under software protection would be desirable. the ec's battle with microsoft was in part an attempt to strengthen the decompilation right with the support of competition law. currently, there is no international or european framework determining jurisdiction in criminal matters, thus, national rules are applicable. the main characteristics of the legal provisions in this matter have already been discussed in chapter , section . . . ; however, it seems useful to refer here to some of our earlier conclusions. the analysis of the connecting factors for forum selection (where a case is to be heard) shows that it is almost always possible for a judge to declare himself competent to hear a case. certain guidelines have already been developed, both in the context of the cybercrime convention as well as the eu framework decision on attacks against information systems on how to resolve the issue of concurrent competences. according to the cybercrime convention, "the parties involved shall, where appropriate, consult with a view to determining the most appropriate jurisdiction for prosecution." the eu framework decision on attacks against information systems states, "where an offence falls within the jurisdiction of more than one member state and when any of the states concerned can validly prosecute on the basis of the same facts, the member states concerned shall co-operate in order to decide which of them will prosecute the offenders with the aim, if possible, of centralizing proceedings in a single member state." legal experts and academics should follow any future developments in application of those rules that might indicate whether more straightforward rules are needed. the discussion on the green paper on double jeopardy should also be closely followed. scenario ("a crash in ami space") turns on an accident involving german tourists in italy, while travelling with a tourist company established in a third country. it raises questions about how ami might fit into a legal framework based on territorial concepts. clear rules determining the law applicable between the parties are an important guarantee of legal certainty. private international law issues are dealt at the european level by the rome convention on the law applicable to contractual obligations as well as the rome ii regulation on the law applicable to non-contractual obligations, the brussels regulation on jurisdiction and enforcement of judgments. the regulation on jurisdiction and enforcement of judgments in civil and commercial matters covers both contractual and non-contractual matters. it also contains specific provisions for jurisdiction over consumer contracts, which aim to protect the consumer in case of court disputes. these provisions should be satisfactory and workable in an ami environment. however, provisions of this regulation will not determine the forum if the defendant is domiciled outside the european union. also, the provisions on the jurisdiction for consumer contracts apply only when both parties are domiciled in eu member states. although the regulation provides for a forum if the dispute arises from an operation of a branch, agency or other establishment of the defendant in a member state, a substantial number of businesses offering services to eu consumers will still be outside the reach of this regulation. this emphasises again the need for a more global approach beyond the territory of the member states. clarification and simplification of forum selection for non-consumers would also be desirable. the complexity of the business environment, service and product creation and delivery would justify such approach. it would be of special importance for smes. currently, the applicable law for contractual obligations is determined by the rome convention. efforts have been undertaken to modernise the rome convention and replace it with a community instrument. recently, the commission has presented a proposal for a regulation of the european parliament and the council on the law applicable to contractual obligations. the provisions of the rome convention refer to contractual issues only. recently, the so-called rome ii regulation has been adopted, which provides for rules applicable to non-contractual obligations. the rome convention on law applicable to contractual obligations relies heavily on the territorial criterion. it refers to the habitual residence, the central administration or place of business as the key factors determining the national law most relevant to the case. but it services can be supplied at a distance by electronic means. the ami service supplier could have his habitual residence or central administration anywhere in the world and he could choose his place of residence (central administration) according to how beneficial is the national law of a given country. the habitual residence factor has been kept and strengthened in the commission's proposal for a new regulation replacing the rome convention (rome i proposal, article ). the new proposal for the rome i regulation amends the consumer protection provisions. it still relies on the habitual residence of the consumer, but it brings the consumer choice of contract law in line with the equivalent provisions of the brussels regulation, and broadens the scope of the application of its provisions. the commission proposal for the regulation on the law applicable to contractual obligations is a good step forward. the rome ii regulation on law applicable to non-contractual obligations applies to the tort or delict, including claims arising out of strict liability. the basic rule under the regulation is that a law applicable should be determined on the basis of where the direct damage occurred (lex loci damni). however, some "escape clauses" are foreseen and provide for a more adequate solution if more appropriate in the case at hand. this allows for flexibility in choosing the best solution. special rules are also foreseen in the case of some specific torts or delicts. uniform rules on applicable law at the european level are an important factor for improving the predictability of litigation, and thus legal certainty. in that respect, the new regulation should be welcomed. the regulation will apply from january . some other legislative acts also contain rules on applicable law. most important are provisions in the data protection directive. this directive also chooses the territorial criterion to determine the national law applicable to the processing of data, which is the law of the place where the processing is carried out in the context of an establishment of the data controller. such a criterion, however, might be problematic: more than one national law might be applicable. moreover, in times of globalisation of economic activity, it is easy for an undertaking to choose the place of establishment, which offers the most liberal regime, beyond the reach of european data protection law. in situations when a non-eu state is involved, the directive points out a different relevant factor, the location of the equipment used, thus enabling broader application of the eu data protection directive. as recital of the proposal states, these amendments aim to take into account the developments in distance selling, thus including ict developments. article ( ) of the directive stipulates: each member state shall apply the national provisions it adopts pursuant to this directive to the processing of personal data where: (a) the processing is carried out in the context of the activities of an establishment of the controller on the territory of the member state; when the same controller is established on the territory of several member states, he must take the necessary measures to ensure that each of these establishments complies with the obligations laid down by the national law applicable. the directive stipulates in article ( ) that the national law of a given member state will apply when the controller is not established on community territory and, for purposes of processing personal data, makes use of equipment, automated or otherwise, situated on the territory of the said member state, unless such equipment is used only for purposes of transit through the territory of the community. the article data protection working party interprets the term "equipment" as referring to all kinds of tools or devices, including personal computers, which can be used for many kinds of processing operations. the definition could be extended to all devices with a capacity to collect data, including sensors, implants and maybe rfids. (active rfid chips can also collect information. they are expensive compared to passive rfid chips but they are already part of the real world.) see article data protection working party, working document on determining the international application of eu data protection law to personal data processing on the internet by non-eu based websites ( / /en/final wp ), may . http://ec.europa.eu/justice_home/ fsj/privacy/ docs/wpdocs/ /wp _en.pdf as we see, in all these cases, the territorial criterion (establishment) prevails. we should consider moving towards a more personal criterion, especially since personal data are linked with an identity and a state of the data subject (issues which are regulated by the national law of the person). such a criterion could be more easily reconciled with the ami world of high mobility and without physical borders. the data subject will also be able to remain under the protection of his/her national law, and the data controller/service provider will not have the possibility of selecting a place of establishment granting him the most liberal treatment of law. data transfer is another issue highlighting the need for international co-operation in the creation of a common playing field for ami at the global level. what is the sense of protecting data in one country if they are transferred to a country not affording comparable (or any) safeguards? also, the globalisation of economic and other activities brings the necessity of exchanging personal data between the countries. the data protection directive provides a set of rules on data transfer to third countries. data can be transferred only to countries offering an adequate level of protection. the commission can conclude agreements (e.g., the safe harbour agreement) with third countries which could help ensure an adequate level of protection. the commission can also issue a decision in that respect. however, the major problem is enforcement of such rules, especially in view of the fact that some "safeguards" rely on self-regulatory systems whereby companies merely promise not to violate their declared privacy policies (as is the case with the safe harbour agreement). attention by the media and consumer organisations can help in the enforcement of agreed rules. the problem of weak enforcement also emphasises the need to strengthen international co-operation with the aim of developing new enforcement mechanisms. providing assistance in good practices in countries with less experience than the european union would also be useful. such a solution has the advantage of covering, with the protection of eu legislation, third country residents whose data are processed via equipment in the eu. a broad interpretation of the term "equipment" would help guarantee the relatively broad application of such a rule (see above). as a result, in most cases, application of the domicile/nationality rule or the place of the equipment used as the relevant factor would have the same result. however, we can envisage the processing of data not using such equipment, for example, when the data are already posted online. then the eu law could not be applicable. see chapter , section . . . . biometrics: legal issues and implications, background paper for the institute of prospective technological studies report on the protection of personal data with regard to the use of smart cards biometrics at the frontiers: assessing the impact on society, study commissioned by the libe committee of the european parliament comments on the communication of the commission on interoperability of european databases article data protection working party, working document on biometrics descriptive analysis and inventory of profiling practices, fidis (future of identity in the information society), deliverable d rfid and the perception of control: the consumer's view article data protection working party, working document on data protection issues related to rfid technology ( / /en -wp ) though these are good examples of the involvement of stakeholders in the discussion, the results are not fully satisfactory. as a compromise between the different actors, the guidelines do not go far enough in protecting the interests of consumers huge practical difficulties of effectively enforcing and implementing data protection, more particularly in the field of profiling structured overview on prototypes and concepts of identity management systems code': privacy's death or saviour? the weaker party in the contract is now protected by the general principles of law privacy and identity management for europe report on actual and possible profiling techniques in the field of ambient intelligence, fidis (future of identity in the information society) deliverable d . ami -the european perspective on data protection legislation and privacy policies privacy and human rights , produced by the electronic privacy information center article (d) of directive / /ec safeguards should be provided for subscribers against intrusion of their privacy by unsolicited communications for direct marketing purposes in particular by means of automated calling machines, telefaxes, and e-mails, including sms messages cogitas, ergo sum. the role of data protection law and non-discrimination law in group profiling in the private sector spam en electronische reclame a strict product liability regime based on the directive is the basis of the claims under the general tort regime. see giensen, i the precautionary principle in the information society, effects of pervasive computing on health and environment in such a case, the intelligent software agent's failure and the pet's failure might be covered by the strict liability regime the applicability of the eu product liability directive to software computer software and information licensing in emerging markets, the need for a viable legal framework the contract intended to treat it as such (as opposed to an information service). see singsangob a., computer software and information licensing in emerging markets, the need for a viable legal framework article of the directive liability article of the directive on liability for defective products liability for defective products and services: the netherlands the oecd has treated software downloads as a service for the vat and custom duties purposes the distance selling directive: points for further revision comparative analysis of national liability systems for remedying damage caused by defective consumer services: a study commissioned by the european commission (the scenario script) and section . . . on the legal analysis of private international law aspects of the scenario regulation (ec) no / of the european parliament and of the council of on jurisdiction and the recognition and enforcement of judgments in civil and commercial matters if the defendant is not domiciled in a member state, the jurisdiction of the courts of each member state shall, subject to articles and , be determined by the law of that member state; . any person domiciled in a member state may, whatever his nationality, avail himself of the rules of jurisdiction in force in that state, and in particular those specified in annex i, in the same way as the nationals of that state. by a translation of existing powers and structures the commission has presented the proposal for a regulation of the european parliament and the council on the law applicable to contractual obligations (rome i), com ( ) final it shall be presumed that the contract is most closely connected with the country where the party who is to effect the performance which is characteristic of the contract has, at the time of conclusion of the contract, his habitual residence, or, in the case of a body corporate or unincorporated, its central administration. however, if the contract is entered into in the course of that party's trade or profession the new proposal does not use the presumption that the country of habitual residence is the most closely connected with the case, as it is under the rome convention. in the proposal, the relevant factor of the habitual residence of the directive on liability for defective products provides for a liability without fault (strict liability). as a recital to the directive states, strict liability shall be seen as "the sole means of adequately solving the problem, peculiar to our age of increasing technicality, of a fair apportionment of the risks inherent in modern technological production." we should keep this reasoning in mind since it seems even more adequate when thinking about the liability issues in ami.most of the "products" offered in the ami environment will consist of softwarebased, highly personalised services. we should then think about adjusting the liability rules to such an environment. if it is difficult to distinguish between hardware and software from a technological perspective, why should we draw such a distinction from a legal perspective? an explicit provision providing for strict liability for software can be considered. nevertheless, such a proposal is controversial as it is said to threaten industry. since software is never defect-free, strict liability would expose software producers unfairly to claims against damages. thus, the degree of required safety of the programmes is a policy decision. strict liability could also impede innovation, especially the innovation of experimental and lifesaving applications. others argue that strict liability might increase software quality by making producers more diligent, especially, in properly testing their products. despite these policy considerations, there are some legal questions about the applicability of strict liability to software. the first question is whether the software can be regarded as "goods" or "products" and whether they fall under the strict actions allowing consolidation of the small claims of individuals could be also examined (i.e., group consumer actions). non-discrimination law can regulate and forbid the unlawful usage of processed data, for example, in making decisions or undertaking other actions on the basis of certain characteristics of the data subjects. this makes non-discrimination law of increasing importance for ami. the creation of profiles does not fall under non-discrimination law (potential use), but decisions based on profiling (including group profiling based on anonymous data) that affect the individual might provide the grounds for application of the non-discrimination rules. they apply in the case of identifiable individuals as well as to anonymous members of the group. profiles or decisions based on certain criteria (health data, nationality, income, etc.) may lead to discrimination against individuals. it is difficult to determine when it is objectively justified to use such data and criteria, and when they are discriminatory (e.g., the processing of health-related data by insurance companies leading to decisions to raise premiums). further legislative clarity would be desirable.however, certain negative dimensions of profiling still escape the regime of non-discrimination law (e.g., manipulation of individuals' behaviour by targeted advertising). here no remedies have been identified.the non-discrimination rules should be read in conjunction with the fairness principle of data protection law. the application of the two may have similar aims and effects; they might also be complementary: can the limitations of non-discrimination law be justified if they are regarded as not fair, as in the example of insurance companies raising premiums after processing health data? they can address a range of actions undertaken in ami, such as dynamic pricing or refusal to provide services (e.g., a refusal of service on the grounds that no information (profile) is available could be regarded as discriminatory).non-discrimination rules should be taken into consideration at the design stage of technology and service development. however, such issues might be addressed by the data protection legislation. in the opinion of gutwirth and de hert, principles of data protection are appropriate to cope with profiling. key: cord- -znfo xgv authors: picek, oliver title: spillover effects from next generation eu date: - - journal: inter econ doi: . /s - - -z sha: doc_id: cord_uid: znfo xgv in july , the european commission announced its € billion package to revive the postpandemic european economy, next generation eu. the programme comprises a number of loans and grants that will be funded by taking out european debt. although the rules on liability sharing for next generation eu prevent a significant mutualisation of the debt, european leaders have taken the long-recognised significant first step towards european financial and political unification that stands in stark contrast to the misguided austerity programmes during the european sovereign debt crisis. next generation eu is the european commission's € billion package to revive the european economy after covid- that is funded by taking out european debt. in haggling over payments from the european union, member states like to count and negotiate their net payments -the difference between what is paid into common budgets and received from them. this chauvinistic view leaves out the additional induced growth effects triggered by the package both at home (domestic multipliers) and abroad (spillover effects to other eu countries). by capturing a share of economic output from southern and eastern european member states that receive more grants, the economies of northern and western europe grow by more than the respective portion of their contributions would suggest. the coordinated fi scal impulse ensures that every country receives a sizeable boost in economic output. the european commission's next generation eu package includes € billion. although rightly criticised as not large enough to address the gravity of the economic consequences of the virus, the instrument is nevertheless the fi rst time in recent history that the eu will directly issue a signifi cant form of mutual debt to redistribute and stabilise the economy. in order to fi nance the package, the european commission will take out loans during the budget period - -in addition to the regular eu budget -and repay them with member states' contributions to low interest payments (from onwards) and sizeable principal amounts (not before the next budget period starting in , and no later than ). some commentators have even called this europe's "hamiltonian moment" reminiscent of the federalisation of american states' debt in the th century. the largest and most economically signifi cant part of the next generation eu package (€ . billion) will be paid out as grants to member states by the new european recovery and resilience facility (rrf), a post-covid- eu reconstruction programme. a smaller portion of the grants (€ . billion) will be allocated and implemented through existing eu programmes outside of the new rrf, as shown in figure . around € billion in loans are foreseen, which will yield useful interest rate savings on loans for member states that refi nance themselves above the predicted eu bond interest rates. a small sum of guarantees completes the package. a time horizon of the payments can be found in figure . the largest portion of the actual payments will presumably be disbursed too late to fi ght the acute crisis, as around threequarters of the rrf payouts become effective in or later (see figure ). signed commitments of future payments in the rrf as well as the other grants (non-rrf, e.g. react-eu) are envisaged to take place from onwards in order to frontload as much as possible. the fi nal form of next generation eu, however, is not yet enacted into law. the council will have to get approval for its proposal from the european parliament. while the total sum of the package and the grants and loans shares are unlikely to change much, the shares for individual facilities and programmes for common european goals may increase. in any case, the plan foresees loans provided by the european union only for southern and eastern eu countries, as others are assumed to be able to fi nance themselves at a lower or equal rate than european institutions would receive in fi nancial markets. all countries receive grants fi nanced by a loan breakdown of next generation eu (€ billion) - , at prices sources: european commission ( a , darvas ( d) , author's representation. that is supposed to be repaid starting in and ending in . as a result, the distribution of the funds cannot be considered a zero-sum game because economic stimuli can boost growth and employment for all member states while repayment occurs at a much later stage and over a longer time period. media, policy analysts, and informally the european commission have provided a quite detailed preview of the distribution of the funds by country for the original proposal from may (e.g. kafsack, ; darvas a,b,c) , which was adjusted by the european council ( ) on july. using a set of assumptions on the eventual allocation rules per country by the council, darvas ( d) has divided up the grants by amounts for individual member states, but not updated the loan amounts per country. the precise suggested distribution of the grant amounts is shown in figure . loans are shown in a brighter shade above the darker grant amounts in percent of local gdp. loans have been criticised for not being an effective way to add expenditure to the economy as they merely refi nance national expenditure that would have taken place anywaygiven the accommodating reaction of the european central initially, the data by country was restricted to internal european commission and member state documents, but leaked to the frankfurter allgemeine zeitung (kafsack, ) and later recalculated by darvas ( a,b,c) . it is presented here in percent of expected gdp for (ameco, june ). the amounts are converted from constant prices to constant prices. for lack of a better estimate, their value is calculated indirectly by taking the total values per country of the original commission may proposal and deducting the grant amounts of the council july proposal. bank to the crisis (darvas, c) . in any case, the expected effect of loans and guarantees is unclear and diffi cult to estimate at this point. for the purpose of this paper, we therefore adopt the view that only grants will provide additional future expenditures that increase demand in the crisis-struck economies of the block. after deducting loans and guarantees from the total package sum of € billion, approximately € . billion in grants remain. for the european union as a whole, . % of (the crisis-reduced) expected gdp in is disbursed via grants and . % via loans over a six-year period. the plan provides for a clear division of countries into those that have been hit hard by the crisis and are economically weaker (southern and eastern europe), and those that can easily afford the costs of the crisis due to the unequal fl ow of capital within the single market, as their safe haven status allows them to draw in funds from other eu and euro area countries (northern and western europe). the bulk of spending in grants occurs only after . formulae for splitting the funds among countries include, among others, gdp per capita, the economic impact of covid- (fall in gdp in ) and unemployment rates. they differ depending on the source of funds and concrete programmes. although a fi nal agreement on the plan as well as some details are still missing, it is a huge political step towards sensible and joint european economic policymaking. the longrecognised need for a common fi scal response to economic crises had been acknowledged in academic and policymaking circles for a long time and became mainstream after the austerity disaster starting in . this more than warrants a fi rst analysis despite the need for a few crude assumptions. the implicit geographical distribution as well as the eventual share of grants, loans and guarantees is still subject to an agreement in the european council and the european parliament in autumn . sources: kafsack ( ), darvas ( b), author's calculations. in particular, we focus on two aspects: first, what is the increase in gdp per country due to the plan? second, how do economic structures of member states allow them to take part in the positive effects of the plan through spillover effects from others? a simple static multi-regional input-output model is employed. the analysis follows picek and schröder ( , ) , who use the world-input output database (wiod) to calculate spillover effects with an impact analysis framework (timmer et al., ; miller and blair, ) . the cumulative future grant receipts of member states are used as a positive expenditure shock to estimate the cumulative increase of gdp. to this end, it is assumed that one-third of the eu funds will be used as government expenditure and two-thirds as investment expenditure (public and private). this is an arbitrary, yet inevitable assumption because the distribution of funds per demand category and economic sector is currently unknown. sector allocations may be at best inferred for some parts of individual smaller programmes, but an accurate global allocation is impossible at this stage. therefore, a second crucial assumption is made by allowing the shock to increase spending proportionally according to the current industry structure -in other words, we split each additional euro of spending (as part of the initial shock) along the industry lines that prevail according to the latest industrial (inputoutput) structure of the economy. the third assumption is that grants and loans have an additional effect in the sense that the expenditure would otherwise not have taken place and does not crowd out other expenditure. fourth, there is uncertainty around the precise amount that will be spent. implicitly, a % take-up rate of potential next generation eu funds is assumed. this may not be the case if the absorption rate of eu structural and cohesion funds provides any indication -the median among the member states lingers around % (see figure in ionescu and dietrich, ) . also, we assume that we know the precise amount that € billion (in constant prices) will be able to buy in the future -in other words, we assume a certain trajectory for the infl ation rate and a fi xed disbursement schedule over time. naturally, these assumptions introduce an unavoidable amount of uncertainty that may make the input to the simulation (the shock) slightly imprecise (matthews, ) . however, the qualitative results should hold provided an absence of large cross-country differences among the factors in the assumptions. three models are calculated with different multipliers to distinguish between the fi rst-round effects arising directly from the shock and induced effects that result indirectly from increased economic activity. the direct effect of the initial expenditures plus the required intermediate input increases are depicted in the baseline model only. a keynesian consumption multiplier based on wage income is added in the second model to arrive at an estimation of the economic effects of induced consumption. finally, a channel that links (induced) investment spending to profi ts is added. spillover effects across borders occur in all three models. according to the rules of the multiannual financial framework, a % infl ation rate for each year must be assumed to convert from constant to current prices. with the european central bank unable to bring infl ation to %, however, the real value of the planned spending in current prices could be higher when infl ation remains below the benchmark. furthermore, delays or unplanned frontloading of the disbursements of funds also infl uence their real value. . . . . . . . . . . . . . . . . . . . . . . . . . naturally, a note of caution is required at this point. model uncertainty is always present and arises in input-output models mainly due to the linearity and simplicity of the model. in order to make useful predictions, a crucial prerequisite for input-output models is that the economy must not be constrained by the ability of fi rms to supply goods and services at the prevailing prices. therefore, the results hold less if disrupted supply lines are present in and later. on the one hand, several typical major supply restrictions such as high energy prices, hawkish central bank interest policy and shortage of labour do not constrain the economy during the current recession. on the other hand, impaired equity values in many fi rms may lead to a lower propensity to invest than in normal times. more important, however, is the fact that the aftermath of covid- or a second outbreak thereof may signifi cantly alter the observed consumption and investment behaviour based on historic data embedded in the model -both at the aggregate and sectoral levels. when propensities to consume and invest falter in and fail to recover entirely in and subsequent years, the multiplier implied in the model does not hold up. the growth and employment effect of the programme could then be muted to a certain extent. readers who are inclined to believe that the economic aftermath of covid- will leave permanent marks could therefore primarily look at the results of the model with direct effects (and intermediate inputs) only, and if they do not want to rely on historic (but uncertain) consumption and investment -multipliers for the next years. readers who believe investment will be impaired for a longer time due to the balance sheet problems of many fi rms, but consumption will not be (thanks to e.g. short-term work schemes), should focus on the results of the direct plus consumption-induced model. the total values for each country from the third model including direct, consumption-induced and investment-induced effects probably tell the story of an optimistic future and strong investment dynamics within a powerful rebound and upswing after the coronavirus has been safely contained. a note on the interpretation of the results is in order. the initial stimulus takes the form of a grant payment that takes place once between and and is not renewed or repeated. the following figures therefore show the cumulative effect of a cumulative shock that is nonetheless a onetime event. for each country, the results are in percent of their respective gdp in . however, it does not mean that real gdp will be higher by this amount at the end of a specifi c year compared to dsge or more modern macro models, however, overfi tting of parameters is impossible, leaving the researcher less choice regarding the parameter input values and the subsequent results. for a discussion with more details, see picek and schröder ( ). or at the end of the programme period. instead, it means that over the - period, the differences between a higher gdp (because of next generation eu) and the baseline gdp add up to the amounts shown in the graph. to receive precise yearly numbers, one would have to divide the results by the number of years under consideration weighted by a presumed disbursement schedule. the maximum effect of the programme can be expected in and , when over % of the resilience and recovery facility grants, the main part of next generation eu, are scheduled to be paid out. for simplicity, a division of the cumulative sum by seven (number of years from to ) can provide a quick estimate of the average yearly effect (in terms of gdp in ). another, hypothetical, way to think about the results is the following: if all planned additional expenditures were theoretically spent in (in a non-covid- economy), by how much would gdp increase in ? naturally, as the one-time impulse is withdrawn in and later years, this opens up room for a subsequent (partial) fall in economic activity unless the additional income stream in has laid roots for a self-contained private sector upswing in the years following the impulse. the main result of the present paper is shown in figure (cumulative) and figure (yearly average). the cumulative increase in real gdp over the period - as a result of the additional investment and government spending is presented for all eu countries in figure . during the programme period, additional greek gdp through next generation eu accumulates to . % of gdp in the best case. and dividing over seven years, this amounts to an average yearly increase of . % (in terms of constant prices as shown in figure ). in the worst case scenario (only direct effects), without signifi cant consumption and investment multipliers, a meagre cumulative increase of . % of expected greek gdp is recorded -or . % per year. roughly, but not precisely in line with diminishing domestic grant expenditure, the economic effect decreases successively, iterating through each southern european country and the central and eastern european accession countries. the south-eastern block is followed by france and malta, before germany and austria begin to lead the geographic and political north-west fl ank of the union with smaller grant amounts (and no loans), ending with denmark. the latter face less than a % higher level of real gdp over the time horizon. how do these results compare with those of other models, in particular dsge models? european commission staff has simulated the impact of next generation eu in a stylised version of their quest iii model (see verwey et al., ) . while not providing a detailed analysis, the article reveals that real gdp levels are % higher in and remain elevated by % until compared to a baseline scenario. even higher income member states (in northern and western europe) will experience an increase in real gdp levels of % in . not knowing the precise specifi cations of their model makes it diffi cult to make a sound comparison. notwithstanding slight differences, the results of the commission model appear nonetheless comparable in magnitude to the results presented in this paper. among the four big euro area economies, spain and italy each sustain an average yearly real gdp level of over % above the baseline if investment and consumption follow historic trends. france sustains a yearly gdp level increase of % above baseline and germany of . % in the period from to . in most countries, economic effects are larger than the initial shock plus direct intermediate inputs (the fi rst model with the smallest multipliers). however, this only applies because the simultaneous expansion in all member states 'returns' spillover effects to each country. for instance, if austria alone were to undertake a national expenditure programme equal in magnitude to the austrian share of next generation eu ( . % of austrian gdp in ), the direct effect (including intermediate inputs of the initial shock) within austria would only yield a . % increase in domestic gdp. a large part of the economic benefi t dissipates -from the domestic point of view -towards foreign countries. but due to the coordinated expansion through next generation eu, other countries 'return the favour' and austria manages to capture its share of their spending for an additional . % of austrian gdp -even in the c r o a t i a b u l g a r i a g r e e c e l a t v i a r o m a n i a l i t h u a n i a s l o v a k i a p o r t u g a l p o l a n d s p a i n e s t o n i a c y p r u s h u n g a r y s l o v e n i a i t a l y c z e c h i a m a l t a f r a n c e f i n l a n d b e l g i u m s w e d e n a u s t r i a g e r m a n y n e t h e r l a n d s d e n m a r k i r e l a n d . . . . . . . . . . . . . . . . . . . . . . . . c r o a t i a b u l g a r i a g r e e c e l a t v i a r o m a n i a l i t h u a n i a s l o v a k i a p o r t u g a l p o l a n d s p a i n e s t o n i a c y p r u s h u n g a r y s l o v e n i a i t a l y c z e c h i a m a l t a f r a n c e f i n l a n d b e l g i u m s w e d e n a u s t r i a g e r m a n y n e t h e r l a n d s d e n m a r k i r e l a n d fiscal policy most restricted model. in figure , the total economic effect of the plan per country is decomposed into a domestic part (green shades) and a spillover part from other countries (grey shades). the shading refl ects the three models. the (green) domestic part is defi ned as the gdp growth that results from each country running its own stimulus programme while no other country runs theirs simultaneously. this stand-alone programme is otherwise identical to the level and composition of next generation eu as scheduled for that country. the foreign part for each country is then simply the difference between next generation eu and the stand-alone expansion for that country. this spillover can be interpreted as the price each country would pay for having to go it alone when all neighbours decide to remain uncooperative and not stimulate their own economy as planned for next generation eu. despite the clear value added for all countries that a cooperative fi scal stimulus entails, signifi cant differences prevail across member states. taking the model with the largest effect (including consumption and investment multipliers), greece, croatia, italy, spain and portugal rely on a domestic effect that accounts for more than % of the total gdp increase of the programme, owing to the fact that their domestic stimulus is large compared to other countries. on the contrary, luxembourg, ireland, denmark, the netherlands, austria and belgium receive more than % of the total effect from foreign-induced demand. of course, any keynesian multiplier effect -either through consumption or investment -increases the economic benefi t of the plan massively for each country. then, even the domestic effect only leads to a higher gdp increase than the original outlays. for several countries with large initial domestic spending (south and east), the greater part of the output increase stems from domestic spending and the own multiplier effect thereof. in particular, southern european countries -being rather closed economies in comparison -exhibit high own domestic multipliers (picek and schröder, ) . for northern and western countries in general, most of the total effect originates from outside the country because parts of austria is an interesting case because it has closer geographic and economic ties to the countries of southern and eastern europe than other countries from northern and western europe. before the european council changes to next generation eu, made in july , that gave additional grants to germany and france, austria was set to become the biggest benefi ciary among the northern and western european countries, for which the plan provides only grants but no loans (right-hand section, figure ). there is also a political dimension: austria nets by far the largest gain among the 'frugal four' (denmark, sweden, the netherlands and austria), making its adherence to the group relatively more costly in economic terms (picek, ) . the three models are: direct effect, direct plus induced consumption, direct plus induced consumption and induced investment. in figure , the partial effects of the three models (direct, induced consumption and induced investment) are shown just as in figures and . the increased economic activity in foreign countries spill over. their domestic fi rms and households can secure a disproportionately high share of the induced investment activity and the (resulting) consumption triggered by it. somewhat smaller than in the investment-induced model, the induced consumption effect of grants and loans (second-round effects) increases northern and western eu gdps by sizeable amounts. of all spillover effects, the ones from the direct effect of the shocks are generally the least important for these european states. the comparatively open economies of northern and western europe manage to obtain a large share of the positive economic effects of the reconstruction plan not only through the 'normal outfl ows' from southern and eastern europe via their trade relations, but also through their favourable to give an example using again the case of austria: at the end of the term - , austria's economic output could be up to . % higher than it would have been without the next generation eu package. around . percentage points result from the domestic shock, while . percentage points arise from spillovers. of the latter, . percentage points come from induced consumption and . percentage points are from induced investment (and the additional induced consumption thereof). note: the spillover effect is not negative. both effects are positive and contribute to the total effect. industrial structure that allows them to capture demand for investment goods. this is shown in figure by relating the funds dispensed in the home country (in the form of grants) to the overall initial plus triggered gdp effects (of the whole programme). this multiplier highlights spillover effects from a clearer angle. even disregarding the infl ated value of luxembourg due to its small size and location right between the european economic powers, northern and western countries (from ireland to france) all have a much higher spillover multiplier than the southern and eastern countries (czechia to croatia). an analysis of the economic effects of the reconstruction plan shows above all the importance of a coordinated fi scal policy response to the economic consequences of the coronavirus pandemic. if only one country takes measures to stimulate the economy, part of the effect is 'lost' from a domestic perspective because this part helps foreign economies. if, however, all eu member states take measures to stimulate the economy at the same time, only a very small part is lost to other european countries outside the european union, while the rest benefi ts other member states within the single market. despite their fairly small domestic grants, even the northern and western european countries can thus benefi t from the next generation eu package to a sizeable extent. next generation eu spillovers: multiplier of the total effect relative to domestic grants source: wiod, picek and schröder ( ) , author's calculations. c r o a t i a b u l g a r i a g r e e c e l a t v i a r o m a n i a l i t h u a n i a s l o v a k i a p o r t u g a l p o l a n d s p a i n e s t o n i a c y p r u s h u n g a r y s l o v e n i a i t a l y c z e c h i a m a l t a f r a n c e f i n l a n d b e l g i u m s w e d e n a u s t r i a g e r m a n y n e t h e r l a n d s d e n m a r k i r e l a n d the eu's recovery fund proposals: crisis relief with massive redistribution having the cake, but slicing it differently: how is the grand eu recovery fund allocated? recovery plan for europe proposal for a council regulation establishing a european union recovery instrument to support the recovery in the aftermath of the covid- pandemic, com/ / fi nal/ . european commission ( c), proposal for a regulation of the european parliament and of the council establishing a recovery and resilience facility eu long-term budget - : commission proposal council conclusions, euco / , co eur concl so viel geld erhalten die einzelnen staaten, frankfurter allgemeine zeitung cohesion policy implementation in the eu commission proposes increased agricultural spending in reinforced mff input-output analysis: foundations and extensions koordinierte europäische fiskalpolitik bringt wachstumsschub auch für Österreich spillover effects of germany's final demand on southern europe euro area imbalances: how much could an expansion in the north help the south? an illustrated user guide to the world input-output database: the case of global automotive production next generation eu: a recovery plan for how to spend it: a proposal for a european covid- recovery programme an uncompromising budget next generation eu: % of grants will have to wait until key: cord- - qwbkg o authors: hoddle, mark s. title: biological control of vertebrate pests date: - - journal: handbook of biological control doi: . /b - - / - sha: doc_id: cord_uid: qwbkg o nan several species of vertebrates, especially mammals, have been successful invaders and colonizers of new territories, in particular, insular island ecosystems. others have exhibited high environmental tolerance and adaptability after careful and repeated introductions to new locales by humans. following establishment, several vertebrate species have become important pests. these pests harm agricultural systems by damaging agricultural lands [e.g., rabbits (oryctolagus cuniculus linnaeus) in australia and new zealand], by attacking crops [e.g., european starlings (sturnus vulgaris linnaeus) in the united states], and by acting as sources for communicable diseases [e.g., brushtail possums, (trichosurus vulpecula kerr) , are reservoirs for bovine tuberculosis (mycobacterium bovis karlson and lessel) in new zealand]. other pest vertebrates damage natural systems by threatening the continued existence of endangered flora [e.g., goats (capra hircus linnaeus) on the galapagos islands] and fauna [e.g., brown tree snake (boiga irregularis { merrem }) on guam], and by adversely affecting wilderness areas by changing ecosystem functions and diversity (vitousek et al., ) . the continued relocation of vertebrates exacerbates the ongoing problem of global homogenization of biota (lodge, ) . movement of particular vertebrates into areas where they had not previously existed has, in some instances, occurred naturally without human intervention [e.g., the passerine bird (zosterops lateralis { latham }) arrived in new zealand unassisted from australia]. the vast majority of vertebrate translocations have been human assisted. accidental introduction has occurred as a consequence of human transportation [e.g., brown tree snakes, mice (mus musculus lin-naeus) , and rats such as rattus rattus linnaeus and r. norvegicus berkenhout) ]. some releases have been intentional (but illegal) to serve self-centered private interests [e.g., monk parakeets (myiopsitta monachus { boddaert }) in new york, florida, and texas]. other species have been legitimately introduced to procure public benefit by providing: ( ) new agricultural products [e.g., european wild boars (sus scrofa linnaeus); sheep (ovis aries linnaeus); cows (bos taurus linnaeus), goats and rabbits for meat, and brushtail possums for fur in new zealand], ( ) recreation [e.g., red deer (cervus elaphus linnaeus), fallow deer (dama dama linnaeus), and trout (salmo trutta linnaeus and s. gairdneri richardson) in new zealand]; ( ) companionship [e.g., cats (felis domesticus linnaeus) and dogs (canis familiaris linnaeus)]; or ( ) biological control agents [e.g., the european fox (vulpes vulpes linnaeus), stoats (mustela erminea linnaeus), weasels (m. nivalis linnaeus) , and ferrets (m. putorius furo linnaeus) for the control of rabbits in australia or new zealand; cane toads chemical and cultural control of vertebrate pests is expensive and nonsustainable, and at best provides a temporary local solution to problems (hone, ; williams & moore, ) . biological control of vertebrates, a potentially less expensive and self-sustaining method of population suppression, has focused primarily on mammalian pests. predators, parasites, and pathogens specific to mammals with two notable exceptions (the myxoma and calici viruses that infect rabbits) have failed to provide satisfac-tory control (shelford, ; howard, ; davis et al., ; wood, ; smith & remington, ; whisson, ) . historical records indicate that the majority of attempts at vertebrate biological control have been ad hoc efforts and not the product of careful studies designed to elucidate factors and conditions likely to affect the impact of natural enemy introductions on pest populations. furthermore, failure of biological control of vertebrates by predatory vertebrates has compounded problems associated with exotic vertebrates because control attempts result in addition of new species that cause biological and conservation problems. the level of control achieved by natural enemies is dependent on ratios of natality to mortality of control agents and their host species (davis et al., ) . for vertebrates these ratios are affected by many factors: advanced learning; social, territorial, and breeding behaviors; chemical, physical, and immunological defenses; temporal and spatial escape strategies; and genetic selection in both natural enemy and host populations for persistent coexistence. these complex interrelated factors, coupled with opportunistic feeding habits, have made vertebrate pests difficult targets for biological control with natural enemies. advances in understanding of mammalian fertilization biology have provided molecular biologists with necessary information to develop and investigate the concept of immunocontraception for vertebrate pest control. immunocontraception utilizes genetically modified pathogens that express surface proteins from the target pest's egg or sperm to induce an immune response in the host. antibodies then attack gametes in the host's reproductive tract causing sterilization (tyndale-biscoe, b . computer models indicate that immunocontraception may provide long-term control of vertebrate pests because genetically modified pathogens reduce net reproductive rates without killing hosts (barlow, (barlow, , . in this chapter, i discuss attributes that have aided vertebrate establishment; damage resulting from colonization and uncontrolled population growth; biological control of mammalian pest species with predators, parasites, and pathogens; and future directions that biological control research for vertebrates is taking with genetically engineered microorganisms. permissive to successful introductions of new species. e ton's ( ) predictions have been substantiated in part by paleobiological reconstructions of invasions between newly joined communities (vermeiji, ) and by mathematical modeling describing multi-species interactions in communities (macarthur, ; case, ) . isolated oceanic islands (e.g., new zealand), and insular continents and habitats (e.g., australia and lakes) often have a low diversity of native species. such environments have typically experienced little immigration and are susceptible to invasion by vertebrates (brown, ) . stable, speciose communities with high levels of interspecific competition appear to resist invasion by new species and are sources of successful colonists into less speciose or disturbed communities (macarthur, ; brown, ; lodge ) . this phenomenon has produced asymmetrical patterns of colonization, with successful vertebrate invaders usually being native to continents or extensive nonisolated habitats within continents with more diverse biotas (brown, ; vermeiji, ) . continental herbivores and predators have been very successful in establishing self-sustaining populations in insular habitats, in part because such habitats often lack large generalist vertebrates and essentially have just two trophic levels, producers and decomposers (if specialist herbivores and their predators are excluded) (vitousek, ) . niches equivalent to those on the mainland are largely unoccupied (brown, ) . insular ecosystems therefore often appear to readily accommodate generalist herbivores and predators, perhaps because of low levels of competition for resources that are often inadequately defended chemically or physically (vitousek, ; vermeij, ; bowen & van vuren, ) . organisms from insular habitats that have not coevolved closely with predators or herbivores lack life history features that deter attack or permit survival despite high mortality from predation or herbivory (bowen & van vuren, ) . lack of such biological attributes may increase the competitive advantage of exotics (case, ; coblentz, ; vitousek, ) . introduced vertebrates can also be extremely disruptive in continental regions when habitat disturbance by urbanization or agriculture occurs. european wild boars have detrimental effects on gray beech forests in the great smoky mountains in the southeast united states. habitat disturbance-through pig rooting, trampling, and browsing-and human removal of predators (pumas and wolves) aided pig establishment and spread in this area (bratton, ; singer et al., ; vitousek, ) . communities vary in their ability to accommodate the establishment and proliferation of new species (primack, ) . elton ( ) suggested that species-poor communities (e.g., islands) or highly disturbed habitats are more although there are well-recognized exceptions to general rules that characterize successful vertebrate colonists (ehrlick, ; williamson & fitter, b) , species that establish self-sustaining populations outside their native range typically exhibit some of the following general characters: ( ) short generation times, ( ) high dispersal rates, ( ) high tolerance for varying geographic and climatic conditions, ( )polyphagy, ( ) low attack rates from upper trophic level organisms, and ( ) human commensalism (ehrlich, (ehrlich, , lodge, ; williamson & fitter, a) . the assumption of a high intrinsic rate of increase is generally unnecessary for establishment (ehrlich, ; lodge, ) , although it is important for the establishment of some exotic bird species (veltman et al., ) . the spread of the european rabbit in australia and new zealand, countries that both historically lacked significant eutherian mammal fauna, illustrates some of the preceding points. the european rabbit originated in spain and portugal (corbet, ) and spread through most of europe over years ago following deforestation by humans for agriculture and overgrazing by livestock. animals reared in captivity were carried through europe in advance of naturally spreading populations (flux, ) . the spread of the european rabbit throughout australia following its introduction in varied from km/year to approximately km/year. rate of spread was fastest across dry savannas where conditions were most similar to mediterranean climates and slowest through woodlands (myers et al., ) . in australia, habitat alteration by humans such as conversion of land to pasture, overgrazing of rangelands, and predator eradication [e.g., dingos (canis familiaris dingo meyer)] aided rabbit survivorship and spread. rabbits are polyphagous and feed on grasses and browse shrubs. during severe food and water shortages, bark, fallen leaves, seed pods, tree roots, and termites are consumed (myers et al., ) . rabbits are highly fecund and exhibit rapid population growth under good conditions. individual female rabbits to months of age can produce litters averaging to young, and with abundant food to offspring per year are produced (gibb & williams, ) . behavioral adaptability, sociality, and territoriality, in addition to use of elaborate underground warrens, have also aided rabbit proliferation in marginal habitats (myers et al., ) . assistance in establishment some of the first introductions of exotic vertebrates were those commensal with humans as they colonized new areas. among well-documented early human introductions are dingoes in australia (brown, ) and polynesian rats [rattus exulans (peale)] on pacific islands (roberts et al., ) . as early europeans explored the planet, other commensal species such as r. rattus, r. norvegicus, and mus musculus expanded their geographic range without deliberate human assistance and are now cosmopolitan in distribution (brown, ) . not all vertebrate pests cohabitat with or are associated with human disturbance of the environment. red deer and feral cats, for example, inhabit much of new zealand's pristine habitats with no human management. most vertebrate translocations fail even with human assistance. failure of exotics to establish may depend on life history parameters, responses to abiotic factors, inability to outcompete native species for resources or enemy free space, or chance (cornell & hawkins, ) . deliberate releases of exotic birds have establishment rates of around to % (veltman et al., ) . establishment estimates for intentional vertebrate releases are biased because successes are more often recorded than failures (ehrlich, ; veltman et al., ) . the amount of effort directed toward introduction is an important variable affecting successful colonization by vertebrates, and establishment rates increase with high levels of management and numbers of individuals released (ehrlick, ; griffith et al., ; williamson & fitter, b; veltman et al., ) . in contrast, organisms that are casually introduced into new areas have much lower probabilities of establishing and proliferating. this phenomenon is expressed in the tens rule, a statistical characterization of probability outcomes for different levels of invasion success (williamson & fitter, a , b . for a variety of plants and animals, a general rule holds that in species imported (i.e., brought into new areas intentionally or accidentally) appear in the wild, in of those now found in the wild become established, and that in of those established with self-sustaining populations become pests (williamson & fitter, b) . following establishment, proliferation, and rise to pest status, control of exotic vertebrates is often prompted by economic, environmental, or conservation concerns. several control strategies may be pursued, the most common being chemical control (e.g., poisoning) and cultural control (e.g., trapping, fencing, and shooting). the least used option has been biological control. chemical and cultural control of vertebrate pests has been covered by hone ( ) and williams and moore ( ) . biological control is the intentional use of populations of upper trophic level organisms (e.g., predators, parasites, and pathogens) commonly referred to as natural enemies to suppress populations of pests to lower densities than would occur in the absence of natural enemies (debach, ; van driesche & bellows, ) . biological control programs for vertebrates have employed all three classes of natural enemies: predators, parasites, and pathogens. in contrast to weeds and pestiferous arthropods, however, biological control as a population suppression tactic for management of vertebrate pests has historically received much less attention. predators of vertebrates in the few instances they have been used have not been particularly successful. some vertebrate predator introductions also had severe impacts on nontarget organisms. consequently, early unpromising results discouraged intensive development of this technology (howard, ; davis et al., ; wood, ) . there is a need for increased effort using biological control agents against vertebrates, especially where resistance to toxins has developed, or behavior and terrain makes chemical and cultural control difficult and expensive (wood, ; bloomer & bester, ) . biological control should be fostered internationally because many countries experience similar problems (e.g., rabbits are agricultural pests in argentina, australia, chile, europe, and new zealand; rats, cats, and dogs attack endangered faunas on many oceanic islands; feral pigs and goats in new zealand, australia, and the united states degrade habitat and threaten endangered flora). biological control can be aided by the establishment of institutions to help coordinate regional and international research activities. for example, in , the australian federal government supported the creation of the cooperative research center for biological control of vertebrate pest populations (also known as the vertebrate biocontrol center), an unincorporated collaborative venture between state and federal organizations with international cooperators (anonymous, a) . the principle research goal of this institution is population suppression of noxious vertebrate species by regulating reproductive rates (tyndale-biscoe, a). a fundamental issue that has important implications for biological control is understanding the regulatory effects predators have on prey populations. determining impact of introduced predators on pest and nontarget populations is becoming increasingly more important as public awareness of potential nontarget impacts increases and the impact of past introductions on nontarget organisms has become clearer (howarth, ; van driesche & hoddle, ) . introductions of vertebrate predators as biological control agents against vertebrates have in some instances had disastrous impacts on nontarget wildlife, especially insular communities that have lacked an evolutionary history with generalist predators (case, ) . an example is the impact the small indian mongoose on native rail populations in hawaii following its release in the s for rat control (loope et al., ) . the mongoose has little demonstrable effect on rats (cagne, ) and mongoose populations are now poisoned to protect native birds (loope et al., ) . exotic predators may enhance the success of introduced pest species by moderating the competitive impact of natives on introduced pests should predators reduce densities of native species (case, ) . on the other hand, there is very good evidence that under certain circumstances introduced exotic predators can regulate target vertebrate pests. in such cases, predator efficacy may be affected by ecosystem complexity; by influence of such extrinsic factors as weather, disease, or human intervention on pest population growth; and by the availability of alternative food sources to sustain predators when pest populations are low. in simple ecosystems such as islands, establishment of reproducing predator populations can result in the extinction of target pests. in the absence of alternative prey, releases of cats for rabbit control on berlinger island near portugal by a lighthouse keeper resulted in the eradication of the rabbits, and subsequently cats died from starvation (elton, ) . in complex communities, alternate prey may be taken when primary prey populations are low for prolonged periods, as is the case for some microtine rodent species in europe. european rodent populations exhibit fluctuations in size as a response to variation in food availability. these fluctuations are also influenced by predation. population cycles are not observed in southern hemisphere countries where european pests and predators have been transferred (korpim~iki & krebs, ; sinclair, ) . rabbit populations in australia and new zealand are maintained at low levels by introduced predators, but regulation only occurs after pest numbers have been reduced by other means. poisoning programs in new zealand in the s and s substantially reduced rabbit densities and populations were maintained at low levels by introduced predators, in particular, ferrets and cats (newsome, ) . similarly in australia, european foxes and cats maintain rabbit populations at low densities following population crashes caused by prolonged hot summers that reduce forage and browse (newsome et al., ; newsome, ) . mouse populations are regulated in a similar fashion by predators (raptors and foxes) in australia (sinclair et al., ) . introduction of two foxes dusicyon culpaeus (molina) and d. griseus (gray) native to mainland chile onto the chilean side of tierra del fuego island regulated rabbit populations after rabbit densities were substantially reduced by the myxoma virus (jaksic & yhfiez, ) . the suppressive action of predators on rabbits in australia has been demonstrated through predator removal experiments [referred to as perturbation experiments by sinclair, ( ) ] in which european foxes and cats were shot from four-wheel drive vehicles at night. removal of predators resulted in rapid rabbit population growth compared with rabbit densities in control plots in which predators were not removed (newsome et al., ) . the "predator pit" first conceptualized by may ( ) describes rabbit regulation in australia and new zealand by generalist predators. the model suggests that once prey populations fall below certain densities (i.e., because of culling or disease) predators can prevent recovery to higher levels. generalist predators achieve this by maintaining relatively high numbers by attacking alternate prey species, and low but persistent levels of predation on species in the pit prevents pest populations from outbreaking. for the rabbit-fox system in australia, a predator pit operates at densities of to rabbits per kilometer of linear transect. below these densities foxes utilize alternate food sources (e.g., native animals) and above this critical density rabbit populations escape regulation by predators (newsome, ) . in new zealand, rabbits are contained in the pit by cats and ferrets when densities are . rabbits per hectare (barlow & wratten, ) . the effect of predators on long-term population dynamics of alternate prey species is generally difficult to disentangle from confounding effects of habitat degradation and competition for food and breeding sites from other introduced species (pech et al., ) . foxes regulate mice and rabbits through positive density dependence at low prey densities. increasing pest densities during outbreaks results in inverse density-dependent predation and type iii functional responses (sinclair et al., ; pech et al., ) . rabbit and mouse populations escape predator regulation when favorable weather provides good breeding conditions, or when predators are controlled by shooting or poisoning (newsome, ; sinclair et al., ; pech et al., ) . predator control may be necessary for lifestock protection or for conservation of endangered wildlife and when implemented resurgence of pest populations occurs (newsome et al., ; newsome, ) . predator numbers may increase when primary prey (e.g., rabbits) are abundant. predation by abundant predators on secondary prey (e.g., native animals) result, leading to declines in secondary prey density. under such circumstances native prey species may only persist in refugia or in areas with artificially reduced predator pressure (pech et al., ) . altematively, declining densities of a primary pest prey species (either through management or disease) can intensify predator attacks on nontarget organisms. therefore, when conservation of endangered natives is a concern, culling of predators may be undertaken either concurrently with the decline in prey density or in anticipation of such a decline (grant norbury, personal communication, ) . in such situations an integrated approach to managing vertebrate pests and their predators is necessary (newsome, ) . predator efficacy can be enhanced either through habitat modification or resource provisioning. cats can maintain rat and mouse populations around farm buildings below environmental carrying capacity as long as they are provisioned with additional food (e.g., milk). dietary supplementation prevents rodent extermination and the subsequent extinction of cats. sustaining a cat population prevents uncontrolled invasions by rodents and low pest densities are maintained (elton, ) . provision of nesting boxes for barn owls (tyto alba javanica) reduces crop damage by rats in malaysian oil palm plantations; rodenticide use declined, and in some instances use was eliminated (wahid et al., ) . changes in management practices can improve predator efficacy. rodent control by tyto alba (scopoli) in pinus radiata don plantations in chile was enhanced by clearing -m wide strips between trees (for owls to maneuver in while in flight) and construction of perches in forests (for resting and surveillance). bam owl numbers and predation rates on rodents increased following habitat modification (mufioz & murfa, ) . under increased predation pressure, rodents will modify foraging behaviors by reducing activity when owls are flying or making hunger calls (abramsky et al., ) . learning in this manner produces behavioral adaptations because of strong selection pressures to minimize predation risks on pest populations (davis et al., ) . parasites or macroparasites (e.g., helminths, lice, ticks, fleas, and other metazoans) do not typically kill their hosts as a prerequisite for successful development as insect parasitoids do. they tend to be enzootic (i.e., remain at fairly constant levels through time) and usually must pass through a free-living stage to complete an entire life cycle (anderson, ; mccallum, ) . the potential of parasites to regulate vertebrate host populations was first proposed as early as (lack, ) and later was demonstrated theoretically with lotka-voltera models in which parasites increased host mortality rates may, ) . parasites act in a positive density-dependent manner by adversely affecting host survival or reproduction dobson & hudson, ; scott & dobson, ) . host parasite load also affects the ability of individual parasites to grow, reproduce, and survive in definitive hosts; and the severity of density dependence on host and parasite fitness is affected by patterns of parasite distribution in host populations (scott & lewis, ) . helminths, for example, tend to be aggregated within host populations so that few hosts are heavily burdened while most are lightly infected (scott & lewis, ) . density-dependent constraints on parasite survival and reproduction occur in the few heavily infected hosts, and under such conditions, helminth population stability is enhanced . furthermore, parasites with low-to-moderate pathogenicity exert stronger regulatory actions on populations than highly pathogenic species that cause their own extinction by killing hosts before transmission (anderson, ) . parasite regulation of vertebrate populations has been observed under field conditions. botfly (cuterebra grisea coquillett) parasitism of voles [microtus townsendii (bachman)] in vancouver canada, is inversely density dependent; and botfly infestation significantly reduces vole survival, reproduction, and development (boonstra et al., ) . the parasitic helminth trichostrongylus tenuis (cobbold) is the primary agent responsible for long-term population cycles in red grouse [lagopus lagopus scoticus (latham)] inhabiting scottish heathlands (dobson & hudson, ) . the regulatory effect of t. tenuis has been demonstrated by reducing parasite infestations with helminthicides in experimental birds. treated grouse showed increased overwintering survival, clutch sizes, and hatching rates when compared with untreated birds (dobson & hudson, ) . in the laboratory, introduction of the nematode heligmosomoides polygyrus dujardin reduced mice densities by % in comparison with control populations. reduction of nematode transmission rates and elimination of parasites with helminthicides allowed infested mouse populations to increase (scott, ) . although host and parasite densities in this study were higher than those found in nature, the data showed that introduction of a parasite regulated host population abundance. the potential effectiveness of nematodes as biological control agents in field situations has been evaluated for control of the house mouse, an introduced pest in australia (singleton & mc-callum, ; spratt, ) . mouse (mus domesticus) populations erupt every to years in cereal-growing regions of southeastern australia (singleton & mccallum, ; mccallum, ) and economic losses to mouse plagues exceed $ million (australian) (beckman, ; singleton, ) . outbreaks are associated with high autumn rainfalls following prolonged periods of drought that extend the growing season for grasses that set seeds. this high-quality food source increases high mouse survivorship and breeding throughout winter. population crashes occur when food supplies are exhausted (singleton, ) . saunders and giles ( ) suggested that droughts are necessary to remove the regulating effects of natural enemies, and this removal combined with favorable weather conditions permitted mouse numbers to increase rapidly. capillaria hepatica (bancroft), a parasitic nematode that infests mice, is naturally occurring and widely distributed in pestiferous rodents in coastal areas of australia. it is, however, absent in mouse populations in cereal-growing areas . this nematode is unique because it is the only known helminth with a direct life cycle that requires host death for transmission. female nematodes deposit eggs in the host's liver; and these eggs are liberated by predation, cannibalism, or necrophagy with subsequent digestion of infected liver. unembryonated nematode eggs voided after ingestion undergo embryonation to become infective and are probably consumed when mice preen their fur and feet (fig. ) . ground beetles (carabidae) may vector c. hepatica eggs after they have been eaten (mobedi & arfaa, ). firststage larvae emerge from ingested embryonated eggs and move into the liver through the hepatic portal system (wright, ) . nematode infestation significantly reduces natality and numbers of young mice weaned by infected females (mccallum & singleton, ; singleton & mc-callum, ; . capillaria hepatica is associated with introduced rat and mouse species in urban areas, and naturally occurring infections in native australian mammals are rare probably because of the susceptibility of nematode eggs to ultraviolet radiation and desiccation singleton et al., ) . native australian mice and marsupials are susceptible to experimental infection in laboratories . rats (r. norvegicus and r. rattus) are major reservoirs for c. hepatica in urban areas and infestation rates range from to % (childs et al., ; singleton et al., ) . infestation levels are lower in sympatric mice populations ( to %, singleton et al., ) . low rat numbers in cereal-growing regions of australia may be a factor contributing to the nonpersistence of c. hepatica in these areas . infestation of nonrodent mammals by c. hepatica is rare but has occurred in rabbits (gevrey & chirol, ) , dogs (leblanc & fagin, ) , horses (munroe, ) , and humans (pannenbecker et al., ) . human infections can be treated successfully (pereira & franca, ) . exploratory models investigating the impact of c. hepatica on mouse populations indicated that the requirement of host death for parasite transmission is strongly destabilizing. in the absence of resource limitation mouse densities increase similarly to disease-free populations before parasites have an impact and infected populations decline in density (mccallum & singleton, ) . slow regulation of mouse populations occurs because of the need for host death for transmission. consequently, the nematode's life cycle operates on the same time scale as that of its host instead of being orders of magnitude faster, as is the case with other parasites that do not require host death for transmission (mccallum & singleton, ; singleton & mc-callum, ) . the destabilizing influence of c. hepatica on mouse parasitology today, , [ ] [ ] [ ] [ ] populations may contribute to localized host and parasite extinctions. these extinctions, coupled with very low mouse densities in nonoutbreak years, result in population bottlenecks and may explain why nematodes do not persist in regions where mouse outbreaks occur. soil type, temperature, and moisture content do not affect nematode egg survival and embryonation under favorable conditions in outbreak regions (spratt & singleton, ) . outbreak intensity can theoretically be reduced by c. hepatica if populations are inoculated early, preferably year before an outbreak is expected (mccallum, ) . releases of high doses of nematode eggs in the summer or autumn when mouse densities are sufficient to enable high levels of transmission may offer the best chance for successful control (mccallum & singleton, ; mccallum, ) . field experiments in enclosures and with increasing populations of free-ranging mice have failed to demonstrate long-term regulation on mouse population growth with periodic inoculative releases of c. hepatica eggs. unexpected declines in control populations (i.e., populations not treated with nematodes) have to some degree masked the effect of c. hepatica on mice populations singleton et al., ; singleton & chambers, ) . transmission of c. hepatica in treated populations is not density dependent and can occur at low levels for to months. transmission rates show seasonal trends influenced by soil temperatures and increasing aridity singleton & chambers, ) . improved understanding of the influence of factors (such as temperature and rainfall on nematode persistence, survival, and transmission in field situations) and timing of releases of parasite eggs may improve releases of c. hepatica for control of mouse outbreaks (singleton et al., ; singleton & chambers, ) . vertebrate species that successfully colonize new habitats have reduced parasite loads in comparison with mother populations from which they originated (dobson & may, ) . lower infestation levels probably occur because individuals that make up small founding populations either were uninfected or had only a limited subset of the total potential parasite species found in the area of origin, or intermediate hosts required for parasite persistence were absent in the new range. sparrows and starlings, both successful colonizing species from europe, have two to three times fewer parasites in north america compared with populations from which they originated. populations established outside of europe may have benefited from reduced parasite burdens, although there are no quantitative data to indicate that this aided establishment and proliferation (dobson & may, ) . investigating the role of parasites on population dynamics of rabbits in europe with the view for possible introduction into countries where rabbits are pests is also warranted (boag, ) . introduced mammals such as rats, goats, and cats on oceanic islands exhibit depauperate parasite faunas (dobson, ) . fewer parasites coupled with presumed low genetic diversity of small founding populations, and reduced selection pressures for parasite resistance may make these pest vertebrates vulnerable to introduced host-specific parasites. the ideal parasite introduced into a high-density pest population that originated from a small founding population should have low-to-intermediate pathogenicity, because such parasites establish and maintain themselves in populations at lower densities than more pathogenic species do . macroparasites that reduce both host longevity and fecundity may have the potential to cause sustained reductions of host population densities (dobson, ) . low genetic variability among target populations should theoretically enable introduced parasites to become more evenly distributed among hosts, and reduction in parasite aggregation would increase natural-enemy efficacy (dobson & hudson, ) . the possibility of reassociating parasites with vertebrate pests is not limited to mammals and birds. host-specific parasites may have the potential to reduce reproduction and longevity of pest reptile (dobson, ) and amphibian species (freeland, ) . the brown tree snake is the proximate cause of native bird extinctions on guam following its accidental introduction after world war ii on military equipment (pimm, ; savidge, ; jaffe, ; rodda et al., ) . the snake also has caused declines of native reptile and small mammal populations, and enters houses and attacks sleeping human infants (rodda et al., ) . additionally, the brown tree snake has caused eco-nomic losses by adversely affecting domestic animals (e.g., chickens and pets), and high densities of snakes on power lines regularly cause short circuits that interrupt electrical supplies and necessitate repairs. control of the brown tree snake has been attempted through trapping, but the snake's extreme preference for live bait over artificial lures has made this approach impractical (rodda et al., ) . the brown tree snake mnative to eastern indonesia, the solomon islands, new guinea, and northeastern australiambelongs to the family colubridae. it is the only member of this family on guam. there is one native species of snake on guam, the blind snake, rhamphotyphlopys braminus (daudin), which belongs to the family typhlopidae and is the only snake occurring on many islands in the central pacific region (t. fritts, personal communication, ) . the brown tree snake has extended its range and is now established on the previously snake-free island of saipan, and this snake has been intercepted in hawaii; corpus christi, texas, and spain (rodda et al., ) . given the propensity for the brown tree snake to be dispersed to new habitats within cargo loads on planes and ships, the major social, economic, and ecological problems that are caused on islands after colonization, in addition to its distant taxonomic relationship to snakes common to pacific islands, make the brown tree snake an excellent target for biological control. the taxonomic relationship between colubrids and typhlopids may simplify the task and reduce the cost of finding natural enemies unique to the brown tree snake. parasites or pathogens that are host specific just to the family (i.e., colubridae) or genus (i.e., boiga) level may be safe to nontarget snakes (e.g., typhlopids) because these organisms have not evolved the ability to cause disease in distantly related hosts. extreme caution should be exercised when implementing a biological control program against vertebrate pests with parasites. parasites and pathogens can pose major threats to populations of endangered animals (mccallum, ; mccallum & dobson, ) . the susceptibility of nontarget organisms, especially endemic species, to infection by candidate biological control agents should be investigated thoroughly prior to parasite releases. reassociating parasites that preferentially infect a competitively dominant pest species may increase species diversity of invaded communities by reducing the pest's prevalence. in this instance, the natural enemy would assume the position of a keystone parasite (marcogliese & cone, ) . pathogens or microparasites include viruses, bacteria, and protozoans. pathogens tend to be unicellular and exhibit epizootic (i.e., boom or bust) life cycles due to rapid proliferation in hosts (anderson, ; mccallum, ) . the potential of pathogens to regulate vertebrate population densities by reducing the longevity and fecundity of infected hosts has been demonstrated theoretically with mathematical models and by perturbation experiments using vaccines (smith, ) . as with macroparasites, models indicate that microparasites of intermediate pathogenicity are more effective biological control agents (anderson, ) . highly virulent pathogens kill themselves by destroying hosts before they can be transmitted and avirulent strains are not transmitted because they are removed by the immune system. the immune system is theorized as being responsible for maintaining the intermediate virulence of vertebrate microparasites (anita et al., ) . pathogens that are readily transmitted (i.e., microparasites spread by water, air, and vectors) or have high-density host populations are more contagious than those with low transmission rates (i.e., spread is by host-to-host contact) or low host densities (ebert & herre, ) . new associations between pathogens and novel hosts are generally not more harmful than those that have evolved closely with the host. experimental evidence indicates that novel disease-causing organisms are on average less harmful, less infectious, and less fit than the same parasite strain infecting the host it is adapted to (ebert & herre, ) . also, a microparasite's ability to infect and exploit novel hosts decreases with increasing geographic and presumably genetic distance from the host to which the pathogen is adapted (ebert, ) . exceptions do occur, however, and pathogens can have devastating impacts on hosts that have no evolutionary history with the disease organism. an example is the myxoma virus, the causative agent of myxomatosis in european rabbits. the use of this natural enemy against rabbits in australia and europe has been the most thorough biological control program against a vertebrate pest. the myxoma virus is a member of the genus leporipoxvirus (poxviridae) and originated from south america where it was first recognized as an emerging disease of european rabbits in laboratories in montevideo, uruguay, in . infected laboratory rabbits died of a fatal febrile disease that caused tumors on the head and ears. the tumors resembled myxomas (a benign tumor composed of connective tissue and mucous elements) and the disease was subsequently named infectious myxomatosis of rabbits (fenner & marshall, ; fenner & ratcliffe, ; fenner, ) . the indigenous host for myxoma virus in south america is the forest rabbit [sylvilagus brasiliensis (linnaeus)]. unlike its effect on european rabbits, myxoma inoculum injected into forest rabbits caused benign fibromas at the site of inoculation that persisted for many months, although death did not occur. mosquitoes were implicated in vectoring the disease from forest rabbits to european rabbits being bred in south american rabbitries. another leporipox-virus has been isolated in california from the brush rabbit, sylvilagus bachmani (waterhouse), and is closely related to the myxoma virus (fenner & marshall, ; fenner & ratcliffe, ; fenner, ; fenner & ross, ; ross & tittensor, ) . myxoma virus has been used in australia, europe, chile, and argentina for biological control of european rabbits. the virus was first imported into australia from brazil in and but was not released (fenner & ratcliffe, ) . work by australians with the virus began again in the united kingdom in and continued with caged rabbits on wardang island off the south coast of australia. the virus was successfully established on mainland australia in (fenner, ) and within years it had established itself over most of the rabbit's range (fen-ner& ratcliffe, ) . the virus initially had a major impact on the estimated million rabbits and on the damage they caused, reducing population density by to %. efficacy was dependent on climate and rabbit population susceptibility. populations have subsequently increased and stabilized at around million because of myxomatosis. damage attributable to rabbits still amounts to $ million (australian) annually, including both lost agricultural production and cost of control applications (robinson et al., ) . in addition to agricultural losses, rabbits severely affect native flora by eating foliage and inducing wind and water erosion of soils by overgrazing. native fauna are also affected as rabbits out-compete indigenous herbivores and dense rabbit populations sustain exotic predator populations that feed on native animals (gibb & williams, ; myers et al., ; robinson et al., ) . within a few years of the initial panzootic, field isolates of the virus showed less virulence when compared with the original strain that had been released. the original strain killed > % of laboratory rabbits on average . days after infection, while circulating strains caused % mortality after . days. genetic resistance in rabbits was also detected (fenner & marshall, ; fenner & ratcliffe, ) . dual natural selection had occurred, the virus had attenuated, and rabbits had increased in resistance to the disease. mosquitoes have been responsible for vectoring myxoma virus in australia. the european rabbit flea, spilopsylus cuniculi (dale), an important vector in europe, was introduced into australia in and increased the geographic distribution of the disease. this flea did not persist in areas with rainfall < mm. the xeric adapted spanish rabbit flea, xenopsylla cunicularis smit, was introduced in and active redistribution is still ongoing (fenner & ross, ) . new zealand also has inordinate numbers of rabbits, and attempts to establish the myxoma virus from to failed because of inclement weather and a paucity of suit-able arthropod vectors. further attempts at establishment were not undertaken because poisoning programs had reduced rabbits to very low numbers, additional control expenditure was unjustifiable, and the new zealand public was not in favor of using lethal myxoma virus for rabbit control on humanitarian grounds (gibb & williams, ) . until the s, myxomatosis was the only disease known to severely affect rabbit numbers. a second highly contagious viral disease emerged in the mid- s and was accidentally introduced onto mainland australia (o'brien, ) . it is the first pathogenic natural enemy to have established in new zealand for biological control of rabbits. rabbit calicivirus disease (rcd) [also known as rabbit hemorrhagic disease virus (rhdv)] emerged as a fatal disease in in angora rabbits exported from east germany to jiangsu province of china (liu et al., ) . in , the disease appeared in italy where million rabbits were estimated to have died. the disease spread rapidly through rabbit populations in europe reaching the united kingdom in (chasey, ) . the probable mechanism for dispersal in continental europe was the movement of live rabbits and rabbit products. transmission of rcd from france into coastal areas of southeast england is thought to have occurred by wind-borne aerosols containing virus, birds, and transchannel ferry traffic (chasey, ) . outbreaks of rcd occurred in mexico in and (gregg et al., ) and in rrunion island in the indian ocean in . movement of rcd to these areas probably occurred with imports of frozen rabbit carcasses from china because the virus can survive freezing to temperatures of - ~ (chasey, ) . the rcd virus belongs to the caliciviridae and consists of a positive sense, single-stranded rna genome, enclosed by a sculptured capsid composed of multiple copies of a single major protein of kda, and is to nm in diameter (ohlinger et al., ; parra & prieto, ) . disease symptoms are characterized by high morbidity and mortality in rabbits over weeks of age. younger rabbits often survive infection and may develop antibodies to rcd virus (nagesha et al., ) . clinically, rcd symptoms are expressed after an incubation period of to h in which a febrile response and increasing lethargy are observed. infected rabbits typically die within to h postinfection and % mortality is observed after days. necropsies show a pale swollen friable liver, enlarged spleen, and clots in blood vessels. death is ascribed to acute necrotizing hepatitis and possible hemorrhaging (fuchs & weissenbrck, ; studdert, ) . however, necropsies close to the time of death show an absence of hemorrhaging and inclusion of hemorrhagic in the name of this rabbit disease that indicates the cause of death is misleading (studdert, ) . a different viral disease is responsible for european brown hare syndrome (ebhs) which causes severe hepatic necrosis in hares (lepus europaeus pallas and l. timidus linnaeus). the disease was first recorded in sweden in , and spread through continental europe and reached the united kingdom in (fuller et al., ) . in sweden, losses of hares to ebhs occurred years prior to sympatric rabbit populations developing rcd. similar observations were made in the united kingdom where hares began dying from ebhs years before rcd was observed in rabbit populations (fuller et al., ) . electron microscopy, nucleotide sequencing, and experimental cross-transmission studies have indicated that rcd virus and ebhs virus are closely related (le gall et al., ) but distinct members of the caliciviridae (chasey et al., ; nowotny et al., ) . disease symptoms are generally similar for rabbits and hares but show distinguishing characteristics in necrosis of liver lobules and clotting of blood vessels (fuchs & weissenbrck, ) . serological studies on rabbit sera collected in from czechoslovakia and austria indicate that rcd virus probably evolved from an apathogenic strain endemic to europe from at least this time (nowotny et al., ) . studdert ( ) speculates that the causative agent of rcd probably existed in europe as a quasi species, a collection of indifferent mutants with a variety of accumulated nucleotide changes. in this scenario, mutants occupied a specific ecological niche until one strain better adapted to prevailing conditions became the dominant member of the population. adaptation may have occurred because mutations caused increased virulence in an avirulent rabbit virus or increased the host range of hare-infecting viruses by allowing mutant strains to bind more efficiently to surface receptors on rabbit hepatocytes. given studdert's ( ) speculative scenario, rcd virus may be a highly evolvable organism. european rabbits appear to be the only animals susceptible to infection by rcd virus, and vaccines have been developed to protect domestic animals (boga et al., ) . other rabbits including cottontail rabbits (sylvilagus spp.), black-tailed jack rabbits (lepus californicus gray), volcano rabbits [romerolagus diazi (ferrari-prrez)] (gregg et al., ) , and hares (gould et al., ) are not affected. the limited host range of rcd virus makes it an obvious candidate for use in a biological control program against european rabbits in new zealand and australia. a joint biological control program between these two countries using rcd virus was initiated in and a strain of virus from the czechoslovakia republic was imported into australian quarantine facilities in to test effects on nontarget species (robinson & westbury, ) . host-specificity testing of nontarget species in australia for susceptibility to rcd virus further verified the limited host range of this natural enemy. test subjects in-cluded domestic lifestock (horses, cattle, sheep, deer, goats, pigs, cats, dogs, and fowls) , noxious exotic vertebrates (foxes, hares, ferrets, rats, and mice), native mammals (eight species), birds (five species), and reptiles (one species). there was no evidence for viral replication, clinical signs, or lessions in any organisms tested (gould et al., ) . artificial inoculation of rcd virus in north island brown kiwis (apteryx australis mantelli bartlett) and lesser short-tailed bats (mystacina tuberculata gray), both native to new zealand, also failed to produce disease symptoms (buddle et al. ) . the apparent host specificity of rcd virus to rabbits, rapidity of action, and the capacity to infect rabbits from other rabbits, [through feed and feces, or from a contaminated environment (o'brien, ) ] prompted further evaluation of this biological control agent under field quarantine conditions in australia. studies monitoring the effects of rcd virus on rabbit populations were initiated on wardang island near adelaide off the south coast of australia in (rudzki, ; robinson & westbury, ) . in september , rcd breached quarantine and appeared on mainland australia, possibly carried there by calliphorid flies and onshore winds (cooke, ; lawson, ) . attempts at containment failed (seife, ) . within months of the initial discovery of rcd virus on the mainland, an estimated million rabbits were killed in south australia. in dry areas, to % of infected populations died (anderson, ) with dead rabbits averaging per hectare. elsewhere, fatality rates were closer to % (anonymous, b) . in the period from october to november , an estimated total of million rabbits died from rcd in south australia and the majority of surviving rabbits were less than weeks of age (cooke, ) . the development of resistance in young rabbits may have profound effects on the long-term population dynamics on the rabbit-rcd virus system. ten arthropod vectors of rcd virus have been identified and include flies, mosquitoes, and rabbit fleas (anonymous, b) . rates of spread of rcd are greatest in spring and autumn at to km a day and are correlated with peaks of insect activity. dispersal of the disease probably has been assisted by humans moving contaminated material to new areas (cooke, ) . increased attacks on native fauna by exotic predators such as foxes because of declines in rabbit numbers do not appear to have occurred because predator populations have declined with rabbit numbers (anonymous, b). the virus is now endemic in australia and will probably be officially declared as a biological control agent under the biological control acts of the commonwealth and states (robinson & westbury, ) . rcd virus was smuggled into the south island of new zealand by high country farmers in august and illegally disseminated by feeding rabbits carrots and oats satu-rated with contaminated liquefied rabbit livers. a network of cooperators spread the virus over large areas of the south island and its subsequent spread (human assisted through the movement of carcasses, baiting, and insect vectors) made containment and eradication of the disease impossible. such actions by farmers clearly violated new zealand's biosecurity act, which was enacted in part to protect agriculture from unwanted introductions of pests. the new zealand government has sanctioned controlled virus releases into new areas. the short-term impact of rcd on new zealand rabbit populations has resulted in to % mortality in central otago and large-scale field studies are planned (g. norbury, personal communication, ) . cats on oceanic islands have been subjected to biological control with pathogens. six cats were introduced onto marion island in the indian ocean in (howell, ) ; by , numbers were in excess of and were increasing an average of % per year (van rensburg et al., ) . populations were sustained by consuming approximately , seabirds yearly and cats were probably responsible for the local extinction of the common diving petrel pelecanoides urinatrix (gmelin) (bloomer & bester, ) . surveys of cats on marion island revealed the presence of feline herpes virus and feline corona virus, but the highly contagious feline parvo virus was absent in the population (howell, ) . initiation of a biological control program with feline parvo virus, the causative agent of feline panleucopenia, began in with the release of artificially inoculated feral cats collected from the island (howell, ) . the disease reduced cat numbers by % after years by reducing fecundity and increasing mortality of juvenile cats (van rensburg et al., ) . virions found in high concentrations in feces, urine, saliva, and vomit were transmitted through direct contact between cats or contact with contaminated objects (howell, ) . annual declines of cat numbers were % from to . this rate decreased to % per year from to and was accompanied by lower titers of virus in serum samples collected from feral cats, indicating that viral efficacy was decreasing (van rensburg et al., ) . at reduced densities, hunting and trapping became viable and have been incorporated into an ongoing eradication program that may be assisted by the use of trained dogs (bloomer & bester, ) . sexually transmitted diseases have adverse effects on domestic and wild vertebrates by reducing survival, conception rates, and numbers of offspring born and successfully weaned (smith & dobson, ) . rabbits are susceptible to infections of venereal spirochetosis (treponema cuniculi), which causes sterility (smith & dobson, ) . goats can develop trichomoniasis, a sexually transmitted disease caused by the flagellated protozoan trichomonas foetus (reidmuller). this pathogen has been suggested as a bio-logical control agent for goat populations on oceanic islands that lack this microparasite (dobson, ) . sexual transmission of diseases may further guarantee host specificity in biological control programs. it also enhances the ability of parasites and pathogens to persist in low-density populations or solitary species (e.g., predators). the rate of spread of sexually transmitted organisms is tightly correlated with mean and variance of the numbers of sexual partners per host because of the need for host-tohost contact (horizontal transmission) for transmission. host population density is not important with respect to persistence or rate of spread of sexually transmitted diseases. this property, coupled with asymptomatic carrier states, long infectious periods, or vertical transmission (infective propagules are passed from mother to offspring), greatly enhances the ability of pathogens to persist in lowdensity host populations (smith & dobson, ) . because sexually transmitted organisms can persist in low-density populations or populations of declining density, the potential of genetically engineering sexually transmitted viruses to sterilize infected hosts is being investigated (barlow, ) . viruses that have antigens from the host sperm, or the zona pellucida around host eggs engineered into the genome provoke an immune system response that renders the recipient sterile. immunocontraception (also referred to as immunosterilization) as a means to control noxious vertebrates is being actively pursued by australia and new zealand (mccallum, ) . an alternative approach to immunocontraception is to use genetically modified microparasites to prevent lactation in females so that juveniles are not successfully weaned or to interfere with hormonal control of reproduction (cowan, ; jolly, ; rodger, ) . many species that become pests are distinguished from nonpestiferous species by their higher intrinsic rates of increase (rm). pest vertebrates have high rm values characterized by large litters, and by maturing sexually at young ages. agents that reduce reproductive rates may be more effective for control than mortality-inducing biological control agents are because resistance development should take longer to occur and population recovery would be slower (tyndale-biscoe, b) . resistance development may be further delayed by combined use of multiple agents that affect fertility in different ways [e.g., using agents that cause sterilization, alter levels of reproductive hormones, or affect lactation (cowan & tyndale-biscoe, ) ]. in sexually reproducing vertebrates, proteins associated with male and female gametes are potentially foreign antigens in the opposite sex. exposure to reproductive antigens occurs when females receive sperm and accessory fluids from males during copulation. as a general rule, females do not develop antibodies to these antigens because physiological and immunological mechanisms have evolved to prevent this (robinson & holland, ) . inoculation of sperm into females of the same species either subcutaneously or intramuscularly produces high sperm antigen antibody titers in recipients. in most cases, this causes either permanent or temporary infertility in females. such results indicate that sperm antigens in the reproductive tract are tolerated and that exposure to these antigens by different routes overcomes protective mechanisms, with infertility resulting (robinson & holland, ) . sperm antibodies in females that can arise from either systemic or local immune responses are found in cervical mucus, genital fluids (e.g., endometrial, tubal, and follicular fluids), and blood. antibodies bind to sperm, often in specific locations such as the head, midpiece, tail shaft, or tail tip. once bound to sperm, antibodies cause agglutination (e.g., irreversible binding to cervical mucus that normally aids sperm transport) or immobilization of sperm. antibodies may also interfere with acrosome reactions preventing ovum penetration and fertilization, or they block the binding of sperm to the zona pellucida (shulman, ) . the zona pellucida (zona) that surrounds growing oocytes and ovulated eggs is antigenic and available to circulating antibodies during oocyte growth and ovulation. nonreproductive tract inoculation of females with zona preparations leads to infertility (millar et al., ) . antibodies produced in response to administered zona antigens bind to the zona and prevent sperm penetration (millar et al., ) . zona glycoproteins are highly conserved among mammals, for example, nonspecific pig zona preparations cause infertility in humans, primates, dogs, rabbits, horses, and deer (robinson & holland, ) . a major objective in immunocontraception research is isolation of species-specific zona glycoproteins that do not cause sterility induced by immune response in species from which zona preparations were not derived. low variability among zona glycoproteins may limit the number of species-specific zona preparations for immunocontraception (millar et al., ) . immunocontraception for wildlife population control has been successfully implemented for horses (equus caballus linnaeus) (kirkpatrick et al., (kirkpatrick et al., , . free-ranging feral mares inoculated by dart gun with porcine zona pellucida showed depressed urinary estrogen concentrations and failure to ovulate. zona booster inoculations given years after initial inoculations prevented conception in treated horses for a third year compared with control populations that were not vaccinated. contraceptive effects were reversible after years of consecutive treatment, but prolonged treatment ( to years) with zona preparations caused irreversible ovarian dysfunction and fertility loss (kirkpatrick et al., (kirkpatrick et al., , . similar results have been achieved with porcine zona pellucida inoculations in whitetail deer [odocoileus virginianus (zimmerman)] (kirkpatrick et al., ) . gametic antigens that induce immune response can be administered by baits that are ingested by target organisms or can be inoculated directly into hosts with darts or bullets (tyndale-biscoe, b). injection of foxes with sperm antigens reduces fertility from to %. baits are considered the favorable method for delivering antigens to foxes in australia. potential baits include dried meats that contain microencapsulated antigens. use of recombinant bacterial vectors (e.g., salmonella typhimurium) also are being considered. an orally administered agent needs to reach the lower gastrointestinal tract to stimulate a response in the common mucosal immune system in the gut-associated lymphoid tissue. this in turn induces mucosal immunity in the reproductive tract of female foxes and causes sterilization (bradley et al., ) . at present, an effective bait specific to foxes that is environmentally stable and easy to manufacture has not been developed. nontarget impact is a concern because most antigens exhibit some specificity to the family level only. effective vaccines for rabies have been delivered as oral baits to foxes in europe, demonstrating the baiting technique is an effective dissemination method (bradley et al., ) . models indicate density-independent factors such as drought and rain (which affect pasture growth and rabbit numbers) strongly influence the effectiveness of baitdelivered fertility control in reducing fox abundance . an alternative proposal to deliver antigens orally is to develop transgenic plants to produce and deliver gametic antigens in palatable form to herbivorous pests. plants could be sown over target areas and allowed to become self-propagating vaccines. transgenic seeds, fruits, or leaves (e.g., transgenic carrots or maize) could be harvested and used as oral baits delivered to specific sites such as fenced watering points that allow pest animals access while excluding lifestock (smith et al., ) . baiting is an expensive form of control that requires monitoring of dosage and uptake rates and multiple areawide applications. problems of hormonal modification of behavior and delayed population control are additional drawbacks. one advantage is that baits can be used to treat localized populations that are problematic. similar shortcomings exist with antigen inoculations by projectiles where cost estimates are significant. to control the estimated , wild horses in australia with dart-delivered porcine zona pellucida would cost $ (australian) per horse per year compared with cents for permanent control with a bullet (tyndale-biscoe, ) . lethal methods of control provide immediate impact on pest populations and reduce pest status rapidly, with control being quickly observable. in contrast, fertility impairment is not immediate, population responses are delayed, and large proportions of populations need to be sterilized for this technique to be effective. large-scale distribution of gametic antigens might be possible through releases of host-specific microparasites expressing species-specific antigen genes (tyndale-biscoe, a , b . self-spreading and replicating parasitic vectors that have been genetically engineered and that may require periodic reinoculation into populations are analogous to augmentative biological control programs with traditional natural enemies (e.g., parasitoids, predators, or pathogens) released periodically for the control of pest arthropods. host-specific viruses carrying foreign dna could be cheap and effective biological control agents that have the potential to disseminate widely by sexual transmission, contagion, or arthropod vectors. the selected micro-or macroparasite must be able to carry foreign dna coding for gametic antigens, as well as promoters to express foreign genes and cytokines to enhance effectiveness (tyndale-biscoe, b). such agents must be able to reduce growth rates of infected populations and to maintain reproductive rates at lower levels (caughley et al., ) , and should not interfere with sexual behavior or social organization (caughley et al., ; robinson & holland, ; tyndale-bisoce, b) . with some pests such as rabbits and foxes, dominant members of populations make the main contribution to reproduction and inhibit breeding by subordinate members by occupying prime territories. ideally, a sterilizing agent should not change social hierarchies by allowing individuals with lower social status to successfully rear more offspring because this will cause pest populations to increase substantially (caughley et al., ) . genetically engineered agents should sterilize females because models predict greater population suppression with infertile females than with sterilized males (barlow, ; caughley et al., ) . in the absence of arthropod vectors, sexually transmitted diseases engineered to cause sterilization are superior to nonsexually transmitted ones because multiple matings with sterilized females increases contact rates and the competitive ability of the engineered agent with nonsterilizing strains. the impact of immunocontraception is further enhanced if the sterilizing agent causes limited host mortality and there is low naturally occurring immunity to sexually transmitted diseases (barlow, ) . sexually transmitted herpes-type viruses are being proposed as vectoring agents to induce sterilization in brushtail possums in new zealand (barlow, ; barlow, ) . the recently identified borna disease virus that causes wobbly possum disease in new zealand may be a suitable alternative to a herpes virus (atkinson, ) . the myxoma virus and murine cytomegalovirus are being investigated as gamete antigen delivery agents for rabbits and mice, respectively, in australia (mccallum, ; tyndale-bisoce, b; shellam, ) . four potential insertion sites for genes coding for gametic antigens have been identified in myxoma virus and recombinants have been constructed to express two esherichia coli (escherich) enzymes and influenza virus hemagglutinin genes. the ability of a novel myxoma virus to compete and spread among existing myxoma strains in field situations has been demonstrated by monitoring the spread of virus containing identifiable gene deletions (robinson et al., ) . the myxoma virus that can express foreign genes may operate as a vector for gametic proteins (robinson et al., ) . work is continuing on isolating and inserting rabbit gamete antigen genes into the myxoma virus genome (robinson et al., ) . the responses of experimental rabbit and fox populations in australia to imposed sterility by surgical ligation of fallopian tubes in females have been studied in an attempt to simulate the effects of virally mediated immunocontraception after recombinant virus establishment in wild populations. this technique prevents conception among predetermined proportions of females in populations without interfering with hormones or reproductive behavior (williams & twigg, ) . the dynamics of rabbit populations enclosed by rabbit-proof fencing that exhibited , , , or % sterilization of females were studied in each of two locations in western and eastern australia where climate patterns differed. females born into treatment populations were trapped and sterilized to maintain the same overall sterility levels (williams & twigg, ) . juvenile rabbits born into populations with sterilized females exhibited greater survivorship because of lowered competition for resources. this greater survival compensated for decreased fertility, but recruitment rates were ultimately constrained by environmental factors (e.g., depletion of vegetation). in populations with % sterility, reduced juvenile mortality did not compensate fully for lowered reproduction, smaller numbers of rabbits were recruited into these populations, and numbers subsequently declined. these results indicate that levels of sterilization with a genetically altered micorparasite have to reach at least % to achieve reductions in population density (williams & twigg, ) . surgical sterilization does not affect reproductive behavior in treated populations. sterile dominant female rabbits maintain hierarchical dominance, increased body weight over control females, continued to defend prime territory, and engaged in normal reproductive behavior including breeding burrow construction (tyndale-bisoce, b). birth rates of sexually mature females were in direct proportion to the level of fertility in experimental populations, indicating that fertile females did not respond to female infertility or decreased densities of young by producing larger litters (williams & twigg, ) . sterilized females tended to live longer than unsterilized females. this increased longevity suggests that sterile females may proportionately increase as treated populations reach an equilibrium density. obviously, larger proportions of sterile females reduce population productivity and the numbers of fertile females that a sterilizing microparasite would have to infect and sterilize. higher proportions of sterile females may reduce numbers of infective individuals harboring sterilizing microparasites and numbers of vectors (e.g., fleas that would spread an engineered myxoma virus), and may contribute to decline of transmission rates. these interactions need to be clarified and mathematical models may be of use here (williams & twigg, ) . engineered microparasites that sterilize pest animals offer the possibility of humane control without killing or causing animals to suffer the effects of debilitating disease. as a form of biological control, immunocontraception may also reduce the need for broadcast distribution of toxins for pest suppression, thereby reducing environmental contamination and nontarget mortality. this is of special concern when pests inhabit suburbs, urban parks, government and state campuses, nature reserves, military bases, or other areas where lethal controls may no longer be legal or safe (kirkpatrick et al., ; williams, ) . the concept of virally mediated immunocontraception has generated considerable debate on legal and ethical issues concerning releases of engineered microorgansims into the environment. once contagious recombinant agents that cause permanent sterilization in animals are released into the environment they cannot be recalled (tyndale-bisoce, ) . several potential risks are recognized. first, engineered viruses that are host specific and contain species-specific antigens could mutate and infect and sterilize nontarget species after release (anderson, ) . under such conditions it may be difficult if not impossible to contain and eradicate a mutant virus from an infected animal population that is abundant, secretive, and free ranging. second, sterilizing viruses either might cross international boundaries accidentally or be maliciously moved to sterilize desirable organisms in new areas (tyndale-bisoce, b) . for example, viruses engineered with little host specificity to sterilize widely dispersed marsupial pests in new zealand may enter australia and infect endangered wildlife (rodger, ; mccallum, ) ; engineered myxoma viruses may spread from australia into the americas and sterilize native rabbit species (tyndale-biscoe, ) . third, dart-delivered contraceptives used for wildlife control in the past have had adverse effects on individuals within target populations. changes in morphology of repro-ductive organs, secondary sexual characteristics, and behavior have been observed. viruses that induce sterility could alter genetic profiles of target populations because infectious agents may act as a new reproductive disease and individuals may exhibit differential susceptibility (nettles, ) . fourth, public concerns over the use of viruses and genetic engineering indicate substantial apprehension about the use of sterilizing viruses for pest management, these fears that need to be fully alleviated may delay or prevent field trials and widespread application (lovett, ) . despite potential drawbacks, immunocontraception is a potentially cost-effective method for reducing pest impact on endangered native species (sinclair, ) and on agricultural yields, and is an additional tool for sustainable pest management (williams, ) . a sterilizing agent that does not cause painful disease symptoms is an ethically acceptable form of pest control that is justifiable from animal rights perspectives, because it does not cause the suffering typical of current lethal methods (e.g., trapping, shooting, poisoning, and introduced disease) ( oogjes, ; singer, ) . under certain circumstances, the use of vectors to disseminate genetically engineered viruses is warranted (mccallum, ) . experience with the myxoma virus in australia indicates that it has not been deliberately or accidentally spread to any other country since its introduction in the s because of either the lack of suitable arthropod vectors or the inability of the virus to establish where different strains are already present. this history may indicate possible difficulty for unintentional establishment of genetically engineered microparasites in new areas, and establishment of engineered myxoma viruses may be possible only with carefully timed and repeated releases into rabbit populations (tyndale-biscoe, ) . however, such safeguards may be moot if a highly competitive sterilizing strain is engineered and released. quarantine legislation designed to prevent accidental or intentional but illegal importation of unwanted organisms would be exercised by countries under current international obligations and should impede establishment in new countries if rigorously enforced. however, current legal safeguards may be insufficient. new zealand's experience with rcd indicates it is possible for lay people to illegally import and establish reproducing populations of exotic pathogens. in australia, rcd breached a carefully planned quarantine on an offshore island. unintended establishment and proliferation of engineered viruses may be contained if outbreaks are recognized early, and if proportions of susceptible individuals are removed rapidly from the population either by culling or by immunizing against the pathogen (tyndale-b iscoe, ) . this has never been tried with wild animal populations. the containment of contagious pathogens, such as foot and mouth disease in livestock, indicates such an approach may be possible. highly attenuated forms of myxoma virus are used to protect wild and domestic rabbits in france and the united states, indicating the availability of such technology for this virus at least (fenner & ross, ; tyndale-biscoe, ) . limited field trails with sterilizing microorgansisms are unlikely before (anderson, ). there is abundant evidence that introduced exotic vertebrates that establish feral reproducing populations have disastrous consequences for agriculture and preservation programs for native plants and animals. sources of current vertebrate introductions include sellers and buyers of exotic pets; acclimatization societies that import, establish, and relocate game animals and whose constituents include hunters and fishermen; and farmers and ranchers who import and experiment with novel lifestock (e.g., fitch farming). exotic vertebrates have in some instances great economic importance (as with lifestock and game animals), they also enjoy public popularity because of interest in hunting, fishing, eating, or viewing large and unusual animals in familiar environments. the negative ecological aspects of introduced vertebrates may be poorly understood by the public at large. such limited understanding may hinder control efforts and prevention of importation (bland & temple, ) . legislation has been passed in the united states to minimize risks of importing new and relocating existing vertebrate species. the lacey act passed in and ammended in was enacted to protect certain animals and endangered habitats, and to prevent introduction of noxious pests. under the act, violation of the law can result in fines and imprisonment [see usc w importation or shipment of injurious mammals, birds, fish (including mollusks and crustacea), amphibia, and reptiles; permits, specimens for museums; regulations m for more details]. similar legislation has been developed in new zealand. the biosecurity ( ) and hazardous substances and new organisms (hsno) ( ) acts were devised to protect the environment by preventing or managing the adverse effects of hazardous substances and exotic organisms. campbell ( ) points out that existing laws have many loopholes and are not effective when applied, indicating a need to improve existing regulations and to develop new laws to curtail unwanted entry by alien vertebrates. one proposal is to require importers of exotic organisms to develop "clean lists" and to prove that organisms are not potentially invasive and disrupting to native ecosystems (campbell, ) . legislative approaches limiting imports and exports of organisms may encounter complaints under the general agreement of tariffs and trade (gatt) that stricter quarantine measures are an unacceptable imposition of one country's environmental standards on others (camp-bell, ) . there is an obvious need for greater cooperation among interest groups, scientists, and legislators to devise solutions to problems associated with continuing introductions of exotic species and to provide direction for future action. as an evolutionarily stable control strategy development of resistance (behavioral or physiological) to pesticides (e.g., rodenticides) by vertebrates, and the need for repeated or multiple simultaneous control strategies (e.g., poisoning combined with trapping and hunting) indicate that control of vertebrates is an ongoing endeavor that attempts to reduce agricultural damage and losses (greaves, ) or to protect wilderness areas (cowan, ; morgan et al., ; payton et al., ) from pest damage. biological control has several advantages over chemical and cultural control practices (van driesche & bellows, ) : ( ) it is relatively cheap and biological control programs are often quicker to implement than to develop and to register new pesticides; ( ) use of carefully screened natural enemies increases selectivity of attack toward target pests; ( ) natural enemies in many instances are self-perpetuating and self-distributing; and ( ) development of resistance to natural enemies is extremely rare. one documented case of pests developing resistance to natural enemies is the development of resistance to myxomatosis by rabbits and corresponding attenuation of highly virulent strains of the myxoma virus to strains of intermediate virulence (fenner & ross, ) . the myxoma virusrabbit system in australia and europe is dynamic with increasing rabbit resistance selecting for more virulent strains of virus. this suggests that for the short-term, at least, the system is coupled in an antagonistic coevolutionary arms race (dwyer et al., ) . flexible natural-enemy behavior patterns and physiology have the potential to weaken evolutionary responses that can cause pest resistance to introduced control agents (holt & hochberg, ; jervis, ) . in comparison, pesticides and cultural controls tend to target a fixed physiological or behavioral function or pattern, and the resulting selection regime is constant allowing pests either to increase tolerance to poisons or to learn and develop avoidance behaviors (e.g., bait and trap shyness). spatial heterogeneity of natural-enemy attack limits selection pressure on hosts by natural enemies, thus reducing the rate of resistance development by pests compared with uniformly applied selection pressures such as pesticides. pests that escape attack move into enemy-free areas and continue breeding; thus, the rate of coevolution is reduced by susceptible pests in transient refuges (jervis, ) . at the metapopulation level, natural enemies may be ineffective selection agents because of widespread extinction and establishment of pest subpopulations that maintain pest susceptibility. additionally, resistance development may involve costs leading to a corresponding decrease in fitness. for example, increased tolerance to attack may reduce the pest's reproductive capacity and ability to compete for resources, or may increase susceptibility to other mortality agents (holt & hochberg, ) . there are opportunities to enhance biological control programs against vertebrate pests that cause social, agricultural, and conservation problems. in many instances, biological control offers the best chances for long-term control, particularly in isolated areas with rugged terrain, in suburban areas with high-density human populations, or in places where pests are nocturnal or secretive. biological control will not totally alleviate vertebrate pest problems. it may, however, reduce the vigor of pest populations, thereby reducing damage, minimizing nuisance value, or allowing native species to compete more effectively for food and breeding sites. programs could be initiated to simply reassociate host-specific micro-and macroparasites with pest populations that have depauperate natural-enemy faunas (dobson & may, ) , and there is no shortage of targets as small founding populations of vertebrates continue to invade and proliferate in new habitats. genetically engineered natural enemies are additional tools to aid biological control efforts. research with agents that cause immunocontraception will likely diversify as advances in molecular biology continue, and routes alternative to sterilization may be taken. this area of vertebrate biological control will be tested more thoroughly once small-scale and long-term field trials begin with sterilizing microorganisms. the effect of barn owls (tyto alba) on the activity and microhabitat selection of gerbillus allenbyi and g. pyramidum runaway rabbit virus kills millions alarm greets contraceptive virus regulation and stability of hostparasite population interactions i. regulatory processes parasite pathogenicity and the depression of host equilibria theoretical basis for the use of pathogens as biological control agents within-host population dynamics and the evolution and maintenance of microparasite virulence the cooperative research center for biological control of vertebrate pest populations rabbit virus vectors named can the nematode capillaria hepatica regulate abundance in wild house mice? results of enclosure experiments in southeastern australia predicting the effect of a novel vertebrate biocontrol agent: a model for viral-vectored immunocontraception of new zealand possums ecology of predator-prey and parasitoid-host systems: progress since nicholson modeling immunocontraception in disseminating systems mice on the farm biological pollution: the control and impact of invasive exotic species control of feral cats on sub-antarctic marion island, indian ocean population dynamics of parasites of the wild rabbit a single dose immunization with rabbit haemorrhagic disease virus major capsid protein produced in saccharomyces cerevisiae induces protection impact of botfly parasitism on microtus townsendii populations insular endemic plants lack defenses against herbivores a bait delivered immunocontracpetive vaccine for the european fox (vulpes vulpes) by the year ? the effect of the european wild boar sus scrofa, on gray beech forest in the great smoky mountains patterns, modes and extents of invasions by vertebrates response of the north island brown kiwi, apteryx australis mantelli and the lesser short-tailed bat, mystacina tuberculata to a measured dose of rabbit haemorrhagic disease virus conservation priorities in hawaiian natural systems biological pollution: the control and impcat of invasive exotic species invasion resistance, species build up and community collapse in metapopulation models with interspecies competition global patterns in the establishment and distribution of exotic birds effect of fertility control on a population's productivity european brown hare syndrome in the uk: a calicivirus related to but distinct from that of viral haemorrhagic disease in rabbits possible origin of rabbit haemorrhagic disease in the united kingdom the comparative epizootiology of capillaria hepatica (nematoda) in urban rodents from different habitats of effects of feral goats on island ecosystems. biological conservation field epidemiology of rabbit calicivirus disease in australia the european rabbit, the history and biology of a successful colonizer accumulation of native parasitoid species on introduced herbivores: a comparison of hosts as natives and hosts as invaders the eradication of introduced australian brushtail possums, trichosurus vulpecula, from kapiti island, a new zealand nature reserve possum biocontrol: prospects for fertility control australian and new zealand mammal species considered to be pests or problems. reproduction theory and practice of biological control the scope of biological control patterns of invasion by pathogens and parasites parasites, disease and the structure of ecological communities restoring island ecosystems: the potential of parasites to control introduced mammals population biology of trichostrongylus tenuis in the red grouse lagopus lagopus scoticus a simulation model of the population dynamics and evolution of myxomatosis virulence and local adaption of a horizontally transmitted parasite the evolution of parasitic diseases the use of cats in farm rat control the ecology of invasions by animals and plants which animal will invade attributes of invaders and the invading process: vertebrates parasitic and infectious diseases, epidemiology and ecology a comparison of the virulence for european rabbits (oryctolagus cuniculus) of strains of myxoma virus recovered in the field in australia the european rabbit, the history and biology of a successful colonizer the european rabbit, the history and biology of a successful colonizer the need to control cane toads comparative histopathological study of rabbit haemorrhagic disease (rhd) and european brown hare syndrome (ebhs) rabbit haemorrhagic disease in the united kingdom a propos d'un cas de capillariose a capillaria hepatica observe dans un elevage de lapins croises garenne the rabbit in new zealand the complete nucleotide sequence of rabbit haemorrhagic disease virus (czech stain c v ): use of the polymerase chain reaction to detect replication in australian vertebrates and analysis of viral population sequence variation resistance to anticoagulant rodenticides viral haemorrhagic disease or rabbits in mexico: epidemiology and viral characterization translocation and a species conservation tool: status and strategy when is biological control evolutionary stable (or is it analysis of vertebrate pest control biological control of vertebrate pests classical biocontrol: panacea or pandora's box? an evaluation of the biological control of the feral cat felis catus (linnaeus, ) and no birds sing: the story of ecological disaster in a tropical paradise rabbit and fox introductions in tierra del fuego: history and assessment of the attempts at biological control of the rabbit infestation parasitoids as limiting and selective factors: can biological control be evolutionary stable? biological control of possums long-term effects of porcine zonae pellucidae immunocontraception on ovarian function in feral horses (equus caballus) case studies in wildlife immunocontraception: wild and feral equids and white-tailed deer predation and population cycles of small mammals the natural regulation of animal numbers rabbit virus threatens ecology after leaping the fence capillaria hepatica infection: incidental finding in a dog with renal insufficiency european brown hare syndrome virus: molecular cloning and sequencing of the gonome a new viral disease in rabbits biological invasions: lessons for ecology comparative conservation biology of oceanic archipelagoes birth control for feral pests fluctuations of animal populations and a measure of community stability food webs: a plea for parasites thresholds and breakpoints in ecosystems with a multiplicity of stable states regulation and stability of hostparasite population interactions ii. destabilizing processes depression of host population abundance by direct life cycle macroparasites models to assess the potential of capillaria hepatica to control population outbreaks of house mice evaluation of a nematode (capillaria hepatica bancroft, ) as a control agent for populations of house mice (mus musculus domesticus schwartz and schwartz, ). revue scientifique et technique office international des epizooties quantifying the impact of disease on threatened species immunocontraception for wildlife population control detecting disease and parasite threats to endangered species and ecosystems vaccination with a synthetic zona pellucida peptide produces long-term contraception in female mice probable role of ground beetles in the transmission of capillaria hepatica why do possums survive aerial poisoning operations control of small mammals in a pine plantation (central chile) by modification of the habitat of predators (tyto alba, stringiforme and pseudalopex sp. canidae). acta oecologia pyloric stenosis in a yearling with an incidental finding of capillaria hepatica in the liver the european rabbit, the history and biology of a successful colonizer self assembly, antigenicity, and immunogenicity of the rabbit haemorrhagic disease virus (cezchoslovakia strain v- ) capsid protein expressed in baculovirus potential consequences and problems with wildlife contraceptives prolonged prey suppression by carnivores: predator-removal experiments the control of vertebrate pests by vertebrate predators phylogenetic analysis of rabbit haemorrhagic disease and european brown hare syndrome viruses by comparison of sequences from the capsid protein gene the social and economic implications of rhd introduction identification and characterization of the virus causing rabbit haemorrhagic disease ethical aspects and dilemmas of fertility control of unwanted wildlife: an animal welfarist's perspective schwerer leberbefall durch capillaria hepatica. monatsschrift fur kinderheilkunde purification and characterization of a calicivirus as the causative agent of lethal haemorrhagic disease in rabbits response of selected tree species to culling of introduced australian brushtail possums trichosurus vulpecula at waipoua forest limits to predator regulation of rabbits in australia: evidence from predator removal experiments predation models for primary and secondary prey species can foxes be controlled by reducing their fertility? successful treatment of capillaria hepatica infection in an acutely ill adult the snake that ate guam essentials of conservation biology a primer of conservation biology the effect of habitat on the helminth parasites of an island population of the polynesian rat (rattus exulans) testing the concept of virally vectored immunosterilization for the control of wild rabbit and fox populations in australia progress towards using a recombinant myxoma virus as a vector for fertility control in rabbits. reproduction, fertility and development the australian and new zealand calicivirus disease program the disappearance of guam's wildlife likely targets for immunocontraception in marsupials the establishment and spread of myxomatosis and its effect on rabbit populations escaped rabbit calicivirus highlights australia's chequered history of biological control a relationship between plagues of the house mouse mus musculus (rodentia: muridae) and prolonged periods of dry weather in south-eastern australia extinction of an island forest avifauna by an introduced snake regulation of mouse colony abundance by heligmosomoides polygyrus population dynamics in wild and laboratory rodents the role of parasites in regulating host abundance a harebrained scheme the potential of murine cytomegalovirus as a vital vector for immunocontraception. reproduction, fertility and reproduction immunological reactions and infertility can predators regulate small mammal populations? evidence from house mouse outbreaks in australia mammal populations: fluctuation, regulation, life history theory and their implications for conservation fertility control of mammal pests and the conservation of endangered marsupials effects of wild pig rooting in a deciduous forest the effects of capillaria hepatica (nematoda) on natality and survival to weaning in balb/c mice population dynamics of an outbreak of house mice (mus domesticus) in the mallee wheatlands of australia: hypothesis of plague formation the potential of capillaria hepatica to control mouse plagues the geographic distribution and host range of capillaria hepatica (bancroft) (nematoda) in australia an experimental field study to examine whether capillaria hepatica (nematoda) can limit house mouse populations in eastern australia a manipulative field experiment to examine the effect of capillaria hepatica (nematoda) on wild mouse populations in southern australia neither human nor natural: ethics and feral animals parasitic and infectious diseases, epidemiology and ecology sexually transmitted diseases in animals plant-derived immunocontraceptive vaccines food specificity in interspecies competition studies of the life cycle infectivity and clinical effects of capillaria hepatica (bancroft) (nematoda) in mice, mus musculus experimental embryonation and survival of eggs of capillaria hepatica (nematoda) under mouse burrow conditions in cereal growing soils the role of helminths in the biological control of mammals rabbit haemorrhagic disease virus: a calicivirus with differences fertility control in wildlife the crc for biological control of vertebrate pest populations: fertility control of wildlife for conservation virus-vectored immunocontraception of feral mammals vermin and viruses: risks and benefits of viral-vectored immunosterilization changing social views of the desired degree of host range specificity of natural enemies of arthropods effects of feline panleucopaenia on the population characteristics of feral cats on marion island correlates of introduction in exotic new zealand birds when biotas meet: understanding biotic interchange science biological invasions and ecosystem processes: towards an integration of population and ecosystem studies the extent of biological control of rats with barn owls, tyto alba javanica in malaysian oil palm plantations. the planter biological control of vertebrate pest effectiveness and cost-efficiency of control of the wild rabbit, oryctolagus cuniculus (l.), by combinations of poisoning, ripping, fumigation, and maintenance fumigation response of wild rabbit populations to imposed sterility development and use of virus-vectored immunocontraception the characters of successful invaders the varying success of invaders biological control of vertebrates--a review, and an assessment of prospects for malaysia observations of the lifecycle of capillaria hepatica (bancroft, ) with a description of the adult acknowledgments i thank vincent d'amico, iii of bean's art ink for preparing the states geological survey provided information on the brown tree snake. key: cord- - amhia authors: lidfors, lena; edström, therese; lindberg, lennart title: the welfare of laboratory rabbits date: journal: the welfare of laboratory animals doi: . / - - - - _ sha: doc_id: cord_uid: amhia nan rabbits were the fifth most commonly used mammalian laboratory animal after mice, rats, guinea pigs and pigs in sweden during (cfn . according to the latest statistics for the eu member states, rabbits were used during (commission of the european communities ). both domesticated rabbits and european wild rabbits may be used for experimental research, but there are several problems in keeping and breeding the european wild rabbit (bell ) . today the most common breeds used are the new zealand white (nzw), the dutch and the half lop (batchelor ) . most laboratories buy these breeds as health defined (previously called specific pathogen free) from accredited breeders (townsend , eveleigh et al. . rabbits are used for many different purposes with a large number being used for antibody production, but also for orthopaedics and biomaterials (batchelor ) . the rabbit is especially suitable for studies on reproduction (batchelor ) . rabbits are also used for cardiac surgery, and studies of hypertension, infectious diseases, virology, embryology, toxicology, experimental teratology (hartman ) , arteriosclerosis (clarkson et al. ) and serological genetics (cohen and tissot ) . laboratory rabbits are by tradition kept individually in small cages with restricted food availability. this has led to several physiological problems related to the fact that they move too little, as well as behavioural disorders. over the past - years many laboratories have improved the housing for rabbits, both in cages and also by introducing floor pens and group housing. however, there are still some aspects to be addressed in rabbit housing especially in relation to the fact that they can be very aggressive when kept in groups. the aim of this chapter is to present the most recent knowledge about the laboratory rabbit's biology, behavioural needs, optimal environment, housing, feeding, care, handling, health and experimental techniques, in order to ensure their optimum welfare. the laboratory rabbit originated from the european wild rabbit (oryctolagus cuniculus, figure - ) (harcourt-brown ) . rabbits were already kept in fenced hunting areas by the romans years ago, and domestication appears to have started around the year ad by the french. european wild rabbits were introduced to great britain by the normans soon after the conquest of (meredith ) . the domestication most probably occurred in the monasteries, where several different breeds were developed. figure - . european wild rabbits in a fenced grass area in england used for behavioural ecology research (photograph by lena lidfors ) during the first half of the th century many new rabbit breeds were developed, and today there are recognized breeds of fancy fur and rex rabbits (british rabbit council ) . the new zealand white (nzw) was accepted as a breed in according to american standards (batchelor ) . this rabbit has been bred for fast growth for meat production and their white pelts which can be dyed any colour, but it has also become a very commonly used laboratory breed. the negative impact of this breeding emphasis is that laboratory new zealand white rabbits relatively easily become obese, and food is often restricted in singly housed animals to prevent this. other rabbit breeds more suitable for laboratory research have been developed more recently, taking into account the purpose of a certain research area, for example the half lop, which has long ears suitable for repeated blood samples during anti-body production. the natural diet of rabbits consists of grass and herbs, but also fruit, roots, leaves and bark (cheeke ) . the rabbit needs coarse fibre, not just grass (brooks , meredith . the rabbit is caecophagous (often called coprophagous), i.e. it needs to ingest the smaller, soft and green coated faeces pellets produced by the caecum - hours after feed intake in order to survive (brooks ) . the rabbit takes the soft faecal pellets directly from its anus, whereas the harder pellets are placed on specific latrines close to the territory borders (donnelly ) . the rabbit has a light and fragile skeleton that only makes up - % of the body weight (donnelly ) . rabbits can adjust to different ambient temperatures, but have problems with too high temperatures and therefore seek shade from sun light in their burrows or under bushes. they have a º field of vision, but can not see the area beneath their mouth. they use their whiskers and the sensitivity of the lips as well as scent and taste during foraging (meredith ) . the whiskers are also used during orientation in the burrows and dens. scent and taste is more important in identifying members of the own breeding group than vision. they have good hearing, and their big ears occupy about % of the total body area (meredith ) . reproduction in the european wild rabbit is seasonal and appears to be determined by the interaction of a number of environmental factors, for example daylength, climate, nutrition, population density and social status webb , bell ) . sexual maturity occurs at the age of - months in males and - months in females, depending on climate (myers et al. ) . domestic rabbits are sexually mature at - months of age (falkmer and waller ) , but the small breeds are sexually mature earlier than the medium and large breeds (bennet ) . before mating the rabbits circle around each other, parade side by side, jump over each other and sniff around the genital region (lehmann ) . hafez ( ) found seven degrees of sex drive which ranged from aggressive with immediate mounting and ejaculation to offensive reaction with general smelling of the skin, biting and no ejaculation. the buck mating takes only seconds, after which he dismounts to the side or backwards (bennett ) . when allowing mating the doe raises the hind quarters. the ovulation is induced by the mating and the sperm is waiting until the egg appears after - hours, when conception takes place (bennett ) . the doe can mate immediately following parturition, and if the young are removed after delivery she will be sexually receptive for at least days (hagen ) . the doe is pregnant for between - days (bell ) and - days (batchelor ) . some days before the birth the doe digs an underground nest either within the main warren or in a separate breeding "stop" dug specifically for the purpose (bell ) . these are short, shallow burrows dug at a distance of - yards from the warren. high ranking females have been found to often give birth to their young in a special breeding chamber dug as an extension to the warren, whereas some of the subordinate females are chased away from a warren and forced to drop their litters in isolated breeding "stops" (mykytowycz ). the survival rate is much higher for the high ranking than the low ranking females. the doe collects and carries grass to her burrow and shortly before giving birth she plucks her own fur from the belly, sides and dewlap, and places it on top of the grass (gonzáles-mariscal et al. , bell . the better the construction of the nest the higher the survival rate of the young (canali et al. ) . the rabbit foetuses are born with no or only little hair cover, and are deaf and blind (batchelor ) . they each weigh around g at birth and gain about g per day (falkmer and waller ) . the size of the litter is - young for european wild rabbits, and - young for domesticated rabbits (batchelor ) . at birth the young rabbits are very sensitive to cold, but as their fur grows out within a couple of days they become less sensitive (batchelor ) . the eyes open at - days (kersten et al. , batchelor , or - days (falkmer and waller ) . hearing develops at the same time, but no reference on this has been found. when the young are born the doe leaves the burrow and covers the entrance with soil, urine marks it, and then leaves (mykytowycz ) . she returns to the burrow once a day, and then digs herself into the burrow and nurses her young (bell ) . in a study on dutch belted rabbits the nursing took place in the early morning and lasted for . - . minutes (zarrow et al. ) . when given the opportunity to retrieve their young, does did not perform this behaviour (ross et al. ) . the doe closes the burrow and or breeding "stop" for about days, after which the young emerges onto the surface (bell ) . the young are very mobile at four weeks of age, and leave their breeding stop soon after emergence (lloyd and mccowan ) . at that time they start seeking forage, but they continue to suckle for some more weeks. the does reach maximum milk production two weeks after giving birth, this declines during the fourth week and they may lactate for an additional - weeks. at weeks of age the young are consuming approximately % of their intake in the form of plant proteins (hagen ) . in commercial rabbit breeding the young are usually weaned and separated from the doe at - weeks of age (hagen , bennett . nzw does in semi-natural enclosures behave in a similar way towards their young as the wild rabbit, but when the young are about days old their mother no longer closes the entrance to the breeding stop properly and the young are nursed outside (lehmann ) . the mother-young relationship was quite loose, and the mothers were not preferred social partners for the young except for suckling attempts. nursing was unlikely after four weeks when the doe littered again within a few days, but suckling attempts occurred up to days (lehmann ). the behaviour of the european wild rabbit (bell , figure - ) and the behaviour of nzw rabbits kept in a free-range enclosure have been studied (lehmann , stauffacher . studies of natural, free-ranging and enclosed populations in australia, new zealand and united kingdom have shown that european wild rabbits live in small, stable, territorial breeding groups (parer , gibb et al. , cowan , bell and webb . the breeding group is generally described to defend its territory, a core area with a warren within a larger home range, by patrolling the borders and scent-marking (bell ) . however, in some studies it has been found that breeding groups only occupy single warrens (myers and schneider , bell ) , whereas in other studies of free-living populations of rabbits most breeding groups used multiple warrens (dunsmore , parer , wood , daly , cowan . during the peak feeding periods at dawn and dusk members of neighbouring breeding groups may move out from their territories to forage in communal grazing areas (bell ) . the number of entrances per warren was found to be . ± . sd, with a range of - (cowan ) . the different breeding groups together make colonies of up to rabbits. the social unit is the breeding group, which consists of - males and - females (meredith ) . within breeding groups separate, stable, linear dominance hierarchies are formed within each sex (bell ). the strict rank order is maintained by rabbits keeping a fixed distance from one another and exhibiting submissive behaviour. early studies of the european wild rabbit in australia revealed that a substantial portion of the rabbits' daily activity was filled with direct or indirect aggression rowley , mykytowycz and fullager ) . it was concluded that this behaviour underlies the social and territorial organisation, which are the main factors affecting the numbers of free-living populations of this species (mykytowycz ) . in some studies does have been found to be more aggressive than bucks (southern , myers and poole ) , or found to fight as strongly as males (lockley ) . females mainly fight over breeding burrows (cowan and garson ) , and are more aggressive towards juvenile females ( % of interactions) than juvenile males (cowan ) . males are tolerant to females, young and sub-adults in their breeding group, and have even been observed to interrupt aggressive interactions between females. when the number of rabbits living together increased, fighting increased dramatically (myers ) . in nature young males normally move to a new social group before the start of their first breeding season, while young females stay on to breed in their breeding group (parer , webb et al. . lehmann ( ) has described different social behaviours of new zealand white rabbits kept in a semi-natural enclosure, and how the social behaviours change with age. indifferent contacts were body-to-body, noseto-body, nose-to-nose contacts and anogenital nuzzling. amicable behaviour comprised cuddling up and allogrooming. subdominant behaviour was crouching, retreating and fleeing, whereas aggressive behaviour was circling, nudging, attacking and chasing. actual fights with aggressive leaping and ripping were observed only times. mykytowycz and hesterman ( ) made paired encounters between male and female european wild rabbits and new zealand white rabbits in a home pen, and found that aggression was equally prevalent in both sexes and inter-sexual fighting occurred just as frequently as fighting between members of the same sex. the domesticated nzw rabbits were fighting just as much as the wild rabbits, but paired domestic females and domestic males paired with wild rabbits of both sexes fought less frequently and less viciously than other paired rabbits. rabbits have three specialised scent glands, i.e. in the anal region, in the groin and under the chin (mykytowycz ). the territory of rabbits is scent marked by placing faeces in dunghills, and about such dunghills have been located in a typical warren. the rabbit also marks its territory by pressing the under-chin against structures in its environment so that droplets from its sub-mandibular glands are forced through pores of the skin. scent marking is more intensive by males than females and more in dominant than sub-dominant individuals, and it is correlated with larger anal and submandibular glands in dominant males (mykytowycz ). the most dominant animals of a warren were found to possess the heaviest anal glands. males scent mark females and young rabbits of their breeding group by spraying urine on them. females scent mark their young, attack other young within the same breeding group, and may chase and even kill young from other breeding groups. females may attack even their own young if they have been smeared with foreign urine (mykytowycz ) . the rabbit has a home-range for performing foraging behaviour. the size of the home-range varies depending on food availability, age of the rabbit, the rabbit's status within the breeding group and number of rabbits in the group (donnelly ) . the home-range of wild european rabbits has been estimated to be ha (myers et al. ) , . - . ha (cowan and bell ) , or ha (vastrade ) . cowan ( ) found that that the mean asymptotic male range was . ha and female range . ha. the rabbit is mainly a nocturnal animal, emerging from the burrows in late afternoon (fraser ) . the old bucks emerge first, about four hours before sunset (mykytowycz and rowley ) , and by sunset % of the rabbits have emerged (fraser ) . the rabbits are visible on the ground during - hours of the diurnal cycle (mykytowycz and rowley ) . when the rabbits are above ground they spend about % of their time eating, % inactive, % moving and % on other activities (gibb ) . during semi-natural conditions young nzw rabbits were active for an average of % of the daytime. feeding on pellets and grazing took a third each of the active time, and the remaining third was spent exploring, gnawing, intensive locomotion and, for the older rabbits, engaging in sexual behaviour (lehmann ) . the rabbit's choice of habitat depends on the opportunities to find shelter and protection, and where the soil is loose it digs burrows, and where the soil is more compact it seeks protection in dense vegetation (kolb ) . in case of danger a rabbit can stamp with its hind feet, thus causing the other rabbits to flee underground (black and vanderwolf ). if it is too late to flee the rabbit can freeze, i.e. stop in its movement and be completely motionless. a rabbit can, if caught by a predator, emit a high distress scream (cowan and bell ) , which may cause the predator to release its prey. apart from this and some low sounds during mating and mother-young care rabbits are silent animals. the movement patterns of rabbits consist of hopping, crawling and intensive locomotion, i.e. running, start-and-stop, jumping, double and capriole (kraft , lehmann . hopping is usually performed when rabbits move over longer distances, whereas crawling is performed when feeding on grass or exploring on the spot and during social encounters (lehmann ) . rabbits perform comfort behaviours such as licking and scratching themselves, shaking the body, rubbing against objects and stretching their body. several studies have compared the behaviour of wild european rabbits with those of domesticated strains of rabbits (reviewed by bell ) . both wild rabbits and domestic strains have reproductive seasonality, suckle their young only once every h, show two main feeding periods at dawn and dusk, form breeding groups with separate linear dominance hierarchies among male and female members, and reproduce successfully with the female digging breeding burrows, building nests for their young and covering the entrance to the burrows with soil (bell ) . the difference found is that domestic rabbits rest more above ground during the day (stodart and myers ) and that males chin-mark more often (kraft ) also in unfamiliar territory (mykytowycz ). new zealand white females had fewer days in anoestrus than wild rabbits, and therefore produced more litters with a larger mean size (stodart and myers ) . when european wild rabbits were brought into the laboratory they failed to breed, and females born into the laboratory as a result of egg transfer from wild to domestic mothers retained their nervous disposition and failed to mature sexually (adams ). when rabbits are kept alone in traditional small cages several abnormal behaviours may arise. the most obvious abnormal behaviour is stereotypy, e.g. wire-gnawing, excessive wall-pawing (lehmann and wieser , wieser , bigler and lehmann , loeffler et al. , stauffacher . when many rabbits gnaw on the wire of their cages there might be a high sound level in the animal room ( figure - ). digging may be constrained by the solid floor of the cage (podberscek et al. ) . the stereotypic behaviour substitutes for natural behaviours which cannot be performed in standard laboratory housing (stauffacher ) and may indicate frustration, anxiety or boredom, and develop in stages involving a progressive narrowing of the behavioural repertoire (gunn ) . individually caged rabbits show stereotypies, such as somersaulting, no full hops, less activity than group-penned rabbits, and less marking and investigatory behaviour than in group pens (podberscek et al. ) . social isolation can induce the physiological symptoms of stress, which are relieved by the presence of conspecifics (held et al. ) . there are indicators of boredom in rabbits such as hunched posture (gunn and morton ) , inertia (metz ) , and a staring coat and dull eyes (wallace et al. ). prolonged inactivity associated with unresponsiveness, may, like stereotypies, be associated with brain chemistry changes which make the problem seem less bad (broom ). gunn and morton ( ) mention problems with under-grooming which may lead to a staring coat, and over-grooming which may lead to hair-balls which in turn may cause intestinal stasis (jackson ) and lead to death by gastric trichobezoars (wagner et al. ) . under-eating, causing weight loss and over-eating causing obesity are other behavioural problems (gunn and morton ) . the very limited freedom of movement in caged rabbits leads to changes in locomotion which prevent hopping, thus causing changes in the locomotion apparatus (lehmann , stauffacher . it is mainly in the femur proximalis and the vertebral column that changes of the bone structure occur, and the changes consist of thinner and less strong hollow bones (lehmann , drescher and loeffler a,b) . lehmann ( ) showed that growing rabbits perform almost no hopping and intensive locomotion mainly during playing when kept in cages compared to an outdoor enclosure. crawling, which was the most common movement pattern, was slightly less common in the cages. these rabbits often performed interrupted jumps, where the hindlegs only were lifted slightly and then put down again, thus not being used as in normal hopping. abnormal postures may also be shown due to spatial restriction for lying stretched out during resting or performing stretching behaviour (gunn ) . rabbits may show restlessness, e.g. afunctional bouts of activity with disconnected elements of feeding, comfort, resting, alertness and withdrawal behaviour alternating with locomotion, which leads to space-time organization disorder as well as panic (lehmann and wieser , bigler and lehmann , stauffacher ). an abnormal behaviour related to reproduction is disturbed sexual behaviour which leads to low conception rates ( - %) (stauffacher ) . one reason for this may be that for mating the doe is placed in the bucks' cage a few days to several weeks after she last gave birth. the mating performed then has been described more as a rape than normal mating behaviour (stauffacher ) . however, the doe may be aggressive against the buck if he is placed in her cage (bennett ) . around giving birth the doe may show disturbed nesting behaviour and nesting stereotypies which may lead to rearing losses (wieser , wullschleger , loeffler et al. . the doe may also show disturbed nursing and cannibalism caused by restlessness of the mother and her pups which may also lead to rearing losses (bigler , brummer , stauffacher ). based on the previously presented research on the abnormal behaviours performed by housed rabbits, the behaviour of wild european rabbits and free-ranging new zealand white rabbits many researchers involved in this area have had an insight to the behavioural needs of rabbits. based on this research and on several discussions in international workshops suggestions have been made on the following behavioural needs and behaviours involved in performing them: ) locomotion and exercise: crawling, hopping, running, jumping, quick changes of direction. ) control, "security and safety": withdrawal, digging, burrowing, sit upright, rearing, sniffing, "stamping". ) foraging: eating, drinking, caecophagi, gnawing, manipulation of resources, searching, exploration, exploitation of resources. ) behavioural rhythm: extended periods of rest and grooming, activity. ) companionship and social contact: different social behaviours. ) demonstrate presence; scent marking by anal and submandibular glands. ) female choice in mating; circling, parading, jump over and sniff genital region before mating. ) nest and burrow with constant temperature: digging, carrying nesting material, plucking fur. ) separate from litter: nursing once/twice a day, close nest entrance. ) warmth and protection: huddle in hair-nest for new-born rabbits. ) learn how to cope: play and exploration in young rabbits. as some of the suggested needs lead to both physical and behavioural problems if they are not performed, e.g. foraging, movements, other are more questionable, for example digging. there is a need for more research to establish exactly which behavioural needs are so important that one must provide them in the rabbit housing, i.e. essential needs. laboratory rabbits are recommended to be kept in animal rooms with a regular light:dark cycle and isolated from external lighting fluctuations (batchelor ) . some laboratory facilities have introduced artificial dawn and dusk periods, usually working for minutes before full light comes on in the morning and minutes before the light is shut down in the evening. however, there is still some debate as to the relative merit of creating an artificial dawn and dusk period (batchelor ) . as rabbits are more nocturnal than diurnal a partially reversed lighting schedule can be established to observe their activity (batchelor ) , but rabbits may also switch to a more diurnal pattern of activity due to noise or scheduled feeding in the laboratory (jilge ) . the optimal light intensity in rabbit rooms is recommended to be lux at one metre above the floor (iwarsson et al. ) . too high levels of illumination can result in retinal degeneration in some albino mammals, and this may include nzw rabbits (batchelor ) . solid side caging will reduce the amount of light, and where in the rack a cage is situated may also have an impact on light intensity. if rabbits are housed in floor pens, shelves and boxes may provide hiding places also from high light intensity. the rabbits hearing range thresh-hold level is - hertz with the best hearing at to about hertz (iwarsson et al. ). the rabbit is sensitive to high sound frequencies (milligan et al. ) , but also sudden noise may scare rabbits and lead to injures. background music masks sudden loud sounds, and is claimed by some to result in lower excitability (batchelor ). the mean room temperature for rabbits is recommended to be ºc with a range of - ºc (gv-solas ) . the lower critical temperature is - ºc and the higher critical temperature is . ºc (spector ). rabbits only have sweat glands on their lips, and lower capacity to ventilate through their mouth than dogs (donnelly ) . wild rabbits avoid high temperature and stay away from direct sunshine, and during the day they stay in the cooler burrows. the ears of the rabbit are highly vascular and function as ventilators during high ambient temperature. in the rabbit room a humidity level of ± % is recommended (batchelor ). in the animal room a ventilation rate of - air changes per hour is recommended (iwarsson et al. ) . air changes lower than this may be satisfactory if the cleaning routines is of high standard and the stocking density is low (batchelor ) . ammonia level should never exceed ppm. when measuring ammonia level in a modified farm animal building with group housed rabbits the highest level was - ppm (batchelor ) . high ammonium levels can inactivate the cilia in the air ways of rabbits. high levels of co may become a problem for the rabbits if the ventilation is not working properly. due to normal changes of the fur - times per year rabbits loose hair in the laboratory unit. the hair may fly around in the animal room, and eventually ends on the floor. especially during handling of the rabbits hair may be released. rabbits kept in cages should be moved to a clean cage once a week, but the waste pan under the perforated floor could be changed more often. if rabbits are kept on wire net floor and a band can be moved under the cages this should be cleaned once per day. many laboratories today have dishwashers where the whole rack is washed in warm water with disinfectants and dried at high temperatures thus working as an autoclave. the feed is usually pseudo-pasteurized ( ºc for minutes) at the producers. in order to reduce the risk of bringing pathogens into the laboratory, hay and any environmental enrichment should be autoclaved or irradiated with cobalt, but the latter is not so common. rabbits kept in floor pens should be taken out of the pen after which the pens should be thoroughly cleaned and disinfected. batchelor ( ) suggest that this should be done at least once per month. however, the laboratories we know of carry out this cleaning at least once per week and more often when needed in order to maintain a high level of hygiene. if cleaning is not done often the rabbits can become infected with coccidiosis. cleaning changes the olfactory environment for the territorial rabbits and may actually be stressful for them (batchelor ) . the rabbits should be in good condition before transport. usually rabbits are transported in containers made of strong cardboard, fibreboard, fibreglass or wood with wire mesh windows for ventilation, and equipped with a filter in summer time (swallow , batchelor . the floor of the container should be waterproof and covered with absorbent material, for example saw dust. for more detailed information about the general requirements and species requirements when transporting animals see swallow ( ) . regulations for transport of animals may differ for the eu (according to the convention of europe) and for different countries over the world. in england and sweden, when transporting rabbits of less than . kg the smallest area per animal should be cm² in non-filtered crates (winter) and cm² in filtered crates (summer) (swallow , sjvfs . in sweden minimum container height should be cm, and in england it should be cm. in the guide to the care and use of experimental animals from the canadian council of animal care (olfert et al. ) it is stated that the rabbits should be transported in disposable containers with sufficient space to allow the animals to stand, lie down and turn around. transporting rabbits in compatible pairs, for example siblings, has been practised by a breeder in sweden with good results. the transportation vehicles should be equipped with thermometers and ventilation that can cool down the air during warm weather and provide heating during cold weather. a source of water, for example a gel, and feed, for example a carrot, an apple or straw, sufficient for the duration of the transportation has to be provided (batchelor , olfert et al. ). swallow ( ) suggests that rabbits should not require feeding for journeys of less than h duration, but if unforeseen delays occur rabbits could be given carrots, fruit, hay or grain. after arrival at the new animal housing the rabbits should be checked for any health problems and injuries, which could either be caused by the transport or which may have been acquired before the transport. the rabbits should be given - weeks of acclimatization time after the transport before they are used for research. this will depend on the degree of stress the animals were under at arrival, which in turn may depend on the nature of the journey, its duration and which parameter is being assessed during the recovery period (swallow ) . olfert et al. ( ) state that newly acquired rabbits should be held in quarantine for at least three weeks, and examined regularly for disease. animals dying during this period should be subjected to complete post-mortem examinations. most laboratories we have contact with bring rabbits from suppliers of health defined rabbits bred behind a barrier. often most of the rabbits come from the same supplier, thus there is no need for quarantine. however, when purchasing rabbits from different suppliers we recommend placing them in different animal rooms at least during an acclimatization period of up to three weeks. there has been very limited research on the effect of transport on rabbits. batchelor ( ) found considerable differences in body weight in rabbits that were housed in group pens compared to solitary in cages after transportation. the loss of weight in rabbits after transport is mostly due to a loss of gastrointestinal contents in of about % of the total body weight (swallow ) . these losses are probably maximal after about h of transport, and similar to depriving animals of food and water during the same amount of time. however, the loss in live weight can take up to days to recover (swallow ) . our own research on the effect of providing male rabbits with or without hay and with a change or no change in the feed after a h transport by truck, plane and truck showed that providing hay had a significant effect of reducing the occurrence of diarrhoea (lidfors pers. com.). laboratory rabbits are traditionally housed singly in barren cages over periods of several weeks up to several years, depending on the research purpose. the cages for laboratory rabbits have changed from traditional wooden hutches to galvanized iron, aluminium, stainless steel and now plastic caging (morton et al., ) . today three main types of cages can be found; wire, sheet metal with wire front and plastic with wire front (stauffacher et al. ) . housing in small barren cages can lead to several abnormal behaviours and reduced welfare of rabbits (see section on abnormal behaviours). the cages with solid sides, back and top tend to isolate the animals, and prevent them from seeing the source of disturbance. this may cause the rabbits to be jumpy, and in breeding units lead to higher losses due to cannibalism. in order to allow rabbits to see their surroundings it is recommended that barred "windows" occupy - % of the total wall area (stauffacher et al. ) . in order to allow rabbits to perform normal hopping movements and sit upright the regulations in several countries (switzerland, united kingdom) require that cages for rabbits are much bigger than the recommendations from the european union and the world rabbit science association (table - ). the heights of the cages should be from cm for rabbits < kg up to cm for rabbits > kg in the swiss ordinance of animal protection (stauffacher et al. ). in the suggestions for new rules in the appendix a from the council of europe it is written that cages and pens for rabbits > weeks of age should be cm² ( cm height) for < kg rabbit, cm² ( cm height) for - kg rabbit and cm² ( cm height) for > kg rabbit. the weights are for the final body weight that any rabbit will reach in the housing. the floor area is minimum floor area for one or two socially harmonious animals, and the height is a minimum height. it is also stated that a raised area, i.e. shelf, must be provided within the cage, and if not providing a shelf the cage size must be % larger for a single rabbit and % larger for two rabbits. the suggested new rules in sweden have been agreed since , but so far no decision has been taken by the agricultural board. during the last years there has been a development of new cage systems (figure - ) , using a cage with an increased floor area, a higher cage for upright sitting, a shelf for rabbits to hop up onto or hide under, racks to make hay feeding easier and flexible cage racks so that several cages can be built together. a raised shelf reduces restlessness, grooming, bargnawing, and timidity of being captured (berthelsen and hansen ) . wild rabbits rear when they are looking at their surroundings (lockley ) , and caged rabbits use boxes and shelves provided as sources of enrichment and lockout posts (hansen and berthelsen ) . mirrors as a form of enrichment do not provide an image of a companion, but probably stimulate activity by enriching the visual environment and increasing the amount of movement perceived by the rabbit (jones and phillips submitted). if rabbits are housed in a cage with a living area and darkened resting area, putting mirrors in the living area increases the time rabbits spend there, and especially the time that they spend investigating their environment and feeding (jones and phillips submitted). rabbits can either be group housed in floor pens or in cages. in the latter case the limited area of a cage usually restrict this to pair-housing (bigler and oester , huls et al. , stauffacher . this combines the benefits of cage housing, e.g. hygiene, experimental purposes, with animal welfare interest, and has been established in switzerland, germany, united kingdom and canada (stauffacher et al. ) . group housing of rabbits in floor pens has been introduced in many laboratories and countries during the last - years (figure - ) . this is beneficial for the rabbits because they can express social behaviours and exercise stott , batchelor ) . abnormal behaviours and physiological conditions caused by small barren environments are also reduced, but there is an increased risk of fighting between rabbits (morton et al. ) . the major factors that have to be considered when group housing rabbits are: compatibility of individual animals, size of pens, stocking density, husbandry practices and environmental enrichment (morton et al. ) . rabbits which are not compatible will fight when placed together in a group, and the greatest problems occur when placing adult males together (morton et al. ). there are strain differences in aggressiveness, i.e. dutch rabbits are more aggressive than new zealand white, whereas lops are more docile (morton et al. ) . some of the small strains of rabbits may show more aggression than the larger strains (stauffacher et al. ) . individual animals may be highly aggressive, and fights can occur for unknown reasons, even in groups that have been stable for a long time (morton et al. ) . therefore, groups of rabbits need to be carefully selected and regularly monitored. the best option is to keep litter mates which have been kept together from weaning (zain ) . groups of intact, mature females not intended for breeding can be kept together (morton et al. ) . in periods of rest, does, bucks, and older young kept in mixed groups congregate and snuggle against each other or engage in mutual grooming (stauffacher ) . does have showed a weak preference for a large, enriched, solitary pen over a group pen, but a strong preference for a group pen over a smaller, barren, solitary pen (held et al. ) . fighting can still occur in groups of does, and a dominant female in oestrus can mount and damage the skin on the backs of other females and harass the group (morton et al. ) . the degree of compatibility of grouped rabbits will depend on factors such as strain, individual characteristics, sex, age and weight, size and structuring of pens, methods of husbandry and the interest and ability of the animal technicians (bell and bray , zain , morton et al. , stauffacher . aggression may be seen in group pens even after the establishment of the dominance hierarchy. from around weeks of age it may be necessary to house males individually to avoid fighting (morton et al. ) . groups of males kept in proximity of females tend to fight and urinate more frequently (portsmouth ) . castration of males kept for longer periods in the laboratory may be one solution to be able to keep them in groups. the practical experience from castration of males is that aggression is reduced and stable for a long time afterwards (gunn pers. com. in morton et al. , lindberg pers. com. . castration should be carried out when the males reach sexual maturity at - days of age depending on strain and food composition (stauffacher et al. ) . it should be carried out only by well-trained persons and always before the males start to show aggressive behaviour, - weeks after weaning at the latest (stauffacher et al. , morton et al. . the testicles move down during sexual maturation, but then are withdrawn again. however, one has to consider what type of research the animals will be used for, as castration has an effect on the animal's physiology. an alternative if castration is not advised is to place individually caged male rabbits in an exercise arena at regular intervals ( figure - ) . this allows them to move around on a larger floor surface, to investigate enrichment objects and to get the smell from other males that have been exercised before them. this has not been evaluated in the laboratory units, maybe because it will take time for the animal keepers to take out and return the rabbits according to a routine schedule. however, in the case where castration is not possible it may introduce an important enrichment for the rabbits. rabbits placed in group housing should be of the same sex, of similar size and if possible, related and grouped when young, i.e. around the time of weaning (morton et al. ) . when establishing new groups of rabbits in floor pens the best option is to wean and mix at the same time around weeks of age and to place - rabbits, preferably of the same sex, in one group (morton et al. ) . the ideal situation is when animals are kept in stable groups from birth (zain , stauffacher . small groups may be most stable (love and hammond ) . some animals do not appear to settle well in groups, either because they are too dominant and bully the others or are too timid and prone to be bullied (morton et al. ) . when putting together rabbits that have been caged for six months or more it may be difficult to avoid fighting or self-inflicted injuries (morton et al. ). however, individually-caged adult female rabbits of peaceful strains can be paired successfully, preferably in structured cages (stauffacher , morton pers. com. in stauffacher et al. . it is very important to provide refuge and hiding places for subordinate animals (morton et al. ). the number of rabbits kept in each group pen should not exceed - mature animals (morton et al. , stauffacher et al. . this is recommended so that the rabbits can be adequately monitored for signs of bullying and ill-health. groups of up to laboratory rabbits have been successfully managed as stock and for the production of polyclonal antibodies (stauffacher et al. ). however, more research is needed on optimum and maximum group size (stauffacher et al. ) . when keeping breeding rabbits in groups they should be composed of - females, one male and their offspring until they are weaned at days of age (stauffacher et al. ) . the group breeding housing system and management has been developed in switzerland (stauffacher ) and used in agricultural rabbit farming, but to our knowledge not being used for breeding of laboratory rabbits. the minimum area of a floor pen should be large enough for the rabbits of a particular weight and size to be able to carry out normal behaviour, especially locomotion. one problem with using weight and size as a criterion of determining floor area is that young animals move more and might need more space to carry out play behaviours (stauffacher et al. ). in the swiss legislation each rabbit must be able to hop some steps or to jump up and down onto a shelf. this may help the rabbits to maintain a level of fitness and reduce the occurrence of disuse osteoporosis (morton et al. ). the working group on "refinement in rabbit husbandry" recommend that rabbits kept in groups have a clear area of cm² with an overall minimum floor area of - cm² per rabbit for groups up to rabbits. if more than rabbits are kept in the groups an extra space of cm² per rabbit is recommended (morton et al. ) . the height of floor pens should be . m., as rabbits can jump very high, and enrichment objects should be placed so that they can not be used for jumping over walls. if european wild rabbits are used for research they should be housed in floor pens with sand or straw substrate so that they can hop around and exercise (bell ) . the reason for this is that wild rabbits kept in small cages have developed weak backs probably due to a lack of exercise. young wild rabbits, between and weeks of age, fare better when they are kept in groups (bell ). rabbits are crepuscular and in the wild they usually graze during their active periods at dawn and dusk (lockley ) , or during early morning and at night (cheeke ) . however, feeding laboratory rabbits in cages means that they are almost invariably not fed the diet of grass for which they evolved to utilize. however, there is little evidence that they prefer a grass diet to one based on compound feed (leslie et al. ) . despite this, it is often beneficial to supplement their ration of proprietary compound pellet with dietary enrichment, which as well as providing adequate nutrients (nrc ) , in particular fibre (lehmann ), which will increase the time usefully spent in procuring their food and reduce potentially damaging behaviours such as chewing their cage (leslie et al. ). the visual stimulus of a varied diet is particularly important (ruckebusch et al. ) . a mixed diet is also a feature of natural herbivore feeding behaviour, due to their desire to sample regularly in case of the disappearance of one feed (parsons et al ) . several forms of dietary enrichments have been tried with a significant degree of success, including supplying fibrous food to reduce boredom: hay (berthelsen and hansen ) , grass cubes or hay in a bottle (lidfors ) , and fresh grass (leslie et al. ) . the most useful enrichment is something to chew (brummer , huls et al. , lidfors , berthelson and hansen and high fibre objects are preferred, with hay or straw remaining an effective enrichment for long periods (brummer , lidfors . hay and straw also cause less weight gain than proprietary fibre sticks or compressed grass cubes (lidfors ) . abnormal maternal behaviours and trichophagia or fur-chewing (brummer ) are eliminated in caged rabbits when hay or straw is given. if the supplementary hay is ground, it is ineffective at reducing problem behaviours, demonstrating a need for long fibre . water should always be available ad libitum to rabbits (mader ). the need for water is - g per kg body weight, and rabbits raised on pellets need g per kg body weight. more water is needed for growing animals and pregnant and lactating females. one rule of thumb is that the need for water per h is % of the body weight (meredith ) . it is important that all personnel that handle rabbits have learned how rabbits react to frightening sounds and handling in order to avoid injuries to both the rabbit and human handler. rabbits that have had positive contact with humans and been handled with care previously will come to the front of the cage when opening the door and sit still when being lifted out. similarly rabbits in floor housing will sit still when a person comes to pick them up. rabbits that have been scared by humans or not been handled try to flee, and may injure themselves. the fear and distress may be communicated to other rabbits in the animal room . rabbits should be handled firm and gently, because if they sense insecurity they may struggle (batchelor ) . lifting of rabbits of all ages should be by grasping the scruff of the neck in one hand while the other hand is placed under the rump to support the animal's weight (batchelor ) . some authorities suggest that the rabbits ears should be grasped in addition to the scruff for greater control, but batchelor ( ) have the experience that some rabbits react as if this was painful, which increase the likelihood of struggling in the rabbit. rabbits should never be carried by the ears. during the last thirty years the health situation of laboratory rabbits in general has moved from a position of disease problems that is still prevalent among many pet rabbits to a vastly improved level of health. the laboratory rabbits of today are bred, housed and cared for in such a manner that their overall health situation is much better than that of their ancestors a couple of decades ago. the awareness among researchers and laboratory animal staff of the great importance of a good health situation for the animals has contributed to improvements in hygiene and in health monitoring systems, which in turn have led to a good general health situation among laboratory rabbits. the establishment of spf breeding colonies in the late s and early s gave the rabbits an immensely improved health situation (eveleigh et al. ) . the first guidelines of felasa (federation of european laboratory animal associations) for health monitoring of breeding colonies on a regular basis were also a milestone in the improvement of health for laboratory animals. at this time, in the mid- s, health discussions mainly dealt with hygienic aspects of health such as infectious diseases of different sorts and how the animals and the research results were affected by the infections. clinically healthy rabbits have a well-groomed fur, alert and clear eyes, and quick reflexes of escape if threatened. when picked up and held by the caretaker a healthy rabbit accustomed to handling by human beings will give the impression of strong muscles in a resting state. when checking the health of laboratory rabbits on a daily basis the major points are the posture of the animal, look of eyes, ears and nose for signs of discharge and other abnormalities, the state of the fur (especially in group-housed rabbits since the effects of fighting might only show as a minor flaw in the fur), and the look of faeces and urine. normal faeces should consist of dry fecal pellets of a uniform size and normal urine can vary in colour from yellow to dark red and is often cloudy due to excretion of calcium in the urine. see meredith ( ) for more information. felasa has issued guidelines (nicklas et al. ) on how to monitor the health status of laboratory rabbits and rodents. these guidelines comprise infectious agents, frequency of sampling, sample sizes and preferred methods of analysis for complete monitoring of the hygienic state of a breeding colony. the result of following the felasa guidelines for health monitoring of laboratory rabbits and rodents can be called a health defined rabbit. there are similar terms of earlier origin that are sometimes used for the same purpose, i.e. spf (specific pathogen free) or vaf (virus antibody free). these different terms aim to give us information of the quality of health of the rabbits microbiologically. a special health problem has been introduced along with the use of certain strains of rabbits with a genetically transmitted disease trait such as the whhl (watanabe heritable hyperlipidemic rabbit). this rabbit is characterized by the development of atherosclerotic lesions in arteries. older rabbits tend to accumulate calcium deposits in various parts of the body, which of course affects their well-being. many diseases will not show clinical symptoms unless the rabbits are compromised by a number of different stressors at the same time, where all factors influencing the animals well-being positively and negatively add up (nerem ). bacterial, viral or parasitic agents cause infectious diseases that rabbits can be affected by. many of the infections of laboratory rabbits are subclinical, especially viral infections and may pass unnoticed, mainly affecting the results of studies performed. in the literature, there are plenty of data about diseases of the rabbits, the literature used here include: harkness and wagner , laber-laird et al. , and hillyer and quesenberry bacterial agents including pasteurella multocida and bordetella bronchiseptica are the main cause of respiratory inflammations, which may give symptoms such as sneezing, coughing, nasal discharge and lethargy. pasteurella multocida and staphylococcus aureus may also be involved in formation of abscesses in subcutaneous tissues, behind the eye bulb or in internal organs as well as inflammation of the mucus membranes of the eyes and in the middle ear. other bacterial infections of rabbits are eye infections by moraxella catarrhalis. infections of the gut by bacteria include mucoid enteritis that affects mainly young rabbits. the symptoms are depression, anorexia, diarrhoea and mucus in the stool and the cause is multifactorial with the bacterium clostridium spiroforme being one of the major factors. escherichia coli and other strains of clostridia may also cause enteritis with diarrhoea as main symptom. inflammations of the gut are aggravated by nutritional imbalance with a deficit of dietary fibres. viral infections comprise mainly viruses that affect the digestive tract. rotavirus and rabbit enteric coronavirus may give the rabbits mild diarrhoea. rabbit viral hemorrhagic disease (rvhd) is a feared disease among pet rabbits but it is unlikely to infect laboratory rabbits. this disease affects many organs and the main symptoms are lethargy, anorexia, diarrhoea and haemorrhage from body openings such as the nose and urogenital opening. myxoma virus could be transferred to laboratory rabbits but this requires the presence of a vector, most often fleas but other insects may also act as vectors for myxoma virus. myxomatosis is common in wild rabbits and could possibly infect laboratory rabbits in areas where wild rabbits are common. parasites that infect laboratory rabbits are mainly endoparasites. the largest problems are created by coccidiosis. this disease is caused by different strains of eimeria. one strain, e. stiedae, infects the liver and causes different degrees of symptoms ranging from unapparent retardation of growth to fatal disease. other eimeria strains such as e. perforans and e. magna infect the intestine of rabbits. symptoms depend on the amount of coccidia present in the gut and of the susceptibility of the rabbit. most often, only the youngest animals show symptoms whereas in older rabbits coccidiosis is sub-clinical. the symptoms include weight loss and mild intermittent to severe diarrhoea. pinworms, passalurus ambiguus, colonize the caecum and colon and the eggs are passed in the faeces. pinworm infections are generally without symptoms. encephalitozoonosis or nosematosis is a disease common in pet rabbits and wild rabbit and regularly occurs in laboratory rabbits. it is caused by encephalitozoon cuniculi, an intracellular protozoan that is transmitted from the urine of infected animals via the oral route to the intestine and tissues of susceptible rabbits. the parasite mainly damages the kidneys and the brain of infected animals but most often no symptoms are seen clinically. ectoparasites are uncommon in laboratory rabbits but a few should be mentioned since they are common in other rabbits. ear mites, psoroptes cuniculi, can cause considerable itching and wounds on and around the ears. cheyletiella parasitivorax is the fur mite of rabbits. mites, fleas and lice of rabbits cause considerable suffering since they produce anaemia and/or pruritus and result in generally poor condition in the rabbit. fighting is the most common cause of traumatic injuries. fighting can occur between all sexually mature males, between females that are not acquainted to another, between individuals in overcrowded pens if the feed hoppers and water bottles are not in sufficient numbers. fighting males are very aggressive and may cause considerable damage to each other. wounds may show readily but may also pass unnoticed, concealed by fur. fractures are uncommon and may result from handling cage-housed rabbits unaccustomed to being handled which cause them to struggle forcefully, and it mainly occurs in cage-housed rabbits with a weak skeletal structure in the vertebral column (rothfritz ). housing and hygienic routines have a strong impact on the health of laboratory rabbits. solid-bottom cages and pens need thorough cleaning and regular disinfections so as not to spread intestinal parasites and bacteria back to the rabbits. cage-floors with perforation for droppings onto a pan reduce the number of coccidian spores etc. that the animals can ingest. "sore hocks" is a condition seen in heavier breeds of rabbits kept on solid-bottom cages or pens with inappropriate hygiene and the symptoms are bleeding and infected wounds along the hind feet of the animals. the diet of laboratory rabbits is crucial for the microbial balance in the gut and intestinal morphology (yu and chiou ) and thus the optimal rate of dietary fibres are essential to keep the animals from developing soft stool or diarrhoea. another type of health problem arises from the use of certain types of diets aiming to inflict metabolic changes in the animals, for example high cholesterol diets for the development of atherosclerosis. a side effect of using these diets in long-term studies is fatty liver and deposits of cholesterol in various organs. rabbits are often used for collecting blood in immunisation studies. blood can be taken from the marginal ear vein or by cardiac puncture. in the former case dilation of the ear vein facilitates the removal of blood, and with good blood flow up to ml blood may be obtained (batchelor ) . local anaesthetic facilitates the removal of blood from those animals which are distressed by the insertion of the needle into the ear vein. the cardiac puncture must be carried out under anaesthesia with the animal in dorsal recumbency. rabbits have about ml of blood per kg body weight, and a maximum of ml/kg body weight should be taken at a single sample (batchelor ) . others have suggested that up to ml of blood could be taken per kg body weight at a single sample (iwarsson et al. ) the body temperature of rabbits should be taken by a suitable thermometer which has been lubricated for ease of introduction into the anus (batchelor ) . the rabbit should be gently restrained, the tail lifted and the thermometer gently inserted. the thermometer should never be forced into the anus, and if there are any obstructions to the passage it should be investigated. the thermometer should be left in situ for - minutes, whereafter the temperature is recorded. in rabbits the normal mean rectal temperature is . ºc with a range of . - . ºc. rabbits may be given different substances by oral (p.o.), subcutaneous (s.c.), intravenous (i.v.), intramuscular (i.m.) or intraperitoneal (i.p.) administration. for more detailed description of these techniques see batchelor and iwarsson et al. . for anaesthesia of rabbits there are many different methods and substances (svendsen ) . depending on the purpose of the procedure or operation a specific substance should be chosen, and then the recommended range of doses given in the literature should be followed (olfert et al. , svendsen , batchelor . generally relief of pain should be given to all rabbits during operational procedures, if there is evidence that pain is present in the individual. postoperative care of rabbits includes placing them in a recovery cage, the box they were placed in before surgery or the home cage which has been lined with a tray liner (batchelor ) . the liner should be folded over the animal and a cotton surgical drape placed over it to minimise hypothermia. a 'vet bed', which is commercially available may also be used. the liner or 'vet bed' should be removed about minutes after the animal has regained consciousness and is sitting up. when operating on several group housed animals the last animal operated on must have completely recovered consciousness before all the animals are returned to the pen simultaneously (batchelor ) . if animals are returned to a group pen while they recover consciousness they may be subjected to aggression. incision sites should be covered with a clear plastic dressing spray, but rabbits may occasionally interfere with their stitches. a plastic collar can then be applied to the animal to restrict its access to the operation site after suturing (batchelor ) . euthanasia of rabbits is usually carried out by intravenous injection of an overdose of barbiturates, such as sodium pentobarbitone (batchelor ) . physical dislocation of the neck is another option for rabbits up to one kg in body weight. however, it is recommended that this always is followed by heart puncture or cutting one of the main vessels so that the animal is drained of blood. in order to ensure the welfare of laboratory rabbits there are several aspects to consider. first of all one has to make sure that the rabbits are purchased from a breeder producing health defined rabbits with a controlled genetic background, and with an enriched housing. the transport to the laboratory should be as short and stress-free as possible. placing two rabbits in the transport box could reduce stress. at arrival to the laboratory the rabbits should be checked for any health problems and placed in its new cage. acclimatization should be one to two weeks. young rabbits could be housed pair-wise in cages, or in larger groups in floor pens. cages should be large enough to give rabbits a place to perform hopping movements and to lie in a position fully stretched out. they should be provided with free access to hay and water, and fed at least once daily. floor pens should be large enough to house the maximum number of rabbits planned for, at least one meter high to prevent escape and enriched with refuges, hay, gnawing sticks, etc. regular cleaning of cages and floor pens are important to keep a good hygiene and healthy animals. there has been relatively large amount of research on the welfare of laboratory rabbits over the past years (morton et al. , gunn , stauffacher et al. , hubrecht et al. . both improvements of cage systems and floor housing have been evaluated. there are several companies selling environmentally enriched cage systems and equipment for building floor pens and enrichment items for floor pens. parallel with this development, new regulations have been planned and intensively discussed over the past years (appendix a of the european convention, regulations in sweden), and some have been put into practice (regulations in switzerland and uk). the most important welfare research to carry out for the future would be to verify if some of the behaviours rabbits perform in semi-natural environments, for example digging, is so important that it could be considered an essential behavioural need. alternatives to permanent cage housing of mature bucks, as for example regular exercise in an enriched area, should also be scientifically evaluated. there is also a need for more controlled experiments on the effects of group housing and environmental enrichment on the results of laboratory research. the laboratory rabbit group housing on floor pens and environmental enrichment in sandy lop rabbits (i) the laboratory rabbit aspects of the social behaviour of wild and domesticated rabbits oryctolagus cuniculus l social olfaction in lagomorphs. symposia of the zoological society of london mate choice in the european rabbit the behaviour of rabbits: implications for their laboratory management the european wild rabbit effects of single-and mixed sex caging on post-weaning development in the rabbit effects of climate on reproduction in the european wild rabbit oryctolagus cuniculus storey´s guide to raising rabbits. storey books the effect of hay on the behaviour of caged rabbits (oryctolagus cuniculus) loose grass hay as a supplement to a pelleted diet reduces fur chewing in rabbits mutter-kind-beziehung beim hauskaninchen schlussbericht ueber die pruefung der tiergerechtheit eines festwandkaefigs fuer hauskaninchen-zibben paarhaltung nicht reproduzierender zibben im käfig breed standards - nutrition and gastrointestinal physiology the scientific assessment of animal welfare symptome des wohlbefindens und des unwohlseins beim kaninchen unter besonderer beruecksichtigung der ethopathien trichophagia: a behavioural disorder in the domestic rabbit rabbit nest construction and its relationship with litter development använda försöksdjur i sverige år -statistik rabbit feeding and nutrition arteriosclerosis research serological genetics third report from the commission to the council and the european parliament on the statistics on the number of animals used for experimental and other scientific purposes in the member states of the european union aspects of the social organisation of the european wild rabbit (oryctolagus cuniculus) variations in the social structure of rabbit populations: causes and demographic consequences leporid social behaviour and social organization effects of social organisation and environmental diversity on determining the genetic structure of a population of the wild rabbit (oryctolagus cuniculus) basic anatomy, physiology and husbandry einfluss unterschiedlicher haltungsverfahren und bewegungsmöglichkeiten auf die kompakta der röhrenknochen von versuchs-und fleischkaninchen einfluss unterschiedlicher haltungsverfahren und bewegungsmöglichkeiten auf die kompakta der röhrenknochen von mastkaninschen the rabbit in subalpine south-eastern australia, . population structure and productivity the production of specific pathogen free rabbits emergence behaviour of rabbits, oryctolagus cuniculus, in central otago försöksdjursteknik-en praktisk handledning time and space in a sparse population of rabbits (oryctolagus cuniculus) natural control of a population of rabbit, oryctolagus cuniculus l. for ten years in the kourarau enclosure maternal behaviour in new zealand white rabbits: quantification of somatic events, motor pattern, and steroid plasma levels evaluation on welfare in the husbandry of laboratory rabbits in environmental enrichment information resources for laboratory animals planung und struktur von versuchstierbereichen tierexperimentell tatiger institutionen th edn sex drive in rabbits. southwest vet colony husbandry the effect of environmental enrichment on the behaviour of caged rabbits (oryctolagus cuniculus) textbook of rabbit medicine the biology and medicine of rabbits and rodents the foetus in experimental teratology housing rabbits the unconventional way choices of laboratory rabbits for individual or grouphousing rabbits and rodents refining rabbit housing, husbandry and procedures: report of the ufaw/rspca rabbit behaviour and welfare group meeting responses of adult new zealand white rabbits to enrichment objects and paired housing common non-surgical techniques and procedures intestinal stasis and rupture in rabbits the rabbit: a diurnal or nocturnal animal? mirrors in rabbit cages: what does the rabbit see and how do they affect behaviour? submitted kraft r. vergleichende verhaltensstudien an wild-und hauskaninchen effects of early handling on later open-field behaviour in rabbits the use of cover and burrows by a population of rabbits (mammalia: oryctolagus cuniculus) in eastern scotland handbook of rodent and rabbit medicine indikatoren für mangeinde tiergerechtheit sonie verhaltensstorugen bei hauskaninchen das verhalten junger hauskaninchen unter verschieden umgebungsbedingungen. dissertation activity requirement for young domestic rabbits: raw fibre consumption and animal welfare social behaviour in young domestic rabbits under semi-natural conditions the preference of domestic rabbits for pasture and concentrate feeds behavioural effects of environmental enrichment for individually caged rabbits some observations on the breeding burrows of the wild rabbit on the island of skokholm social structure and stress in the rabbit warren einfluss unterschiedlicher haltungsverfahren auf das verhalten von versuchs-und fleischkaninchen group-housing rabbits basic approach to veterinary care the rabbit. in online information for veterinary students at royal school of veterinary studies general biology and husbandry effects of early handling in the domestic rabbit sound levels in rooms housing laboratory animals: an uncontrolled daily variable supplementary hay reduces fur-chewing in rabbits the effects of density on sociality and health in mammals a study of the biology of the wild rabbit, oryctolagus cuniculus (l.), in confined populations ii. the effects of season and population increase on behaviour observations on reproduction, mortality and behaviour in a small, free-living population of wild rabbits the rabbit in australia social behaviour of an experimental colony of wild rabbits, oryctolagus cuniculus (l.), iii second breeding season territorial marking by rabbits continuous observations of the activity of the wild rabbit, oryctolagus cuniculus (l.) during -hour periods effect of social environment on reproduction in the rabbit, oryctolagus cuniculus (l.) an experimental study of aggression in captive european rabbits, oryctolagus cuniculus (l.) nutrient requirements of rabbits social environment as a factor in diet-induced atherosclerosis felasa recommendations for the health monitoring of rodent and rabbit colonies in breeding and experimental units guide to the care and use of experimental animals the population ecology of the wild rabbit dispersal of the wild rabbit, oryctolagus cuniculus, at urana in new south wales diet preference of sheep -effects of recent diet, physiological state and species abundance the behaviour of group penned and individually caged laboratory rabbits commercial rabbit keeping, rd edition genetic, physiological and behavioural background of reproduction in the rabbit. v. nonretrieving of neonates einfluss unterschiedlicher haltungsverfaren und bewegungsmöglichkeiten auf die spongiosastruktur der rippen sowie brust-und lendenwirbel von versuchs-und fleischkaninchen electrical activity of the intestine and feeding associated with a visual conditioning in the rabbit sexual and aggressive behaviour in the wild rabbit handbook of biological data kaninchenhaltung in zucht und mastgruppen -ein neues tiergerechtes haltungskonzept fuer hauskaninchen behaviour ontogeny and the development of abnormal behaviour group housing and enrichment cages for breeding, fattening and laboratory rabbits tierschutzorientierte labortierethologie in der tiermedizin und in der versuchtstierkunde -ein beitrag zum refinement bei der haltung und im umgang mit versuchstieren the accommodation of laboratory animals in accordance with animal welfare requirements. o'donoghue pn a comparison of behaviour, reproduction and mortality of wild and domestic rabbits in confined populations laboratory animal anesthesia transporting animals the grading of commercially bred laboratory animals spacing behaviour of free-ranging domestic rabbits spontaneous deaths in rabbits resulting from trichobezoars the assessment and control of the severity of scientific procedures on laboratory animals natal dispersal and genetic structure in a population of the european wild rabbit (oryctolagus cuniculus) funktionale analyse des verhalten als grundlage zur beurteilung der tiergerechtheit. eine untersuchung zu normalverhalten und verhaltensstoerungen bei hauskaninchen-zibben the demography of a rabbit population in an arid region of new south wales, australia nestbeschaeftigung bei saeugenden hauskaninchenzibben effects of crude fibre level in the diet on the intestinal morphology of growing rabbits effects of early social environment on physical and behavioural development in the rabbit rabbit: frequency of suckling in the pup we would like to thank clive phillips for commenting on the first draft of this chapter and for adding important information to this text. many thanks to astrazeneca for taking the initiative to research into improving the housing of laboratory rabbits, which made it possible for lena lidfors to collect references and get knowledge about the welfare of rabbits. we also want to thank the swedish agricultural board for taking the initiative to improve the regulations for laboratory animals and thus involving lennart lindberg and lena lidfors in giving suggestions on new regulations. key: cord- - if qquj authors: nan title: perspectives on the economics of the environment in the shadow of coronavirus date: - - journal: environ resour econ (dordr) doi: . /s - - - sha: doc_id: cord_uid: if qquj nan the environmental and resource economics special issue "economics of the environment in the shadow of coronavirus" comes at a hugely critical time for environmental economists and policy makers alike. we are in a situation of significant social change, a change that could potentially lay the foundation for mankind's future in the years to come. as part of this special issue, ere is trialling a novel, experimental form of article, drawing together short, focussed pieces from a wide group of authors addressing the plethora of issues which such a fundamental challenge as the coronavirus pandemic generates. these provide critical and reflective perspectives on the environmental, socio-economic and policy paths that may be taken in the near and further future-strategies that could lead mankind either on roads to a much more sustainable development, or along paths that could bring about more instability, inequality and further environmental pressures. this innovative article combines short, policy-relevant and less technical papers that deal with specific aspects and provide clear recommendations for policy makers and suggestions for future research alike. the target audiences are policy makers and companies, but also researchers who want quick yet sufficiently detailed knowledge about particular analyses relating to covid- and issues in environmental economics. we hope that the articles contained within this perspectives collection provide the necessary information for policy makers to take wise decisions for our future, and for researchers the knowledge to help guide policy makers in their decisions. humankind has been very fortunate to have lived through a period of sustained economic growth pretty much since the agricultural revolution, with especially high rates of growth starting from the second half of the twentieth century. this economic progress has allowed us to make unprecedented improvements in consumption, in health, in education and in addressing inequality. many of us have been fortunate enough to have lived without a war for the past years, which is widely believed to be due to the development of international institutions and a deepening of international trade that led to widespread cooperation and, with it, it brought a new era of global stability. at the same time, the rapid increases in humankind's population, from around billion in to . billion in , coupled with an increase in real-world gdp by a factor of roughly during the same period, have led mankind to progressively push closer to the boundaries of planet earth. to provide additional food for the surge in population, agricultural land use has increased by %; to provide goods and services for the surge in demand, the material footprint of our production increased by an estimated factor of ; and to provide energy for our lifestyles, our use of non-renewable and polluting resources (coal, oil, gas) increased by a factor of . this increase in consumption coupled with a similar increase in input use has transformed the face of the planet earth and has given rise to unwanted side effects and new challenges. some of these challenges are well known, such as local and global pollution, problems of waste and certainly climate change. another, often neglected challenge, has been a consistent pressure on biodiversity due to our increase in land use. a mixture of burgeoning population and increasing resource use that carved deeply in nature's pristine areas has led to species conflict manifested not only in the rapid loss of other species, but also in a much ignored yet increasingly visible negative feedback in the form of viral crossovers (smith et al. ). the linkages between economic development, viral crossovers in the form of communicable diseases and environmental issues in particular have, up to now, seen little attention from environmental economists. as we have now seen, it was worryingly neglectful on our part to not consider these feedbacks more seriously. the greater interconnectedness via global trade and international migration, air travel for both tourism and business purposes, as well as the ongoing growth of large city hubs, have made it easy for communicable diseases to transcend local spaces and quickly make their appearance in even remote corners of the world. while the black death and the spanish flu have been among the worst communicable disease outbreaks in recent history, in late a new virus was detected in wuhan, china. identified as a new member of the coronavirus family and subsequently called covid- , within the course of half a year this virus has spread out from the huanan wet market in wuhan across the whole world. even inhabitants from otherwise remote places such as villages in timbuktu, the korubo and yanomami tribes of the amazon, the navajo nation of north america and the arctic inuit have already tested positive for due to initial uncertainty surrounding both the impact of covid- and its spread through society, many policy makers quickly decided to shut down interactions among individuals by restricting local, national and international mobility. these "lockdowns" had pervasive impacts on economic activity across the globe, with significant reductions in production, increases in unemployment, falls in international migration, diminished levels of international trade, significant increases in bankruptcy filings and large ripple effects down supply chains. relative impacts between developed and developing countries are still very much developing. globally, herd immunity is expected to take some time to develop if indeed it ever does. a vaccine that has the potential to be potent and widely available may need at least another or years for development and broad deployment. some countries are already close to a second wave-this pandemic is here to stay for a while. the question is as to how we shall deal with it. while reducing physical contacts to "flatten the curve" of disease and death has been the preferred policy to slow down the spread of the were advocating for. it is at this point where the contributions selected for this special issue provide first thoughts, first answers and first suggestions for policy makers from the cuttingedge research of environmental economists. in particular, the arguments forwarded support a strengthened focus on economic recovery that, first and foremost, should not undermine the green transition, while also, if possible, provide measures to advance the green transition. the articles then discuss the approaches and potential difficulties that policy makers will be faced with when being confronted with the precise means to implement these green recoveries. as a first step, due to unprecedented levels of unemployment in places such as the usa and significant contractions to economic growth in most countries of the world, an important consideration is that the focus of the stimuli packages should be the economic recovery, i.e. to predominantly deal with the direct impact of the lockdowns on economic activity. once the virus is contained and the short-run recoveries are under way, then it is, however, important to quickly integrate longer-term factors into policy making (borghesi and co-authors). here, it is vital that, in contrast to the stimuli in the aftermath of the financial crisis, policy makers also address inequality (koundouri and co-authors). a more specific focus on furthering a green transition should only be placed once a certain level of economic recovery has been achieved. this is especially vital as the disruptions to supply chains can have fundamental and unpredictable consequences, as often even companies themselves are not fully aware of their complete supply chains. cazcarro and co-authors estimate some of the impacts of these trade-related supply chains and, for example, show that the european demand changes due to covid- have, in total, larger impacts on the rest of the world than on europe itself. requirements for successful green stimuli are that these policies are implemented in a clear and transparent manner (rickels and peterson). in this regard, ing and nicolai argue that companies are likely to prefer stimuli packages that are tight to some environmental efforts rather than to new environmental regulations. on the converse, linking stimuli with environmental efforts is more costly for policy makers and likely to be less efficient. several of the articles in this special issue draw particular attention to the fact that the green stimuli are not enough to successfully further a green transition. what is also necessary is to couple this with a price on carbon and a restructuring of the subsidies paying attention to both the green and fossil industry (gawel and lehmann). a stronger social contract with a higher degree of citizen involvement will furthermore help gain public support but also strengthen social norms and thus decentralized internalization of externalities. lopez-feldman and co-authors discuss the policy responses to covid- with a focus on latin america and argue that, to minimize the likely rebound effect, policies need to be much better coordinated. we have seen that international cooperation quickly breaks down when a crisis looms, so that it would make sense to design international institutions with binding laws and penalties in case of non-compliance. what we have seen so far is that covid- has the potential to become a game changer when it comes to combining stimulus packages with the green transition. that this is a sensible strategy derives from the observation that restricting global warming to . °c requires efforts that go beyond what countries were willing to do so far, and that the stimuli provide the needed opportunity. while the articles contained in this special issue already provide many reasons for policy makers to push for green stimuli, they also clearly point out the difficulties associated with implementing these well. some articles in this special issue also show limitations of current policies or research approaches. for example, borghesi and co-authors discuss that during the covid- crisis the market stability reserve helped to stabilize the eu ets price, but imperfectly. it is, therefore, important to consider ways in which these imperfections can be redesigned. on a different topic, laude explains how the covid- crisis has brought to light both advantages and problems with having local, short supply chains for food, and that there is a substantial lack of research directed towards the impact of crises on the agricultural sector. another example is a more cautionary tale and deals with the covid- cases data. here, cohen and co-authors show very clearly that researchers must be careful with simply using these data as there are many problems in the data collection processes, which differ across countries and also time. as a final remark, we would like to observe that this special issue not only comes at a very turbulent time for mankind in general, but it also comes at a special time for environmental economists in particular. the covid- crisis gives the opportunity to invest significant amounts of money towards aiding the green transition, and the widespread public support is there. we need to now be able to advise policy makers on efficient, reasonable and relevant policies that they may implement as part of the green stimuli packages. however, these policies also need to be well structured and grounded in good research. the peer-reviewed articles in this special issue provide suggestions and articles with these features and will thus, hopefully, serve as a first benchmark in this endeavour. in this paper, we use a multisectoral and multiregional model of the world economy to evaluate the short-term effects that the covid- crisis may have on environmental pressures and resource consumption (measured in terms of water, air emissions and materials extraction). specifically, we focus on the relationship between current and forecast changes in demand and mobility patterns in the eu + uk for and its effect on global resources through global supply chains. this integrated analysis could answer urgent research questions: what are the short-term impacts on emissions and resource consumption of the current and predicted declines in final demand? do the impacts differ among environmental pressures? how elastic are environmental responses to demand drops? on the basis of these short-term responses, what is the relationship between economic growth and environmental pressure? do these effects go beyond european countries through global supply chains? in order to evaluate the short-term effects of changes in aggregated sectoral consumption and demand on environmental pressures, and their diffusion through global supply chains, we develop an environmentally extended multiregional input-output model (see miller and blair , or recently hubacek et al. ; guan et al. ), using the exiobase . database (stadler et al. ) . we focus on water consumption (blue and green), mineral extraction and emissions (co eq , sox, nox, nh and co). we rely on the estimates from the eurostat spring forecast (ec a) which estimate for the eu + uk a change of − % in consumer expenditure, . % in government expenditure and − . % in investment for . sectoral changes in private consumption are estimated based on ec ( b) and oecd ( ), assuming different sectoral sensitivities to the covid- crisis. specific details and the complete matching process can be found in online appendix. our results are as follows: figure shows the eu + uk (light blue bars) and global (dark blue bars) change in different environmental pressures associated with the shock to final demand and household mobility in eu + uk, as a consequence of the covid- lockdown. the expected percentage impacts are much larger in the eu + uk, as the simulation of final demand and mobility directly affects these countries, given the high level of intra-eu dependence. however, the european lockdown also impacts foreign resources due to the interlinkages throughout the production chain. in general, the largest decline happens to gas emissions, both in eu + uk and worldwide. the average . % fall in the aggregate of the three cited components of european final demand (households, government expenditure and investment, representing a . % fall in total final demand) that globally represents a . % decline, would involve a . % decline in global water consumption and . % in mineral extraction. the global fall in emissions from the slowdown in economic activity and european mobility restrictions would be around . % (co eq ). comparing the effect in eu + uk with the global impact, the percentage change runs from four times larger in europe, in the case of water, to times greater for co eq emissions. we find a drop of more than % for mineral extraction and emissions in eu + uk. emissions are reduced, and by more than water and minerals. also, even when no direct electricity demand change is assumed, given the reduction in sector activity, indirect energy demand diminishes and so do emissions. specifically, our model indicates that the production and supply of electricity explain approximately % of the greenhouse emissions fall worldwide and % in the eu + uk. in line with the results of le quéré et al. ( ) , our estimates confirm that the reduction of emissions associated with private mobility restrictions would account for % of the total fall in emissions in europe, whereas the reduction linked to air travel reaches . %. worldwide, these drops would be % and %, respectively. the european lockdown affects the iron and steel industries globally, representing % of the global fall in greenhouse emissions. several other resources are less dependent on the most-affected sectors. the supply chain of food clearly depends on the supply of water. it is difficult to estimate the reductions in the horeca sector based on the available data. that sector, as well as many others that indirectly require these natural factors throughout their supply chains, exhibits significant reductions in resource use. our data show that the largest decline in water consumption (both blue and green) is associated with the primary sectors, which, as expected, experience slight negative growth rates around − . %, given the relatively stable and anticyclical nature of food demand. analysing the changes induced by the different components of final demand, the decrease in household consumption and investment drives environmental impacts to a lesser extent. worldwide, the fall in investment mainly affects environmental impacts (except for water consumption). reductions in investment have a larger role in reducing emissions in small countries like croatia and malta, and also in eastern europe, such as in hungary, latvia, and slovenia. in the eu + uk, household consumption is the most significant element of final demand driving the changes in environmental pressures (with the exception of mineral extraction), explaining % of the european greenhouse gas emissions fall. as for minerals, their strong dependency on gross capital formation destinations, such as construction, manufacture of machinery and equipment, computer and related activities, explains that the . % fall in investment worldwide (- . % in eu + uk) triggers a . % drop in their global extraction (− . % in europe). these changes are distinct by country, with implications for different areas of the world. the intra-eu trade is revealed to be highly important, showing that the main changes occur within the eu + uk countries (even if the individual shock of a given country is not so significant). figure displays the percentage falls in co eq , so x , no x and nh emissions by eu + uk country. according to the projected changes, the greatest cuts are expected in co eq emissions. almost a quarter of the estimated reduction in co eq emissions occur in germany (figure si ) . the reductions in co eq emissions in smaller economies, such as ireland, austria, greece, and cyprus exhibit the largest percentage falls. reductions in mediterranean and eastern countries are high, mainly in large countries like france, romania, portugal, and spain. we note the potential reduction in no x emissions as a consequence of the production shutdown in france and germany (as it occurs with nh emissions), but the largest percentage drops occur in smaller economies, like austria, ireland, and greece. france also shows the highest reductions in so x emissions, together with poland and germany, ireland and smaller countries. our model also allows us to evaluate how the reduction in the eu + uk demand modifies the pressures on environmental resources outside europe, by considering transmission effects through global supply chains. the largest percentage reduction in domestic impacts arising from the lockdown and subsequent situations in europe occurs in non-european developed countries, with smaller effects in developing economies (fig. ) . we find considerable indirect declines in mineral extraction (larger than %) and gas emissions (between − . and − . %) in the usa and japan. these are mostly associated with linkages with italy, germany, uk and france. in countries such as china, brazil and india, none of the falls in environmental pressure exceeds . % (with the exception of a % fall in mineral extraction in india). again, we find important declines related to the commercial linkages of developing countries with european powers, such as france, germany and the uk, and with other european countries severely affected by the pandemic (and thus their final demand), such as italy and spain. these results support the statements of baldwin and mauro ( ) on the existence of a "supply-chain contagion" related to the covid- lockdown (in europe in this study). however, the "environmental supply-chain contagion" is modest outside the eu + uk. one of the main implications of our work is that the changes occurring in in the eu + uk are not, in and of themselves, able to sufficiently reduce global environmental pressures. these changes have entailed reductions in domestic activities, which have also affected other eu partners, given the high level of intra-eu trade. although some of those changes do not have strong impacts on domestic environmental pressures, transport restrictions within the eu have notably reduced co eq emissions and, even more positively, reduced other pollution-induced health damage. changes outside europe occur due to spillover effects, being relatively more notable for minerals. however, the demand for goods, which ultimately depends on resource use and pressures external to the eu + uk, has not fallen so clearly, and the lion's share of the pressures has not been reduced as much as final demand. in short, we have shown the importance of intersectoral relationships and multipliers to understand demand changes, which are uncertain along , and which we anastasios xepapadeas on december , the european commission presented the european green deal (egd )-a roadmap for making the eu's economy sustainable by turning climate and environmental challenges into opportunities across all policy areas, and making the transition just and inclusive for all. the central objective of the egd is to attain a climate neutral eu by , which means that the eu will aim to reach net-zero greenhouse gas emissions by that year. the actions required to reach this target include decarbonizing the energy sector, which accounts for more than % of the eu's greenhouse gas emissions; renovating buildings to help reduce energy use which currently accounts for % of the eu's energy consumption; supporting industry so it can innovate and become a global leader in the green economy; and promoting cleaner transport, which constitutes an important source of the eu's emissions. in terms of resources needed, at least € trillion (european commission ) are projected to be necessary over the next decade according to the european commission, with sources including the eu budget, national budgets and the private sector. furthermore, the egd encompasses the so-called just transition mechanism whose objective is to help reduce the negative impacts on coal mines or steel factories associated with decarbonization. the roadmap for the egd includes actions related to climate ambition, clean energy, circular economy, smart mobility, greening the common agricultural policy, preserving and protecting biodiversity, and attaining a toxic-free environment and was planned to commence during the spring of . the covid- pandemic which appeared in europe in early , and the containment measures taken in order to control the pandemic and reduce the transmissibility of the virusthe r -below , have had a profound impact on the economy. in terms of macroeconomics, the covid- shock on the economy can be regarded as a keynesian supply shock in a multi-sector economy which triggers shortages in aggregate demand larger than the shocks themselves (guerrieri et al. ) . policies to deal with immediate impacts of covid- are aimed at a fast recovery from the recession, so that the policies are designed and implemented in a short-run context. the question which this note seeks to explore is whether the policies undertaken as a response to covid- could have long-run implications-positive or negative-in terms of sustainability and the objective of carbon neutrality. arrow et al. ( ) state that economic development is sustained at a given point in time if intergenerational well-being is non-declining at this point in time. intergenerational well-being is non-declining if the comprehensive wealth of the economy is non-declining. comprehensive wealth is the value of the assets of an economy, with the asset base or productive base consisting of reproducible capital, natural capital, human capital and health capital. social capital can also be included in the productive base. thus, the issue of sustainability can be analysed in terms of non-declining comprehensive wealth or productive base. the implications of the covid- shock on sustainability should therefore be examined in the context of its impacts on the productive base of an economy. the impact of covid- on the productive base is realized directly through morbidity and mortality and indirectly through the recession that is induced by policies implemented to contain the pandemic and the positive effects of public spending aimed at recovery. r is the basic reproduction number which is defined as the average number of secondary infections produced when one infected individual is introduced into a host population where everyone is susceptible. in a fully susceptible population, an infection can get started if and only if r > . if r < , a typical infective replaces itself with less than one infective, and the number of infectives tends to zero with the passage of time (e.g. hethcote ) . see, for example, the world economic outlook (imf, ) in which the projection for the percentage change in output in the euro area in is − . % with a rebound to + . % in . it should be noted, however, that in the current world of deep uncertainty, these predictions could be inaccurate. greenstone and nigam ( ) indicate that moderate distancing policies have substantial economic benefits in terms of mortality benefits and avoided hospital intensive care unit costs. the valuation is performed in shadow prices, with the shadow price for an asset being the present value of the contribution to well-being from one additional unit of the given asset. sustainability can also be defined in a similar way in terms of comprehensive investment. thus, the virus does not destroy reproducible capital per se, but it reduces its utilization through containment policies. however, the pandemic could affect capital accumulation if the recession slows down investment in reproducible capital. covid- seems to be beneficial for natural capital, at least in the short run. coal use fell by % at china's six largest power plants between the last quarter of and march , while in europe satellite images showed nitrogen dioxide (no ) emissions fading away over northern italy, with a similar picture in spain and the uk (henriques ) . according to a recent international energy agency ( ) report, global co emissions are expected to decline during to . gtco , which is . % lower than in ( . gtco ) (global carbon project ). this would be the lowest level since , and six times larger than the previous record reduction of . gt in due to the financial crisis. however, the international energy agency ( ) report also indicates that if efforts to contain the virus and restart economies are more successful, the decrease in energy demand could be limited to less than %. since the international energy agency's ( ) april report, data show signs of a recovery in carbon emissions. a very recent reportin june -states that new data show a v-shaped recovery in carbon emissions, with carbon emissions declining from february , reaching a minimum in april and then recovering slowly towards the february levels (domjan ) . regarding human capital, defined in terms of changes in the work force and education level, the pandemic is having a short-term negative effect on work force, but in the long term it might change educational patterns and the geographical structure of the supply of education if extensive on-line teaching is established. health capital is, of course, negatively affected through the value of statistical life. the above discussion suggests that the pandemic has a profound impact on the factors characterizing sustainability. thus, policies to start up the economy after appropriate containment of the pandemic and return to long-term desired growth paths will be beneficial for productive-based sustainability components such as health, human and social capital and reproducible capital through the increase in its utilization. the issue of primary interest is the impact of recovery-related policies on natural capital and climate change. this issue relates to the way in which the egd might be adjusted during the start-up period and whether the current beneficial impact of the virus-induced recession on emissions signals a long-term impact. recent results in climate science have established an approximately proportional relationship between the change in the global average surface temperature relative to the preindustrial period (the temperature anomaly) and cumulative co emissions relative to the same base period (e.g. matthews et al. ). with cumulative emissions since being approximately ± gtco (friedlingtein et al. ) , the expected reduction of approximately . gtco relative to due to the covid-related recession will have negligible effects on the temperature anomaly. furthermore, if the recovery projections in are realized, then it is reasonable to expect that the pre-pandemic situation will reemerge with regard to greenhouse gas emissions, unless the pandemic continues in strong recurring waves which make extended and persistent lockdowns necessary. this situation, however, cannot be regarded as the most likely scenario. if emissions recover in the short or even medium term, the projected time period for crossing the . °c threshold-according to the business-as-usual or alternative emissions paths-will not be affected in a significant way (ipcc ) . in this context, the covid- event is expected to have a negative impact on the global productive base (health, human, social and reproducible capital) and therefore on the global sustainability conditions, while the seemingly beneficial impact on natural capital, and especially climate, will be temporary and a return to pre-pandemic paths is most likely. this means that in the post-pandemic world, risks related to climate change damages, including risks from tipping elements and crossing of climate thresholds, are not expected to change. how does this picture fit with the european response to covid- in the context of the egd? the eu is currently developing and implementing a number of policies and stimulus packages to address the recession. the purpose of this note is not to analyse these measures, but rather to explore induced adjustments to the egd. it has been reported that there will be some reprioritization of edg initiatives as a result of the eu response to the pandemic (involved in europe ). some of the initiatives such as the renewed sustainable financial strategy, which aims to increase private investment in sustainable projects, or the "renovation wave", will remain since they are expected to stimulate economic activity; others such as "offshore renewable energy" or "the biodiversity strategy for " might be delayed, but initiatives such as the new eu strategy on adaptation to climate change and the new eu forest strategy will be delayed to . it is clear that the covid- shock and the need to start up the economy will make substantial policy changes necessary. looking at sustainability in terms of natural capital and the environment, it should be clear that any short-run improvement is transient, while looking at sustainability in terms of climate change, it is most likely that there will be no change in the long-term trends. what is important is that, after the shock, the start-up will be based on environmentally friendly policies. thus, maintaining initiatives such as the renovation wave, or promoting the pillar of cleaner transport, is important. on the other hand, delaying initiatives like the strategy on adaptation to climate change may need to be reconsidered. this is because the rationale behind delaying such strategies seems to be that recovery from the recession is expected to be rapid and therefore the delays will be of short duration and no significant time will be lost. however, recovery might be impeded or delayed by issues such as new, possibly weaker waves of the pandemic or more technical issues such as fiscal multipliers. fiscal multipliers during a recession when shocks are concentrated in certain sectors are not expected to be operational, with the multiplier for government spending being around one and the multiplier for transfers likely less than one. these factors might result in the delays-initially projected to be of short duration-extending for a much longer period. the important point here is that the covid- shock will have a negligible effect on the evolution of the temperature anomaly. thus, if adaptation activities and decarbonization do not proceed rapidly, the risks of a climate shock will not be sufficiently mitigated. the need for strong action now is exemplified by the fact that the emissions gap in between current policies and the emissions necessary to keep the temperature anomaly below . °c in is approximately - gtco (unep ). the covid impact on emissions in is expected to reduce the gap by approximately gtco , which falls very far short of the gap that needs to be closed. the important aspect of a climate shock is that in addition to the negative impacts on the productive base of the economy-human, health and social capital-it will have a much more serious negative impact on the reproducible capital relative to the pandemic. this is because the climate shock will not just reduce the utilization of this type of capital, it will destroy part of the capital stock, since it will affect infrastructure, equipment, buildings and so on. recovery in such a case will clearly be slower and more difficult. this creates a serious argument against delaying adaptation programmes which could provide substantial benefits in the presence of climate shocks. typical adaptation projects (e.g. early warning systems, water resource and flood-risk management, sustainable agriculture, strengthening the resilience of existing infrastructure making new infrastructure resilient) are characterized by high benefit-cost ratios (fankhauser ; gca ) . under the resource constraints imposed by pandemic containment policies and the deep structural uncertainty characterizing the situation, the prioritization of these adaptation policies could necessitate the use of max-min expected utility criteria in decision making. as second-round benefits, adaptation programmes which involve investment could stimulate the economy and provide new jobs. to summarize, three important points should be taken into account: (i) that the indications thus far suggest that the benefits associated with the reduction of greenhouse gas emissions because of the covid- are transient, which means that the long-term trends associated with climate change are not expected to change, and therefore, mitigation and adaptation strategies should be strongly pursued in the post-covid- era ; (ii) that there is a need for an ambitious and comprehensive european economic recovery plan from the covid- crisis (european economic and social committee b); (iii) that climate-change-related investments, in particular in adaptation, are expected to deliver significant economic benefits. based on these points, it becomes clear that a green recovery plan with resources directed towards achieving the combined objective of both providing the necessary economic stimuli for recovery and also promoting the transition to a low-carbon economy and adaptation to climate change along with investment in natural capital and increase in comprehensive savings could be a feasible and efficient plan. the egd is an important strategy for securing the sustained development of the eu and protecting climate as a global public good. at the same time, it is clear that addressing the pandemic requires action now in the form of new policies and changes in priorities. however, although covid- is of necessity in the spotlight at present as a major global threat, it should not displace action aimed at an equal or greater global threatthat of climate change-under the misconception that the temporary drop in emissions during the pandemic allows us to delay climate change action now. a green recovery plan could realistically provide the double dividend of helping the eu economies to recover from the covid- crisis and, at the same time, promote the attainment of a climate-neutral eu. according to the gca ( ) study, adaptation investment of $ . trillion in the areas mentioned will provide total net benefits of $ . trillion by , with benefit-cost ratios ranging between and . see, for example, the recent eu european economic and social committee ( a) opinion about the transition to a low-carbon eu and the financing of adaptation to climate change. involved in europe ( ), leaked: full list of delayed european green deal initiatives-euractiv.com. available at https ://europ e.vivia nedeb eaufo rt.fr/leake d-full-list-ofdelay ed-europ ean-green -deal-initi ative s-eurac tiv-com/. ipcc ( ), global warming of . °c. an ipcc special report on the impacts of global warming of . °c above pre-industrial levels and related global greenhouse gas emission pathways, in the context of strengthening the global response to the threat of climate change, sustainable development, and efforts to eradicate poverty. masson-delmotte, v., zhai, p. rebound effect studies have been generally focused on energy use (sorrell ) , although some studies for other natural resources have recently emerged (freire-gonzález and font vivanco ). rebound effect occurs when the use of resources is not reduced as expected after a resource efficiency policy or a specific behaviour. empirical rebound studies aim at capturing the secondary effects of policies and behaviours in order to obtain more adjusted assessments of policies and actions. it is well known in the rebound literature that, counterintuitively, resource efficiency may not reduce the use of these resources, but the contrary. this extreme case is known as backfire, khazzoom-brookes postulate, or jevons' paradox. rebound effects are not usually observed by policy-makers, as it requires different perspectives and approaches coming from social, behavioural and environmental sciences. environmental and social sciences show us that human-environment systems are deeply interconnected. this way of thinking has, however, still not fully permeated in mainstream policy decision circles, which are largely rooted in old intellectual paradigms and other short-term interests. the pandemic has caused many abrupt changes in production and consumption, transport patterns, working conditions, social interaction and many other aspects. most of these changes have been triggered by the policies implemented to contain the pandemic. overall, they have translated into improvements in most environmental indicators, such as carbon emissions, air quality and biodiversity loss (saadat et al. ) . while some authors claim that such changes will not have a lasting impact when the epidemic subsides (mccloskey and heymann ), others argue that aspects related to urban planning, micro-mobility, sharing economy, public transportation, teleworking, tourism, etc., may change for good (honey-rosés et al. ). an important question is, thus, whether covid- will reduce environmental impacts in the future, when economic activity returns to "normality" (in terms of pre-covid conditions). rebound literature shows the importance of considering behavioural and systemic responses to answer this question. beyond other considerations, the pandemic has accelerated some already observed trends, like the pace of implementation and use of digital technologies. one of the most remarkable changes are those related to the impulse of information and communications technologies (ict), due to imposed social distancing rules. there already was a tendency towards an increased use of ict, but its use has been dramatically accelerated due to the pandemic. this acceleration can be observed in many areas, such as teleworking, e-commerce, remote social relationships, virtual sightseeing, surveillance technologies, and other online areas and events (cultural, academic, leisure, educational, etc.) . for instance, in many countries, nonessential workers have been legally obliged to be confined during the pandemic to stop the contagion of the virus, thus promoting telework. despite the potential advantages of teleworking in increasing labour productivity in many industries (harker martin and mcdonnell ) , rigidities in corporate culture and other legal and cultural restraints were hindering and adjourning its consolidation. the use of ict is thought to be environmentally beneficial, largely due to decreased transport, but this premise has been challenged by rebound effect studies. gossart ( ) shows that existing evidence suggests that ict are subject to important rebound effects, mainly because it is a general-purpose technology, and so prone to backfire (sorrell ) . takahashi et al. ( ) calculated the rebound effect of ict services in a case study on videoconferences and found that rebound can reduce up to % of carbon savings. joyce et al. ( ) recently found for sweden strong environmental rebound effects associated with ict use, in most cases far above % (more resources use than before). this backfire effect is strongest for energy use and total material footprint, both close to %. another change may take place in land use and the housing sector, as initial evidence suggests that attributes such as floor space and outdoor space will have elevated importance (mikolai et al. ) . the potential re-distribution of time and expenditures towards resource-intensive sectors, such as construction, water and energy services, will likely cause material, water and energy rebounds. however, the expansion of teleworking can, at the same time, reallocate space and incomes in office rental market. city centres will not need to concentrate workspaces, changing mobility patterns and urban structures in the long term (elldér ). public transport may also be negatively impacted in the short and mid-term, leading to increased private transport (honey-rosés et al. ), another resource-intensive activity. other structural changes may also take place, such as changes in sufficiency measures and broader productivity, leading to macro-economic rebound effects (lemoine ). the pandemic may increase the social acceptance of sufficiency measures such as working less time, spending more time with family and friends, or connecting with nature. these measures have long been proposed to reduce consumption and associated environmental impacts (hayden and shandra ). these measures have, however, been associated with macro-economic price rebound effects as the decreased demand for some products can lower their price and induce additional demand (sorrell et al. ) . moreover, the postpandemic society may likely be a more productive one in labour and capital terms. for example, teleworking (harker martin and macdonnell ) and increased spending in research and development have been associated with productivity growth, which boosts economic growth and resource use. the covid- pandemic will likely cause a range of changes in society, but their permanence and effect on the environment are unclear, especially if we contemplate the secondary effects of behaviour, measures and policies. a key question is whether they will acquire a certain level of permanence, even modifying the mindsets of agents. given the high uncertainty around this aspect, its real dimension could only be assessed ex-post. however, due to confinements, the pandemic has greatly accelerated the expansion and use of general-purpose technologies, like ict. as this has been a long-observed trend, before the irruption of the virus, they have probably come to stay to a large degree. the pandemic offers a great potential to improving (and consolidating) environmental conditions. but beyond what conventional environmental indicators show, additional measures would be needed to counteract hidden rebound effects and, therefore, take full advantage of potential improvements. recent literature shows that different economic instruments like environmental taxation, resource pricing or setting limits to resource use, can be effective for this purpose. this is particularly necessary in this case, given the high risk of backfire due to the high expansion of general-purpose technologies observed. the global financial and commodity markets are facing economic distortions caused by the coronavirus (covid- ) pandemic (fernandes ; irwin ). covid- acts as a negative shock to overall demand of goods and services, resulting in aggravated shortrun volatility in prices (albulescu ). among these products, oil has been dramatically affected due to community lockdown regulations, shutdown of car factories, decline in energy use and increase in unemployment (reed a; the associated press ). however, a decrease in oil consumption may lead to reductions in carbon dioxide emissions (peña-lévano et al. ). data recorded by epa ( ) during march-april show an improvement in overall air quality, especially in high-density populated cities (regan ). air pollution is considered by many scientists as a negative contributor in the coronavirus situation by worsening the susceptibility of infection. a decline in emissions somehow may help prevent mortality temporarily, especially among more vulnerable individuals with underlying health conditions, such as heart and respiratory diseases (conticini et al. ; dutheil et al. ; mooney ; ogen ). thus, in this short article, we discuss the interaction between fossil fuels, air pollution and health risk under the coronavirus pandemic and the lockdown regulations during the period of march-may . the oil market has been dramatically affected by several exogenous factors during the pandemic period: ( ) "shelter in place" mandates aimed at curbing the spread of the coronavirus have decreased people's social mobility and transportation activities. households leave their houses just for short travels for some essential errands, such as shopping groceries and/ or medicine. only essential personnel are required to be physically present at their jobs, whereas the majority are working from home (reed a); ( ) commercial flight demand plummeted globally (reuters ). in the usa, the passenger volume dropped % from a year ago (compared to april ) as estimated by airlines for america, with an overall reduced accommodation of passengers per carrier (rappeport and chokshi ); ( ) tourism activity slumped as many governments banned international travel, heightened border and immigration controls and barred the entrance of foreign visitors to decrease risk of infection. these policies further decreased the demand for taxis and cruise services. for regions such as the caribbean islands (where tourism is a primary economic staple good), this also represents a significant decline in their gross domestic products (semple ); ( ) unemployment has pervaded among the national economies (mazzei and tavernise ). more than million people have filed for unemployment in the usa in a -week period (u.s. department of labor ). this condition is mirrored in other countries. worsening unemployment conditions have substantially reduced consumers' purchasing power. among its consequences is an overall decline in car sales (reuters ; the associated press ); ( ) many industries, restaurants and buildings have been shut down during the lockdown. these closures reduced energy consumption, especially petroleum use, which is a large energy input in the usa and other countries (eia ). meanwhile, global oil supply responded slowly to the decline in demand as refineries cannot abruptly halt production (caldara et al. ; peña-lévano ). the russia-saudi arabia oil price war in march worsened the situation by oversupplying the market and consequently dropping oil prices (irwin ; reed a). in april, the major petroleum exporting countries agreed to decrease the world oil output by % during may-july, which is equivalent to . million barrels a day (krauss ; reed b). two recent studies published in geophysical research letters validate the realized reduction in nitrogen dioxide (no ) pollution in several regions. bauwens et al. ( ) compared no levels in the atmosphere recorded in january to april and for the same period in . notably, no , which is produced by emissions from vehicles and industrial operations, can cause serious lung ailments. their estimates indicate a significant reduction of % in china and - % drop in the usa and western europe. shi and brasseur ( ) estimate a % reduction in no pollution in northern china in january and february . in the same period, they also found a % reduction in particulate matter pollution (particles smaller than . µm). while such cleaner air conditions may persist only temporarily, these trends indicate that the desired environmental gains are feasible and realizable if stringent emission regulations, perhaps mirroring to some extent the pandemic's enforced limits on social mobility and industrial activities, are enforced in the future. several studies relate ambient air quality to mortality and morbidity conditions caused by covid- . this contention echoes an earlier correlation applied to the sars virus outbreak in china in the early s. researchers from ucla's school of public health analysed air pollution levels and sars fatality rates among chinese residents. their results indicate that sars patients' probability of dying would be doubled among residents in areas with high air pollution indexes (cui et al. ) . as applied to the current pandemic, the center for disease control and prevention (cdc) explains that covid- causes a respiratory illness with a heightened risk among people who are years and older as well as those with certain underlying health conditions. latest cdc statistics on the pandemic's severity and fatality indicate that persons with heart ailment, diabetes and chronic respiratory diseases could be at a higher risk of being severely infected by the virus (vogel ). wu et al. ( ) further clarify that pre-existing health conditions identified as relatively more susceptible to contracting covid- are similar to those normally affected by air pollution in the usa. their study estimates that an increase of g m − in long-term exposure to particular matter (pm . ) increases the coronavirus mortality rate by %. isaifan ( ) presents corroborating evidence indicating that % of covid- -related deaths were cases with pre-existing illnesses, with majority of the victims over years of age. conticini et al. ( ) , however, warn that even young and healthy individuals could also be at risk as prolonged exposure to dangerous air pollutants causing chronic respiratory issues could be an additional co-factor that helps increase their vulnerability to being infected by the virus. ogen ( ) establishes that exposure to no may be an important instigator of covid- fatalities according to his research involving four european union countries. these findings are supported by the findings of a study published by the italian society of environmental medicine (setti et al. ) on virus infections in northern italy associated with air pollutants tagged as carriers and boosters. poor air quality has been cited as an aggravating factor in virus transmission. using data from italian province capitals, coccia ( ) notes an accelerated transmission dynamics of covid- leading to his conclusion that the spread of this virus can be considered more as following an "air pollution-to-human transmission" mechanism instead of an interpersonal transmission mode. moreover, fattorini and regoli ( ) analyse long-term air quality data in northern italy and suggest that chronic exposure to a contaminated atmosphere may have created conducive conditions for the spread of the virus. several studies relate the time frame of exposure to toxic air pollutants to mortality and morbidity conditions. lim et al. ( ) establish a significant association between longterm ozone (o ) exposure and elevated mortality risk of certain respiratory diseases. given limited data on the current pandemic, hoang and jones ( ) present emerging evidence on the severity of covid- infection attributed to persistent air pollution conditions, thus suggesting that longer exposure to a polluted atmosphere could aggravate virus infection. zhu et al. ( ) , however, provide concrete evidence suggesting that even short-term exposure to air pollution could increase probability of virus infection. their study analysed daily confirmed cases in cities in china recorded from january to february and found significant relationships between the levels of certain air pollutants and the number of newly identified covid- -infected cases. specifically, their results indicate that a -μg/m increase in the air pollutants' levels was associated with about . % to . % increases in daily new covid- cases. conversely, the health benefits of cleaner air resulting from reduced emissions from fossil fuel during the pandemic's lockdown period deserve attention. several studies recognize that improved air quality during the pandemic temporarily mitigated health risks associated with respiratory illnesses. cole et al. ( ) employed a two-step analytical approach using machine learning techniques and the augmented synthetic control method to estimate possible reductions in death rates in certain regions in china and for the whole country that may be attributed to actual reductions in no concentrations during the lockdown period. isaifan ( ) analyse air quality conditions prior to and during the lockdown period. his results indicate that lives may have been saved due to diminished ambient pollution levels, although eluding possible virus infection still is not necessarily guaranteed. this contention is corroborated by another china-based study conducted by dutheil et al. ( ) . these studies' assertions imply that air quality improvements realized even in such a shorter period of time (spanning less than half of a year) already could have some health benefit potentials, especially in relation to respiratory ailments. the health benefits of the current pandemic's notable environmental gain in air quality, however, will be optimized only if such favourable conditions are sustained over the longer term. the recent lockdown has been short-lived as some communities nowadays have started to revert to normal social routines and regular users of fossil fuels among industries have resumed operations. a brief respite from a usual contaminated atmosphere does not ensure an effective permanent eradication of chronic health conditions. the pandemic experience, however, demonstrates that better health is maintained and ensured not only through medical remedies and but also through more favourable environmental conditions, if sustained over a much longer period. current restrictions on social mobility and economic flexibility under covid- pandemic conditions have actually produced important economic and environmental repercussions that are interestingly contrasting. a general economic slowdown overtly reflected in, among others, reduced consumer demand, spiralling unemployment figures and significant drop in oil consumption causes heightened fears of an imminent economic recession. however, in spite of all the negativity surrounding the pandemic, its environmental consequence of improved air quality is a highly positive note. interestingly, the global community has been trying to accomplish such feat of attaining better air quality over many years of discussions, policy making and policing each other. unexpectedly, it took a serious pandemic to realize such feat. this article traces the interplay of reduced oil consumption with economic issues as well as environmental consequences under pandemic conditions. the more imperative issues now lie on the severity of a looming recession and the global economy's resiliency in transcending the difficult challenges it may bring. should that happen, will the economic cost burdens be outweighed by the realized environmental gains? experts may be quick to assert that improved environmental conditions actually may be short-lived as expected resurgence of resumed economic activities may only quickly bring back pre-covid air conditions. however, proponents of a cleaner world can always draw some inspiration from recent successes in air quality control, especially with the assurance that cleaner air is not necessarily a lofty goal. the challenge in the future lies in achieving such environmental benefit without the need to sacrifice the economic health of the global community. on february , , the brazilian ministry of health confirmed that a -year-old man was positive for sars-cov- : covid- had arrived to latin america. as of july , there have been . million confirmed cases in latin america, compared to . million in the european union and . million in the usa (jh-csse ). furthermore, so far there have been more than , deaths in the region and the trends show that the first wave of the pandemic is far from over (ibid.). there is of course no suitable time for a pandemic to arrive, but these are especially complicated times for latin america. the region is in the midst of a difficult economic situation accompanied by rising social discontent (eclac ; oecd ). moreover, it is characterized by high rates of informality, health systems with limited and unequal capacity, and most of the countries have high levels of debt (oecd ). under these circumstances, covid- is having major short-run socio-economic effects with possible serious long-run consequences, including several potential implications for the environment and the management of natural resources. restrictions of free movement and circulation within and across urban areas of latin america have reduced economic activity as well as the use of motorized vehicles. as a result, many latin american megacities have experienced a short-run decrease in air pollution. concentrations of no have decreased considerably in cities all over the region compared to the levels observed prior to the lockdown measures (iadb ). levels of pm , pm . and co have decreased in bogota, buenos aires and quito (bogota's district secretary of environment, personal communication, june ; roa ; rocha ). nevertheless, the pandemia has not had the same effect on air quality in all the major cities in the region. in mexico city, the reductions in so , pm . and pm concentrations have been modest, and there has been no reduction in ozone. in rio de janeiro, ozone concentrations have increased (dantas et al. ) . furthermore, as the virus and its negative consequences spread across rural areas and make its way through the southernmost part of the region, outdoor and indoor pollution might actually increase. in mexico, as well as in other countries in the region, the use of firewood is likely to rise as rural households try to deal with income reductions (masera et al. ). meanwhile, as winter hits central and southern chile, urban households might increase their use of firewood for heating given that, due to the lockdowns, they have to spend more time inside dwellings (encinas et al. ) . this rise in air pollution could arguably increase the risks associated with covid- . it is too soon to do a formal evaluation of the effects of the pandemic on deforestation and land use change in the region. nevertheless, the available information suggests that covid- is likely to have negative effects on forest cover across the region. early deforestation warnings from peru show that, although deforestation decreased between march and april , since then it has increased surpassing the levels observed during the same period in . according to data from the brazilian national institute for space research (inpe), the first quarter of already evidenced a rise of % in deforested hectares compared to last year's figures. the figures for april reinforce this pattern, with a % increase with respect to april (manzano ). from january to april , deforestation alerts in indigenous territory increased % when compared to the same period of the previous year (greenpeace brasil ). although at this point it cannot be claimed that the pandemic caused the observed increase in deforestation, it certainly does not seem to have provided incentives to halt it. in colombia, contrary to other countries in the amazon region, the trend in showed a reduction in deforestation compared to . however, started with an increasing tendency and the quarantine seems to have worsened the situation (fcds ). the absence of environmental monitoring during the pandemic seems to have encouraged illegal armed groups and regional mafias to take advantage of the situation, exacerbating deforestation with the possible intensification of illegal activities from which these actors derive income, such as illegal mining, land grabbing and illicit crops (bbc ). according to the pan american health organization, in may there were already , covid- -confirmed cases in the amazon basin (martín ). the impact of the pandemic in forest-based indigenous communities is an important source of concern. the spread of the virus in these communities could imply a tragedy that, in addition to the human losses, could affect the traditional knowledge, having negative impacts on the governance of natural resources in the region. this could lead to even more deforestation processes in the future. covid- has caused a disruption in the national and international trade of nature-based goods and services. tourism has come to a halt, affecting the economy of almost all of the countries in the region (mooney and zegarra ). in countries like costa rica, where the touristic industry is intertwined with nature, the shock to the sector could have negative effects for biodiversity and forests. without income from tourism, and given that as a slow recovery process is anticipated, the incentives to protect forests are expected to decrease in the short and medium run. fishing and aquaculture are other industries that have been negatively affected. information for the case of the chilean salmon aquaculture industry suggests that there has been a reduction in demand from international markets (chávez et al. ) . the effect of the shock is being transmitted through the value chain, affecting processing plants and farming facilities. the economic crisis can end up having long-run negative consequences for the environment if, as a result, regulations and environmental policies are relaxed or if institutions are weakened. although at this point there is no evidence of any country in the region purposefully relaxing environmental regulations to promote growth, it is certainly a possibility. what has been observed is that, in order to fund measures to reduce the economic and social impacts of the pandemic, some countries have decided to reallocate funds across the public administration. ecuador, for example, announced cuts affecting the ministry responsible for enforcing environmental regulations (bbc news ). something similar is happening in mexico, where the plan announced by the president is to reduce the operational budget of almost all government entities by % (d.o.f. ). even if countries have a relatively strong environmental legal framework, without a budget to monitor and enforce the regulations, this framework is worthless. countries in the region will very likely incur fiscal deficits and increase their debts in order to fight the crisis. it remains to be seen how the service of the increased debt will impact economic growth and the environment. in the meantime, it seems that the short-run legitimate demands to recover employment levels and improve the health systems might very well push aside the necessary investments to successfully tackle climate change and biodiversity loss. if this indeed happens, it could be the most serious effect of the covid- pandemic in both environmental and social terms. the economic projections suggest that the region will experience a crisis whose magnitude has no precedent in modern history (eclac ). in order to overcome this apparently insurmountable challenge, latin american countries will need well-designed policies that should reconcile economic objectives with social and environmental goals. the social unrest manifested recently in the social mobilizations in the region, should make clear that the apparent trade-off between economic, social and environmental objectives is the result of a false dichotomy between shortand long-run objectives. if environmental objectives are put aside, as has so often happened with social objectives, the economy might recover in the short run but at a very high price. the lockdown measures seem to be having a temporal positive effect on reducing urban pollution in some latin american cities. the challenge now is how to intervene to prevent a return to the same or even higher pre-quarantine emission levels. this is an opportunity to rethink the urban environmental policies while trying to recover from an unprecedented social crisis. at the same time, the observed increase in deforestation reopens political and academic debates about the role of national parks, indigenous reserves and other protection categories in a context of deteriorated livelihoods, illegal economies and a lack of state presence. latin american countries could see this moment as an opportunity to improve regional cooperation in order to design and implement coordinated policy responses not only to the economic crisis but also to the challenges of mitigation and adaptation to climate change. furthermore, countries should coordinate efforts to increase monitoring and presence in the region to effectively reduce deforestation. we have presented an account of some of the most evident environmental effects that the covid- pandemic is having in latin america at this point. considering that we are in the midst of the health crisis and in the beginning of an economic one, it is natural to expect that the trends that we see now will change in the near future and that other environmental impacts will become evident. research that contributes to a better understanding of the environmental impacts and the effectiveness of different policy responses to the pandemic in latin america will be invaluable. there are many potential paths for future research; here, we mention just a few. the consequences of the interactions between poor air quality and covid- on human health are clearly worth studying. this is particularly relevant for the latin american context, characterized by health systems with very limited capacity and high numbers of population without formal employment. results from studies in this area could help us provide better guides to set environmental quality goals, as well as to implement policy interventions that can reduce pollution in the region's context of income inequality and spatial segregation. the short-run environmental effects of covid- show early warnings of an increase in the pressure on forest and other ecosystems across latin america. understanding the impacts of the pandemic on terrestrial and marine ecosystems, as well as on livelihood opportunities for local communities, has the potential to contribute to the design of policies which can improve management and conservation. the pandemic is opening new research questions regarding the impacts of global shocks on natural resource-based industries that participate in international markets. furthermore, the paths that different countries take to get out of the economic crisis might have profound impacts on international trade. if, for example, the world transitions to more reliance upon local production, or if emissions-related tariffs are imposed, large exporters of commodities in the region will be highly affected. the impacts that these potential trade changes could have on the environment are unknown. furthermore, if developed countries implement recovery plans that include provisions to reduce emissions in significant ways, as has been discussed in the european union, will latin american countries be able to respond in the same way? in any case, latin american countries are highly vulnerable to the effects of climate change and some of these effects (e.g. migration) could result in future health crises. a better understanding of the ways in which individuals might adapt to a changing climate, as well as of the barriers that they face to adopt adaptation measures, will be a valuable tool for the design of adaptation policies that prevent future health crises in the region and elsewhere. the distributional and gender-differentiated impacts of the pandemic, and the related environmental policy responses, is another area that deserves attention, especially because early evidence shows that the more vulnerable segments of the population in the region are the ones that are being hit hardest. finally, as has been recently pointed out by , the experience of the pandemic might lead to changes in behaviour and personal choices. it remains to be seen if this is in fact the case, and if so, how are these changes in behaviour modulated by the local context. an even more important issue to consider is what would these changes imply for the design of behaviour-based policy instruments aiming to change consumption and production patterns, as well as transport and land use decisions in latin america. bbc ( ) the place where nature isn´t healing. https ://www.bbc.com/reel/video / p bd wfc/the-forgo tten-peopl e-of-the-globa l-pande mic accessed june bbc news ( ) coronavirus en ecuador: las multitudinarias protestas por las drásticas medidas económicas y recortes de lenín moreno. https ://www.bbc.com/mundo / notic ias-ameri ca-latin a- accessed june chávez c, salazar c, simon j ( ) efectos socioeconómicos y respuestas públicoprivadas de corto plazo ante la crisis del covid- en el sector salmonicultor, una fotografía de la experiencia internacional. interdisciplinary center for aquaculture research-incar, fondap-anid, center, chile. https ://www.incar .cl/wp-conte nt/ uploa ds/ / /pb .pdf accessed june dantas g, siciliano b, frança b. b, da silva c. m, arbilla g ( ) the impact of covid- partial lockdown on the air quality of the city of rio de janeiro, brazil. sci total environ . https ://doi.org/ . /j.scito tenv. . . d.o.f. ( ) decreto por el que se establecen las medidas de austeridad que deberán observar las dependencias y entidades de la administración pública federal bajo los criterios que en el mismo se indican. de abril de . presidencia de la república de méxico. eclac ( ) report on the economic impact of coronavirus disease (covid- ) on latin america and the caribbean: study prepared by the eclac, santiago, p encinas f, truffello r, urquiza a, valdés m ( ) covid- , pobreza energética y contaminación: redefiniendo la vulnerabilidad en el centro-sur de chile. centro de investigación e información periodística. https ://ciper chile .cl/ / / /covid - pobre za-energ etica -y-conta minac ion-redefi nien do-la-vulne rabil idad-en-el-centr o-surde-chile /. accessed june fcds. ( ). cifras deforestación en el bioma amazónico, enero-abril .https :// fcds.org.co/site/wp-conte nt/uploa ds/ / /defor estac ion_ .pdf accessed june greenpeace brasil ( ). desmatamento em terras indígenas aumenta % em . https ://www.green peace .org/brasi l/press /desma tamen to-em-terra s-indig enas-aumen ta- -duran te-a-pande mia-da-covid - / accessed june helm d ( ) . contributing to a global effort. oecd. roa s ( ) medidas para enfrentar al covid- mejoran calidad del aire en dos ciudades ecuatorianas. mongabay-latam.https ://es.monga bay.com/ / /menor -conta minac ion-del-aire-por-coron aviru s-en-quito -y-cuenc a-ecuad or/ accessed jun rocha l ( ) por la cuarentena, la contaminación del aire bajó a la mitad en la ciudad de buenos aires. infobae. https ://www.infob ae.com/socie dad/ / / /por-lacuare ntena -la-conta minac ion-del-aire-bajo-a-la-mitad -en-la-ciuda d-de-bueno s-aires / accessed june resource exporters are now facing new urgent economic policy challenges due to covid- . these challenges are aggravated due to their dependence on finite commodities with volatile prices and demand. in response to the pandemic, resource exporters (such as botswana and saudi arabia) announced cuts in expenditures along with large fiscal (tax relief) and consumption-focused macroeconomic stimulus packages. critically, the novel issues raised by the coronavirus pandemic bring new trade-offs of energy policy between shortterm gains and long-term sustainability, creating an urgent need for critical, quantitative, policy-focused research in the resource exporters-energy policy nexus. a few novel issues emerged in resource exporters during the pandemic that give rise to short-term economic challenges. first, price shocks of unprecedented magnitude for commodities (deutsche bank ; world bank ), from coffee (hernandez et al. ) to hydrocarbons (iea b), causing a large drop in energy investment (iea a). for oil, prices dropped initially due to travel bans and economic activities hiatus, and further with oil price wars following the collapse of opec + agreement. by april , with oversupply, the rise of stockpiles, and the saturation of available oil storage, oil prices reached the lowest level in more than years, and west texas intermediate (wti) reached negative levels for the first time. prices have subsequently partially recovered but are expected to remain low with continuous fears of new covid- waves and uncertain demand. second, unprecedented economic contraction, because the pandemic-triggered decline in economic activity was significantly exacerbated by declines in export revenue. third, unprecedented fiscal pressure, resulting from costs of fiscal and economic stimulus packages plus a simultaneous rise in domestic expenses (especially healthcare and unemployment benefits) and sharp declines in resource export revenue. the effects have been so colossal that states like kuwait are considering halting legally-mandated contributions to the future generations sovereign wealth fund (swf) to ease fiscal pressures (al-zo'bi ). fourth, record and continuous withdrawals from swfs to fund post-pandemic recovery along with reallocation of funds and increased government debt (examples in arabian business ; holter and bloomberg ). even states with the largest swfs such as norway and kuwait have been affected, with the latter expecting depletion of its fiscal stabilization swf (al-zo'bi ). fifth, in oil-exporting gulf states, the unusual stay of millions of citizens and guest workers in the upcoming scorching summer in lieu of usual tourism travel or home-country visits, pressuring existing energy capacity. critically, in oil exporters, these novel issues and policy responses to them further expose existing economic fragilities and challenges, threatening long-term economic and environmental sustainability. for distorted oil economies in urgent need for economic diversification, said diversification and reverse dutch disease are impeded by their existing high distortions in labour, fiscal, industrial and energy markets (shehabi ) . oil exporters suffer from economic inefficiencies resulting from pervasive oligopolies (shehabi ) and market failure in long-run contracts for exploration and development of natural resources (ruta and venables ). fossil fuels are the primary source of energy in most oil exporters. in developing oil exporters, the dominant political economy undergirding policy making is a welfare rentier state, in which maintaining a political equilibrium is central. balancing spending-saving decisions for sustainable resource rents management depends on the sustainability of windfall expectations (gelb and grassmann ). reforming energy policy-which drives fiscal, industrial and environmental policies-is critical for these states' long-term development. yet the novel issues create new trade-offs of post-pandemic energy policy: achieving short-term gains will be at the expense of longterm gains in resource economies. thus, it is critical for fast policy-focused research to address the resource exporters-energy policy nexus, especially in the following two areas. although investments in renewables have been more resilient than in fossil fuels, and while we have seen an increased share of global energy spending on clean energy technologies in (iea a), the reality in hydrocarbon-exporting developing states is different. global oil market supply and demand dynamics have raised their opportunity cost for transitioning away from fossil fuels and for investing in green technology. accordingly, post-pandemic economic stimuli might achieve short-term recovery but harm long-term energy transition. first, the rise in opportunity costs of energy transition will facilitate the reallocation of funds away from renewables projects towards post-pandemic economic stimuli (similar to "green tape" cuts in australia and canada). second, recovering lost investments in energy transition projects is unlikely because the resource rents that fund them are likely to continue to be low in the future, given low resource prices and demand pressures of climate change mitigation. third, continuous withdrawals of diminishing swfs will entail limited resources for future energy transition projects. this is especially so as recovering swfs' withdrawals is unlikely in the light of expected low resource export revenue and the collapse in financial and commodity markets accompanying the unprecedented global recession. finally, economic stimuli expand consumption and welfare redistributive measures, which increase greenhouse gas (ghg) emissions ) and exacerbate existing distortions that have been shown to prevent economic and energy diversification (shehabi ) . new research should investigate the design and implementation of new economic solutions that have at their forefront long-term energy transition goals along with short-term economic recovery. three policy solutions are suggested at the consumer, the energy industry and economy-wide levels. a policy solution to target consumers is technological advancements (both private and public funded) in energy efficiency coupled with economic incentives to rationalize energy consumption. this policy combination can achieve energy transition goals despite expected delays in renewables and without requiring large multiyear investments. it is especially important in gulf oil states where skyrocketing cooling and desalination needs in the summer will be met using fossil fuels (renewables contribute less than % of power generation). a policy suggestion at the sectoral level is investing in clean energy technologies to decarbonize the energy sector itself, namely through carbon capture and storage as well as hydrogen. beyond short-term economic and environmental benefits, these investments can keep oil exporters relevant in a future with a diminishing role of hydrocarbons and collapsing commodity prices. the final policy suggestion is microeconomic and energy policy reform that can moderate effects of export price declines on the economy without the need for additional cuts in renewables investments or further withdrawals from swfs. examples include microeconomic reforms of labour and human capital to increase long-term productive capacity and oligopoly regulation in non-tradables and energy sectors which can increase efficiency and welfare gains that translate economywide (shehabi ) . consequently, resource rents could be salvaged for swfs resources or investments necessary for future development. in the light of the aforementioned novel issues in resource exporters, the postpandemic energy policy and economic stimulus packages are likely to have negative long-term effects on the environment. the reasons are as follows. first, while lockdown measures that minimized transportation and human activity reduced short-term emissions, they also increased power demand which in resource exporters is met mostly through fossil fuels, especially as international demand and prices remain low. second, short-term improvements will be negated upon the resumption of human activity, absent changes in energy policy regimes. importantly, as domestic energy prices remain low and highly subsidized in developing resource exporters, the extent to which covid- restrictions would shift energy consumption habits and behaviours of agents (households and institutions) remains very doubtful. third, there will be limited resources in the future to dedicate towards environmental regulation and ghg emissions reduction. the reason is that funding the post-pandemic recovery will reallocate funds away from environmental projects and savings in swfs. fourth, ghg emissions will increase due to consumption-focused economic stimulus packages and expanded use of fossil fuels to meet rising energy demand. this is particularly problematic because even prior to the advent of the pandemic, resource exporters were among the highest energy consumers and carbon emitters globally. indeed, the ten highest per capita carbon emitters are all resource exporters, with emissions ranging from tonnes (t) per capita (qatar) to . t per capita (kazakhstan), more than . times the global average of . t per capita (ritchie and roser ) . finally, and most importantly, the implementation of policy instruments-namely energy subsidy reform and carbon taxes-to achieve resource exporters' intended nationally determined contributions (indcs), is rendered significantly more difficult post-pandemic. it is a consequence of the magnitude of novel economic contraction and fiscal pressures in welfare-based states. these additional political constraints exacerbate environmental laws' enforcement, for which there already was a widespread failure in resource exporters (unep ). therefore, new research must quantify effects of proposed economic stimuli and of rising use of fossil fuels on resource exporters' environment and economy and accordingly evaluate and design new alternative policies that can achieve short-term recovery and national climate target goals. to that end, four policy suggestions are offered. first, reducing emissions in ways other than the politically difficult tax instruments, mainly through enhancing energy efficiency that reduces emissions at a given consumption level coupled with economic incentives that reduce energy consumption. second, implementing carbon tax instruments in ways that do not harm the most vulnerable of populations and reduce resource exporters' fiscal distortions, thus achieving a "double dividend". third, in the light of the novel issues, a key policy solution is adopting "green welfare expansion": including green options in the typical post-pandemic welfare packages at lower governmental budgetary requirements. examples include expanding subsidies for green technologies (such as solar panels, public transportation or electric cars), as well as excluding from subsidies non-essential high-carbon-emitting products for households above a certain income level. fourth, increasing the political viability of green recovery packages (such as clean physical infrastructure and natural capital investments), including engaging the private sector in initiatives that are typically public-led-such as climate finance, renewables expansion and resource mobilization. this policy is critical because, although green recovery packages may boost economic growth while helping climate change , they are politically contentious in resource exporters where they compete with welfare distribution. critically, applicable to the aforesaid two research areas, the pandemic offers resource exporters an opportunity to engage in a comprehensive reform agenda that addresses short-term pandemic effects while advancing long-term energy transition, economic and resource sustainability. the challenge for policy makers is to avoid implementing policies in haste. properly coordinated policy reforms offer an avenue to address inefficiencies and underlying structural distortions in a way to realize mutual gains and multiple policy objectives at the lowest cost. in designing these reforms and addressing the two areas above, the most suitable research methods are economy-wide general equilibrium models that can quantify effects of shocks and policy solutions in a "second-best" environment. this feature is necessary given the large existing economic distortions in resource economies. these models represent economic linkages and agents and include a wide range of policies (energy, carbon, fiscal, labour or industrial). as such, they can inform evidence-based policy making that accounts for political economic considerations. this research will be critical for filling gaps in the literature on long-term economic and resource sustainability in resource exporters. the current global greenhouse gas (ghg) emissions trajectory indicates that the world is likely to experience catastrophic consequences due to climate change, unless swift action is taken towards funding green solutions and the defunding of fossil fuel activities ( given the ambition of the european union to become a net zero-carbon economy by and the numerous calls to avoid the bailout and stimulus packages towards fossil fuel companies , we examine whether the features of the european central bank's (ecb) € billion pandemic emergency purchase programme (pepp) encourages the resilience of the incumbent fossil fuel sector, or whether it promotes the growth of the emerging low-carbon energy sector during the covid- pandemic and beyond. we draw on a novel dataset of corporate bonds issued in the european energy sector between january and june in combination with the european central bank's (ecb) purchases under the pandemic emergency purchase programme in response to covid- . we show that the likelihood of an energy company bond to be bought as part of the ecb's programme increases with the ghg intensity of the bond issuing firm. we also find weaker evidence that the ecb's pepp portfolio during the pandemic is likely to become tilted towards companies with anti-climate lobbying activities and companies with less transparent ghg emissions disclosure in the event of increased euro-denominated bond issuances in the following months, or re-denominations of non-euro bonds already issued by european energy companies. kainen et al. ). that does not mean, however, that the aim of central banks to remain sector neutral is achievable in practice, as the implementation of ecb's post- quantitative easing shows that assets purchased by central banks to stimulate overall economic growth are benefitting more from the policy than assets which are not purchased by the bank (haldane et al. ; matikainen et al. ) . this means that the choice of asset class through which asset purchasing programs are implemented matters. this is particularly important in the low-carbon economy context, as the fossil fuel energy sector is largely financed through bonds and syndicated bank loans (cojoianu et al. ) , whereas much of the emerging clean technology companies are financed through private equity, equity issuances and asset financing (cojoianu et al. ; gaddy et al. ) . given that the ecb has chosen to enact its asset purchasing program post- crisis predominantly through bonds, this has been shown to favour the incumbent fossil fuel industry (battiston and monasterolo ; matikainen et al. ) , as % of ecb's corporate bond purchases (out of a total of € billion) are in ghg intensive sectorsthough they make up only % of the eurozone area economy and produce % of ghg emissions. the criteria for the corporate bonds bought under the pepp are that: (i) the company must be incorporated in the eurozone and its bond issuance denominated in euro, (ii) the firm cannot be a financial corporation (or a credit institution supervised by the ecb), (iii) it cannot be a public entity, (iv) the bond issuance has to be endorsed by one positive credit rating by an external credit assessment institution accepted within the eurosystem credit assessment framework and (v) they have a maximum maturity of up to years and a minimum maturity of months. in order to understand whether the ecb's bond buying activity during the covid- pandemic has been tilted towards less transparent, more fossil fuel intensive as well as anticlimate lobbying european energy companies, we undertake the following steps. first, we collect all the bonds issued by european energy companies during the period january to june from bloomberg. these span the following energy subsectors as classified by bloomberg industry classification system (bics): power generation, renewable energy, integrated oil and gas companies, oil and gas exploration and production, oil and gas services and utilities. this results in bonds. we then match each bond with ecb's bondholding portfolio, the borrower's record on pro/anti-climate lobbying from influencemap, the ghg intensity of the borrower (collected from bloomberg and measured as thousands tco -e/million eur revenue) and the ghg reporting completeness of the borrower (which is assessed by bloomberg and quantified as if the company is transparent about the organisational boundary it chooses to quantify its ghg emissions and otherwise, bloomberg terminal code es ). we further collect the borrower's revenue (million eur), bond amount issued (million eur) and coupon rate for each bond, also from bloomberg. our resulting dataset with complete data across all variables of interest is comprised of a cross-section of bonds issued across several currencies, and eurodenominated bonds. our dependent variable quantifies the likelihood that the bond of a european energy company is bought by the ecb during the first months of and coded as if it has been bought by the ecb, and if it has not. for our model, we employ a binary logistic regression model with robust standard errors. the full model specification is the following, where ε i is the stochastic error: we show that after controlling for the revenue of the issuer, the bond amount raised and the rate of the coupon, the ecb is statistically significantly more likely to buy the bonds of more ghg intensive european energy companies (models - , table ). on average, a one standard deviation increase in the ghg intensity of an energy company results in a % increase in the likelihood that its bonds are bought by the ecb (β = . , p < . , odds ratio: . , model ). when we consider only euro-denominated bonds (models and ), which are directly under the remit of the ecb, ghg disclosure completeness and pro-climate lobbying are statistically insignificant, yet negative, which suggests that the ecb may be likely to tilt its portfolio towards companies with poorer ghg emission disclosures and less responsible climate lobbying activities. subsequently, we include the bonds issued by european energy companies in denominations other than euro, to account for potential sample selection bias due to the choice of energy companies to abstain from issuing euro-denominated bonds as they may have received discouraging signals from the ecb. in other words, analysing the bond issuance of european energy companies in all currencies considers signals that the ecb may have given to the energy companies prior to issuance, while analysing only euro-denominated bonds only considers the observable decision of the ecb to purchase the bonds of specific energy companies post-issuance. when we do so (model ), it emerges that considering the entire universe of bonds issued by european energy companies, the ecb's portfolio is tilted not only to those energy companies that are more ghg intensive, but also to companies which are less transparent on their ghg performance as well as those companies who are more likely to oppose progressive climate action. having established statistical significance, we investigate the economic and statistical relevance (brooks et al. ) . inspecting the economic relevance of the ghg intensity variable in model , we find ghg intensity to have the largest marginal effects. in terms of statistical relevance, we find ghg intensity to have by far the largest shapley r-squared value, contributing more than % to the overall explanatory power of model . in conclusion, the importance of ghg intensity is underlined by its marginal economic effects and its statistical relevance, as it explains more variation in the dependent variable on its own than all other variables taken together. we also conduct further robustness tests controlling for ecb bond i = + * pro-climate lobbying activities score i + * ghg emissions intensity i + * ghg reporting completeness i + * borrower revenue i + * bond issuance amount i + * bond coupon rate i + i . bond maturity, bond rating and interactions of key variables and find our results to remain statistically significant. in conclusion, drawing on a novel dataset of corporate bonds issued in the european energy sector since january and the database of ecb's purchases under the pepp in response to covid- , we find evidence that the likelihood for a bond to be bought by the ecb increases with the ghg intensity of the bond issuing firm. we also find weaker evidence that the ecb's pepp portfolio during the pandemic is likely to become tilted towards companies with anti-climate lobbying activities and companies with less transparent ghg emissions disclosure. our findings imply that, at later stages of the covid- recovery, an in-depth analysis may be necessary to understand if, and if yes, why the ecb fuelled the climate crisis. even if one accepts that fossil fuel companies were eligible for pepp, then our preliminary evidence still raises the significant question, why the ecb was more likely to directly finance those fossil fuel firms that are likely more harmful to the planet (i.e. have a higher ghg intensity)? the coronavirus covid- pandemic is the defining global health crisis of our time, causing over half a million deaths to date ( july ). but covid- is much more than a health crisis, it has tremendous socioeconomic impact, the scale of which is still hard to assess. measures to address the health crisis generate economic impacts (and vice versa). social isolation measures to "flatten the curve of the pandemic" buy time to increase capacity in the healthcare sector but inevitably deepen the macroeconomic recession. the world bank expect the global economy contract by . % in , approximately three times the size of the - great financial crisis (gfc) and far more widespread. emerging markets and developing economies (emdes) have been severely hit by massive capital outflows, reducing debt servicing abilities (especially for dollardenominated debt) and generating long-term challenges; "with more than % of emdes expected to experience contractions in per capita incomes this year, many millions are likely to fall back into poverty" (world bank ). the economic consequences of the adverse coronavirus shock are: (i) an elevation of uncertainty, which increases precautionary savings, thus reducing consumption and also curtails the appetite for productive investments; (ii) a rise in unemployment, part of which is likely to be permanent; (iii) a decline in the volume of international trade and disruptions in global supply chains; (iv) falls in commodity prices (especially the price of oil), making current account financing of traditional commodity exports challenging; (v) a sharp increase in the required risk premia for holding risky assets. this initially resulted in a plunge in prices of risky assets (e.g. stocks or high-yield bonds) and a sharp increase in financial volatility. however, central bank interventions (especially by the federal reserve of the united states of america-us fed) have seen recoveries from march . extraordinary macroeconomic policy response, on both the fiscal and monetary fronts, has slowed the rates of economic decline. in many countries, fiscal measures have replaced a proportion of lost incomes and mitigated default risk, loan guarantees have helped keep businesses afloat, and liquidity provision by central banks have kept the financial system functional. fiscal authorities in european and the usa took measures that can be classified within three broad categories: immediate fiscal measures and direct transfers to households; tax deferrals and liquidity measures; and loan guarantees. the scale of intervention is truly unprecedented reaching almost % of gdp in germany and % of gdp in the usa. on the monetary front, the fed cut interest rates to zero, announced unlimited purchases of treasuries and mortgage backed securities and started buying corporate debt. moreover, the fed opened debt swap lines with foreign central banks to provide dollar liquidity to the international financial system. the value of fed measures to date exceeds . trillion dollars. the european central bank ecb "has offered low-interest loans to banks, significantly boosted asset purchases, and allayed fears of member-country defaults by lifting distributional restrictions on its bond-buying program" (world bank ). moreover, the eu's recovery plan (proposed may ) mobilizes investments through a recovery instrument of € bn for the period - and a reinforced a long-term budget of € . trillion for the period - . despite these unprecedented measures, the course of the pandemic and its developing economic impact remains uncertain. how can long-term economic dislocation be avoided? a first priority should be to ensure that the work force remains employed. second, governments should channel financial support to public and private institutions that support vulnerable citizen groups. third, small medium enterprises (smes) should be safeguarded against bankruptcy. (the need for taxpayer money to support large nonfinancial corporations is much less obvious.) fourth, policies will be needed to support the financial system as nonperforming loans mount. fifth, fiscal packages, comparable to the loss of gdp, will have to be financed by national debt. while this should be structured to avoid another debt crisis, finance should be directed to investments with positive social, economic and environmentally sustainable profiles, as discussed in the following. there is widespread scientific speculation that economic growth has pushed humanity into new ecological niches wherein humans and animals exchange novel, infectious viruses of which covid- is just the latest in a considerable list of examples. furthermore, the intergovernmental panel on climate change (ipcc) has warned that global warming will likely accelerate the emergence of new viruses. overall, climate change has the potential to end up killing more people than covid- , although this is obtusely referred to as an "increased frequency and severity of natural disasters". such language, the delayed, cumulative nature of the threat and the necessity of coordinated international response, all mitigate against the urgent action that is required. timing is also important. as ipcc ( ) reports, the level and speed of change needed to successfully tackle the climate crisis, is unprecedented; incremental changes will not be enough. that said, there are aspects of the climate change crisis which are less challenging than the covid- pandemic. as sterner ( ) notes, the climate crisis requires policy changes that are less disruptive, economically, socially and culturally, than the measures being taken right now to tackle covid- . for the climate, we do not need to close down the economy. on the contrary, we need a transition to a low-carbon economy that supports public and private investments in renewables, energy efficient and circular, technologies and infrastructure. these technologies exist (wind, solar, etc.) and are becoming consistently cheaper than fossil fuels, while energy storage installations are increasing exponentially. there is growing evidence that green stimulus policies have advantages over traditional fiscal stimulus and that climate-positive policies also offer superior economic characteristics. for example, hepburn et al. run a global survey to assess stimulatory fiscal recovery policies implemented in response to gfc. the economists from countries were asked to ascertain their perspectives on covid- fiscal recovery packages according to: "speed of implementation", "long-run economic multiplier", "climate impact potential" and "overall desirability" according to social, political and personal factors. the responses indicate that green stimulus policies deliver higher multiples due to reduced long-term energy costs and flow-on effects to the wider economy. "relaunching the economy does not mean going back to the status quo before the crisis but bouncing forward. we must repair the short-term damage from the covid- crisis in a intergovernmental panel on climate change, . way that also invests in our long-term future". this code explains the strategy of the eu recovery plan based on three axis: the european green deal, adaptation to the digital age, and a fair and inclusive recovery for all. we must start investing in what makes our socio-economic system resilient to crisis. now is the time to usher in systemic economic change and the good news is that we have our blueprint: it is the combination of un agenda ( sdg) and european commission's european green deal. the european green deal (egd) announced december is the new growth strategy of the european union and is based on four principles: (a) climate neutrality by , (b) protection of human life and biodiversity by cutting pollution, (c) world leadership in clean technology, (d) leave no one behind. the egd investment plan amounts to eur billion per year by . the egd just transition mechanism will help mobilize at least € billion over the period - in the most affected regions. the - multiannual financial framework (mff) allocates an overall target of % for climate mainstreaming across all eu programs. the proposed european climate law aims to turn the egd's political commitment of climate neutrality, into a legal obligation. based in un environment program (unep) emissions gap report , global emissions need to be reduced by % by . unfortunately, the proposed law does not include an ambitious goal with regards emissions, nor does it address the legislative interventions required to achieve climate neutrality by . together with the egd, the sustainable development goals (sdgs) and the paris agreement call for deep transformations. while significant progress is being made on some goals, no country is currently on track towards achieving all sdgs. sachs et al. ( ) identify the major interventions needed to achieve each sdg and group them in six sdg transformations, which operationalize the sdgs at government level and can prove instrumental for egd implementation. the success of the join implementation of the sdgs, the egd and the eu recovery plan will depend on the eu's capacity to engage with its citizens in codesigning the pathways that will allow them to reach the vision, hence the introduction of the european commission climate pact. https ://ec.europ a.eu/info/live-work-trave l-eu/healt h/coron aviru s-respo nse/recov ery-plan-europ e_en. european parliament, committee on environment, public health and food safety draft report on the proposal for a regulation of the european parliament and of the council establishing the framework for achieving climate neutrality and amending regulation (eu) / (european climate law), / (cod). the review of the climate law summarizes the review of the climate change committee of the greek ministry of energy and the environment, the first author of this paper is a member of this committee and co-author of this greek review. united nations environment programme ( ). emissions gap report . unep, nairobi. the proposed climate law does not allow the european commission to impose sanctions on member states (ms), while there is lack of reference to the financial mechanisms that will be required to achieve the goal of climate neutrality. moreover, the law does not consider (a) the well-documented heterogeneity in area-specific climate vulnerability among ms, (b) an eu wide carbon tax and (c) strengthening the european ets. sachs et al. ( ) : the sustainable development goals and covid- . sustainable development report . cambridge: cambridge university press. ( ) education, gender and inequality; ( ) health, well-being and demography; ( ) energy decarbonization and sustainable industry; ( ) sustainable food, land, water and oceans; ( ) sustainable cities and communities; and ( ) digital revolution for sustainable development. https ://ec.europ a.eu/clima /polic ies/eu-clima te-actio n/pact_en. recent generations, including the present, have experienced at least three global crises: the financial crisis - , the ongoing covid- pandemic and the developing climate crisis. if we continue attempting to address the latter two with the same socio-economic model that gave rise to the former crisis, we will fail to find a sustainable and resilient socio-economic-environmental pathway. we believe that we can even do better than just react to crises by adapting to the new crisis-born reality. we can use the integration of scientific, economic and socio-political knowledge to design policies which not only address the immediate impacts of the covid- pandemic, but mitigate the existential threats of future pandemics and the ongoing and unfolding disasters of climate change, biodiversity loss and planetary boundary exceedance. what is needed now is a fundamental transformation of economic, social and financial systems that will trigger exponential change in strengthening social, economic, health and environmental resilience. we need big thinking and big changes. system innovation and transitions thinking can help but calls for intense public participation. now is the time, in addition to directing funds to the control of the epidemic and relevant biomedical research, as well as investing in border security, safe travel and safe trade, now is the time for financial institutions and governments to embrace eu taxonomy for sustainable investments ( ) to phase out fossil fuels by deploying existing renewable energy technologies, eliminate fossil fuel subsidies and redirect them to green and smart climate mitigation and adaptation infrastructural projects, invest in circular and low-carbon economies, shift from industrial to regenerative agriculture and invest in food security, promote european supply chains, reduce transportation needs and exploit the limits of the digital revolution, while ensuring secure information and communication technology networks. a decisive march along this sustainable pathway will enhance economic and environmental resilience, create jobs, and improve health and well-being. the transition should be inclusive and "leave no one behind", hence the need of transforming citizens into codesigners and co-owners of the sustainability transition pathways. kiel institute for the world economy, kiellinie , kiel, germany. wilfried. committing to rigid shutdown measures to contain the spreading of the corona virus has been undertaken on the tacit assumption that these measures will be temporary and can be loosened when the covid- infection rates decrease and discontinued altogether once vaccines are available. mitigating climate change and achieving ambitious temperature targets as set out in the paris agreement require a long-term structural change taking us away from our current carbon-intensive economy to a zero-carbon and then net-negative carbon economy. as current research holds out little hope that a "perfect" vaccine in the form of solar climate engineering will be available in the future, the measures and efforts required must translate into a permanent, ongoing form of commitment. while progressive climate change and the spread of the coronavirus operate on very different timescales, impatience about the duration of corona lockdown has indicated once more a fundamental problem for (long-term) environmental concerns. clearly, the economic and social costs associated with the emergence of the virus and the shutdown are significant , oecd . but any serious cost-benefit analysis would need to take into account not only the fact that different degrees of lockdown are available but also that the overall cost is affected by the expectations of agents regarding possible future re-lockdowns due to insufficient containment of the virus. seen thus, it is anything but clear at which point in time the actual cost of lockdown would have exceeded the economic cost of the virus spreading in an unmitigated (or insufficiently mitigated) way. during the course of lockdown measures, voices calling for a "green" recovery stimulus package centring around low-carbon investments in the aftermath of the corona crisis have make themselves heard. by contrast, advocates of postponing climate mitigation-related taxes, levies, and regulations have also entered the fray, claiming that timely recovery should not be jeopardized by any additional economic burdens. the debate on the relation between (economic) recovery and climate policies has been conducted from three major perspectives. the first of these is largely notable for general statements of intent recommending that the recovery should be "green" and sustainable, that eu climate targets should be supported, and that other environmental targets (maintaining biodiversity, etc.) need to be taken into account when designing recovery measures. such well-meant counsels as the statement issued by the german national academy of sciences leopoldina ( ) are useful in reminding us that recovery from the corona crisis should not come at the expense of neglecting other objectives and that climate policy should not be backburnered, as was the case after the financial crisis in . otherwise, they are of little practical value. the second approach has involved rather detailed proposals calling either for a "greening" of recovery by foregrounding measures to support renewable energies, public transport, energy efficiency, etc. or for a "blackening" of recovery by postponing and/or abandoning climate measures and environmental regulations. predominantly, these proposals are representing the positions of the various interest groups involved. for example, representatives of the aviation industry try to prevent the harmonization of carbon prices on fuels with respect to kerosene and argue against the introduction of kerosene taxes. this idea resurfaces in the discussion on recovery measures by, say, the austrian aviation association ( ). on the other hand, in its comprehensive list of (recovery) demands, the ngo german environment action ( ) urges for example for the abandonment of blue hydrogen projects (though not explaining why this is likely to stimulate economic recovery). various other interest groups are in favour of postponing, suspending or even abandoning existing environmental and climate regulations. for example, janusz kowlaski, the polish deputy minister of state assets urges "…[that] the ets [european emissions trading scheme] should be removed from january , or at least poland should be excluded from the system". clearly, there is no point in discussing nonsensical ideas of this kind. but some of these proposals also make sensible suggestions like adjusting the german cap on renewable energy installations or abandoning the eu average fleet-consumption regulation because the former contradicts german renewable energy targets and the latter is an inefficient instrument for regulating vehicle emissions. however, these suggestions do nothing to provide stimulus for a quick recovery. while specific processes and regulation timelines for regulations may need to be adjusted in the context of the corona crisis, sensible measures of this kind should be discussed and decided upon in the regular political process. confining potential stimulus and recovery measures to their proper purpose does not mean imposing a ban on meaningful (climate or environmental) policies that are not associated with the corona crisis. the third and most sensible perspective replaces specific proposals with (sustainability) assessment guidelines like those suggested by the world bank ( ). while hardly any possible recovery measure would perform well against the comprehensive list of criteria provided by the world bank, such guidelines are helpful in arguing against interest group driven proposals. the world bank has suggested that potential measures up for consideration as part of a recovery strategy need to be assessed against both, short-and long-term criteria, an example for the former being the expected economic multiplier associated with certain measures. bayer et al. ( ) suggest that income transfers (as planned under the us cares package) perform well against this specific short-term criteria: they could help to stabilize private-sector spending and the multiplier could increase to in the case of transfers being conditional-but not related to emissions but to the propensity to consume, i.e. conditional on being unemployed. however, privatesector spending like this should not imply any unintended adverse effects on essential long-term structural change that might arise from such things as (temporarily) adjusted risk preferences. once postponed or stimulated demand and investment take place during the recovery process, carbon-price signals are vital in providing technology-neutral incentives for low-co purchasing and production decisions. overloading stimulus or recovery packages with too many (emission-related) conditions performs poorly against the short-term criteria with respect to a timely recovery. even worse, the inclusion in recovery packages of various detailed suggestions from the various interest groups usually results in a non-transparent, rent-seeking, and political bargaining process in which it remains unclear whether (sensible) individual emissionrelated decisions are being prioritized at the expense of a more challenging long-term climate policy. accordingly, accounting for the long-term criteria requires that existing or planned climate policies providing incentives for emission reductions and technological innovation should remain in place and not be postponed, let alone weakened. otherwise, uncertain (short-term) recovery impulses most likely come at the cost of less efficient emission-reduction paths in the long term. the coronavirus pandemic is having a serious impact on the economy. eurostat estimates that seasonally adjusted gdp decreased by . % in the eu during the first quarter of , compared with the previous quarter. in certain sectors, the covid- crisis has led to the temporary or even permanent closure of sites, a collapse in demand and an increase in production costs linked to the fight against the spread of the virus. this decline in activity can trigger bankruptcies, which result in unemployment and the destruction of physical and human capital, leading in turn to a loss of specific knowledge and skills and to market concentration and the partial relocation of production activities. from an environmental point of view, bankruptcies and relocation may generate higher emissions by increasing production in more polluting sites abroad and by increasing the transportation of goods. in addition, the decline in economic activity may have a negative effect on research and development spending, which is crucial for future growth and the development of more environmentally friendly technologies. many measures have been put in place to limit the economic effects of the covid- crisis. at the european union level, a number of measures such as direct subsidies, selective tax benefits, advance payments, state guarantees for loans and subsidized public loans to companies have been implemented. in addition, many large companies are being bailed out and there is a debate on whether conditions should be attached to this state aid. these conditions could relate to various commitments such as relocating activities, paying taxes, safeguarding employment, and protecting the environment. on may , the eu confirmed that large companies receiving emergency cash during the covid- crisis will not be obliged to devote funds to "greening" their operations. rescued firms will only have to report on their use of the aid and in some cases the aid could be attached to conditions such as a ban on dividends and management bonus payments. nevertheless, member states are free to design national measures in line with additional policy objectives, such as further enabling the green transformation of their economies. many stakeholders are still pushing for conditions to be attached to the bailouts. for instance, the eg think tank proposes making the rescue of airlines conditional on the use of less polluting fuels and tying aid to car manufacturers to the development of electric vehicles. a group of german companies has also requested that state aid be linked to climate actions. this article examines the merits of making aid conditional on environmental efforts. we focus on the rescue plans for companies rather than on recovery plans or the european green deal, which was launched before the covid- crisis. we show that tying aid to environmental efforts is difficult to implement and requires both controls and sanctions, as well as a large amount of information. we also discuss the merits of tying bailouts to environmental efforts compared with implementing more stringent environmental policies (presuming that companies will be bailed out in any case). since environmental efforts can lead to higher product prices and lower profits, two dimensions of acceptability must be taken into account: acceptability from the point of view of companies and social acceptability. acceptability of environmental policies by companies is a necessary condition for their implementation. in the midst of the current crisis, many polluting industries are lobbying to weaken and delay environmental regulations. for instance, the environmental protection agency issued a sweeping suspension of its enforcement of environmental laws, allowing companies to breach environmental standards during the coronavirus outbreak. in indonesia, the trade ministry revoked rules requiring basic certification that wood exports were legally produced, in response to lobbying from the furniture and logging industries. lobbyists may also encourage legislators to compensate companies for losses related to environmental regulation. the implementation of rescue plans, by providing aid and guarantees, could be a tool to make environmental efforts more acceptable for firms. acceptability by society is also central. it depends on the redistributive effects of environmental policies, especially since they are often regressive. in france, the "red caps" and "yellow vests" movements illustrate the difficulty of implementing such policies. both of these movements led to the cancellation and delay of environmental policies (eco-tax applied to heavy vehicles and carbon tax, respectively). the covid- crisis may affect societies' support for more ambitious environmental policies. several observations can be made. first, it appears that there is a confirmation bias, as countries and ngos that had already committed to fight against global warming want to intensify efforts, while countries that were already recalcitrant are calling for a decrease in efforts in the light of the economic crisis. second, the covid- crisis makes the risk of disaster more salient and vivid and may therefore increase the demand for stronger environmental protection. as explained by sunstein ( ) , if a particular risk is cognitively "available" then people will have an increased fear of the risk in question. finally, the crisis has revealed that populations will accept drastic measures (lockdown, wearing face masks) and a rapid change in social norms (social-distancing, for instance), which shows the ease with which individuals could adapt to more ambitious environmental policies. see oates and portney ( ) for a literature review of interest groups and environmental regulation. "polluter bailouts and lobbying during covid- pandemic", the guardian, april , . burkey and durden ( ) and joskow and schmalensee ( ) have detailed how firms can influence the regulator's decisions in the context of pollution rights markets, while bovenberg and goulder ( ) , hepburn et al. ( ) and nicolaï ( ) have shown that few permits are sufficient to neutralize the losses in profits and make the implementation of pollution permits acceptable. on march , the czech prime minister, andrej babis said that the european green deal should be put to one side. during the same period, the green coalition of environmental organizations organized an appeal urging lawmakers to design a green, healthy and just recovery. we study two possible ways of increasing environmental efforts: making aid to companies conditional on environmental efforts or making existing environmental policy more stringent. making aid conditional on efforts to protect the environment can take different forms. commitments may relate, for example, to reducing pollutant emissions, using less polluting production processes, increasing expenditure on research and development, and producing less polluting products. for instance, in france, renault's bailout requires it to increase the share of electric vehicles, while the billion euro air france bailout imposes the use of at least % of alternative jet fuel by , a target of % emissions cuts by , and a % decrease in domestic flights by , especially those that compete with high-speed trains. conditional aid as currently envisaged is equivalent to negotiating and implementing individualized standards on technology, final products, research and development efforts, emissions and performance for each company. the first question that arises is how to determine the conditions. conditional bailouts allow the regulator to individualize the standards applied to a particular firm, making it possible to set the most appropriate instrument and the optimal level of severity for each firm. when making its decisions, the regulator should consider the possibility of a rebound effect as explained by saunders ( ) . an emissions standard may avoid such an effect, but a standard on technology may increase emissions. furthermore, the regulator should take into account its access to information. if the firm commits to finance research and development in green technology in exchange for being bailed out, it will be hard for the government to determine whether the state aid was actually allocated to research. for example, since the results of research are uncertain, a lack of innovation may be explained either by the firm's characteristics (anti-selection problem) or by the misallocation of public funds (moral hazard problem). finally, the regulator should take into account competition, since firms receiving bailouts usually have market power. a government will mainly intervene to save a firm if its bankruptcy would have a significant effect on the economy. the government should hence ensure that the conditions do not further distort competition. a firm with a competitive advantage, for example with cleaner technology, may have an interest in advocating more ambitious environmental efforts for itself but also for its competitors in order to increase its market share by preventing the entry or inducing the exit of such competitors. the effectiveness of conditions will therefore depend on the type of commitment, competition in the market, and the demand elasticity of the final good. another question relates to the duration of the contract. some actions are reversible and after a certain period of time companies can cancel or amend them. for example, the use of cleaner but more expensive inputs is an easily reversible strategy, while a shift in production, for example from internal combustion vehicles to electric vehicles, is a more expensive choice to change. in addition, research and development efforts produce long-term effects with the creation of innovations and possible technological spillovers. the duration must therefore be individualized in function of the company's characteristics. it should also be noted that conditional aid can lead to windfall gains. some companies had already planned emission reductions and changes in strategy. these decisions were aligned with the companies' interests. if the company benefits from the environmental efforts put in place, these can no longer be considered as a counterpart to the company's bailout and the company should then make greater efforts. it seems clear that the success of such schemes lies in companies' compliance with the commitments. companies may nonetheless have an interest in not complying with them. it is therefore necessary to monitor companies' implementation of the commitments and to sanction those that fail to honour them. supervision costs will thus be incurred. one possibility for alleviating control costs is to publicly announce the commitments, which facilitates the monitoring of companies' commitments by society (e.g. by journalists, politicians and non-governmental organizations). this also gives the firm a further incentive to respect the agreement in order to maintain its reputation and to retain its customer base (heyes et al. ( ) and heyes and kapur ( ) ). we now focus on the sanctions to be applied in the event of non-compliance with the commitments. various penalties could be considered, such as financial penalties, a ban on applying for the attribution of public contracts and the state's participation in the company. the sanctions must be explicit, credible and sufficiently significant to discourage companies from breaking their contracts. however, given the current economic situation and the fragility of companies, governments should be cautious when using financial penalties or bans from participating in public contracts since, if effective, they could be fatal to companies. an alternative could be for the state to take a share in the company in the event of non-compliance with commitments. such shareholdings would make it possible to influence companies' strategic choices, but they raise efficiency problems. cavaliere and scabrosetti ( ) , schmidt ( ) and de fraja ( ) , for instance, highlight two effects: production is more efficient in privatized firms because better incentives can be given to managers and employees (productive efficiency), while public firms are more socially efficient because the government cares about social welfare and internalizes externalities associated with firm liquidations (allocative efficiency). existing environmental policies inevitably generate monitoring costs; however, these costs will be borne in any case. increasing the stringency of environmental policy involves determining new levels for existing instruments, which generates costs, but there will be no additional monitoring costs. moreover, opting for a more stringent environmental policy means that market-based instruments can be employed, which is not feasible in the first strategy. due to the principle of non-discrimination, it is unlikely that individualized taxes could be introduced under the negotiated bailout conditions. environmental economists agree that these instruments are more effective than command-and-control instruments since they have the potential to generate revenue and allow for a double dividend, provide incentives to invest in clean technology and require less information to be designed. nevertheless, making aid conditional on environmental efforts makes it possible to individualize the regulatory instruments for each firm, which is not possible in the context of marketbased instruments. the positive effect of the instrumental individualization induced by conditionality only materializes if the existing environmental policy is based on commandand-control instruments. with market-based instruments, individualization is not required to achieve an efficient outcome, whereas it is necessary in the case of command-and-control instruments. companies' negotiation power is an additional argument for increasing the severity of environmental policies rather than conditioning bailouts on environmental efforts. regardless of the policy, firms may use their bargaining power to mitigate the stringency of the regulation or the effort they commit to make. it is clearly easier for a firm to influence its specific negotiated conditions, which are one-on-one relationships, than to influence environmental policies, which would require coordination and a common interest among firms. furthermore, making environmental policy more stringent will affect all companies and not only those receiving bailouts, which increases the attractiveness of this strategy. since they apply to a larger number of companies, these more restrictive policies will have a greater effect on the environment. to conclude, this note analyses the advantages and disadvantages of making bailouts conditional on environmental efforts. we show that such a system could be beneficial for the environment and would be a counterpart to bailing out companies. however, it is costly, requires a large amount of information, and is less efficient than increasing the stringency of existing environmental policies. on the other hand, making aid conditional on environmental effort is more acceptable from the companies' point of view than increasing the severity of environmental policies. companies would rather commit themselves to environmental efforts than pay taxes or buy permits in order to minimize compliance costs. to succeed in making environmental policy more ambitious, the regulator needs the support of society to compensate for companies' reluctance. if there is strong public support, governments should not negotiate conditional aid but should increase the severity of environmental policies. they can integrate such actions into recovery plans or, in the case of the european union, into the revision of the green deal. it is therefore particularly important to study how the covid- crisis has changed society's perception of the need for more stringent environmental policies. for instance, a recent survey by ipsos shows that three out of four people in major countries expect their government to make protection of the environment a priority when planning a recovery from the coronavirus pandemic, although there is considerable heterogeneity across countries. the post-covid- reality is changing the context of most policies, including those in the fields of energy and the environment. the eu green deal objectives should be maintained but concrete policies may have to be adjusted to this new reality. a major energy transition is under way, shaped by political will to tackle climate change. policies have been defined under the paris agreement and geared to a number of targets. the eu has decided to reduce its greenhouse gas emissions by by at least % compared to and has agreed to continue the path towards climate neutrality by . the immediate recession following the covid- crisis drastically reduced energy consumption and greenhouse gas emissions (le quéré et al. ) . the sudden decline in greenhouse gas emissions is the opposite of what a meaningful response to climate change should be, in terms of both quality of life and economic efficiency. the challenge is to structurally decouple economic growth from emissions, not to have them both going down. the lockdown may have some lasting effect in changing some of our habits ), e.g. a generalized familiarity with telework. one can also expect a more general reflection on whether all business and leisure travel, not least by air, is necessary. nevertheless, many changes may not be as extensive and lasting as some would wish. for this reason, it is important to integrate a climate check into the stimulus packages that governments and european institutions are currently designing. the current health crisis and the likely economic downturn could be seen as an unsolicited-and much regrettableopportunity towards a carbon-neutral future koundouri ) . the agreements reached in the context of the eurogroup, the latest turn in the european budgetary discussions, as well as the statements by the commission president all point into this promising direction. the eu green deal has confirmed the eu emissions trading system (ets) as a key element and the price of eu allowances serves as a key indicator worldwide. as could and should be expected, carbon prices have fallen during the lockdown reflecting reduced demand for allowances in line with the drop in power and industrial production. this temporary drop from the € to a € - range, acts as an important automatic stabilizer for businesses in distress. a repetition of the experience of a sharp fall in the see on this issue the important draft report prepared by the european parliament committee on the environment, public health and food safety (european parliament, ). https ://lifed icetp rojec t.eui.eu/ / / /covid - -clima te-polic y-and-carbo n-marke ts/. https ://www.consi lium.europ a.eu/en/press /press -relea ses/ / / /repor t-on-the-compr ehens iveecono mic-polic y-respo nse-to-the-covid - -pande mic/-point : "work is ongoing on a broader roadmap and an action plan to support the recovery of the european economy through high quality job creation and reforms to strengthen resilience and competitiveness, in line with a sustainable growth strategy". https ://ember -clima te.org/carbo n-price -viewe r/. price of eu ets allowances after the recession and a long period of very lowcarbon price levels should be avoided. there are two reasons to believe that this could happen: (i) the eu ets now has a market stability reserve (msr) operating in the short-medium term and (ii) long-term market expectations could be shaped by the carbon neutrality objective. the msr started to operate last year and absorbs a potential oversupply in the market in the short and medium term. if the allowances market is "long" beyond a threshold set in legislation ( million tons, which is approximately % of ets emissions in ), the msr intervenes by withdrawing allowances equal to a percentage ( % up to , % thereafter) of the excess. at the end of , the allowance market was . billion tons "long" and, therefore, the supply of allowances for the period september to august has been reduced by million tons (these allowances are placed in the msr). some question whether the msr, with its delayed adjustment mechanism, will be sufficient to avoid a sharp fall in the allowance price (cf. flachsland et al. ) . in fact, any reduction in the supply of allowances through the msr for the period september -august will still be based on the excess supply of allowances in , i.e. before the current crisis started. in case the msr is unable to absorb the surplus, the eu commission should propose changes in the planned review of the ets in . long-term expectations might overtake this short-term risk. in its european green deal, the commission already indicated its intention to tighten the greenhouse gas emission reduction target from % to %, or even % (with respect to levels). today, the ets sectors face a mandatory emission reduction of % by compared to , and revised targets will likely increase that obligation by at least another or %. such a reduction in the supply of allowances would likely push their prices up, or at least reduce the extent of their decline, in the face of lower demand due to the economic downturn. therefore, carbon market participants as well as those developing innovative clean technologies and products of the future are likely to expect significantly higher carbon prices in the future. this raises the question about a possible loss of competitiveness in europe's traditional industries vis-à-vis competitors located in countries where no similar burden is imposed. some proposed a border adjustment, which would imply both a levy on imports and possibly a rebate on exports. however, such mechanisms are not easy to implement and are subject to criticism on both analytical and political grounds. therefore, it is of crucial importance to realize that a wto-compatible border adjustment mechanism will take some time before it can be implemented. meanwhile, at least equal attention needs to go to domestic policy reinforcement, such as support for innovation and a less rigid interpretation of the state aid rules. governments and european institutions are currently developing a major investment stimulus package. the commission's impact assessments on climate and energy policy serve as useful guidance on where a major surge in investments is needed. we know that energy efficiency requires a major push in the construction sector, both for new buildings and renovations in social housing, hospitals and schools. we know that the energy transition requires more investments in renewable energy, digitised grid infrastructure and energy storage. the significantly reduced electricity demand of today indicates that much more attention should go towards managing flexibility in real time instead of increasing baseload capacity. in transport, electrification is on its way, but investment in charging facilities, traffic management, clean public transport and long-distance rail needs to be ramped up. in industry, not least in power generation and carbon intensive industrial sectors, major efforts are being undertaken to develop new technologies based on hydrogen and carbon capture, use and storage, for example. such investments can create the jobs we need in the post-covid- era and allow to realise the eu green deal objectives at the same time. apart from carbon prices and investment support schemes, it remains of equal importance to maintain a consistent energy price signal throughout the economy. however, the recent fall in the prices of fossil fuels is upsetting the incentives that should support a transition towards sustainability. a reference criterion for economic decisions has been lost and the consequence may be a paralysis, or even a "comeback" of fossil fuels, mothballing or abandonment of investment in renewables. consequently, the business case of the green transition may appear to be weakened in the near term, but a clear policy response may be politically more acceptable because of the price drop in fossil fuels. this raises the question about the other half of europe's emissions, namely those not covered by the ets, in particular the sectors of transport and buildings. the pricing of these emissions can be much improved through the planned review of the eu's energy tax directive. this offers a major opportunity to remove the all too generous exemptions, such as on maritime and aviation fuels, or by adding a co element to the harmonized minimum tax rates. in such a manner, prices of fossil fuels could be stabilized at the pre-covid levels via temporary taxes, whose revenue could help finance the various income-support policies adopted and/ or an acceleration of investment projects for sustainability. in this way the eu should better prepare for a long period of low oil and gas prices and seize the opportunities these can offer. public measures to combat the covid- pandemic have led to a severe economic crisis. in order to cope with this crisis, comprehensive government aid is being requested. accordingly, governments across the world have pledged billions of euros for extensive recovery programs. one of the main questions debated in this context at the moment is how "green" these recovery programs should be ). the expectation of huge amounts of public money being distributed at short notice brings interest groups of every shade to the scene-preferably with old wish lists on hand. consequently, there is a big risk that recovery programs will be captured by interest groups (for an overview of the literature on regulatory capture, see dal bo ). on the one hand, climate change mitigation is put under pressure as being an "extra burden" for industries. for example, european car manufacturers have called for postponing the upcoming tightening of eu emission standards for car fleets (topham and harvey ) . some eu member states call for stalling the eu commission's plan of a european green deal (simon ). on the other hand, many recommend spending the public money mainly on measures that also help mitigating climate change-among them frans timmermans, executive vice-president of the european commission (schulz ), or fatih birol, head of the international energy agency (birol ) . there is one thing that must not be overlooked in this politico-economic competition: public funds are still short and must be used reasonably. otherwise ill-designed (green) subsidies can quickly turn into a part of the problem instead of being the solution. previous "cash for clunkers" programs warn as an example of a misguided recovery measure. these programs were introduced in many countries after the financial crisis and provided financial incentives to trade old, less fuel-efficient cars for new, more efficient ones. empirical analyses have shown very mixed results regarding both the economic and the environmental stimulus effects of these measures (grigolon et al. ; li et al. ; mian and sufi ) . at the beginning of every discussion about (green) recovery programmes, it is therefore important to develop transparent and sensible criteria based on which public aid should be allocated. after the initial bail-out programmes, public recovery programmes to stabilize the economy are now debated politically. certainly, this generates an unprecedented window of opportunity for structural transformation. moreover, the distribution of public aid may also justify committing beneficiaries to public interests to a certain extent. consequently, the currently available political degrees of freedom should be used to promote the transition of society towards sustainability. subsidies to branches like tourism, aviation and agriculture-which are particularly hit by the crisis and are lagging behind in terms of sustainability-should be paid conditional on meeting minimum environmental standards. new investments into long-lived, fossil-fuelled assets must be avoided. a recovery program cannot only be about re-establishing the status quo ante by assigning large public funds, possibly creating new barriers for sustainability transitions. in this respect, it makes sense to implement recovery programs that are in line with the objective of mitigating climate change-as called for by many at the moment. however, such green recovery programmes must not be arbitrary. green recovery programmes must go beyond green subsidies. first of all, it is also important to reduce unnecessary barriers for green investments, for example by revising legal constraints to the expansion of renewable energies like solar photovoltaics of wind power. moreover, any green recovery program can only effectively and efficiently spur decarbonization if it combines with a carbon price and the abolition of environmentally harmful subsidies. the direction of recovery must be crystal clear. otherwise green subsidies risk being ineffective and costly approaches to mitigating climate change (kalkuhl et al. ; palmer and burtraw ) , while imposing additional burdens on public budgets and reducing political degrees of freedom in the future. for subsidies to be economically justified, they need to meet clear criteria. for green recovery programs to succeed in the competition for public funds with other important policy fields (such as health or digitalization), they must help stabilize the economy. moreover, policy makers need to be aware that some of the currently observed economic problems might even resolve without any government aid. it can expected, for example, that global supply chains will resume and that people will catch up on purchasing durable goods like cars, at least partly. it is exactly (the maintenance of) environmental regulation that may help steer this consumption towards more sustainable modes. government interventions must take effect where permanent disruptions are looming. one example: innovative green business models may particularly be at risk if banks limit loans in the presence of the current uncertainties (lehmann and söderholm ) . in this case, government loans may provide direct assistance. in contrast, attempts to lower prices for goods and services-e.g. for cars (vat reduction, purchase premiums) or electricity (reduction of energy levies)-are rather inappropriate means to stabilize the economy. such measures fail to address the actual sources of insufficient investments or reduced purchasing power, and are therefore inefficient ways of spending public budgets. furthermore, it is unclear whether and to what extent such discounts will be passed through to final consumers by market prices (peltzman ) . green recovery programs should focus on government interventions that would also have been economically reasonable without the covid- crisis-for example, to correct market failures next to the co externality (bennear and stavins ; fischer and newell ; lehmann )-and that have the highest priority for climate policy. moreover, those measures should be implemented for which rational concepts have been drafted already and that can be realized promptly. positive examples of such "no-regret measures" can be found in the transport sector, for instance. this sector is severely lagging behind in terms of climate change mitigation, and economic rationales for public expenditures exist at least partly (briggs et al. ; low and astle ). in addition to that, numerous actors have already developed elaborated programmes of measures. those measures that can be implemented quickly, should now be launched-for instance to electrify the transport sector or to strengthen public transport. (green) recovery programmes must not only address the expenditure side. a currently still disregarded issue is the question how the required billions of euro could be raised. public expenditures for a green recovery program should at least partly be funded by polluters by implementing a carbon price and abandoning ecological harmful subsidies. such policies internalizing environmental costs would not be an extra sacrifice-but rather part of the solution both for revenue problems and for the redirection towards sustainability (for a review of the double-dividend hypothesis, see goulder ). the coronavirus crisis has opened up a window of opportunity for transformation. this should be used without getting off the regulatory track. green recovery programs must not be reduced to a mere competition for green subsidies. abandoning barriers to green investments and imposing a carbon price are equally important. where economically sensible, green subsidies should contribute to both stabilizing the economy and mitigating climate change. moreover, smart green recovery programs may contribute to raising revenues for the additionally necessary public expenditures. the world is currently facing the largest pandemic since spanish flu in . this has led to a lockdown policy on an unprecedented scale and measures of social distancing that are expected to continue. in france, as elsewhere, the "great lockdown" has disrupted food production chains through simultaneous shocks to demand and supply. populations have seen their food consumption habits be severely modified (e.g. closure of restaurants and markets). fearing food shortages, consumers have often stockpiled basic necessities, which has led them to actually provoke shortages, albeit temporarily. food production has also been disturbed, most notably by the reduction in the available workforce, whether domestic (due to the lockdown, illness or childcare) or foreign (temporary closure of borders). the issues at stakes are numerous, including deglobalization and environmental impacts, in both the long and short terms . the pandemic will lead to unprecedented uncertainties in food supply chains. in addition to the health crisis, a dramatic drop in worldwide gdp is anticipatedapproximately %-with a partial catch-up in , according to the international monetary fund (imf ). the reduction in income will soon impact food consumption. some authors are already pointing to an increase in social inequalities. the most fragile populations could slide into severe food insecurity, including in developed countries, as shown by deaton and deaton ( ) for canada and power et al. ( ) for the uk. however, the fao is rather reassuring about the total volume of food (cereal) at the global level. it also notes that the world price index has fallen in recent months. this does not necessarily mean that this decline is effective regardless of the type of agricultural production. in particular, fruits and vegetables could become more expensive. agriculture is by nature a risky activity, and the adoption of eco-environmental practices can only increase this risk. as a consequence, organic farmers, for instance, could be particularly affected by the combination of health and economic crises. the cost of organic production is therefore structurally higher, which could pose a problem in the event of an income shock. to add value to their production, many small farmers combine organic farming with short food supply chains. this makes it possible to regain a more comfortable margin than that allowed by mass distribution. however, here again, the health crisis has profoundly changed marketing channels. while supermarkets were stormed with shoppers at the beginning of the crisis, some short channels were closed, such as markets and restaurants, to preserve social distancing. this article therefore considers the consequences of covid- for sustainable agricultural practices, particularly for farmers who have chosen organic farming in short supply chains. to do so, we use the multi-level perspective (geels ) approach to conceptualize socio-economic transitions. the multi-level perspective (mlp) approach analyses transitions as mutation processes from one socio-technical regime to another under the pressure of macrolevel forces and the emergence of market niches that could provide the basis for a new regime (geels ) . we show here why local food supply chains are perceived by some farmers-especially the smallest-as a way to enhance the economic value of eco-friendly agricultural practices, such as organic farming. in france, the current dominant socio-technical regime based on conventional agriculture emerged gradually in the s. conventional agriculture diffuses slowly at first because of the high investment costs. however, the french state and farmers' unions support it as a means of increasing production and improving living and working conditions. farmers are then inclined to take on more debt and become dependent on their suppliers (phytosanitary products, seeds, tools, etc.), the food industry and supermarkets. as a result, this modernization of agriculture has been controversial from the outset, at least with regard to farmer autonomy and the country's food sovereignty (levidow et al. ). the early development of organic farming in france began in the s due to the effects of three negative effects of conventional agriculture, namely pollution, soil impoverishment and the lack of autonomy mentioned above. the organic label has become a marketing argument that is profitable for large farms, since they achieve significant scale effects on these standardized products. large scale organic farming is therefore less demanding and, by its nature, allows to benefit from the effects of scale. this is new competition for small farmers struggling to differentiate themselves. the organic label itself does not allow farmers to free themselves from the pressures upstream and downstream in the production chain. to differentiate themselves from large organic farms, some farmers are starting to sell in open-air markets again. at the same time, the idea that fair trade can concern north-north relations and not only north-south relations is beginning to emerge, which favours the development of community-supported agriculture. a new initiative is gradually emerging: collective farmers' shops. these initiatives began, for example, in the south of france in the mid- s and represent a more important restructuring of the farmers' market. in the medium term, the drop in income could increase the consumption of basic necessities (giffen goods) to the detriment of organic products. everything will depend on the elasticity of the demand for these products. until now, organic products have been extremely popular. it is possible that "industrial organic" will take market share from a "more artisanal organic" if consumers still want to consume organic products but cannot afford to spend a large part of their budget on them. farmers in short supply chains often have several distribution channels simultaneously, especially market gardening farmers. this allows them to be more responsive to demand. thus, the closure of farmers' markets and restaurants can be compensated for by farm shops, community-supported agriculture (csa), fixed point or home delivery, or collective farmers' shops. regarding hygiene measures, farmers are faced with two main types of strategies: receiving customers in the original locations or digitizing the process, including the more marginal case of automatic food dispensers (e.g. for eggs). selling in physical places must respect social distancing, which discourages consumption. however, conserving this method makes it possible to maintain the relational proximity between farmers and consumers. initially, this relational proximity is one of the main arguments in favour of short supply chains, as it is supposed to allow better traceability of products and to fight against social isolation (especially of the elderly). the other strategy, therefore, is to limit direct contact between human beings by means of computer tools. internet platforms already existed before the crisis and are expected to develop. some were public, such as those set up to supply school canteens. the pandemic has accelerated this process, and some regions have launched their own platforms. this solution is very time-consuming and seems difficult to sustain, unlike the others, which should have an impact in the long term. another practice has been reinforced: farm shops. during the lockdown, some consumers presumably had more time to cook and go to farms. others fled from supermarkets, considering that the products were handled by a large number of people and that there was too much traffic. it is difficult to predict whether these changes in behaviour will have a long-term impact. in addition, the pandemic has raised concerns about the reliability of international distribution channels. it is therefore possible that some policies may be sensitive to the has been growing at a very fast pace (see an indicative list of references in supplementary material ). however, little attention has been paid to the reliability of this type of epidemiological data to make statistical inferences. our initial aim was to produce a detailed statistical analysis of the relationship between weather conditions and the spread of covid- . this question has attracted significant attention from the media (e.g. ravilious ; clive cookson ) and the research community (e.g. araujo and naimi ; carleton et al. ; see a wider list in supplementary material ) due to the possibility that summer weather might slow the spread of the virus. after going through all the steps of such an analysis, we reached the unexpected conclusion that the limitations of the available covid- data are so severe that we would not be able to make any reliable statistical inference. this applies, for example, to the data provided by the john hopkins university (dong et al. ) and the data collated by xu et al. ( ) . this is a concerning and very important finding considering that such data are being widely used to make crucial policy decisions on a wide range of topics. since invalid causal inferences could be made with the publicly available covid- data, and then enter policy-making discourse, there is an urgent need to raise awareness among the scientific community and decision makers regarding the limitations of the information at their disposal. the elements discussed in this paper are also likely to be applicable to other epidemiological datasets obtained with insufficient testing and monitoring, either during exceptional epidemics or seasonal outbreaks. several challenges could undermine any causal statistical analysis of the influence of a potential determinant, such as the weather, on the spread of covid- . to start, confounding variables are likely to pose a significant problem: many factors (e.g. changes in policy or social interactions) are simultaneously influencing how the disease spreads. in addition, significant challenges come from the limitations of the covid- case count data itself. firstly, testing capacity has been a major issue in most countries. before march , very few countries had sufficient testing capacity. by april , highincome countries had significantly increased their testing capacity, but testing remained critically infrequent in most low-and middle-income countries. figure , panel a, illustrates the effect that insufficient testing capacity has on the number of confirmed cases. it distinguishes between three phases of limited (i), intermediate (ii) and widespread (iii) testing. in phases i and ii, there is a risk that the number of confirmed cases depends more on the number of tests available than on the actual number of people who have covid- , questioning the validity of any analysis relying too heavily on these data. moreover, there have been numerous concerns regarding the accuracy of the covid- tests performed so far (ai et al. ; apostolopoulos and tzani ; hu ; hall et al. ) . figure , panel b , illustrates the effects of both false-negative and false-positive test results on the number of confirmed cases. false-negative results would imply that the number of confirmed covid- cases is underestimated. false-positive results would imply that people who do not have covid- are included in the number of confirmed covid- cases. concerns regarding test accuracy create an additional problem of measurement that might affect statistical analyses. the two above-mentioned challenges are inherent in all current datasets of covid- confirmed case count and mortality. in addition, specific datasets may have imperfect geographical or time coverage. to look at the impact of the weather on the spread of covid- , we initially used a well-established approach, similar to the ones used previously to look at the impact of the weather on other diseases (e.g. deschenes and enrico ; gasparrini et al. ) (see details in supplementary material ). however, the fundamental measurement issues associated with the covid- case count data cannot be corrected by statistical techniques, as we outline in the following. the main problem is that the weather could be influencing the number of tests carried out and the segment of the population tested. for example, other respiratory diseases are often similar to covid- in their symptoms (e.g. who ) and are more common during cold weather (e.g. deschenes and enrico ; gasparrini et al. ) , which could influence the number of tests performed on people displaying symptoms of respiratory infection. therefore, even if the model correctly identified the impact of the weather on covid- case counts, it cannot distinguish between the impact of the weather on the spread of the disease and its impact on testing. table provides a non-exhaustive list of elements that could undermine any analysis of the impact of the weather on the spread of covid- using data on confirmed cases. the evidence suggests that the weather may correlate with the number of tests conducted and who gets tested. we have not been able to find any specific covid- -related evidence that the weather could impact test accuracy (e.g. the weather affecting the nasopharyngeal or oropharyngeal swabs used in the pcr analysis), even though this could be possible. other points of concern include: the fact that there may be indirect effects of weather conditions on other factors that could have an impact on the spread of covid- (such as social interactions or air pollution); the heterogeneity of impacts across populations and subgroups within a population; and the fact that some people may have travelled and therefore been infected in a different place from where the cases are reported. we ran our model (as detailed in supplementary material ) and provide results and robustness checks in supplementary material . the model would technically suggest a negative correlation (e.g. colder days would be associated with more confirmed covid- cases, and hotter days with fewer cases). yet, these results could be highly misleading since these estimates are likely to be substantially biased because of the aforementioned reasons. figure , panel a, provides an illustration of how we could have obtained a negative correlation even if temperature had no impact or a positive impact on the spread of covid- in our sample. the total number of estimated cases is given by the size of the circles as a function of temperature (x-axis). the circles in green correspond to the effects we are interested in-those that explain the influence of temperature on the spread of covid- . if temperature has no effect on the spread of covid- , then the green circles should be the same size at low and high temperatures. the pink circles represent the possible effect of temperature on testing (as reported in table ) under the illustrative assumption that high temperatures reduce testing frequency. in this case, the overall result is a negative correlation between temperature and confirmed covid- cases, even if temperature has no effect on the spread of the disease. in practice, we naturally do not know the direction of the bias caused by the effect of temperature on testing when using standard statistical methods. there is also no way for us to evaluate the contribution of each of these effects (green table non-exhaustive list of reasons why weather conditions could affect the number of covid- tests carried out and who gets tested potential reason potential implication unrelated respiratory diseases are weather sensitive (e.g. deschenes and enrico ; gasparrini et al. ) and can be confused with covid- (e.g. ai et al. ; chen et al. ) more patients with symptoms of unrelated respiratory diseases could be tested during cold weather the prevalence of other weather-sensitive respiratory diseases might make false-positive results more likely, especially if only radiographic imaging is used, since it is possible to confuse these diseases for covid- (e.g. ai et al. ; chen et al. ) the incidence of other pathologies (e.g. cardiovascular diseases) is influenced by the weather (e.g. deschenes and enrico ; gasparrini et al. ) hospital capacity and the workload of medical staff and testing structures are affected by weather conditions, with potential implications on the number of tests conducted at-risk individuals suffering from unrelated conditions are more likely to be tested for covid- , even if they only have mild symptoms for covid- people may be more inclined to seek medical attention depending on the weather (e.g. norris et al. ) due to weather conditions, people may or may not decide to seek medical attention, affecting the number of patients going to the hospital with covid- , and the workload of medical staff or pink) in our estimate. we arrive at the final size of the circles and cannot be sure if the association that we are interested in is either negative, null or positive. figure , panel b, focuses on the risk that effects could be different across different samples. the circles in blue capture other underlying factors that are influenced by temperature (such as acclimatization or the level of social interactions in the population), as well as other socioeconomic factors (such as the demographic characteristics of a population). these factors could be radically different in different regions but may also evolve over time (e.g. between winter and summer seasons). there are strong reasons to be concerned with the scenario illustrated in figure , panel b. in our sample, for example, we only have data from the start of the pandemic until end of april ; some countries (e.g. china) may be over-represented in the dataset; and the average daily temperature is relatively low at . °c. furthermore, many countries have implemented a stringent containment policy during the period covered by the sample. containment policies may have heightened (or lowered) the sensitivity of the spread of the disease to the weather because social interactions are limited. we are not able to observe how for example, sars-cov- is more infecƟous; the weather affects people's immune systems; people-to-people transmission is higher. for example, changes in the number of people with respiratory diseases other than covid- , affecƟng the number of tests performed. for example, social interacƟons evolve; or samples are for different countries. example temperature (a) esƟmates are likely to be biased because the weather influences data collecƟon (b) esƟmates could vary between samples fig. effects potentially captured by our estimate. the size of the circles represents the estimated number of cases at different temperatures. these are examples that do not correspond to actual data. in these examples, we assume no correlation between temperature and the effects in green (see legend), a negative correlation with the effects in pink (example ) and a positive correlation with those in blue. (color figure online) the impact of the weather on covid- might change at different gradients of social interaction. finally, our estimate is based on small, observed changes in temperatures, and not on radical increases or reductions in temperatures. the spread of covid- may respond differently to large variations in temperature, e.g. by °c or °c across seasons, making seasonal predictions even more unreliable. strong precautions need to be taken before using covid- case count datasets for inference. the results of our model using existing covid- data would seemingly imply a negative association between temperature and confirmed covid- cases. any projection of covid- cases with such estimates could conclude that, during the upcoming months of june to september , southern hemisphere countries would be exposed to higher risks of covid- spread and northern hemisphere countries to lower risks. these types of unsubstantiated results could be used as a misinformed justification for an early relaxation of effective social distancing measures in the northern hemisphere. these findings have equally strong implications for statistical analyses focusing on other questions that rely on covid- confirmed case count and/or mortality count data. even though the exact nature of the effects may change, such studies are also at risk of capturing the effect that their parameters of interest have on tests and test results. for example, studies interested in the effect of containment policies may have to consider that these policies substantially affect testing because they change the awareness of the disease in the population, political demands for more testing or the risk of contracting other respiratory diseases. other studies may also produce estimates that are very specific to the current circumstances in the development of the pandemic and are, therefore, not suitable to use for forecasts of what could happen in the coming months. in the medium term, more reliable data need to be gathered, for example through experimental studies that randomly test a sample of the population for covid- . in the short term, we are in a situation of fundamental uncertainty about how different factors affect or are affected by the widespread societal changes we see with the covid- pandemic. therefore, scientists, policy makers, journalists and the general public need to be very cautious when discussing how the spread of covid- correlates with the weather or any other factor. in the long term, this paper suggests that more attention should be given to how epidemiological data is recorded and used during exceptional epidemics and seasonal outbreaks, since insufficient testing and monitoring can undermine essential statistical analyses. this article calls for the complementary use of different methods for data collection, such as random testing in samples of the population. sars to novel coronavirus-old lessons and new lessons global rise in human infectious disease outbreaks carbon emissions come roaring back: will the economy, too? 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( ) consulted on th april use of radiographic features in covid- diagnosis: challenges and perspectives an empirical investigation into factors affecting patient cancellations and no-shows at outpatient clinics cohen is the first author. he had the original idea, wrote most of the paper and the code to produce the econometric analysis. he also coordinated the team. jani ensured the material was consistent with the epidemiological evidence. li produced the required climate data for the statistical analysis. lu helped on literature review, on coding the econometric analysis and on producing the tables. schwarz helped on data coding and matching and created the projections. all authors contributed to the text. data and materials availability all data and software are publicly available at https :// githu b.com/morit zpsch warz/covid - -weath er-oxfor d. key: cord- - viq ak authors: vogler, sabine; fischer, stefan title: how to address medicines shortages: findings from a cross-sectional study of countries date: - - journal: health policy doi: . /j.healthpol. . . sha: doc_id: cord_uid: viq ak shortages of medicines have become a major public health challenge. the aim of this study was to survey national measures to manage and combat these shortages. a questionnaire survey was conducted with public authorities involved in the pharmaceutical pricing and reimbursement information (ppri) network. reponses relating to measures as of march / april were received from countries ( european countries, canada and israel). in countries, manufacturers are requested to notify – usually on an obligatory basis – upcoming and existing shortages, which are recorded in a register. further measures include a regular dialogue with relevant stakeholders ( countries), financial sanctions for manufacturers in cases of non-supply and/or non-compliance with reporting or stocking requirements ( countries) and simplified regulatory procedures ( countries). for defined medicines, supply reserves have been established ( countries), and legal provisions allow the issuing of export bans ( countries). some measures have been introduced since the end of and countries are planning and discussing further action. while governments reacted by taking national measures, the covid- crisis might serve as an opportunity to join forces in cross-country collaboration and develop joint (e.g. european) solutions to address the shortage issue in a sustainable manner. a practical first step could be to work on a harmonisation of the national registers. medicines shortages have become a major issue globally [ ] , as they affect numerous countries, including canada [ , ] and european countries [ ] (e.g. belgium [ , ] , finland [ ] [ ] [ ] , france [ , ] , germany [ ] , italy [ , ] , the netherlands [ , ] and poland [ ] ). shortages relate to temporary (and sometimes permanent) disruptions of the supply of medicines. the national definitions of a shortage of medicines differ, and some countries do not have an official definition at all ( [ , ] , for examples see supplementary materials ). a common feature of a shortage is that the supply of a medicinewhich has been brought onto the marketis discontinued. there are other circumstances when medicines are unavailable for patients but they are not called shortage: medicines are not launched if a pharmaceutical company does not perceive a country's market to be sufficiently attractive (e.g. in the case of small markets or lower-income countries with limited ability to pay [ , ] ). medicines may be launched with a delay of some months and even years; this delayed availability is frequently caused by the widespread use of the external price referencing policy (for instance, apart from sweden and the uk, all european countries with price regulation use external price referencing, at least as supplementary pricing policy [ , ] ). external price referencing incentivises pharmaceutical companies to bring medicines to market later in lower-priced countries so as not to reduce the benchmark price across countries (strategic launch of pharmaceutical industry) [ ] [ ] [ ] . shortages limit patient access to medicines which likely results in increased negative health outcomes (e.g. no or delayed therapy, risks for patient safety due to switches and medication errors [ , ] ), increased workload for health professionals (e.g. searching for therapeutic alternatives) [ , , ] and economic consequences (e.g. need to procure higher-priced alternatives) [ , ] . to respond to the public health threat resulting from shortages, governments started taking action to improve the management of these disruptions and to reduce and ideally prevent them. measures reported in the literature include the obligation for industry to report shortages to authorities (e.g. france [ ] ), the netherlands [ ] ) and the creation of national reporting systems (several countries [ , ] ), supply reserve stocks (e.g. finland [ ] ), notification of parallel exports [ ] and collaborative projects with stakeholders such as representatives of the pharmaceutical industry, wholesale and pharmacies (e.g. italy [ ] ). most scientific articles related to single countries and reported measures as background information while the j o u r n a l p r e -p r o o f research question had a different focus (e.g. workload for pharmacists to manage shortages, types of medicines subject to shortages). acosta et al. [ ] presented some national approaches to manage shortages in their scoping study but their narrative review was limited to published articles. to the knowledge of the authors, no up-to-date overview of government policies to address shortages of medicines is available. therefore, this study aimed to present up-to-date information about the actions taken in different countries to manage, reduce and prevent shortages of medicines. to ensure the collection of the most current information and inclusion of a high number of countries, including those that tend to be less covered in scientific research (e.g. smaller and/or less resourced countries), a primary survey (using a questionnaire) with public authorities was carried out. the authors benefited from an existing collaboration with the pharmaceutical pricing and reimbursement information (ppri) network. this network comprises public authorities for pharmaceutical policy, with a focus on pricing and reimbursement, in countries (at the time of the study). most ppri member countries are from the who european region [ ] . given an ongoing exchange of information to update each other, ppri network members are used to receiving and responding to requests, and involving further colleagues and institutions in their country if needed [ ] . a questionnaire was developed based on a taxonomy of possible measures to manage, reduce and/or prevent shortages of medicines (implemented as well as under discussion), which were known from the literature [ , , , , , ] and policy debate [ , ] . the authors tested the questionnaire by pre-filling information for five pilot countries (austria, finland, italy, the netherlands and sweden). the information for these countries was sought in an unsystematic literature review that considered peer-reviewed articles and grey literature (e.g. media reports, websites of the public authorities and private actors in the medicines supply chain) in national j o u r n a l p r e -p r o o f languages as well as in english. additionally, the authors contacted national stakeholders (usually the medicines agency and the community pharmacy association) in the pilot countries (except austria) and asked them in a telephone interview to identify further measures, including those already implemented or under discussion. the research for the pilot countries was conducted in january . countries were asked to validate the pre-filled information. the questionnaire was sent to the members of the ppri network on march , with the request to respond within two weeks. the invitation to participate in the survey was accompanied by information on the authors' intention to publish the findings. in march and april two rounds of reminders were sent to those network members that had not yet responded. in addition, during the compilation of answers in april some ppri network members were contacted on an individual basis to clarify some ambiguous answers and ask for missing information. a summary table of the synthesised results was shared with all ppri network members that participated in this study, thus allowing them to comment on possible errors. in august , as part of the revision of the paper, the authors analysed publicly accessible registers for notification of shortages with regard to the type of information provided (e.g. cause of shortage, information on alternative medicine for substitution). a frequently used measure to manage shortages was a national register to which suppliers report current and upcoming shortages: in of the responding countries, regulatory authorities (usually the medicines agency or the ministry of health) ran a shortages register. in all countries with such a register, with the exception of malta, suppliers were obliged to report to the register, while austria and germany had made their voluntary registers obligatory only on april ( table ). the majority of these registers were publicly accessible. the outlines of the registers and the kinds of information collected differed from country to country. while several registers provided for the possibility to include relevant dates (expected and/or actual start and end dates of the shortage), only few registers published information on causes and solutions (e.g., existence of alternative medicines) for managing the shortage situation (supplementary materials ). to cover shortages, countries had simplified regulatory procedures, in particular related to the import, marketing authorisation and also dispensing of medicines procured on the world market. usually, exceptions were granted with regard to labelling requirements of the packages and product information leaflets in other languages were permitted. banning exports of medicines that are critically needed for the supply in a national market was a less frequent measure, but it was increasingly being used at the time of the survey. of the responding countries, five countries provided for the possibility to impose export bans on defined medicines by the end of , seven countries by the end of q / and ten countries by mid-april . medicines eligible for an export ban were usually listed (e.g. the "ex-ante notification list" in portugal). three further eu member states considered introducing export bans, and israel discussed an extension to further medicines. in addition to export bans for some medicines, four countries asked for export notification of further selected medicines (czech republic, norway, portugal) or all medicines (latvia), respectively. romania, which requested distributors to notify the export of reimbursed medicines, had planned to introduce an export ban in , however, it had withdrawn this measure due to changes in the political environment. insert table around here in countries (including the netherlands which planned to implement this measure in the course of ) marketing authorisation holders and/or wholesalers have the obligation to keep a stock of defined medicines for a certain period (usually at least three months). such stocking requirements were also built into tender contracts (e.g. albania, denmark). most of the surveyed countries ( countries) reported regular meetings with relevant stakeholders, in particular marketing authorisation holders, wholesalers and community pharmacists and, but less frequently, patients and the public (e.g. in finland). this exchange usually took place in a rather formal setting of a working group or task force. in germany, the stakeholder dialogue was officialised as an advisory board to the medicines agency ("jour fixe") in a law that came into effect on april (table ) . norway was the sole country to offer financial support to economic operators. in a covid- related voluntary agreement of march between the authorities and wholesalers, the latter were asked to extend their stock to a wider range of medicines (this request added to a previously defined supply reserve obligation). the additional costs for wholesalers would be j o u r n a l p r e -p r o o f covered by the state. in several countries, financial sanctions were in place to penalise noncompliance (in cases of non-supply - countries, of non-compliance with reporting requirements - countries and of non-compliance with stocking requirements - countries plus the netherlands in the planning phase). nine countries did not report the use of financial sanctions (table ) . insert table [ ] ). in addition, the covid- pandemic that hit european countries in the course of march , prompted further measures (e.g., the "covid- ad-hoc agreement" as of march in norway, which asked wholesalers to stock more, and export bans imposed in israel and russia in march and in latvia in april , respectively). however, the legal changes in austria and germany, which made reporting to registers mandatory in april , had been decided at earlier times, and it was a coincidence that they entered in force in covid- times. third, in addition to obligations and regulatory measures, supplemented by sanctions in some countries, public authorities tend to seek collaborative action with stakeholders. canada started its stakeholder dialogue already in . italy opted for a large stakeholder platform that involved law enforcement institutions besides national and local authorities and associations (industry, wholesale, pharmacy). in a joint project ("inspection campaign"), first done as a pilot in one region and later repeated at national level, economic operators responsible for illegal transactions and breaches of the good distribution practice were identified and penalised (over withdrawals of licences, financial sanctions of nearly € , ) [ ] . it is interesting to note that in most countries marketing authorisation holders are the sole economic operators that have to report to the register (in of countries with a register). in this respect, the introduction of the notification obligation in germany that is also targeted at wholesalers may serve as a good practice example. despite obligatory notification to the register in all countries except malta, not all countries impose financial sanctions in case of non-compliance (e.g. norway: obligatory, but no sanctions). however, a few countries (e.g. cyprus and finland) were discussing the introduction of sanctions at the time of the survey. fourth, access to information related to the shortage situation plays a major role, not only for public authorities, but also for health professionals such as pharmacists who are involved in the management of shortages [ , ] . in several countries, public authorities responded to this need for information for health professionals by offering publicly accessible registers. for health professionals it is particularly important to learn about the expected duration of the shortages and possible approaches on how to ensure access to (similar) treatment. however, j o u r n a l p r e -p r o o f in only few countries do the regulatory authorities publish on alternative medicines or other solutions. in some countries (e.g. italy, the netherlands), pharmacists have established their own registers to cover for missing data of the authorities' registers [ ] . fifth, differences exist with regard to the medicines that are subject to shortages-related measures. most countries apply a rather broad approach for the registers ( of the countries with a register report shortages on any authorised medicine; countries on prescription-only medicines or prescription-only and selected non-prescription medicines, respectively, and countries on reimbursed medicines), while three countries (denmark, germany and switzerland) limit the reporting requirements to medicines that are considered critical to ensure supply and treatment of patients. medicines supply reserves and particularly export bans are usually more focused and often concern considerably fewer medicines compared to the medicines requiring notification. sixth, the measures comprise different types of action, with different purposes. registers and stakeholder dialogues are, in principle, measures to generate information before and during the shortages management. but they are not necessarily policies to prevent or combat shortages. preventive measures and those to fight the shortages include export bans and reserve stocks, which aim to protect the supply of the national markets, as well as imports from other countries, with the intention to cover gaps in national supplies. having a focus on the national markets, these preventive measures compete with similar initiatives of other countries that struggle with the same challenges. anecdotal evidence on trucks and airplanes with covid-related medical supply goods being stopped at borders or being redirected to other countries that were willing to pay higher prices as reported in the media [ , ] confirm conflicting national interests. more than five years ago, researchers called for collaboration among countries to tackle the shortages challenge: "currently, the member states of the european union are striving to resolve the problem very much on their own, although a far more focused and dedicated j o u r n a l p r e -p r o o f collaboration may well prove instrumental in coping with drug shortages throughout europe more effectively" [ ] . the situation appears not to have changed considerably though some initiatives were launched and the topic has meanwhile become high on the political agenda [ , ] . in [ ] . but the jpa framework was not intended to be extended to medicines in general [ ] . in recent years, cross-country collaborations (such as the beneluxa initiative or the "valletta declaration") were established between european countries. their key aim is to ensure sustainable patient access to new innovative medicines [ ] . shortage management appears to be of lower importance for these collaborations, except for the baltic procurement initiative of estonia, latvia and lithuania. for the latter, managing shortages based on a "lending agreement" is a major collaborative action. in case of a shortage in one of the countries, another country lends the needed medicines or medical devices and does not charge any fees for this service. once the beneficiary country receives its supply, it will return the products it borrowed to the lending country [ ] . major causes of shortages are production and quality problems [ , [ ] [ ] [ ] . in our research, two responding countries mentioned national production to overcome medicine shortages (switzerland reported a discussion on strengthening the domestic production base, and italy j o u r n a l p r e -p r o o f mitigates some shortages by using military sites for production). bringing the pharmaceutical production back to europe has been proposed and discussed in the political debate, including discussions initiated by the german presidency of european council in the second half of [ ] . this longer-term endeavour would require joint efforts of several european countries, based on strong political will. it is beyond the scope of this paper to assess the feasibility and effectiveness of this measure. however, the covid- crisis could offer an opportunity to use the momentum for change. in the meantime, european countries could start collaborating in the more technical area of information sharing. despite the above-mentioned agreement on a definition of a shortage between ema and national regulatory agencies, the national legal specifications continue to differ among european countries ( [ , ] , see also supplementary materials ). these differences are also reflected in the variation in the deadlines for the shortage notification. as shown in this study (table ) , deadlines range from "immediately" to "at least six months in advance". in addition, the structure and the contents of the national registers (e.g. frequency of updates, information on start and end of the shortages, causes) also vary among the countries ( [ , , , ] governments have increasingly been using a mix of measures to address and combat medicines shortages. in the months and even weeks preceding and even during this survey, some countries implemented additional measures. regarding the design of policies, voluntary tools tend to have been substituted by obligatory mechanisms linked to sanctions. high-income countries were found to apply a higher number of measures, but the introduction of new policies in this field was also observed in some lower-income countries in europe. the measures appear to be rather reactive with a focus on managing existing or upcoming shortages and not addressing the causes of shortages. for the time being, measures to manage and combat shortages of medicines were predominantly taken at national level, though the need for a multi-country approach to identify global or at least european answers has been repeatedly voiced. the covid- crisis that will likely aggravate the shortage situation could give new momentum to jointly search for solutions that go beyond national measures and could also address the causes of shortages. the authors declare that they have no conflict of interest. this research did not receive any specific grant from funding agencies in the public, commercial or not-for-profit sectors. however, the authors acknowledge to have drawn from knowledge that they have gained in a previous research. that project investigated medicines shortages in finland, italy, the netherlands and sweden and was funded by the national association of statutory health insurance funds ("gkv-spitzenverband") in germany. furthermore, the pharmaceutical pricing and reimbursement information (ppri) secretariat that maintains and coordinates the ppri network of competent authorities, whose delegates were key respondents in this study, has been financially supported by the austrian federal ministry of social affairs, health, care and consumer protection. no funding was received for designing the survey, analysing and compiling the data and writing the manuscript. further information on the accessibility of the shortages registers maintained by public authorities and their contents is provided in supplementary materials . shortages of medicines: a complex global challenge drug shortages in canada and selected european countries: a cross-sectional, institution-level comparison. the canadian journal of hospital pharmacy drug shortages in canada: data for - and perspectives on the problem european drug shortages: a call for action! chemotherapy drug shortages in paediatric oncology: a -year single-centre experience in belgium time spent by belgian hospital pharmacists on supply disruptions and drug shortages: an exploratory study medicine shortages -a study of community pharmacies in finland the reasons behind medicine shortages from the perspective of pharmaceutical companies and pharmaceutical wholesalers in finland a regulatory perspective on the availability of medicines and medicine shortages in outpatient care: case finland the new regulatory tools of the health law to fight drug shortages in france. health policy a qualitative approach to a better understanding of the problems underlying drug shortages, as viewed from belgian, french and the european union's perspectives drug shortages may compromise patient safety: results of a survey of the reference pharmacies of the drug commission of german pharmacists oncologic drug shortages also in italy tackling distribution-related shortages of medicines: an italian case study evaluated in the european union framework drug shortages from the perspectives of authorities and pharmacy practice in the netherlands: an observational study dsl- drug shortages in the netherlands: monitored by farmanco drug shortages as a result of parallel export in poland -pharmacists' opinions. health policy medicine shortages: gaps between countries and global perspectives toward a european definition for a drug shortage: a qualitative study challenges to orphan drugs access in eastern europe: the case of bulgaria how can pricing and reimbursement policies improve affordable access to medicines? lessons learned from european countries. applied health economics & health policy pharmaceutical pricing and reimbursement policies in ppri network member countries vienna: who collaborating centre for pricing and reimbursement policies practice of external price referencing the impact of price regulation on the launch delay of new drugs-evidence from twenty-five major markets in the s pharmaceutical price controls and entry strategies. the review of economics and statistics evolution of average european medicine prices: implications for the methodology of external price referencing the impacts of medication shortages on patient outcomes: a scoping review medicine shortages took an extra hour per week of pharmacist time in , survey finds. the pharmaceutical journal clinical, economic and policy implications of drug shortages in the european union. applied health economics and health policy results of eahp's survey on medicines shortages systemic measures and legislative and organizational frameworks aimed at preventing or mitigating drug shortages in european and western asian countries gesundheit Österreich the pharmaceutical pricing and reimbursement information (ppri) initiative-experiences from engaging with pharmaceutical policy makers report from a multi-stakeholder meeting held at ema on th addressing the root causes of medicines shortages. supply chain stakeholders' views on root causes and solutions tekorten in cijfers internationale evidenz und empfehlungen für deutschland [medicines shortages: international evidence and recommendations for germany die schweiz liegt mit deutschland im offenen streit. nzz am sonntag us accused of 'modern piracy' after diversion of masks meant for europe. the guardian pharmaceutical strategy for europe -timely patient access to affordable medicines. draft for consultation deutsche eu-ratspräsidentschaft (german presidency of european council guidance on detection and notification of shortages of medicinal products for marketing authorisation holders (mahs) in the union (eea) joint procurement of medical countermeasures medical countermeasures that could be procured in common under the joint procurement agreement beneluxa initiative -vision cross-country collaborations to improve access to medicines and vaccines in the who european region. word health organization regional office for europe causes of drug shortages in the legal pharmaceutical framework drug shortages: root causes and potential solutions drug shortages in european countries: a trade-off between market attractiveness and cost containment? the authors are highly grateful to the representatives of public authorities for pharmaceutical pricing and reimbursement involved in the pharmaceutical pricing and reimbursement information (ppri) network who responded to the survey of this study. in line with the ppri policy, their names and affiliations are not mentioned. further thanks go to nina zimmermann, ppri coordinator, for keeping this network active, and to manuel alexander haasis, member of the ppri secretariat, for his support in the data collection. since a legal change on april , which introduced the obligation to notify the register c in addition, financial sanctions for the export of medicines which are prohibited to be exported key: cord- - hih qoy authors: crosier, david; haj, cezar mihai title: evolving social dimension of the european higher education area date: - - journal: european higher education area: challenges for a new decade doi: . / - - - - _ sha: doc_id: cord_uid: hih qoy the social dimension is a term coined in the early years of the bologna process (bp). although mentioned in the early ministerial communiqué texts, the term itself was not clearly defined until . looking back, this could perhaps be considered an oversight. however, the more likely explanation is that the term was chosen intentionally, leaving open possibilities for national and institutional action while, at the same time, committing countries to nothing precise. this definition continues to be a valid and actual reference point. it is a definition that embraces a clear, aspirational objective-the representation of the diversity of the population. it also entails a process to reach the goal-i.e. the development and implementation of policies and practices (bologna process b) . most importantly, establishing this definition brought clarity to discussions. everything that has been written in subsequent communiqués and ehea policy texts derives from and sometimes repeats the working group report and the definition in the london communiqué. thus "student body within higher education should reflect the diversity of europe's populations (bologna process )","the student body entering and graduating from higher education institutions should reflect the diversity of europe's populations (bologna process )", "….the student body entering and graduating from european higher education institutions should reflect the diversity of europe's populations…(bologna process )" some subsequent communiqués focus on commitment to develop and implement national strategies (bologna process , while others give particular emphasis to other aspects. for example, the higher education role in building inclusive societies is a focus of the yerevan communiqué. in this document, ministers agree to 'enhance the social dimension of higher education, improve gender balance and widen opportunities for access and completion, including international mobility, for students from disadvantaged backgrounds.' (bologna process ) . the communique further emphasises flexible learning paths and the inclusion of different types of learners by promoting life-long learning and recognition of prior learning. while responsibility lies with countries and institutions, there have nevertheless been important developments in european-level discussions throughout the bologna period. despite each country being left to determine the particular parameters of under-representation in its specific context, a european discussion has required some broad categories of under-represented groups to be identified. these categories may also correspond to notions of disadvantaged or vulnerable students in national contexts and commonly include people from a low socio-economic background (identified through low income or low educational background of parents), gender, disability and immigrant or minority status. mature students are also often included. european discussions also recognise that students from particular geographical regions may be disadvantaged-particularly those in isolated rural areas or areas of high urban social deprivation, and that under-representation is also associated with insufficient formal education qualifications for entry into higher education. these categories of under-represented, vulnerable or disadvantaged groups are not separate, discreet entities. in lived reality, they combine as characteristics of individual citizens, and under-representation may be apparent in a combination of these different factors. thus, a person from a low socio-economic status background will also have a gender, may come from an area of social deprivation, may be a migrant or have a disability, and all of these factors will combine to play a role in shaping or limiting opportunities in a particular context. within the bologna process, the main discussion focuses on the need to develop strategies and actions to improve access to, participation in and completion of higher education for members of these groups. this paper aims to present an overview of the ways in which the issues under the umbrella term of "social dimension" have been conceptualised and addressed by higher education policy throughout the bologna process. it also considers the challenges that lie ahead in improving social reality in european higher education and introduces the papers that have been selected to provide in-depth research on different aspects of the social dimension. despite discussion and commitment for national strategies and action plans (bologna process a) , there are few countries that have actually made a conscious effort to develop genuine social dimension strategies in the european higher education area. ireland is perhaps the most notable exception to this general rule. here, a succession of higher education policy strategies has consistently featured inclusion and widening participation as high-level policy goals throughout the bologna period. the country has also developed support measures focused at students from under-represented groups-particularly mature students, students from the traveller community and students with disabilities. ireland has also established good quality monitoring systems to track the impact of policy-measuring completion and drop-out rates for specific under-represented groups. (department of education and skills, dublin ) the focus on social inclusion has not inhibited other policy initiatives. on the contrary, ireland's strategy has engaged fully with other trends in learning and teaching, research and innovation, community engagement and internationalisation. indeed, the country's approach offers a good model for the wider ehea commitment being prepared for the rome communiqué to make the social dimension a core commitment on which to build all future policy. other countries also offer interesting models. in particular, the nordic countries demonstrate a model of social equity that permeates higher education. this can be most clearly seen through the type of financial measures that are used to support students. in contrast to nearly all other european systems, no tuition fees are charged to national and european students. this policy also extends in some countries to all international students, but in recent years, countries such as denmark and sweden have introduced fees for students from outside europe. (european commission/eacea/eurydice ) in addition to the absence of fees, a very high proportion of students receive grants, with amounts usually adjusted according to the individual student's socioeconomic situation. the public investment is, therefore, considerable and provided in a mainstream form. one important aspect of this nordic model is that no specific groups are identified. through providing generous conditions and support to all students, disadvantaged or vulnerable students also receive sufficient support and are not stigmatised. while one major benefit of this model is its inherent inclusivity, its most significant disadvantage is that there is no way of knowing how many students from under-represented, disadvantaged or vulnerable groups actually benefit from the support. as the notion of groups is anathema to a system designed to support everyone, there remains a potential for hidden inequity to continue without specific action being taken to address it. almost all other european higher education systems take a different approach to student financing. most commonly, countries charge tuition fees to some studentsbut provide fee reductions or exemptions to students who are considered as 'disadvantaged'. this refers to students with low socio-economic background and those belonging to various under-represented groups, such as students with disabilities, ethnic minorities, orphans or, in a few countries, children of war victims. fee waivers or reductions based on socio-economic criteria generally concern a relatively small proportion of students. a number of transition countries such as latvia, lithuania, hungary, romania, bosnia and herzegovina, serbia and north macedonia use merit-based criteria in differentiating fees paid by full-time higher education entrants. in these countries, based on performance in upper secondary education and/or results of admission tests, students start their higher education studies in one of two groups: a 'state-funded' group, including better-performing students who do not pay fees or pay only small administrative charges, and a group of 'self-financing' students who cover (fully or partially) their tuition expenses. depending on the country, self-financing students may comprise between % and % of students. even more countries use merit-based criteria for allocating grants. according to eurydice data, in higher education systems, grants are awarded either based on educational outcomes during higher education studies or based on secondary school results or performance in admission tests. this merit-based approach to student funding appears to be blind to the reality that it rewards previous social and educational advantage. yet research consistently shows that students coming from families where parents have themselves benefitted from higher education have greater opportunity to access higher education. providing fee waivers and grants on the basis of merit-based criteria thus shores up social advantage and inequity. one of the reasons why merit-based student funding may not be a high profile social justice concern for citizens or policy makers is that almost all countries also have parallel support that is attributed on a needs basis. indeed, eurydice shows that in higher education systems, grants exist that prioritise socially or economically disadvantaged students. while student financing is widely understood as an important aspect of the social dimension, there is little evidence that specific funding approaches are effective in widening participation. the united kingdom provides an interesting case study in this respect, as england (and to a lesser extent wales and northern ireland) operates a system where high fees (capped at £ /e per year) are charged to all students. however, these fees are only paid after the student graduates, and when she or he is earning over a specific income threshold. in combination with these fees, students may take out loans which also have to be repaid after graduation. most prospective students, therefore, begin their study programmes in the knowledge that they will accrue high levels of debt. while this system transfers some of the cost of higher education from state to individuals who participate, it has also been designed with features to support underrepresented groups. indeed, higher education institutions have to sign access agreements that aim to ensure equality of opportunity for students from low-income and other under-represented groups and to increase retention rates. some of the fee funding is thus redistributed to low income and other under-represented groups through specific measures at institutional level, while higher education institutions also take other action in terms of service provision to support disadvantaged students. scotland, which has full responsibility for its education system, takes a radically different approach to student funding. no fees are charged in short and first cycle higher education to scottish and european students-except to those from the rest of the united kingdom. meanwhile, a combination of grants and loans support students from disadvantaged backgrounds. interestingly, there is no clear evidence that one approach is having a more positive impact than the other on widening participation to students from under-represented groups. the evidence suggests rather that the factors affecting participation of underrepresented and disadvantaged groups run deeper than funding. in other words, if funding at higher education level is the main mechanism used to attempt to widen participation, then success is likely to be limited. austria and romania are both countries which have made attempts to take a broader strategic approach to social dimension issues in recent years. these countries are thus the subject of a comparison of experience in the paper by torotcoi. austria has established long-term targets for increasing participation from under-represented groups and has set multiple goals for different societal groups. (bmwfw ) stakeholder representatives have all been involved in defining and developing the objectives, and there appears to be a widespread understanding of the underlying rationale for action. torotcoi demonstrates, however, that other national strategies and policies are in contradiction with the objectives of higher education social dimension strategy, and hence the likelihood of deeply-rooted change is diminished. in contrast, romanian universities have worked individually to define their social dimension strategy and have then been brought together through stakeholder interaction. however, so far improving the social dimension has not proved to be a sufficiently strong motivating factor for universities to form a true critical mass. a broader debate and greater societal engagement will be needed if there is to be agreement on the direction of such social inclusion policies. this debate needs to engage actors in social policy across a number of sectors and involve different responsible government ministries. one of the main challenges for research in this field is that it has often been limited to exploration of policy measures taken at higher education level. there are many rational reasons for this research choice. among them is the fact that policy-making is often differentiated by sectors in national systems. while some governments have a ministry responsible for all aspects of the education system, many countries separate ministerial responsibility for school and higher education. moreover, even in more unified systems, administrative divisions may facilitate separate spheres for action in higher education. one important study (orr et al. ) examined the impact of admission systems on higher education outcomes-including opportunities for access as well as successful completion of degree programmes. this study showed that admission is best conceptualised as a process that is developed through the relationship between different parts of the education system. thus in some countries, the fact that a significant share of lower secondary education pupils may be directed into educational paths that do not provide access to higher education is a stage of defining who can and cannot be admitted to higher education. the implication is that, if the social dimension agenda is to be coherent, it must consider how this kind of system feature can be reformed to ensure that equity runs throughout the education system. this approach would apply from early childhood education and care, through primary and secondary education as well as higher education. gender is another aspect of the social dimension that illustrates the difficulty of taking action only at higher education level. the bologna process implementation report shows that fields of study are highly differentiated by gender. women make up almost % of first cycle students enrolled in subjects related to education, health and welfare, but less than % of the students in information and communication technologies. (european commission/eacea/eurydice ) this reality is shaped by our societies from early childhood education onwards. without awareness of gender differentiation and action to address it coherently throughout society, there is little hope that the pattern of participation in higher education programmes would change. indeed, it is a moot point whether this is an issue that can be significantly affected by higher education policy reform. higher education is also a sector that often seems to be more reactive than proactive in the face of certain societal developments. one example is demographic change. since , many ehea systems, particularly ukraine, kazakhstan, estonia, poland, romania, slovenia, slovakia and lithuania, have been experiencing a decline in the numbers of secondary school leaving students seeking access to higher education. (european commission/eacea/eurydice ) this demographic decline could have been a catalyst for policy makers and higher education institutions to innovate in adjusting their offer. such innovation could, for example, have comprised the exploration of targeting new potential learners in higher education. admission through the recognition of prior learning, or other criteria than achievement of upper secondary education could have been explored. alternatively, countries could have invested in preparatory programmes to develop a baseline of knowledge, skills and competences in more mature returners to higher education. it is noticeable, however, that none of the countries mentioned above has taken steps to make this kind of adaptations to their system. (european commission/eacea/eurydice ) rather, they remain steadfast in the model of higher education on offer. while there may be discussion to broaden access among the typical higher education age group, there has been little or no reflection on changing the nature of the programme offer. two of the papers in this section consider the way in which social dimension policy is having an impact on under-represented groups in different parts of europe. torotcoi et al. explore the question of whether existing policy measures have demonstrated success in increasing participation of under-represented groups. through analysis of research in the field, it becomes clear that this question has no simple answer. just as different characteristics combine in the single identity of any given citizen, so too do different factors affect the impact of actions and measures in any particular context. guidance and counselling services, for example, may be highly effective to support certain students in accessing higher education and studying. however, the quality of these services, their ability to identify and target students in need of support and myriad other factors may affect their impact. similarly, funding support to disadvantaged students can intuitively be recognised as a necessary measure to support those students with few financial resources to participate in higher education. however, if this is an isolated policy action that is not complemented by other elements of a widening participation strategy, then it is very unlikely to stimulate significant change. a further problem identified in this paper is that policies may often be developed without sufficient attention to the monitoring required to assess their impact. the outcomes of this work emphasise the complexity of understanding reality, of developing effective policy, and of assessing impact. the research confirms the enormous need for quality data and research to improve understanding of social reality and to develop and refine more effective policy interventions. brooks et al. approach the issue of policy needs in this area from the perspective of learners and staff. the research explores how staff and learners perceive the impact of social characteristics on higher education experience. they do this through a research project focused on subjective perception of staff and learners' experience of social characteristics in seven european countries. the research uses focus group discussions to reveal a number of interesting findings. notable among them is that staff and student perceptions of social reality differ significantly. while staff may be conscious of certain characteristics in the student population-particularly characteristics often featured in national policy and media discussion-they are often seemingly unaware of others. students, on the other hand, appear to have a higher level of awareness of the different social characteristics of their peers, as well as of the impact that these characteristics may have. discussions with students thus tended to consider a broader range of factors than those with staff. the research also revealed significant national differences in the perception of impact of social characteristics on opportunities to access higher education. in particular, in poland-which was the only example of a post-soviet transition country in the sample-there was a widespread view that there is no issue of under-representation to be tackled. these findings give further arguments to pursue some of the challenges that the european higher education area faces. firstly, there is clearly a need to engage students in policy discussions-particularly in relation to social dimension issues if these are to be tackled effectively. secondly, it is essential to provide training to staff to raise awareness of how social characteristics may affect student learning and the overall higher education experience. there is also a need to improve understanding of how education provision could better respond to a broader range of needs in a more diverse student body. finally, it will be important to assess the impact of measures taken to address these challenges. effective monitoring, therefore, needs to be built in to all policy actions. although the social dimension has remained a constant discussion on the bologna process agenda, it is difficult to find evidence that the process itself has been an effective vehicle for addressing social dimension challenges. nevertheless, the bologna process implementation report shows that most countries have some measures in place designed to improve inclusion. as already discussed, financial support for students from under-represented groups exists almost everywhere, and access in some systems has also been facilitated by preferential treatment and opening alternative entry routes. outreach programmes and information campaigns directed for under-represented groups are also commonly used, and counselling and guidance services are widespread. performance indicators for institutions often now include financial incentives for institutions to improve access and completion for underrepresented groups. however, there is no macro-level correlation of the existence of these measures with any improved outcomes. the main aspects of inequity have remained constant throughout the bologna period and across geographical boundaries. under-representation persists in all countries, despite a variety in policies and diversity in the underlying socio-economic conditions of countries. there remains a clear correlation between high educational background of parents and the chance of their children entering higher education. (european commission/eacea/eurydice ) gender differences in certain study subjects persist and are strengthening. students with an immigrant background are also clearly under-represented in most ehea higher education systems. meanwhile, although the numbers of mature students vary between countries, life-long learning is often not a significant reality. few inroads have been made to any of these aspects of reality. one important aspect of social dimension policy is that impact-positive or otherwise-can only be assessed if policy is underpinned by effective monitoring. all countries monitor the composition of the student body to some extent. however, when monitoring practice is examined more closely, gaps are revealed. the bologna process implementation report shows that ehea countries report that the student body is monitored usually at entry and during studies, but to a lesser degree at graduation and least often after graduation. this means that, even if information on the social composition of the student body can be provided at entry and during studies, comparable information is not collected at the point of graduation, and thus the effectiveness of measures and services in supporting underrepresented groups through higher education is impossible to determine. even less is known about what happens to graduates beyond higher education and whether social factors continue to have an impact on their opportunities in the labour market. the lack of systematic monitoring at key stages of higher education is compounded by two other issues. the first is that monitoring tends to be undertaken as a snapshot-revealing a picture of current reality. it is rare to find systems that monitor the development of cohorts or track individual students. this type of tracking, however, is needed to allow a clearer picture to emerge in relation to the particular characteristics of the student population. the second problem is that, even when administrative monitoring is undertaken, many relevant social characteristics are commonly overlooked. across the ehea, it is unsurprising to find that gender and age are routinely monitored. beyond that, the picture varies. many systems collect data on disability, but this is likely to be far from comprehensive as many students with impairments may choose not to disclose them for fear of stigma. (e.g. eurostudent) data on socio-economic background is also often collected in relation to student support systems. however, it may not be used for policy reflection. data on migrant or ethnic minority status is much less often gathered. overall, according to the bologna process implementation report, only ehea countries have reliable data on completion rates of studies for students from under-represented or disadvantaged groups. the other countries, despite monitoring overall completion and drop-out rates, do not collect information in relation to specific groups. the only conclusion that can be drawn from this information is that this rather ineffectual action in monitoring means that social dimension policy implementation cannot be considered as a great priority. if it were, there would be available data gathering systems in place to understand the impact of policy. the issues already outlined in this paper provide a massive policy agenda implying, as they do, a major effort at holistic education system reform. nevertheless, the reality that we face today is fast-moving and rapidly changing, as exemplified by the sudden onset of the covid- pandemic. the closing of higher education institutions is being accompanied by a surge in online provision, and this temporary reality will no doubt drive a shift in attitudes towards digitalisation that will have some permanent repercussions. the social dimension, therefore, needs to take account of the changes in the global context as well as the potential opportunities provided by new developments. the social dimension is inextricably related to sustainable development, another paradigm that must guide future policy-making. the objective must be to harness new technologies, as well as new ways of thinking and collaborating to face the challenges in our world. this is the logic that has been followed within the bologna process by moving the social dimension to a core commitment for future development. it implies that other emerging challenges-such as the harnessing of digital technology to support inclusive teaching and research, or the integration of micro-credentials into the higher education framework-are understood as part of a new social dimension paradigm. from , europe was faced with what at the time was often described as a "migration crisis". this followed the significant increase in numbers of asylum seekers and refugees seeking to enter european union countries as a result of war in syria, and continued post-conflict societal degeneration in a number of other states-with the most significant refugee numbers coming from afghanistan. mass migration is not a new phenomenon: while the factors behind mass movements of people change, the reality of migration has been with us throughout the modern era. so while the increase in numbers of asylum seekers to european shores was an unusual and unexpected event, it is inevitable that migration will continue to be a feature of our lives in the future. we may not be able to predict the precise causes of future mass migration, but we can prepare for such events knowing that they will occur at some point. two of the papers in this volume-by berg and erdogan-consider the emergence of refugees and asylum seekers as a group affecting the social dynamics of higher education systems in recent years. while media reporting tended to focus attention on the potential impact of asylum seekers and refugees in each individual country, in reality, the only european union countries that responded through opening up their countries to refugees to any significant extent were germany and sweden. it was the neighbouring countries to syria that accepted the vast majority of asylum seekers and refugees. foremost among these countries is turkey, which in has a population of more than million asylum seekers and refugeesthe largest population of displaced people in the world. overall, among all syrian asylum seekers fleeing the country in and , more than % were in the age category - . among these, many have been seeking to develop their educational skills since leaving syria. this provides challenges as well as opportunities for the host country in developing skilled and educated citizens. the paper from erdogan is based upon survey research with syrian refugees studying in the turkish higher education system. the research explores a variety of aspects of the refugees' experience of turkish higher education, and the findings provide an important overview of the extent to which this vulnerable population has been supported into higher education institutions. the findings are relevant for improving turkish action and measures. however, they are equally relevant for wider european policy-thinking-especially if european countries are to prepare better to integrate asylum seekers and refugees in the future. it is, of course, impossible to predict when a significant new wave of refugees will be in need of asylum nor in relation to what particular conflict or natural disaster. however, it is entirely possible to predict, on the basis of past experience and known realities about climate emergency, that the future will see significant numbers of new refugees arriving at some point. it would, therefore, be sensible to prepare for such an eventuality. berg consider the integration of syrian refugees into germany-the european union country that also welcomed a large number of refugees in and . the research explores the transition into higher education, focusing on refugee students, and highlighting the ways in which the experience for refugees may differ from ideal or typical transitions. the paper illustrates how professionals working in the higher education system may need to think carefully about their routine practice. while "typical" native students may have acquired knowledge of the transition process, many of these aspects cannot be assumed for refugee students. hence there is a need to think ahead of potential barriers and the support that refugees may need. to do this effectively, it is important to find out directly from refugee students how they experienced the process. this research focuses on how often the provision of information is constructed with a particular idea of a student (coming out of secondary education) in mind, and not considering the needs of vulnerable students. it also shows how counselling services may carry unconscious or conscious biases that mean that sometimes refugee students are not encouraged to develop their full potential but rather to enter lowlevel education programmes. both papers on refugee students point to the need not only for the potential student to adapt to the higher education system, but also for the system, institution and support services to adapt to the needs of the student. the capacity of a system to do this effectively should be considered as a measure of success in the social dimension. perhaps because only a small minority of students studies in third cycle programmes, these studies have often been neglected in considerations of the social dimension. the research undertaken by o'regan addresses this cycle, focusing in particular on differences between full and part-time doctoral candidates in ireland in access to programme-based information and academic and personal support networks. in the particular sample selected, part-time candidates have a higher share of mature, female students. the findings suggest that awareness and consideration of the needs of this group of students would enable information and support to be more effectively delivered. while this research draws on information related to a specific geo-cultural reality in irish higher education, the outcomes are worthy of more general reflection. indeed, they reinforce points revealed in the research by brooks et al. and by berg that academic staff often appear not to be sufficiently aware of students' social characteristics, or of the impact that these characteristics may have on their learning experience. the social dimension, despite being a broad concept, has been clearly defined in the european higher education area since . the definition of the london communiqué responds to a basic question for citizens and policy makers alike, "what kind of higher education system do we wish to develop?" the response contained in the definition is that we are striving for a higher education that is open and equitable, and where the diversity of our populations is reflected in higher education. although the goal has been acknowledged throughout the bologna process, it has largely remained an aspiration. no country has achieved the objective, and most countries have taken little action even to move towards it. indeed, the social dimension has remained a largely peripheral concern of the ehea, with periodic reminders in subsequent communiqués to develop strategies to address the topic. while the social dimension has been largely a neglected aspect of ehea attention, there are now signs that this is set to change. recent work undertaken by the bfug advisory group for the social dimension has produced a document that aims to push the social dimension agenda to the forefront of policy discussion in the bologna process. the principles and guidelines to strengthen the social dimension of higher education in the ehea will be a highly beneficial tool for any government willing to address this area seriously. they provide a clear and concise set of issues that require reflection and debate (bologna process ). the principles and guidelines document establishes ten principles, accompanying each principle with guidelines on how they could be implemented. it should be adopted by ministers in rome, november , alongside a strong statement on why the social dimension should be at the core of the bologna process. another reason to hope that the principles and guidelines may stimulate positive change is that it is a very comprehensive overview of the issues that need to be considered. it provides the perspective of long-term commitment, with governments mainstreaming social dimension, accessibility, equity and inclusion in all aspects of their education systems. if implemented seriously, it may act as a catalyst for the type of holistic thinking around education systems that is essential for social dimension challenges really to be addressed. the principles and guidelines also consider the main areas of action required from higher education institutions and the fact that they need support to strengthen their capacity to respond to the needs of a more diverse student body. the document highlights the need for effective counselling and guidance and raises questions about the type of student funding required to strengthen the social dimension. these actions will require good systems for monitoring, and this task is also fully incorporated within the principles and guidelines. the social dimension clearly requires major policy attention. european higher education serves many purposes, but until now, it is not a strong force in redressing societal inequity. indeed, admissions systems, curriculum organisation and resource distribution are currently more likely to contribute to and strengthen inequity than to tackle it. greater understanding of social reality is clearly needed, and better data gathering systems must be developed. however, it would be wrong to wait for better data before taking policy action. the process of addressing social dimension challenges needs to begin now as a matter of urgency. open access this chapter is licensed under the terms of the creative commons attribution . international license (http://creativecommons.org/licenses/by/ . /), which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons license and indicate if changes were made. the images or other third party material in this chapter are included in the chapter's creative commons license, unless indicated otherwise in a credit line to the material. if material is not included in the chapter's creative commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. london communiqué key issues for the european higher education area -social dimension and mobility bucharest communiqué yerevan communiqué european principles and guidelines to strengthen the social dimension of higher education, draft submitted to the bologna board meeting in zagreb on nationale strategie zur sozialen dimension in der hochschulbildung [national strategies in the social dimension of higher education national strategy for higher education to -report of the strategy group luxembourg: publications office of the european union national student fee and support systems in european higher education / . eurydice -fact and figures. luxembourg: publications office of the european union study on the impact of admission systems on higher education outcomes. volume i: comparative report key: cord- - i tw authors: richau, lukas; follert, florian; frenger, monika; emrich, eike title: the sky is the limit?! evaluating the existence of a speculative bubble in european football date: - - journal: j bus econ doi: . /s - - - sha: doc_id: cord_uid: i tw in light of increasing salaries and transfer fees, the present study investigates the existence of a speculative bubble in european football. by applying the kindleberger–minsky model to football, we show that developments in recent years do not meet the criteria of a classic bubble. although transfer fee spending in recent years does meet the typical pattern seen in historical bubbles, the case of football rather resembles an atypical bubble. this is because the rise in transfer fees for most clubs is largely backed by cash inflows, prompting an elevator effect for transfer fees. typical bubbles, on the other hand, contain heavy debt-financing in the absence of respective and sustainable cash inflows. nevertheless, despite the absence of a speculative bubble on the aggregated league level, some individual clubs seem to “live in a bubble”. furthermore, the french and especially italian leagues should be cautious about overspending. we further discuss the main risk factors that can lead to a turning point in european football’s constant revenue growth, including potential implications of a financial downturn. speculative bubbles are not a new phenomenon despite the new economy and housing bubbles being two prominent examples in recent years. indeed, cases such as the tulip mania indicate that market bubbles have existed in the markets hundreds of years ago. historical examples further illustrate that speculative bubbles are not limited to a few industries but can occur in all markets that feature floating prices (kindleberger and aliber ) . in such cases, markets show a stable pattern of prices rising sharply to a high, followed by a subsequent turnover and a collapse to the extent of a bubble-and-crash pattern. this price movement pattern is of interest to economists since such (extreme) price movements are problematic for economies as a whole. market prices influence the investment decisions of market participants and have a direct impact on companies' financing costs. as a result, extreme price movements in financial markets such as those observed in bubble-and-crash patterns can (negatively) affect the so-called "real economy" through spillover effects (for an evaluation of whether it is possible to design markets that preclude such bubble-and-crash patterns, see palan ). as a recent example, european football saw astonishing price developments (relative to historic prices) in the past years, which are garnering considerable public attention. although discussions concerning the risk of an emerging bubble in football already occurred in (lill ) , recently the subject has attracted renewed interest-further fueled by the covid- pandemic and its associated uncertainty for the football industry (follert and daumann ; klosok ) . in past transfer periods, clubs have repeatedly set new transfer fee records, at both aggregated and individual-player levels. although the english premier league drives this development to some degree, the pattern is also present in the other four top european leagues, which include spain (laliga), germany (bundesliga), italy (serie a), and france (ligue ). based on this trend, transfer fee developments in the football market have been compared to a speculative bubble (e.g., brennan ; smith ) . the football market is of particular interest since it is traditionally characterized by an overinvestment environment with low profitability and high debt levels (franck a) . in this sense, the present covid- crisis can be seen as a kind of stress test for the financial stability of european football showing the weaknesses of the current system. because of mounting criticism of exorbitant transfer fees, the current study investigates whether there are indicators of a speculative bubble (in an economic sense) in european football to assess the financial stability of european football. by applying the kindleberger-minsky model to european football, the study is conceptual in nature, which is complemented by quantitative key indicators used to assess the risk of a speculative bubble in other industries (e.g., the housing market). the findings of this paper are, on the one hand, of practical relevance for football governing bodies and club managers. in the case of a speculative bubble, it could be an important implication for the international and national governing bodies to enforce stricter financial regulations beyond those existing (e.g., uefa's financial fair play) . moreover, club managers should be interested in this question to restrict their investment behavior in case of an emerging bubble and to ensure the club's financial stability. on the other hand, the results of our analysis should contest the public discussion about a speculative bubble in european football from an analytical perspective and should thus reduce the debate's emotional factor. furthermore, the paper contributes to the large academic research stream focusing on financial stability and insolvencies in football. the paper is structured as follows. first, we provide a short description of the football labor market and the clubs' financing, including an overview of the related literature (sect. ). we then present a theoretical framework and general definition for speculative bubbles alongside a description of the kindleberger-minsky model, which explains the different phases of a typical bubble. next, we apply the model to the current situation in european football (sect. ). this is followed by a discussion of the main risk factors for the recent transfer fee growth (sect. ). finally, we conclude our analysis in sect. . every market (e.g., stock exchanges, housing, arts, etc.) is defined by the existence of buyers and sellers who meet to trade goods and services (mankiw and taylor , p. ) . despite its peculiarities and restrictions (e.g., specific transfer periods), the football labor market is no exception (franck ; frick ) . as a result, players can be seen as assets from an economic point of view (morrow ) comparable to tulips, houses, or company shares, and this aligns with previous analyses (tunaru and viney use the term "vulnerable assets"). however, according to a more nuanced and technical definition, "it is not […] the player himself who is the asset, but rather the provision of services" by a player in the future (morrow , p. ) . football clubs, on the other hand, act as buyers and sellers who trade the football players' registrations. clubs spend money to purchase player registrations to improve their roster and increase the club's utility. by owning player registrations, a club's utility can increase in two ways: either through a player's contribution to the club's sporting and commercial success during the contract period or by making a profit after selling the player at a price exceeding the initial acquisition costs, which is comparable to other investments for example in the stock market. thus, a club does not necessarily look to sell a player registration at a higher price for a financial profit at a later date. if a club is interested in buying a player from another club, the purchasing club has to contact the selling club. transfer fees stem from negotiations and therefore constitute an agreement between the purchasing and selling clubs about the appropriate price. like other labor markets, a crucial determinant of transfer fees and wages is the player's marginal revenue product, which represents the marginal utility a club can generate by signing a player (daumann , p. ; lucifora and simmons , p. ) . consequently, clubs will be willing to pay a higher transfer fee in case of a higher marginal revenue product of the player. previous literature agrees that transfer fees (and, therefore, the players' marginal revenue products) are based on both club and player-related factors (e.g., carmichael and thomas ; speight and thomas ) . frick ( ) provides an overview of the early empirical studies that have evaluated the determinants of transfer fees and market values, whereby the latter are often used as proxies for actual transfer fees. club-related factors include gate attendance and the club's sporting success (e.g., speight and thomas ) . player related factors have been categorized into four principal dimensions among other player characteristics: the player's human capital, performance, popularity and effort (müller et al. ; weimar and wicker ) . examples of the player's human capital used in previous studies are the player's nationality and age (e.g., carmichael et al. ; muehlheusser et al. ; dimitropoulos et al. ) . in recent years, with an increasing number of companies focusing on the data gathering of football statistics, contemporary researchers can include a diverse set of player performance measures (the second dimension) such as passes, shots and tackles, including the respective success ratios (e.g., herm et al. ; richau et al. ) . the third dimension (player popularity) refers to the superstar literature largely based on rosen ( ) and adler ( ) and examines the relationship between talent and popularity. rosen ( , p. ) defines superstars as "relatively small numbers of people [who] earn enormous amounts of money and dominate the activities in which they engage". he claims that small talent differences lead to considerable earning differences due to a lack of adequate substitutes for great talent. adler ( ) , on the other hand, says that superstars emerge even in the case of equal talent because positive network externalities associated with popularity create a snowball effect. by analyzing the superstar phenomenon and the effect of popularity in football, previous studies have shown a positive effect of popularity-for example, operationalized as press citations (franck and nüesch ) , google hits (herm et al. ) or social media followers )-on the players' market values. as a fourth dimension, recently added to the discussion, wicker et al. ( ) analyze the effect of effort (e.g., running distance) on market values, but find only insignificant or negative effects. an overview of the studies on market values and transfer fees mentioned above is provided in table . still, despite numerous studies evaluating the determinants of transfer fees, the final price (so-called arbitration value, matschke ) is the outcome of the negotiations ) between the club that would like to buy a player and the selling club. if the negotiating parties respect their individual agreement conditions, both clubs increase their welfare. it is important to note, though, that the football labor market is judgment-based. although various (objective) player statistics are available as shown above, the question of how to combine these statistics into a single number (i.e., market value or transfer fee) is disputed. managers might assign a different weight to the various performance measures, which increases the extent of subjectivity in player valuations and transfer fee negotiations (for further information about judgments in markets that lack objective criteria, see karpik ; maurer and mikl-horke ) hence, the selling and purchasing clubs might differ in their judgments regarding a fair price. further, it is difficult to measure the individual marginal revenue product (i.e., payment streams) that a player will provide (e.g., through his contribution to the sporting success and his marketability) to the club since the value differs among clubs (ackermann and follert ) . given the absence of objective market values in the inherently imperfect football transfer market, past transfer fees of comparable players and other indicators (e.g., performance indicators, expert valuations) can be used as reference points in the negotiations (for market values as an argument in negotiations, see ackermann and follert ) . furthermore, it is worth noting that the inherent bidding process on the football labor market might come with a winner's curse whereby the winning bidder overinvests (based on capen et al. ) . previous research argues that this phenomenon exists also in the sports industry (andreff ) such as baseball (cassing and douglas ) . in the case of player trading, a winner's curse can exist if multiple clubs want to sign a player. this can force the winning club to acquire the respective player by spending more than the player's marginal revenue product (andreff ) . however, among other motives, a club might be willing to overinvest to prevent other teams from benefiting from the player's marginal revenue product (i.e., weakening other teams). to finance transfers and meet other financial obligations, football clubs rely on internal and external financing. their internal financing consists primarily of broadcasting, matchday, and sponsorship revenues whereby the major part of their revenue comes from broadcasting and sponsorship/commercial revenues in the major five leagues dworak ) . on the external side, football clubs can rely on multiple financial instruments, including bank loans, bonds, and money injections by external corporate or private partners (berenberg ) . for external suppliers of financial resources (e.g., banks), loans provided to sports clubs are especially risky since the outcome and respective returns of sports events resemble "a mix of predictability and gambling including a systematic tip" (emrich et al. , p. , translated) . hence, the importance of bank loans for football clubs has decreased in past years due to the clubs' limited credibility. as an alternative, clubs, particularly in germany, have tried to use other forms of external financing such as fan bonds (weimar and fox ) . previous literature points out that the clubs' team investments (e.g., wages, transfer fees) have a positive effect on their sporting success (e.g., forrest and simmons ; frick ) . thus, the more (internal and external) capital football clubs can generate, the higher their spending power on the labor market, which increases the probability of sporting success. often akerlof's ( ) metaphor of a "rat race" is used to explain the clubs' investment behavior where clubs overinvest to increase their chances of sporting success (e.g., rohde and breuer ) . previous literature supports that this investment behavior leads to an overinvestment environment with low profitability and the clubs' high debt levels (e.g., dietl and franck ; franck a franck , . it is further argued that football clubs face a soft budget constraint (following kornai kornai , ) that drives the overinvestment environment since, from a social perspective, clubs are too big to fail and stakeholders will bail out those clubs in financial distress (andreff ; storm and nielsen ) . a related topic discusses the circumstances for insolvencies in football, the key drivers for which are shocks (e.g., player injuries, relegation) that affect the clubs' financial stability szymanski ; szymanski and weimar ) . beech et al. ( ) name ( ) relegation, ( ) outstanding tax payments, ( ) soft debt turning into hard debt, ( ) loss of stadium ownership and ( ) repeat offenders as the main reasons for insolvency. in light of the low profitability in european football, another recurring topic concerns the objective function of football clubs that discusses if football clubs act as win-or profit-maximizer. sloane ( ) was first to note that european football clubs try to maximize wins under a financial break-even constraint, which aligns with today's principal view (e.g., garcia-del- barrio and szymanski ; kesenne ; sloane ) . besides the scientific contributions mentioned above, football-governing bodies (uefa) and corporations (e.g., deloitte) regularly publish studies that provide overviews of the financial state of (european) football. these are frequently cited in the existing academic literature (e.g., storm and nielsen ; szymanski ) and thus underscore their relevance and reliability for academic and public discussion. in economic terms, a speculative bubble exists if "assets trade at prices that are far in excess of an estimate of the fundamental value of an asset, as determined from discounted expected cash flows […]" (vogel , p. ) . thus, the key element of a bubble is the unsustainability of a given price structure that will eventually result in an implosion of the existing system (kindleberger and aliber , p. ) . minsky ( , p. ) describes the characteristics of bubbles and financial instability, stating that "the critical element in explaining why financial instability occurs is the development over historical time of liability structures that cannot be validated by market-determined cash flows or asset values. thus, in a free market, wide and spreading bankruptcies can occur […]". in particular, taking into account that the view of fully informed market participants is a fiction and that significant asymmetries exist among market participants, liabilities must be backed up with constant and proportionate revenue streams if financial instability are to be precluded based on minsky's argumentation. one framework that explains the pattern of speculative bubbles is the kindleberger-minsky model. besides its application to several historical bubbles (e.g., saqib ) , the work of charles kindleberger-who built on a model by hyman minsky-drew renewed attention following the new economy and housing bubbles (e.g., gray ; rosser, rosser and gallegati ) . the kindleberger-minsky model takes a conceptual approach by separating the rise and burst of a speculative bubble into five phases, which characterize the typical pattern of such events: ( ) displacement, ( ) boom, ( ) euphoria, ( ) financial distress, and ( ) revulsion ( fig. ). the first phase (displacement) involves some regulatory or other externally driven changes to the existing system that provide investors with new business opportunities. these opportunities are often promoted by the mass media and trigger the demand for these business opportunities. with the boom comes a price increase that starts slowly but accelerates quickly as the investment draws public attention; this results in a growing number of market participants. the price increase is driven by a growing and easier supply of (external) financial resources, which lead to higher investor indebtedness. sources of financial resources can be financial institutions or private lenders, inter alia. the euphoria stage is accompanied by a strong belief that prices will rise indefinitely-although the inherent risk is acknowledged, individuals feel compelled to take part in these opportunities if they do not want to leave the competition. in this way, investments that fueled the price increase in the first place and brought high returns now attract even more investors. prices become disconnected from established valuation concepts, given the belief that the asset can always be sold at a higher price while financial resources, as a prerequisite for the investment, can be accessed easily (kindleberger and aliber , pp. - ). thus, the success of the investment becomes a self-fulfilling prophecy (merton ) . then, abruptly, following a supposedly unimportant event, prices begin to drop (financial distress). market participants who rely heavily on debt financing for their investments find themselves in trouble, as the assumption of rising prices no longer holds. investors need to off-load their assets to generate liquidity, which further erodes the price level. in the final phase (revulsion), prices drop rapidly to extremely low levels-until, for instance, a "lender of last resort" steps in and prices stabilize (kindleberger and aliber , pp. - ). according to kindleberger and aliber ( , pp. - ) , although speculative bubbles feature a similar pattern, their causes and origins can vary. from a financial perspective, however, the authors suggest that ease of access to financial resources (e.g., through low interest rates), which can result in unsustainable indebtedness, is a common element to speculative bubbles. from one viewpoint, speculation can be considered the core of economics. as well as from a financial perspective, research has examined extreme price movements from a psychological perspective (for an overview of various psychological aspects, see vogel ) . in past discussions of extreme price movements, economists often assumed that investors behave rationally in their investment decisions. blanchard and watson ( ) , for example, describe the concept of rational bubbles, whereby deviations from fundamental values can be deemed rational from an investment perspective. this view of rationality, however, contrasts with observations made by studies that focus on psychological aspects. among other findings, studies that focus on behavioral finance have found that people often adhere to their established beliefs and assumptions and thus oversee warning signals (vogel , pp. - ) . furthermore, bubble-building is often associated with the investors' herding behavior when they react to market signals and actions by other investors (for more details on signaling, see bikhchandani et al. ). in the great crash , galbraith ( explains this behavior: based on first successes with an investment, other (institutional and private) investors follow, which leads to a further (exponential) price increase. external pressure on all investors not to miss out on this business opportunity accompanies the herding behavior (galbraith ) ; additionally, successful investors who jump on the bandwagon early signal the prospect of further rewards. a differentiation is made between rational and non-rational herding: while non-rational herding implies following other investors without considering personal information, rational herding means that investors lack personal information and must rely on information from other investors (aytaç et al. ; for further details on rational herding behavior, see bikhchandani and sharma ) . in the current debate, the question of a speculative bubble in football is based largely on the recent rapid increase in transfer fees (fig. ) . a time trend analysis confirms a statistically significant increase of the aggregated transfer fee spending for all leagues ( table ). the premier league stands out with a trendline that shows an annual increase of . €m, which is almost three times higher than the increase of the other leagues. a time lagged regression further indicates that the transfer spending in the premier league, serie a and bundesliga is to a large extent driven by the previous year's spending with an adjusted r between . and . for these leagues (table ). in line with this discussion, we use transfer fees as a price indicator to examine the existence of a speculative bubble, by applying the kindleberger-minsky model to the football market. in so doing, we divide the development in european football into the first three kindleberger-minsky stages (i.e., displacement, boom, euphoria). based on the kindleberger-minsky model, it appears that a structural or regulatory change in the market opens or creates new business opportunities. in football, the development within the last decade can be traced to three main "exogenous shocks", besides some overarching developments. as such, although the individual shocks did not automatically lead to an immediate jump in revenues, the changes laid the foundation for recent-year developments and can thus be considered "displacements" in the sense of the kindleberger-minsky model. while revenues and transfer fees in european football have been growing for several decades (deloitte , p. ), the past decade (especially the last years) have shown increases beyond levels seen in previous decades. because these recent changes have triggered the public discussion of whether there is a speculative bubble in european football, we focus on the revenue growth in the ten most recent years in this study. the first "shock" concerns the transfer system in european football. although they had only an indirect effect on today's growth in transfer fees, the regulatory changes regarding the transfer system laid the foundation for today's trading of player registrations. starting with the bosman ruling in , players whose contracts had expired were allowed to move to a new club without the previous club being compensated by a transfer fee (büch ; feess and muehlheusser ) . this change, representing the first liberalization of the football labor market, was followed by the monti system in (as well as by later adjustments), which set the contract duration to a maximum of years. in , the uefa limited transfers to two transfer periods-one in the summer and one in the winter (warshaw ). a second regulatory intervention, driven by a government investigation, changed the bidding process for (domestic) live tv broadcasting rights in europe. besides limiting the duration of tv broadcasting contracts to years in the united kingdom, the "no-single-buyer-rule" came into effect starting with the sale of tv rights for the / - / seasons in the english premier league. the intention of the rule was to ensure fair competition in the football broadcasting rights market by guaranteeing the sale of live packages to more than one bidder (connolly ) . the entry of (foreign) private financial investors, as the third "shock", started in with mohamed al-fayed taking over fc fulham. the actual starting point of an increased number of (foreign) financial investors, however, was the entry of roman abramovich in at chelsea london. following this prominent example, european football has seen an increasing number of non-european investors taking over football clubs (rohde and breuer ) . despite different investor types and motives, it appears that media-attracting examples such as paris saint-germain and manchester city have had a considerable impact on the economic development of european football. as a consequence of these displacements, more money than ever began to flow into the market that the clubs could spend for example on player transfers; these inflows primarily comprised broadcasting revenues and money injections by financial investors. to understand the drivers of rising cash inflows, we now examine the economic processes behind the boom phase. by evaluating the increase in tv broadcasting revenue, we find that on both the demand and supply sides, some changes occurred based on the regulatory changes. we illustrate the rise by focusing on the english premier league, as the most prominent example. however, other leagues-such as the german bundesliga-have shown similar trends (albeit to a lesser degree) and comparable underlying reasons for the increase (despite some local peculiarities and different starting points). as described, one major driver in the increase in domestic and international broadcasting income (fig. ) has been intensified competition in the bidding process. the joint selling approach to the sale of broadcasting rights seen in most leagues-in which there is only one seller (i.e., the respective league)-prompted regulatory intervention that sought to prevent collusion and ensure innovation as well as a fair and competitive bidding process in the upstream market of tv broadcasting rights. following an intervention in england as the first major european league, england, italy, france, and germany now all have in place the "no-single-buyer-rule" (ramcke ). on the supply side, to allow more broadcasting companies to tender a successful bidding attempt, the leagues increased the number of packages offered at the auction. the premier league, for example, offered four live broadcasting packages in the bidding process for / - / ; this increased to six in the following period and seven for the / - / auction (connolly ). one peculiarity of the english market is that not all games are included in the live broadcasting packages. thus, to attract more bidders and increase prices, the league increased the number of live broadcasting matches in previous years to (kalenberg ) . in germany, for example, to increase the exclusivity of single matches for bidders and establish a second bidder besides sky, the number of simultaneously played matches was reduced (ramcke ) . the regulatory requirements and changes on the supply side also affected the demand side of the bidding process. in particular, it seems that new (international) competitors were challenging the incumbents. given the necessity for (at least) two successful bidders, the chances of a successful bid for the individual companies increased, resulting in intensified competition from an economic perspective. the most relevant new competitor in the united kingdom was british telecommunications (bt), which entered the market before the / auction and led to a considerable revenue increase in the following two periods. before bt entered the scene, sky faced rather weak competitors that struggled to hold the rights for one period. other new entrants in the premier league in recent years have been espn, bein, and amazon. markets in the other top leagues (e.g., orange, telefonica, and vodafone in spain) saw a similar increase in competitors (connolly ; ramcke ) . questions, however, arose concerning the profitability of the broadcasting companies, especially following the reduction of individual package sizes. indeed, feuillet et al. ( ) find a "winner's curse"-not for the holder of the majority of the packages but for that with the smaller offering ("second winner"). from a financial perspective, broadcasting companies might nonetheless follow additional strategic objectives. one such objective could be to leverage the sports offering (e.g., football) to attract customers to their full product portfolio. in cases where such a strategy is used, broadcasting companies might accept low stand-alone profitability as a form of internal subsidization (feuillet et al. ) . beyond the regulatory requirements and increased competition, some complementary developments have been driving broadcasting revenues. one such development is the ongoing internationalization strategy of the european clubs, wherein the clubs especially focus on china and the united states as two large markets with high growth potential (deloitte , p. ) . by holding organized practices and matches in these countries as well as opening local offices, the leagues and clubs increase public awareness and their foreign fan base and thus increase demand for foreign broadcasting rights. one tangible result was the revenue increase from broadcasting rights, for domestic and international packages alike (uefa , p. ) . moreover, the growing importance of digital media channels and social media provided clubs and leagues with new opportunities to promote the product internationally (mccarthy et al. ) . besides football, prices have increased for broadcasting and media rights for many other sports events worldwide (e.g., north american sports, olympics), which underscores the great global marketing potential of major sports events (chapman ; crupi ) . the second driver to increase the financial means available to clubs is the entrance of financial investors, particularly in england, italy, and france. based on the increasing importance of international markets for the leagues, club ownership became more and more attractive to foreign investors (deloitte , p. ) . today, club owners, especially in england but increasingly in italy and france, come from all over the world. as for bubble-building and foreign cash inflows, it should be noted that past bubbles were further fueled by foreign investors (sornette and zhou ) . from the financial side, investors in football do not necessarily expect a financial return; in this sense, football clubs differ from typical investments (for a literature overview on club ownership and financial investors in european football, see rohde and breuer ) . however, some investors might try to create direct financial gains by trading players or by increasing the value of the club and sell it at a higher price later (bergin and bryan-low ) . in the literature, us-american investors in particular are frequently considered profit maximizers (nauright and ramfjord ) . a different category of investors follows strategic objectives, such as being involved in the business affairs of their club's home city (mcgeehan ). further, some investors look to generate some kind of positive externality through their club investment by promoting their main business or other investments (franck b) . in addition to these business-oriented motives, there is a reputational component that makes sports investments attractive to external individuals. investors such as chelsea london's owner roman abramovich seem to care most about the positive recognition about the good they are doing and the sporting success they bring to the club. especially for investors looking for positive recognition, the football market (which is inherently emotional) is more promising than investments in other industries. this is because investments in football allow investors to be associated directly with the club's results in public discussions (franck a ). in the case of institutional investors and investors connected to (foreign) governments, investments in sport (and in football clubs particularly) can serve as parts of a reputation management strategy and nation branding (buschmann and winterback ) . furthermore, football club ownership might also be a hobby for rich business people (franck b) , as a kind of "conspicuous consumption" (veblen ) . besides the entry of financial investors, income from merchandising and advertising has grown; this, again, refers to the increased professionalization and commercialization that has taken place in european football and opened new revenue potential (berenberg ) . as a further driver for the revenue increase at least for the most successful teams, prize money in international competitions has increased considerably in recent years (rohde and breuer ) . although it remains unclear whether the european football market has reached the kindleberger-minsky "euphoria" stage, several key components of that stage indicate that euphoria is very unlikely in the football market, given its peculiarities. as kindleberger and aliber ( , p. ) note, price increases are often "accompanied by an increase in trading volume." fig. , however, shows that there has been only a slight increase in trading volume (i.e., the number of new arrivals) in european football (except spain), despite some variations. most aspects that favor speculative trading-a characteristic of the euphoria stage-are regulated in the football labor market. first, the fixed and rather short transfer windows largely restrict the possibility of large fluctuations in, and speculations on, player values. moreover, players can be registered for only up to three clubs per season (fifa , p. ). hence, football player registrations are not assets of a speculative nature, like company shares or other financial products with frequent trading and price changes. further, it is not necessarily the club's objective to make a profit with player trading. as described, clubs can benefit from player registrations through their contribution to the team's sporting or commercial success. unlike many other markets, only a small group of club managers can trade players, preventing those who lack the required expertise (e.g., hobby managers) and just follow others from investing in player registrations. as a consequence, only a small group of people drives the transfer fees (i.e., the price), which restricts excessive (irrational) herding behavior and the belief in ever-growing prices. the recent development of transfer fees in european football-in particular the rapid increase from / to / -has followed a pattern resembling historical speculative bubbles as described by the kindleberger-minsky model. in particular, the premier league stands out in both recent revenue and transfer fee increases. following minsky ( , p. ), a bubble exists if the price increase is based on debt-financing and is not backed up by a sustainable cash inflow. thus, the development in football is not critical as long as higher transfer fees are backed by a respective cash inflow (i.e., broadcasting income, investors) that allows clubs to meet current and future obligations. additionally, in contrast to previous bubbles, the respective cash inflows presented in chapter . . primarily represent actual revenues instead of external financing. to complement the findings from the kindleberger-minsky model, we analyze typical indicators for bubble-building used in other industries. in the housing market, the price-income ratio is a well-established indicator for testing speculative bubbles and financial stability (rosser et al. , p. ); deloitte uses the wagerevenue ratio in its yearly football report (deloitte , p. ) . as an extension, we check for the relation between club revenue and transfer fee spending, both on an aggregated level. we set revenues (excluding player trading) from t − in relation to transfer fee spending in t, assuming that clubs can spend their revenues in the following transfer window at the earliest. this indicator allows us to check the sustainability of the current transfer fees, in the sense that a rising ratio indicates that the growth in transfer fee spending is exceeding revenue growth. figure shows a mixed picture by leagues. while a trend analysis (table ) shows no statistically significant increase for laliga, a small (significant) financial investors provide equity or soft loans. however, usually, these loans are not expected to be paid back (beech et al. ) and are thus not comparable to typical loans known from historical crises. time trend analysis transfer fee/revenue ratio * p < . ; **p < . ; ***p < . ; standard error in parenthesis dependent variable: transfer fee/revenue ratio increase can be observed for the premier league and bundesliga. the serie a and ligue , however, show the highest annual increase of . in the trend analysis (statistically significant). furthermore, the transfer fee/revenue ratio to a large extent depends on the ratio of the previous year in the serie a compared to the other leagues (bundesliga as an exception as shown in table ). this trend raises the question of the financial sustainability of the french and italian leagues, since transfer fees constitute only one part of the expenses, while (player) wages, signing bonuses and other expenses (e.g., agent fees) also need to be paid. as fig. shows, according to the latest figures, wage costs in italy and france are also particularly high, relative to their revenues (i.e., % and %, respectively). the wage situation in italy and france needs particular monitoring, as % is considered the critical wage-revenue ratio threshold, assuming that no considerable income is generated from transfer activities. while italy has significantly reduced its wage costs relative to revenues in recent years (table ), the latest (deloitte , p. ) . furthermore, the trend analysis shows that the premier league is the league with the highest decrease of the wage/revenue ratio in the past years. the time lagged analysis again indicates that the wage-revenue ratio of the previous year has a higher influence on the indicator in the serie a than in other leagues (indicated by r in table ) . other important indicators are the club's debt-revenue ratio and (bottom-line) profitability. the debt-revenue ratio is presented in fig. and shows a stable development for the bundesliga, laliga and ligue despite transfer fee increases in the past years. while the increase in fy for the premier league can be attributed chiefly to tottenham's stadium investments (uefa , p. ) , the serie a stands out in two ways: it shows a considerably higher ratio than other leagues and has experienced a further increase beyond the levels of the other leagues (except for the premier league). looking at the profitability by considering both sides of player trading (i.e., buying and selling), the italian clubs additionally reported a bottom-line loss of € m for fy . in france, the bottom-line loss of € m in fy was followed by a small profit in fy . similar to the previous years, england, spain, and germany were the most profitable european leagues, with profits of € m, € m, and € m, respectively for fy (uefa , p. ) . overall, despite some watch-outs in france and particularly italy, the indicators suggest that, rather than a speculative bubble, there has been an "elevator effect" (beck , pp. - ) in european football. in addition to (pretransfer) revenue increases, football clubs in the five major european leagues not only buy player registrations at higher prices but also sell them at a higher level to mitigate or preclude bottom-line losses. moreover, in looking at aggregated numbers, it becomes clear that a cumulative effect has further fueled the development of transfer fees. since monetary circulation in football is partially closed, money injected into the market largely remains in the system by being circulated multiple times with each transfer among the clubs. hence, at this point, there seems to be no league-wide speculative bubble in economic terms based on recent transfer fee increases in any of the major european leagues, especially since cash inflows are largely based on economic processes rather than on speculative behavior. in this sense, the situation in european football can be defined as an atypical bubble: while showing a similar pattern to other bubbles, typical growing indebtedness without proper cash inflows is missing. however, france and especially italy need to be careful not to further outgrow their cash inflows and slide into financial instability or even a bubble. furthermore, accounting for player transfers incorporates some peculiarities that need to be considered when interpreting today's bottom-line figures: while transfer income is effective immediately after the sale, costs are distributed over a player's contract duration (uefa , p. ) . while looking at aggregated data earlier, a deep-dive on the profitability indicates that some clubs might "live in a bubble" despite the absence of a league-wide bubble. however, with some clubs living beyond their means, this does not mean that the entire league is subject to a speculative bubble. football club insolvencies are not uncommon (szymanski ) , and thus an individual insolvency does not automatically lead to a "bursting bubble," in the sense that the entire market will collapse. however, a chain reaction following multiple (and almost simultaneous) insolvencies among football clubs could entail just that. for the premier league, bundesliga, laliga and ligue , a great majority of the clubs reported a bottomline profit for fy . however, the club-level overview confirms that especially the italian league should carefully monitor the investment behavior in order to avoid the metaphor of an "elevator effect" refers to the increasing (absolute) wealth for the entire society in western germany following the second world war and thus moving up with the elevator (beck ) . based on the paradox of competition (see grass and stützel ) , clubs can only improve their position if they increase their team investments more than other clubs (sentence about the mechanics of size). if, however, all clubs increase their team investments, relative sporting differences will remain. the result is just a higher spending behavior for all clubs (global sentence). except for fy in england and italy. slight increase in number of clubs with moderate bottom-line losses compared to fy in the premier league and bundesliga. a potential bubble-building as the majority of the clubs reported a loss in fy , which represented a further worsening of the situation compared to previous years (uefa ). in the past, the leagues and the uefa have already implemented several measures, such as the financial fair play regulations (schubert and lopez frias ) , to ensure the financial stability. should the situation become more challenging, these regulations might be extended in the coming years. the results of this study show that especially the french and foremost the italian league should carefully monitor the financial development and could consider to enforce stricter financial regulations if the situation does not improve. similar to the rapid revenue increase that drove the clubs' team investments in recent years, an opposite trend can lead to lower revenues. two of the main associated risks are decreased broadcasting income and the exit of wealthy club owners. however, the risk of an immediate drop in revenue because of decreased broadcasting income is limited since contracts are negotiated or years before coming into effect. thus, even in cases featuring lower contract values, clubs can adjust their team investments accordingly. furthermore, it is unlikely that a rapid decrease will occur although the interruption due to covid- caused some uncertainty regarding the broadcasting income for the / season. the premier league, for example, experienced a slight decrease in its latest domestic broadcasting deal ( / - / ) rather than a drastic one; furthermore, this was offset by an increase in the international broadcasting deal (deloitte , p. ). still, these events might indicate that some leagues have reached a turning point, or at least a plateau, concerning domestic broadcasting contracts. however, given the lead time in advance of a drop in cash inflows, cash inflows from financial investors are more risky revenue sources. if the investor is heavily engaged in the club, such an exit can result in severe financial problems if there is no appropriate replacement. if these risks were to occur and resulted in lower revenues, the risk of a leaguewide bubble would increase in case clubs do not adjust their spending behavior accordingly. the potential consequences of a bursting bubble would affect the sporting as well as financial dimensions of european football. from a financial perspective, pending payments for transfer fees, wages and other obligations would occur at a time of financial distress. ultimately, the number of insolvencies among football clubs might increase, especially in the lower divisions. this raises the question if managers should adjust their investment behavior in order to prepare for a potential (long-term) downturn. from the managers' perspective, however, the observable investment behavior seems rational. as managers are often judged according to the club's short-term success, they are incentivized to improve team quality, which requires higher investments instead of greater profitability. this aligns with the predominant view that clubs are win maximizers subject to a breakeven constraint. manager behavior similarly aligns with the expectations of fans, who are generally more concerned with sporting success than with the club's financial stability-unless the club's financial stability is in question (tunaru and viney , p. ) . this view is supported by previous research on insolvencies (e.g., szymanski ) that indicates that often negative shocks rather than irrational behavior lead to insolvencies. furthermore, higher profit requirements are seen as a competitive disadvantage in football's arms race (franck a) . thus, managers are not incentivized to prevent overspending and potential bubble-building. nevertheless, from an economic perspective, it would certainly be recommended to use at least part of the additional cash inflow in a less risky way. examples are investments in the club's long-term infrastructure, support for the local community, or increases in the club's equity as a capital buffer to prevent financial distress in case of decreasing revenues. as shown in this study by applying the kindleberger-minsky model to football, the pattern of current developments in european football resembles that of an atypical bubble, with the characteristic shape of bubbles; however, unlike historical bubbles, there are proportionate cash inflows that justify the recent transfer fee increase for most of the clubs in europe's five top divisions. from a contemporary perspective, the developments in football seem to be a continuation of the overall growth that the football market has experienced for decades, though fueled and accelerated in the last decade. although some experts discuss the presence of a speculative bubble in football, it can also be argued that the football market was undervalued in previous decades and that better fundamentals (i.e., higher revenues) show higher marginal revenue products of the players and thus led to higher team investments in recent years. a study by the berenberg bank ( , p. , translated) sums up the developments in football: "higher revenues land in the pocket of the players due to intensive competition for these best players." when this thinking is brought to bear on the current study, it appears that this war for talent increases not only player wages but also transfer fees. from today's perspective, it appears there are three possible scenarios for the future that ensure a sustainable development. first, revenues continue to grow, and this is accompanied by a further increase in transfer fees. second, the current revenue and transfer fee levels become the "new normal" and remain at those same levels. finally, the revenue drops, which would require the clubs to lower transfer fees. especially with respect to the current covid- crisis-which can be seen as a negative shock (following szymanski ) not only for football but for the whole economy-decreasing revenues and hence decreasing transfer fees can be anticipated at least in the short-term. thus, the crisis represents a stress test for the financial stability of european football and reveals the financial instability of some clubs. although our study indicates that there is no bubble on an aggregated league level, the pandemic reinforces the financial problems especially for those clubs with unsustainable spending before the crisis. the main limitation of this study is that it provides no final assessment. one peculiarity of bubbles is that they are impossible to predict in advance; thus, this paper provides only a conceptual evaluation that uses available parameters and considers the experiences of past bubbles and financial crises by applying the kindleberger-minsky model. only future developments can conclusively indicate whether there is a bubble in european football. hence, even the use of a more sophisticated approach would not yield a clear answer to the question of whether a speculative bubble exists. other limitations concern the scope of this study. while focusing on the european top leagues, the financial stability in lower divisions might be more uncertain (e.g., szymanski and weimar ) . based on the observed tendency to gamble for success (franck a) , it might be tempting for lower-division clubs to overspend to benefit from the considerably higher broadcasting income in the first division. similarly, the risk of a bubble-building in the top divisions of other european countries that generate lower revenues or rely on selling home-grown talents might be higher if they simultaneously try to compete in the european "rat race". the findings of this study suggest several promising areas for future research. first, the individual components of this study (e.g., economic mechanisms affecting broadcasting income and empirical studies on financial investors) can be further investigated. a second research stream relates to the role of agents in football contract negotiations. this is especially relevant because agents receive a provision for every transfer, based on its value, and therefore have a pecuniary interest in concluding as many transfers as they can. third, in light of increasing transfer fees, loans are becoming more popular in football; this represents a different contract construct that deserves a closer look in future research. finally, a troubling implication of transfer fee developments is that clubs are starting to scout players at even younger ages, to avoid high transfer fees, which opens new areas of strategic and ethical issues that should be addressed in future research. einige bewertungstheoretische anmerkungen zur marktwertanalyse der plattform transfermarkt stardom and talent the economics of caste and of the rat race and other woeful tales french football: a financial crisis rooted in weak governance the winner's curse: why is the cost of sports mega-events so often underestimated? the winner's curse in sports economics herding behavior among wine investors risikogesellschaft: auf dem weg in eine andere moderne _beren berg% m akro_trend s_finan zieru ng_im_profi fussb all_final .pdf. accessed wie investoren am transfer-boom im fußball verdienen wollen learning from the behavior of others: conformity, fads, and informational cascades herd behavior in financial markets in: wachtel p (ed) crises in the economic and financial structure european soccer has an inflationary bubble that will eventually burst 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no conflict of interest. key: cord- - ezfke z authors: baruah, h. k. title: an empirical inference of the severity of resurgence of covid- in europe date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: ezfke z in europe the corona virus spread had started to retard months ago, but after some time it has started to accelerate again. in this article, we are going to analyze the current covid- spread patterns in italy, the uk, germany, russia, spain and france. we have found that the current spread has perhaps been underestimated as just the second wave. as per our analysis, as on october the resurgence is much more vigorous than the first wave of spread of the disease. it is going to be most serious in russia, followed by italy, germany and the uk, while in spain and france the patterns are yet to take inferable shapes. in the european countries the second wave of covid- has been observed to be much more serious than the first one. the data clearly reflect resurgence of the disease, but how serious the matter is can be judged only after a proper data analysis. in this article, we are going to show how to infer empirically the severity of the spread. we shall show that the covid- situation in europe is indeed very serious. a look at the daily increase in the cumulative totals of the covid- cases in the european countries mentioned above and the world gives us an apparently clear picture how much europe is contributing to the daily increase in the total number of cases in the world. for example, on october the increase in the world total were out of which the uk contributed ( . %) cases, italy contributed ( . %) cases, germany contributed ( . %) cases, france contributed ( . %) cases, russia contributed ( . %) cases and spain contributed ( . %) cases. the total increase on that day in these six european countries were ( . %) to the total increase in the world. on that same day, india which is currently contributing the maximum to the daily increase in the world contributed ( . %) cases while the usa on that day contributed ( . %) cases. therefore it can be seen that on october , these six european countries together had contributed more to the world total than the usa although the contribution was less than that from india. however, we have to note that in india and the usa, the spread is retarding while in these six european countries that kind of retardations took place months ago, and thereafter it has started to move towards resurgence -the so called second wave. indeed the figures for one single day may actually be misleading. we shall put forward a numerical analysis of the spread data for a period of days from september to october. we would like to mention at this point that the current growth curves of the disease in these six european countries are obviously not like the curve assumed in the compartmental epidemiological models [ , . ] . at the start of an epidemic, the classical compartmental models do not presume a change from a retarding state to an accelerating state. it is apparent from the graphs published by worldometers.info [ ] that the spread patterns are of the exponential type in the six european countries mentioned above. we shall consider the total number of cases in these six countries from september to starts to follow a reducing trend. it was earlier observed that a change from a nearly exponential pattern to a nearly logarithmic pattern took place in italy [ ] months ago. in the other five european countries too such retardation must have followed the nearly logarithmic pattern. it was observed [ , , , , is really useful. we now proceed to analyze the recent data on the cumulative total number of covid- cases in the six european countries mentioned above. we shall now put forward our empirical analysis of the covi- spread patterns in the six european countries concerned. in table- showing an unusual increasing trend, and such a thing is not assumed in any classical epidemiological mathematical model. . cc-by-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted october , . ; https://doi.org/ . / . . . doi: medrxiv preprint . cc-by-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted october , . . cc-by-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted october , . ; https://doi.org/ . / . . . doi: medrxiv preprint in russia, have shown a straight increasing trend which is very unusual in the growth of an epidemic. how long it would continue to have such an increasing trend is not possible to judge. a diagrammatic representation has been shown in fig. . . cc-by-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted october , . ; https://doi.org/ . / . . . doi: medrxiv preprint in spain, table- , the situation is different from that in italy, the uk, germany and russia. in spain, no increasing trend could be seen, but the values are not showing any constancy and they are not showing any decreasing trend as well. the diagrammatic representation has been shown in fig. . . cc-by-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted october , . ; https://doi.org/ . / . . . doi: medrxiv preprint in france too, have been found to be nowhere near any constant value. the values are not increasing, but they are not showing any decreasing trend also. the diagrammatic representation has been shown in fig. . . cc-by-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted october , . show a linearly reducing trend with respect to time until a retarding phase starts and in that phase the spread pattern becomes nearly logarithmic. in the standard epidemiological models also this kind of a growth pattern is assumed. however, the pattern in the second wave of the epidemic is different in the sense that ∆ ‫ݖ‬ ሺ ‫ݐ‬ ሻ in these six european countries has been showing an increasing trend instead of a decreasing trend as was observed in earlier studies. there can be only one explanation of this phenomenon. in the first wave, zሺ‫ݐ‬ሻ was of first degree but in the second is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. (which was not certified by peer review) the copyright holder for this preprint this version posted october , . ; https://doi.org/ . / . . . doi: medrxiv preprint a contribution to the mathematical theory of epidemics contributions to the mathematical theory of epidemics ii. the problem of endemicity contributions to the mathematical theory of epidemics iii. further studies of the problem of endemicity total corona virus cases in india, publishing date the covid- spread patterns in italy and india: a comparison of the current situations a simple method of finding an approximate pattern of the covid- on reliability of the covid- forecasts , but this kind of an exponential growth was not seen in the first wave which is why it was observed that the values ofwere showing a linearly decreasing trend earlier.therefore we can empirically infer that as long as unusual. in effect, the resurgence is following a much faster growth pattern different from what was seen to be followed when covid- had appeared. the covid- situation in europe is showing an unusual trend. how long would the situation remain in this way cannot be said now. but one thing can be said with certainty that the situation has become explosive. in russia in particular, the growth is not following the standard epidemiological growth pattern. in italy, germany and the uk too, the growth is very unusual. in spain and france, the situation is uncertain as yet. we conclude that the growth pattern of the epidemic in the current resurgence in the six european countries concerned is of a type very different from the nearly exponential type followed in the accelerating phase in the first wave. key: cord- -ig nwtmi authors: nan title: th european conference on rare diseases & orphan products (ecrd ) date: - - journal: orphanet j rare dis doi: . /s - - - sha: doc_id: cord_uid: ig nwtmi nan theme: when therapies meet the needs: enabling a patient-centric approach to therapeutic development. background: rare diseases (rd) often result in a wide spectrum of disabilities, on which information is lacking. there is a need for standardised, curated data on the functional impact of rd to facilitate the identification of relevant patient reported/patient centered outcome measures (proms/pcoms) as well as for the use of validated quality of life instruments based on functional outcomes. to address these issues, orphanet is partnering with mapi research trust (mrt) in order to connect orphanet to proqolid ™ , mrt's proms/pcoms database, through disease codes. visit orphanet at www.orpha .net. methods and materials: the orphanet functioning thesaurus (oft) is a multilingual controlled vocabulary derived from the icf-cy. a subset of rd present in the orphanet nomenclature is annotated with the oft, with the addition of attributes for each functional impact (frequency, severity, and temporality) for each specific rd. annotations result from structured interviews with clinical experts, medical-social sector care providers, and patient organisations. in order to link proqolid ™ data with orphanet disability data, the taxonomy used to qualify rds in proqolid ™ was reviewed and mapped to orphanet's. all proms developed for rds were identified, and all products approved by the fda and ema from to with an orphan drug designation (odd) and a pro claim were listed. results: the orphanet knowledge base contains over rd, of which rd have been assessed for their functional consequences, of which rd have been annotated: the remaining rd were annotated, after discussions with medical experts, as either being highly variable, non-applicable or resulting in early-death. of the most prevalent rare diseases, have been annotated according to their functional consequences. rds had a prom (n = ) in pro-qolid ™ and . % of odd included a pro claim. the rds with the most prom were sickle cell anemia, spinal cord injuries, cystic fibrosis, all forms of hemophilia a and b and duchenne muscular dystrophy. prom used in labels were primarily focusing on symptoms ( %), rarely on functioning ( %) or health-related quality of life ( %). conclusions: linking these two databases, and providing standardised, curated data, will enable the community to identify proms/ pcoms for rd, and is the first step towards validated quality of life instruments based on functional outcomes. and eurordis. some patient organisations distributed the survey too more common disease patients as well, e.g. hashimoto's disease, and these responses were excluded. the survey asked patients to give suggested topics (i.e. fertility, heritability, tiredness, daily medicine intake, sleep quality, physical discomfort, and ability to work, partake in social life, and sports) a priority score and to suggest their own topics for research in open fields. open field responses were analysed with topic modelling and klipp-analysis. results: after exclusion of responses from more common endocrine disease patients, survey responses were analysed. most responses were received from northern ( %) and western europeans ( %), while southern ( %) and eastern europe ( %) were underrepresented. of the suggested topics respondent were most interested in research concerning the ability to work and participate in social life, and on tiredness. when patients were open to suggest their own topics, common responses included long-term side effects of drugs and quality of life. however, priorities differed between disease groups. for example, adrenal, pituitary and thyroid patients were more interested in research concerning tiredness than others. conclusion: with this survey endo-ern is provided with a large sample of responses from european patients with a rare endocrine condition, and those patients experience unmet needs in research, though these needs differ between the disease groups. the results of this study should be incorporated by clinical experts in the design of future studies in the rare endocrine disease field. purpose: when developing a health technology that requires clinical studies, developers institute working relations with clinical investigators. patient representatives can also create and manage advisory boards with product developers. this was of high utility in the s, in the development of products to treat hiv infection. inspired by this model, the european organisation for rare diseases (eurordis) proposes the eurocab programme to facilitate a two-way dialogue between patient representatives and medicine developers. as of , disease-specific cabs exist of approximately members each and others are being formed. methods: eurordis invites developers to sign a charter for collaboration with patients in clinical research, and provides guidelines together with a mentoring and training programme for patient networks. cabs help set the agenda with the developer, work on topics as diverse as study design, feasibility, informed consent and site selection, qol and proms, and organize the meetings. discussions also cover compassionate use, pricing, relative efficacy, etc. meetings last for to days with sessions with different developers, all under confidentiality. there are regular between-meeting teleconferences for trainings and action plan updates, and some cabs have instituted working groups on access, psychological support, etc. the collaboration is evaluated via a post-meeting survey send to both cab members and medicine developers. in addition, cabs have recently started to monitor outcomes of the meeting and progress towards their goals with a tracker tool. results: the results of the first surveys from distinct cab meetings with companies show that this form of shared decision-making is valuable as well as ethical for both parties. we have seen that working relations always continue, even when discussions become heated. all involved show interest in the co-creation possibilities of such collaboration and we look forward to seeing progress and change via the tracker. conclusions: monitoring and evaluation are crucial to understand whether and how the cabs are making an impact on medicine development. demonstrating impact is challenging because of the contextualized nature and complexity inherent to patient engagement collaborations in research design. eurordis is working within para-digm on our monitoring and evaluation strategy, focusing on improving its comprehensiveness and including multi-stakeholder perspectives. our current experiences show that the eurocab programme, with collective thinking and exchange between patients and a collaborative mentality from both sides, ensures high-quality and constructive dialogue with researchers and developers and can eventually inform both hta and regulatory decision-making (fig. ) . we have started to work on the metrics of markers of success. purpose: the french national network for rare sensory diseases sensgene launched in a -min motion design video ( fig. ) aiming at guiding healthcare professionals to welcome visually impaired patients in the hospital. this educational video was created to address patients' expectations and improve their experience in the network's hospital. the european reference network for rare eye diseases (ern-eye) collaborated on the project and created an english version of the video in order to distribute it widely in europe. method: sensgene worked on this project with two big french associations of visually impaired people: fédération des aveugles et amblyopes de france and fédération des aveugles alsace lorraine grand est. more than french patients' associations actively contributed to the project through five focus groups (workshops) which collected testimonies and gathered the needs of visually impaired persons and health-care professionals (fig. ). an evaluation was made by the independent body ipso facto: health professionals answered to a survey before and after viewing the video. results: the video deals with common situations in the delivery of care activities for different types of visual impairment: reception in a hospital center, consultations, moves and orientation in a hospital room (fig. ) . this fits perfectly with the needs of the patients reported in the focus groups. besides, the evaluation showed that % of them improved their knowledge on the topic. background: traditional appraisal and reimbursement approaches such as cost/qaly are increasingly recognised as being potentially unsuitable for rare disease treatments (rdts). approaches to appraising rdts vary across countries, from the same processes used for all medicines, to those completely separate from the standard, to adapted standard processes with greater willingness to pay (wtp). this study examines the impacts of standard versus special appraisal processes for specific rdts in selected countries. methodology: a case study analysis was conducted in which countries with a variety of rdt appraisal processes were selected, along with two rdts representative of the following criteria: rare/ultra-rare treatment, affecting child/adult, cancer/non-cancer, life-threatening/disabling. public hta reports for each country's appraisal of the selected rdts were retrieved and used to extract information into predesigned templates, which allowed for systematic comparison of the rdt processes across countries to compare and exemplify the impact of the different processes. results: reports from belgium, england, france, germany, u.s., italy, lithuania, netherlands, poland, romania, scotland, slovakia, and sweden were selected for spinraza and voretigene. characteristics of each country's process were extracted, including special reimbursement for rdts, special rdt committees, economic evaluation modifications, greater wtp, quality of evidence flexibility, additional considerations, etc. special and standard processes seemed to have different impacts on the appraisal of rdts. special processes more consistently managed rdt issues such as evidential uncertainty and higher icers. standard processes sometimes informally applied some of the characteristics included in special processes, such as broader consideration of value. conclusions: comparing case study country examples of rdt appraisal exemplified the complexity of these processes. special processes were more consistent in managing the challenges in rdt appraisal than standard processes. practical application: findings suggest a need for adapted approaches for rdt appraisal, to facilitate management of associated challenges and more consistent decision-making. estimating the broader fiscal impact of rare diseases using a public economic framework: a case study applied to acute hepatic porphyria (ahp) mark p. connolly , background: the aim of this study was to apply a public economic framework to evaluate a rare disease, acute hepatic porphyria (ahp) taking into consideration a broad range of costs that are relevant to government in relation to social benefit payments and taxes paid by people with ahp. ahp is characterized by potentially life-threatening attacks and for many patients, chronic debilitating symptoms that negatively impact daily functioning and quality of life. the symptoms of ahp prevent many individuals from working and achieving lifetime work averages. we model the fiscal consequences for government based on reduced lifetime taxes paid and benefits payments for a person diagnosed aged experiencing attacks per year. materials & methods: a public economic framework was developed exploring lifetime costs for government attributed to an individual with ahp in sweden. work-activity and lifetime direct taxes paid, indirect consumption taxes and requirements for public benefits were estimated based on established clinical pathways for ahp and compared to the general population (gp). results: lifetime earnings are reduced in an individual with ahp by sek . million compared to the gp. this also translates to reduced lifetime taxes paid of sek . million for an ahp individual compared to the gp. we estimate increased lifetime disability benefits support of sek . million for an ahp individual compared to gp. we estimate average lifetime healthcare costs for ahp individual of sek . million compared to gp of sek . million. these estimates do not include other societal costs such as impact on caregiver costs. conclusions: due to severe disability during the period of constant attacks, public costs from disability are significant in the ahp patient. lifetime taxes paid are reduced as these attacks occur during peak earning and working years. the cross-sectorial public economic analysis is useful for illustrating the broader government consequences attributed to health conditions. ethics approval: the study results described here are based on a modeling study. no data on human subjects has been collected in relation to this research. the european cystic fibrosis (cf) society patient registry collects demographic and clinical data from consenting people with cf in europe. the registry's database contains data of over , patients from countries. high quality data is essential for use in annual reports, epidemiological research and postauthorisation studies. methods: a validation programme was introduced to quantify consistency and accuracy of data-input at source level and verify that the informed consent, required to include data in the registry, has been obtained in accordance with local and european legislation. accuracy is defined as the proportion of values in the software matching the medical record, consistency as definitions used by the centre matching those defined and required by the registry. data fields to verify: demographic, diagnostic, transplantation, anthropometric and lung function measurement, bacterial infections, medications and complications. the number of countries to validate: % of the total countries/year. in the selected country ≥ % of the centres should be visited and - % of the data validated. results: in , ten out of centres ( %) in countries (austria, portugal, slovakia, switzerland) with ≥ % of all patients in their countries were selected. in a day visit, the data of the registry were compared with the medical records, the outcomes and recommendations discussed, and a final report provided. demographic, diagnostic and transplant data were checked for patients ( %*), clinical data for patients ( %*) ( data). challenges were: informed consent, mutation information (genetic laboratory report missing), definitions interpretations. see fig. for the results. conclusion: the registry's data is highly accurate for most data verified. the validation visits proved to be essential to optimise data quality at source, raise awareness of the importance of correct informed consent and encourage dialogue to gain insight in how procedures, software, and support can be improved. *of the total patients in these countries. background: people with duchenne muscular dystrophy (dmd) adopt compensatory movement patterns to maintain independence as muscles get weaker. the duchenne video assessment (dva) tool provides a standardized way to document and assess quality of movement. caregivers video record patients doing specific movement tasks at home using a secure mobile application. physical therapists (pts) score the videos using scorecards with prespecified compensatory movement criteria. objective: to gather expert input on compensatory criteria indicative of clinically meaningful change in disease to include in scorecards for movement tasks. approach: we conducted rounds of a delphi panel, a method for building consensus among experts. we recruited pts who have evaluated ≥ dmd patients in clinic and participated in ≥ dmd clinical trials. in round , pts completed a preliminary questionnaire to evaluate compensatory criteria clarity and rate videos of dmd patients performing each movement task using scorecards. in round , pts participated in an in-person discussion to reach consensus (≥ % agreement) on all compensatory criteria with disagreement or scoring discrepancies during round . results: of the pts, % practiced physical therapy for ≥ years, % provided physical therapy to ≥ dmd patients, and % participated in ≥ dmd clinical trials. of version compensatory criteria, ( %) were revised in round . of version compensatory criteria, ( %) were revised in round . the pts reached % agreement on all changes made to scorecards during the in-person discussion except the run scorecard due to time restrictions. a subset of the panel ( pts) met after the in-person discussion and reached consensus on compensatory criteria to include in the run scorecard. conclusion: expert dmd pts confirmed that the compensatory criteria included in the dva scorecards were appropriate and indicative of clinically meaningful change in the disease. introduction: fifty percent of rare disease cases occur in childhood. despite this significant proportion of incidence, only % of adult medicines authorised by the european medicines agency (ema) completed paediatric trials [ , ] . as a result, many clinical needs are left unmet. various factors compound the development of treatments for paediatric rare diseases, including the need for new clinical outcome assessments (coas), as conventional endpoints such as the minute walking test ( mwt) have been shown to not be applicable in all paediatric age subsets, [ ] and therefore may not be useful in elucidating patient capabilities. coas are a well-defined and reliable assessment of concepts of interest, which can be used in adequate, well-controlled studies in a specified context. coas capture patient functionality and can be deployed through the use of wearable sensor technology; this feasibility study presents data obtained from patients with paediatric rare diseases who were assessed with this type of technology. methods: niemann pick-c (np-c) (n = ) and duchenne muscular dystrophy (dmd) (n = ) patients were asked to wear a wrist-worn wearable sensor at home for a minimum of weeks. feasibility was assessed qualitatively and quantitatively, with data captured in minute epochs, measuring the mean of epoch's with the most steps over a month (adm), average daily steps (ads), average steps per minute epoch (ade) (table ) and reasons for non-adherence (table ) . no restriction in the minimum number of epochs available for analysis were applied, and all patient data analysed. results: discrepancies in ambulatory capacity were observed between np-c and dmd patients overall, with np-c patients covering greater distances and taking more steps daily. qualitative assessment of both patient groups highlighted their relationships with the technology, which in turn detailed adherence. some patients exhibited behavioural issues which resulted in a loss of data and low engagement. conclusions: the wearable sensor technology was able to capture the ambulatory capacity for np-c/dmd patients. insights into disease specific parameters that differed were gained, which will be used for developing the technology further for use in future trials. additional work is required to correlate the wearable device data with other clinical markers, however the study displays the feasibility of wearable sensors/apps as potential outcome measures in clinical trials. background: neonatal surgery is decentralized in germany. in there were departments of pediatric surgery that treated % of the abdominal wall defects with an average case load of less than per unit [ ] . patient organizations stress the importance of quality measurements for the care of children with rare diseases. study plan: currently, there is no nationwide data collection regarding the short term and long term care of patients with congenital malformations, who often need surgery during the first weeks of life. the german society of pediatric surgery, which covers almost all of the german pediatric surgical units, has initiated the work of creating a national patient registry (kirafe) for the following congenital malformations: malformations of the gastrointestinal tract, the abdominal wall, the diaphragm, and meningomyelocele. the development of the registry involves three different patient organizations and health care professionals from all over germany. the registry will be set up in based on the open source registry system for rare diseases (osse). the primary objective of the registry is the measurement of quality attributes of rare congenital malformations. furthermore, the registry will facilitate recruitment of patients to clinical trials. it will also serve as a basis for policy making and planning of health and social services for people with rare disorders. informed consent will be obtained from the participants. the registry will include core data, mainly comprising information on the set of malformations of each patient. each malformation will then prompt further different modules for data collection. this modular structure offers the greatest possible flexibility for the documentation of patients with more than one congenital malformation. data will be collected by health care professionals. results: since the start of the preparation individuals, either working in one of hospitals or being member of one of the three patient organizations, have contributed in the ongoing activities. the registry is listed in the european directory of registries (erdri) [ ] . ethical approval was obtained, financial resources were secured. in , german hospitals and three non-german hospitals confirmed their intention to document their patients within the registry. conclusion: the registry is an example for a nationwide collaboration with the goal to optimize the quality of care for a patient group with rare diseases. is a collaboration between cf europe and five pharmaceutical companies (to date). through biannual meetings, we aim to institute a longterm educational collaboration with companies with an interest in cf. membership of industrial partners is dependent upon adherence to the cfrtoc code of conduct and a financial contribution for cf europe to fulfil its missions. common objectives include access to information. one strong example, applicable even beyond rare diseases, is the need for improved communication regarding clinical trials (cts) which has been inconsistent and often difficult to understand. from , the new european ct regulation / will oblige sponsors to share ct results through lay summaries. to help move this initiative forward, cf europe, with the active support of the cystic fibrosis trust, is collaborating with the european cystic fibrosis society-clinical trials network (ecfs-ctn) and cfrtoc members to establish a glossary of relevant cf terms. it will be freely available so that all stakeholders can systematically use it in patient-friendly scientific summaries and wider communication. in a pilot project, people with cf and patient associations, together with industrial partners will shortlist terms. these will be defined by lay members and subsequently subjected to the study and approval of the legal department of participating companies. provided this process is successful, we aim to create approved definitions by the end of . cf europe and ecfs-ctn intend to advertise the use of this glossary online and through communications at scientific events. national patient organisations will be further encouraged to provide translations in their national language. alkaptonuria (aku, ochronosis) is an inborn metabolic disease, resulting in the accumulation of the metabolic intermediate homogentisic acid (hga). oxidation of hga by air or within connective tissue causes darkening of the urine, pigmentation of eyes and ears, kidney-and prostrate-stones, aortic stenosis, but most severely an early onset of arthritis called ochronotic arthropathy (ochronosis) due to deposition in the cartilage. ochronosis is very painful, disabling and progresses rapidly. starting in the thirties with the spine and affecting large joints in the forties, patients frequently require joint replacements in their fifties and sixties [ , ] . like many of the rare diseases, aku-patients undergo a long odyssey of several years until their diagnosis. the german aku-society "deutschsprachige selbsthilfegruppe für alkaptonurie (dsaku) e.v. " was founded in and became subsequently registered as a non-profit patient organization. first of all, the dsaku identified aku-patients, set up a homepage [ ] and designed flyers with information for patients, their families, medical professionals and healthcare services. second, it offered workshops on aku-related issues and enabled personal exchange. third, it raised awareness of aku, both nationally and internationally by information booths, presentations and posters at scientific congresses as well as rare disease days (rdd). fourth, in response to the needs of patients, it established collaborations and built up national networks for a better health care accordingly. thus, patients were encouraged to visit the centers for metabolic diseases at the charité (berlin), hannover medical school (mhh), university of düsseldorf and institute of human genetics at the university of würzburg to bundle knowledge and expertise. the dsaku is member of achse e.v., nakos, eurordis and metabern and registered in the databases se-atlas, zipse and orphanet. finally, the dsaku is nationally and internationally active in health politics regarding training in drug safety and evidence-based medicine. introduction: autoinflammatory diseases are rare conditions characterized by recurrent episodes of inflammation with fever associated to elevation of acute phase reactants and symptoms affecting mainly the mucocutaneous, musculoskeletal or gastrointestinal system. these diseases affect the quality of life of patients and their families. objectives: aim of this project is to develop a tool able to ameliorate patients' management of the disease and to enhance patientphysician communication. to develop a tool based on real-life needs, we involved patients and caregivers since the initial phase of the project. a first workshop designed to capture their needs was organized. innovative co-design activities were performed through "legoserious-play ™ " (lsp) methodology [ ] [ ] [ ] . during a first phase of "divergence" patients (from teen-agers to adults) affected by different aids (fmf, trap, caps, mkd) and physicians where involved in the lsp activities. participants were asked to describe, through lego and metaphors: • the disease • themselves in comparison with the disease • solutions and supports which could help them in managing the disease after each step the participants presented their models, and everyone was engaged in the discussion. the ideas collected during the three phases allowed to make a list of functionalities identified as necessary for the app to be developed. due to the actual sars-cov- sanitary emergency the second phase of the project, aimed at presenting the participants the results of the first meeting and proceed with the app finalization was performed through web-based meeting and surveys in which the patients and caregivers actively participated. results: in the first phase patients and caregivers participated actively expressing various needs, that we subsequently summarized in main areas (table ) . participants were then further involved and their opinion taken into consideration for the user experience and interface definition for the development of the mobile app including the required functionalities (after a further activity of prioritization). introduction: gaps in communication and education are becoming one of the biggest key pain points for patients that are suffering rare diseases. due to the limited resources and the misleading information on the internet we wanted to test the poc systems to deliver more efficiently the information to our patients and their relatives. userfriendly information at the point of care should be well structured, rapidly accessible, and comprehensive. method: we implemented a specific poc channel using several touchpoints to deliver the right content at the right time. we created and selected the video content that will be most helpful to our patients. later on, we analyzed the patient journey and we decided to use a mobile app where the patients could search for information when they are at their home. at the medical practice, we use the waiting room and exam room as learning areas through monitors and tablets. moreover, healthcare professionals are prescribing content to their patients that they reviewed when they are home. results: thanks to the use of the poc channel and technologies related we were able to reduce the time needed to perform an explanation by %. furthermore, our healthcare professionals reported that their conversations with the patients improved % and patient satisfaction increased by %. conclusion: poc channel created a positive impact on our patient experience allowing us to be more efficient delivering the information to our patients and their relatives. [ , ] , realworld safety and efficacy data are limited -particularly for patients who receive > treatment. we report initial data from the restore registry, including cohort clinical characteristics, treatments received, and outcomes. materials and methods: restore is a prospective, multicenter, treatment-agnostic registry of sma patients. the primary objectives include assessment of contemporary sma treatments; secondary objectives include assessment of healthcare resource utilization, caregiver burden, and changes in patient functional independence over time. planned follow-up is years from enrollment. as of january , data were available for patients, all from de novo clinical sites in the united states; information on treatment regimens was available for patients (table ) . disease-modifying treatments were administered sequentially or in combination. % of treated patients showed symptoms at sma diagnosis, with the most common being hypotonia and limb weakness ( table ) . Ågrenska, a swedish national centre for rare diseases, has for thirty years arranged courses for families of children with rare diagnoses and has experienced that the conditions often have complex and varying consequences in the children ś everyday lives. knowledge of these consequences and of how to adapt the treatment, environment and activities to create the best possible conditions for participation and learning, is often lacking. many professionals also report lack of sources of knowledge. knowledge formation and dissemination are thus of outmost importance. in order to aid knowledge formation and dissemination Ågrenska has developed an observation instrument for children with rare diagnoses, identifying both abilities and difficulties on a group level. the instrument consists of quantitative and qualitative items and covers ten areas: social/communicative ability, emotions and behaviours, communication and language, ability to manage his/her disability and everyday life, activities of daily life, gross and fine motor skills, perception and worldview, prerequisites for learning and basic school abilities. observations are made during the children ś school and pre-school activities during the Ågrenska course. teachers and special educators, working with the children, are responsible observers. some school-related abilities are difficult to observe during the five-day stay. this information is instead collected through a telephone interview with the children ś home teacher. the instrument was content validated against a number of existing instruments. the items were considered relevant as they, with few exceptions, appear in well-known assessment tools. to test interrater reliability observations of six children were performed. each child was observed by two educators. interrater reliability was calculated for the quantitative items usually observed during the course. interrater reliability reached . %. background: sma is a neurodegenerative disease caused by survival motor neuron gene (smn ) deletion or mutation [ , ] . disease severity (sma type) correlates with smn copy number [ , ] . gene therapy with onasemnogene abeparvovec provides sustained, continuous production of smn protein, and is fda approved [ ] , with ongoing trials for sma type (sma ) and sma , and presymptomatic treatment for all sma types. with treatment options available, many states in the united states (us) are implementing newborn screening (nbs) to detect smn deletions and smn copies, providing early diagnosis and the option of pre-symptomatic treatment [ ] . we examine the economic consequences of implementing nbs for sma and pre-symptomatic treatment with onasemnogene abeparvovec gene therapy among newborns in the us. a decision-analytic model was built to assess the cost effectiveness of nbs in , hypothetical newborns from a us third-party payer perspective. the model included separate arms, each allowing for a different treatment strategy. model inputs for epidemiology, test characteristics, and screening and treatment costs were based on publicly available literature (table ) . inputs and assumptions of lifetime costs and utilities for sma types were obtained from the institute for clinical and economic review sma report [ ] ; other values were sourced from published literature. model outputs included total costs, quality-adjusted life years (qalys), and incremental cost-effectiveness ratios (icers). scenario and sensitivity analyses tested model robustness. park's programme, particularly across education and engagement and prioritisation and development of research. in addition to representation on governance structures, wales gene park (wgp) collaborates with patients and the public to involve them in rare disease and genetic research. wgp has co-produced a rare disease research gateway following consultation with patients and the public from its networks. the gateway hosts relevant studies in genetic and rare disease research on the wgp website. it promotes involvement opportunities in addition to signposting to studies that patients and other members of the public can participate in. it also links to training opportunities for ppi representatives. consultation with patients and the public regarding the usability, design and development of the gateway was undertaken. feedback has enhanced the user experience and it was launched in october . there are currently over studies featured, and the gateway is searchable according to condition or key word. impact will be monitored through online usage and website analytics. engagement with researchers through a professional network enables opportunities to be advertised from all areas of genetic and rare disease research and ensures that patient and public representatives are involved in the design and development of research from its inception. wgp were invited to present at the welsh health and care research wales conference in as the gateway was highlighted as an exemplar of good practice. specialist visit, medications) and non-medical resource use (lost productivity and homecare or caregiver's time). outcomes of interest for treatment options assessed the efficacy and safety of treatments for rett syndrome. results: the search on economic burden yielded articles; intervention type and costs were extracted from , representing studies. in the economic burden studies, enteral feeding and assisted walking increased the risk of respiratory-related hospital admissions, while length-of-stay was lower in younger patients. mean recovery-stay after scoliosis-correcting surgery was . days and . days in each of studies. care integration improved outcomes and reduced costs. the search on clinical trials yielded articles; efficacy and safety were extracted from , representing studies ( randomized controlled trials, single-arm; n = - ; follow-up - months). of these, focused on pharmacological symptom treatment; examined environmental enrichment effects; none targeted the underlying cause. the most common primary endpoints are stated in table . naltrexone, trofinetide, and mecasermin demonstrated clinical benefits versus placebo, but most treatments yielded no significant improvement ( table ) . the cml advocates network (cml an) is an active network specifically for leaders of chronic myeloid leukemia (cml) patient groups, connecting patient organisations in countries on all continents. it was set-up and is run by cml patients and carers. its aim is to facilitate and support best practice sharing among patient advocates across the world. the cml community advisory board (cml-cab) is a working group of the cml advocates network. since its inception the cml-cab has met on nineteen occasions with five sponsors. the cml-cab is comprised of two chairs and cab-members. cml-cab organisation, sustainability and follow-up is supported by a part-time cml-cab officer and the cml-an executive director. the principles of leaving no one behind are essential to the goals of world health organization (who) and united nations (un). in , an ambitious objective to ensure that billion more people will benefit from universal health coverage (uhc) until was entrenched in the who th general programme of work [ ] . all un member states have agreed to try to achieve universal health coverage by , as part of the sustainable development goals [ ] . however, it is essential, that rare disease (rd) patients are not left behind on our trip to uhc. in , un declared that rd are among the most vulnerable groups that are still on the fringes of uhc [ ] . the first step on a way to the full uhc cube [ ] for rd is an identification of root causes of health inequities. health determinants of rd fundamentally differ from those for common diseases. some of them are unavoidable: up to % of rd have a genetic basis (individual or genetic determinants). although socio-economic factors are highly important, in contrast to common diseases, they are a consequence rather than a cause of rd. meanwhile, one of the major root cause amenable to change are health system determinants: organization of services for rd requires unique solutions in our health systems that are mostly adapted for common diseases. political and legal determinants also play a key role: while rd is an explicit example of an area, loaded orphanet j rare dis , (suppl ): with needs for pan-european solutions, relative "weakness" of eu legal powers to regulate and have an impact on implementation of pan-european policies in health results in vast inequities among and inside member states and lack of engagement at a national level. health activism that includes strong advocacy and a loud voice of patient organizations has also been ascribed to health determinants and may have a crucial role in rd [ ] to improve the situation, we already have some powerful tools at hand including national plans for rd, european reference networks [ ] and european joint programme on rare diseases [ ] . however, to reach the full potential of these, multiple obstacles have to be removed and full implementation ensured. since march , there has been an explosion in digital health adoption as people look for remote ways to manage their health and wellbeing. national government covid- strategies, local authorities and consumers, have all turned to health apps, both as a potential means of slowing the spread of the virus, and a method of allowing people to self-manage their own health. in the first few weeks of the covid- pandemic, orcha worked with app developers to build a dedicated covid- app library full of evaluated apps. free to use for all, it included relevant, quality assured apps that had been through orcha's rigorous review process. to build such a tool in such a short space of time is testament to the speed of this market. more consumers have been using health and care apps. in just one week, orcha saw an increase of . % in app downloads from its app libraries, and a , % increase in app recommendations from health and care professionals. orcha can see from the data across its app libraries that the most popular search terms since the pandemic began have included: mental health, physiotherapy, fitness, anxiety, rehabilitation, diabetes, respiratory, and sleep. whereas 'covid' was initially the most searched term at the beginning of the outbreak, people have since searched for specific condition areas. this indicates a shift in focus to actively self-managing health and wellbeing, and a desire for knowledge about particular health areas. the recent increase in digital health adoption has highlighted that the challenge remains of helping consumers to understand which apps are potentially unsafe to use, and ensuring that consumers are armed with the full facts about the strengths and weaknesses of an app, before it is downloaded. while considerable progresses have been made in the last years in research on innovative medicinal products for adults, children have not benefited from progresses to the same extent as adults in terms of appropriate treatments and advanced tools. it is well known that the availability of drugs for paediatric use still represents a challenging issue, since research and development in this field is characterized by many that range from methodological, ethical and economic reasons, especially when neonates and rare diseases are involved. moreover, even when industry has the capacity to perform a paediatric drug development plan, there are many economic reasons limiting the commercial sponsors' interest (the paediatric population is a small population; paediatric diseases often concern rare disorders with unknown mechanism; it is very difficult to perform preclinical and clinical studies; ethical concerns are still relevant and additional regulatory requirements have to be considered). in this scenario, eptri can make the different in closing the gap between innovative technologies and paediatric drug development processes. it is a eu-funded project that arises from the need to find answers to the serious lack of medicines for children in eu and worldwide, and aimed to design the framework for a european paediatric translational research infrastructure dedicated to paediatric research. an high interest is tailored on rare diseases (rd) as they affect mainly children and genetic rd start early in the prenatal/childhood life with an high frequent use of medicines not specifically tested (off-label, unlicensed). eptri will work to accelerate the paediatric drug development processes from medicines discovery, biomarkers identification and preclinical research to developmental pharmacology, age tailored formulations and medical devices. this will allow is to facilitate the translation of the acquired new knowledge and scientific innovation into paediatric clinical studies phases and medical use. neonatal screening started in many countries around - after phenylketonuria turned out to be a treatable condition. if diagnosed early, a diet could help to avoid impaired brain development. public health programmes were developed to offer all newborn children the possibility to be tested. screening always has benefits and disadvantages, and only rarely pros outweigh cons at reasonable costs. the world health organization in published criteria to evaluate benefits and disadvantages, concerning amongst others ( ) important health problem ( ) treatment ( ) suitable test and ( ) appropriate use of resources. pku was mentioned as an example of an important health problem [ ] . neonatal screening is more than a test. information to parents, communication of results, ict infrastructure, follow-up of affected infants, reimbursement of test and treatment and governance all need adequate attention [ ] . around the number of diseases covered in european countries in neonatal screening programs was very diverse: from zero in albania to more than in austria, hungary, iceland, portugal and spain [ ] . many countries have seen an increase in the number of diseases covered because of new tests and treatments becoming available. health authorities were almost always involved in changes in the programmes, hta experts and parents organizations sometimes. half of the countries had laws on nbd, and half had a body overseeing nbs programs. less than half of the countries informed parents of the storage of dried blood spots [ ] . after the eu initiated "tender nbs" had provided advice to eu policy makers [ ] , little initiatives for harmonization were taken, because health is the mandate of member states. from the perspective of newborns this implies that early diagnosis and adequate treatment for nbs conditions may differ very much for children being born in one or another eu country. with more tests and more treatments becoming available, this makes it even more urgent to attune the perspectives of different eu stakeholders for the benefit of all newborns. background: as genome sequencing is rapidly moving from research to clinical practice, evidence is needed to understand the experience of patients with rare diseases and their families. in the presentation, we discuss families' experience of receiving, making sense of and living with genomic information. the presentation includes video-clips from two short films from families' narratives. specifically, families struggled with the lack of information on the course of the disease, the difficulties to access support and navigate health and social care services, and the challenges related to making sense of the implications of genomic information for other family members. despite these issues, families identified a wide range of benefits from taking part in genome sequencing, which were broader than the clinical utility of the diagnosis. the findings raise questions regarding how to talk about 'diagnosis' in a way that reflects families' experience, including their uncertainty but also their perceived benefits. they also have implications for the design and delivery of health services in the genomic era, pointing to the need to better support families after their search for a diagnosis. saluscoop [http://www.salus coop.org] is a non-profit data cooperative for health research that aims to make a greater amount and diversity of data available to a broader set of health researchers, and to help citizens to manage their data for the common good. data heals. health research is data-driven: the larger the universe, the greater the quantity, quality, and diversity of the data, the more potential the data has to cure. in our european context, it is clear: data belongs citizen. gdpr regulates ownership and our rights over data that include portability. data protection laws rightly consider that health data deserves the maximum protection. however, the only truth, we note every day: in practice citizen often cannot access their data or control its use. the future of our health depends significantly on the ability to combine, integrate and share personal health data from different sources. the only one who can integrate all your information (public, private, clinical, personal, habits, genetics) is the citizen himself. using data well, it is possible to obtain more and better health for all. we are a cooperative that works to facilitate the transformation process towards this goals doing: -dissemination, awareness, communication -studies, manifestos. -licenses to facilitate it -salus common good license - it is necessary to dissociate the provision of services, of the possession of the data. the accumulation and centralization of the data is not necessary. blockchain and the like allow the certification of transactions without the need for intermediaries. the need for the existence of new social institutions for the collective management of data for the common good is much clearer today: so that these citizens have the technological and legal tools effectively manage their data. so that health research can address the real problems of our societies. the abstract is being presented on behalf of a saluscoop management board group. the region of murcia, located on the southeast of spain, has . million inhabitants. in , approximately % of its population was identified with a rd, based on the regional rd information system, which showed a public health problem requiring an integral and coordinated approach. results: in , after years of participative work (interdepartmental government representatives, patients associations and professionals) the regional plan for rd integrated (holistic) care was approved, for a period of years ( - ) and a budget of millions euros; with the goal of improving health, education and social care through interdisciplinary coordination and placing patients and families in the center of the actions. the plan includes ten different strategic areas related to information, prevention and early detection, healthcare, therapeutic resources, social-health care, social services, education, training of professionals, research, monitoring and evaluation a regional rd coordination center, linked to the medical genetics unit in the tertiary reference hospital, is connected to the health areas, educational and social local services, through a case manager integrated in the multidisciplinary team. this was our building experience presented in the innovcare project, co-funded by the eu. to design a holistic care plan for rd we need to know the prevalence based on rd registries, available and needed resources and an interdisciplinary participative action approach with the appropriate government and financial support with periodic evaluation. case management has an important role. the recognition of clinical genetics as health specialty is also urgent in spain to provide equal access to rd patients and families all over the country. [ ] . these policies have served us well, but it is essential that the policies guiding us towards the future we wish to see are equipped to address the needs of the future rd population. the rare project [ ] is working towards precisely this goal, and has identified over a hundred future-facing trends likely to impact on the field. some of these trends concern demographic changes about which we can be reasonably certain: whilst overwhelmingly positive, changes such as ageing rd populations will bring new challenges in managing comorbidities. they will also create new opportunities as well as risks in areas such as reproductive choice; however, these choices incur major ethical, legal and social concerns, and it is unclear how many countries really have robust frameworks in place to cope with this. besides the fairly certain demographic changes, there are many topics -and many needs-for which the future is not clear. will there be easier access to expert multidisciplinary teams? what will be the role of technology in care delivery? these fundamental issues are here debated in interview format [ ] . adrenoleukodystrophy, or ald, is a complex x-linked genetic brain disorder which mainly affects males between the ages of four and -males who are previously perfectly healthy and 'normal' . ald damages the myelin in the brain and spinal cord, and those with cerebral symptoms become completely dependent on their loved ones or carers. this usually involves patients becoming wheelchair or bed bound, blind, unable to speak or communicate and tube fed. it is a difficult disorder to diagnose with behaviour problems usually the primary indicator. in males, cerebral ald is a terminal illness with most dying within one to years of symptoms developing. if diagnosed before symptoms become apparent, usually through identification of a family member, the condition can be successfully treated through bone marrow transplant. some adults (males and females) develop a related condition called adrenomyeloneuropathy, or amn. symptoms include difficulty walking, bladder and bowel incontinence and sexual dysfunction. tragically, around one third of males with amn go on to develop cerebral ald. initial behavioural symptoms often have an impact on the individual's professional and personal lives -their capacity to work, maintain relationships and family ties -over time, they can become isolated and socially unacceptable. commonly, those individuals without supportive family structures are missed or misdiagnosed. the presentation presents a personal case study detailing the impact of an ald diagnosis on the whole family, moving on to alex tlc's experience in applying to add ald to the uk's new born screening programme. the conclusion includes next steps following an initial negative response, and thoughts on the methods used to assess decisions on the prevention and treatment of rare disease. the rare foresight study gathers the input of a large group key opinion leaders through an iterative process to propose recommendations for a new policy framework for people living with rare diseases (rd) in europe. since the adoption of the council recommendation on european action in the field of rd in , the european union has fostered tremendous progress in improving the lives of people living with rd. rare will recommendations for the next ten years and beyond. the rare foresight study includes major stages (fig. ) . the european conference on rare diseases and orphan products (ecrd ) marked the occasion to present four proposed future scenarios (fig. the market-led approach first creates the technology innovation, then seeks out its market. deep understanding of needs as the starting point of the innovation process. with symptoms and being suspected of having a rare disease can be the longest in many steps to getting a diagnosis. this is something we have the power to change now by providing content tailored to medics, early in their careers that will equip them to #daretothinkrare. to prepare for delivery of gene therapies, companies typically focus on four key areas: patient identification & diagnostics; treatment centre qualification; manufacturing & supply and market access. timely diagnosis of patients is important as with progressive disorders, the earlier patients are treated, typically the better their long-term clinical outcomes will be. targeted tools and resources are used to educate clinical specialists on the early symptoms of the disease. improving access to the appropriate diagnostic tests is essential. if newborn screening is considered, validated assays and pilot studies are required. gene therapies have to be administered in qualified treatment centres. after regulatory approval, treatment centres are relatively few so patients may need to cross borders and work is required to expand the recognition of patient rights to be treated in another eu country (e.g. through the s mechanism). many companies partner with contract manufacturing organisations and are developing ways to preserve gene-corrected stem cells to enable their transportation from the manufacturing site to treatment centres. the final area is market access, whereby it is vital to evolve the way healthcare systems think about delivery, funding and value determination. manufacturers have the responsibility to generate health economic evidence. recent research [ ] in metachromatic leukodystrophy showed that caregivers (n = ) spend an average of hours a day caring for their child. % of parents were forced to miss work with % of this being unpaid leave. in addition, it is recommended to have the optionality of payment models that allow the sharing of risk between the healthcare system and manufacturer (e.g. annuity or outcomes-based payments). orchard has developed a holistic value framework as gene therapies are expected to benefit patients, families, communities, healthcare systems and society reference background: employment has always been one of the fundamental human rights. it is important for people with rare diseases, because it helps to stay connected to the community and to continue professional development. equal access to job employment can help to overcome the consequences of the condition and to gain financial independence. on the other hand unemployment can increase the social exclusion. in the last few years there is an improvement in the european policies about job employment. in spite of this, people with rare diseases still have to overcome discrimination in this field. as a proof of this statement is the recent online survey, conducted by eurordis. according to it, % of the respondents admitted they had to reduce their professional activities after they were diagnosed with rare condition. this means that more than half of the people with rare diseases in europe face employment challenges. the analysis of this survey was important input to the presentation of the epf youth group project -ways. results: this is the abbreviation of work and youth strategy and it is a two year project, disseminated among young patients with chronic conditions. the main purpose was to increase the awareness about positive and negative practices for young patients on the labour market and to develop recommendations to employers and decision makers. that is why epf youth group conducted an online survey and provided different deliverables like factsheet with recommendations to employers and video about young patients' rights on the work place. the results of both survey provided important insight about the challenges people with rare diseases face in job employment. it proved the fact that only if we work together as a community of patients, we will be able to provide better opportunities for national and international inclusion. paul rieger , eberhard scheuer centiva health ag, zug, switzerland correspondence: paul rieger -paul@centiva.health orphanet journal of rare diseases , (suppl ):s the lack of access to research participants is the number one reason why medical studies fail [ ] . real-world data is often difficult to get despite usd billion costs of patient data intermediation. therefore, a new model for patient access is necessary where patients get paid fairly for their data, retain control over their data, and drive citizencentered research. on the other hand, researchers and industry must be enabled to access patients directly without violating their privacy, while reducing time and costs of data access at the same time. current patient registries facilitate patient access and match patients with a centralized data flow while giving little to no incentives. whereas, a decentralized patient registry allows for direct and confidential matchmaking between patients and organizations looking for data through the use of blockchain technology. it lets the patient decide with whom they want to share their data. on such a platform, patients can receive incentives in the form of digital currency. currently, centiva health [ ] is used in the context of rare diseases and population health, i.e., outbreak monitoring. in the area of rare diseases centiva health cooperates with patient advocacy groups by enhancing existing registries with the ability to collect real-world data. the access to patient via a decentralized registry leads to aligned incentives, real-time access to data, improved disease visibility while preserving patient privacy. orphanet j rare dis , (suppl ): the united kingdom and in the czech republic, to co-design optimal methods/services for the communication of genomic results. methods and results: using a methodology called experience-based co-design (ebcd) , we supported families and health professionals to shared and discuss their experiences, identify priories for improvement and then work together to prototype and test out interventions to address these. the process involved observations of clinical appointments (), interviews with families () and health professionals and a series of workshops and remote consultations at both sites. results: five shared priorities for improvement were identified by participants at the two sites, and eight quality improvement interventions were prototyped/tested to address these ( table ) . discussion: the findings clearly indicate the need for improved follow-up care to support families in the short, medium term after the sharing of the results, including when a diagnosis is confirmed. different service models were prototyped, including follow up consultations with clinical geneticists and a dedicated role to facilitate co-ordinated care. the findings also demonstrate the need for continued workforce development on the psychosocial aspects of genomic and genetic communication, specifically on families' needs regarding genomic consent and the experience of guilt and (self-) blame. to use technology has been used in the home to provide objective seizure data prior to upcoming clinic appointments. the covid- pandemic has prompted an acceleration in telemedicine and epihunter has improved the effectiveness of virtual consultations bringing opportunities for both diagnostics and informed changes in treatment. epihunter is an example of technology repurposing to create a new normal for people with hidden disabilities such as those living with absence epilepsy. the rare disease patient community tried to get this well detailed plan to be transferred to regulation which usually means an adequate financial substitution of those expert services. the patients should benefit from a centralized expert treatment/care pathway. esophageal atresia (ea) is a rare congenital condition with an estimated prevalence of to in , live births. esophageal atresia patients require life-long attention. ernica has developed a 'patient journey' for ea patients, under the leadership of patient representatives from the international federation of ea support groups (eat). in germany, patients with congenital malformations which need surgery in early life are treated in hospitals with (very) low experience. how can we as patient representatives get the fruits of the erns into the national health system? we don't have public money. we have no official contract and no political support. keks e.v., the german ea support group together with other support groups (e.g. soma e.v.), and with surgical expert teams across germany, some of them members in erns, started to organize monthly virtual boards for those patients. a self-commitment on ethical and medical standards following the ern-criteria, and a collaborative attitude within the group, help us to get step by step the first ernica results to the bedside of ea patients. springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. delays in completion and results reporting of clinical trials under the paediatric regulation in the european union: a cohort study new strategies for the conduct of clinical trials in pediatric pulmonary arterial hypertension: outcome of a multistakeholder meeting with held at the european medicines agency on monday decentralized rather than centralized pediatric surgery care in germany erdri.dor -european directory of registries natural history of alkaptonuria recent advances in management of alkaptonuria when you build in the world, you build in your mind white paper on lego serious play articulation of tacit and complex knowledge zolgensma (onasemnogene abeparvovec-xioi) childhood spinal muscular atrophy: controversies and challenges spinal muscular atrophy bannockburn, il indirect estimation of the prevalence of spinal muscular atrophy type i, ii, and iii in the united states pilot study of populationbased newborn screening for spinal muscular atrophy in new york state presymptomatic diagnosis of spinal muscular atrophy through newborn screening one year of newborn screening for sma -results of a german pilot project correlation between sma type and smn copy number revisited: an analysis of unrelated spanish patients and a compilation of reported cases available from: www.ibm.com/produ cts/micro medex -red-book available from: www.ibm.com/produ cts/micro medex -red-book references . united kingdom national health service international rett syndrome foundation presented at aacap's th annual meeting placebo-controlled crossover assessment of mecasermin for the treatment of rett syndrome cerebrolysin therapy in rett syndrome: clinical and eeg mapping study effects of acetyl-l-carnitine on cardiac dysautonomia in rett syndrome: prevention of sudden death? rett syndrome: controlled study of an oral opiate antagonist, naltrexone pharmacologic treatment of rett syndrome with glatiramer acetate safety, pharmacokinetics, and preliminary assessment of efficacy of mecasermin (recombinant human igf- ) for the treatment of rett syndrome effects of ω- pufas supplementation on myocardial function and oxidative stress markers in typical rett syndrome thirteenth general programme of work - . promote health -keep the world safe -serve the vulnerable principles and practice of screening for disease newborn screening programmes in europe; arguments and efforts regarding harmonization. part -from screening laboratory results to treatment, follow-up and quality assurance newborn screening programmes in europe; arguments and efforts regarding harmonization. part -from blood spot to screening result a framework to start the debate on neonatal screening policies in the eu: an expert opinion document communication from the commission to the european parliament, the council, the european economic and social committee and the committee of the regions on rare diseases: europe's challenges on an action in the field of rare diseases on the application of patients' rights in cross-border healthcare available from: https :// drive .googl e.com/file/d/ sfe xp deisc ogrbw swht uznx erj/ view?usp=shari ng . which scenarios are most preferred by the rd community? . which scenarios are most likely to happen? . how do we achieve the scenarios we prefer and avoid those we don factors associated with clinical trials that fail and opportunities for improving the likelihood of success: a review s background: to help inform cross-national development of genomic care pathways, we worked with families of patients with rare diseases and health professionals from two european genetic services bringing user experience to health care improvement: the concepts, methods and practices of experience-based design department of medical informatics correspondence: info-rdsgofair@go-fair.org (marco roos -m.roos@ lumc.nl, gülçin gümüş -gulcin.gumus@eurordis.org) in practice, it can take months of searching data, understanding the sources, mapping to consistent standards, and negotiating how one might use the data. many assume that for sharing and analysis, data need to be moved between sources. this can lead to sharing only minimal, non-sensitive data: a fraction of global rare disease data. alternatively, data elements and local access conditions can be described by globally agreed, computer understandable standards conform fair principles. this enables analysis at each source, while sharing only the analysis results. fair prepares data for rapid discovery, access, and analysis, also when data remain at source. projects such as the european joint programme for rare diseases work on the technical infrastructure to support this. adopting fair principles requires culture change. fair advocates working on rare diseases have organised the 'rare diseases global open fair implementation network rds go fair prioritizes patient representatives for their capacity to reshape current practices, welcoming them to organise their own network within rds go fair to foster fair for patient priorities (registration for follow-up meetings is possible via eucerd recommendations on quality criteria for centres of expertise for rare diseases in member states joint action rd-action (european union's health programme european commission website pdf s patient's view on disruptive innovations in clinical research elizabeth vroom we would like to thank the j-rare patient organization groups and the asrid research ethics committee. consent to publish: informed consent to publish has been obtained from patients. we thank all the families who took part in the interviews, the staff of the health services and charities who collaborated to advertise the study to eligible participants and the members of the family advisory groups who reviewed the interview schedule and provided invaluable feedback on the preliminary findings. the work has been presented on behalf of the study "improving the communication of genomic diagnosis results using experience based co-design (ebcd)", which is part of the solve rd project. the solve-rd project has received funding from the european union's horizon research and innovation programme under grant agreement no . we thank all the families who took part in the interviews, the staff of the health services and charities who collaborated to advertise the study to eligible participants and the members of the family advisory groups who reviewed the interview schedule and provided invaluable feedback on the preliminary findings. the work has been presented on behalf of the study "improving the communication of genomic diagnosis results using experience based co-design (ebcd)", which is part of the solve rd project. the solve-rd project has received funding from the european union's horizon research and innovation programme under grant agreement no . acknowledgements: we would like to thank all seed group members of the rare diseases global open fair implementation network, the go fair office, eurordis, the european union's horizon research and innovation program under the ejp rd cofund-ejp n° , the rd-connect community, the lumc biosemantics research group, simone louisse (guardheart epag), and the many patients and patient representatives that inspire us. recent advances in next-generation phenotyping (ngp) for syndromology, such as deepgestalt, have learned phenotype representations of multiple disorders by training on thousands of patient photos. however, many mendelian syndromes are still not represented by existing ngp tools, as only a handful of patients were diagnosed. moreover, the current architecture for syndrome classification, e.g., in deepgestalt, is trained "end-to-end", that is photos of molecularly confirmed cases are presented to the network and a node in the output layer, that will correspond to this syndrome, is maximized in its activity during training. this approach will not be applicable to any syndrome that was not part of the training set, and it cannot explain similarities among patients. therefore, we propose "gestaltmatch" as an extension of deepgestalt that utilizes the similarities among patients to identify syndromic patients by their facial gestalt to extend the coverage of ngp tools. methods: we compiled a dataset consisting of , patients with , different rare disorders. for each individual, a frontal photo and the molecularly confirmed diagnosis were available. we considered the deep convolutional neural network (dcnn) in deepgestalt as a composition of a feature encoder and a classifier. the last fully-connected layer in the feature encoder was taken as facial phenotypic descriptor (fpd). we trained the dcnn on the patients' frontal photos to optimize the fpd and to define a clinical face phenotype space (cfps). the similarities among each patient were quantified by cosine distance in cfps. results: patients with similar syndromic phenotypes were located in close proximity in the cfps. ranking syndromes by distance in cfps, we first showed that gestaltmatch provides a better generalization of syndromic features than a face recognition model that was only trained on healthy individuals. moreover, we achieved % top- accuracy in identifying rare mendelian diseases that were excluded from the training set. we further proved that the distinguishability of syndromic disorders does not correlate with its prevalence. conclusions: gestaltmatch enables matching novel phenotypes and thus complements related molecular approaches.an audience of over delegates voted on the rare scenarios and discussions throughout the sessions of ecrd indicated the following opinions:-if we continue as we are we will find ourselves in the "fast over fair" scenario which forecasts high collective responsibility but an emphasis on market-led innovation -the majority of the audience preferred a future scenario with continued high collective accountability but more of an emphasis on needs-led innovation, "investments for social justice" -a significant portion of the audience agreed that a balance must remain with the market led attractiveness of the "technology along will save you" scenario -a scenario where "it's up to you to get what you need" was least preferred by all the diagnostic pathway in rare disease has a number of bottlenecks that can result in the pathway becoming an odyssey. while some barriers are being removed through remarkable innovation, there is one story of diagnostic delay that is echoed by rare disease patients across the globe and across thousands of different rare diseases: doctors failed to suspect something rare. however we cannot expect doctors to suspect rare diseases when they haven't been trained to or, in some cases, have been trained to do the exact opposite with the mantra "common things are common". without appropriate training 'rare' can be mistaken for 'irrelevant' when in reality million european citizens live with a rare disease [ ] . medics rarediseases is driving an attitude change towards rare diseases in the medical profession. this begins with explaining that rare diseases are collectively common and all clinicians should expect to manage people with diagnosed and undiagnosed rare disease regularly during their careers. this attitude change is called #daretothinkrare. secondly m rd is suggesting a new approach to educating about rare disease for trainers and training institutes. this approach tackles rare disease as a collective and focuses on patient needs rather than details of individual diseases. this not only solves the impossible challenge of covering over rare diseases during medical training but also provides some equity between different diseases. lastly, m rd promotes the use of rare disease specific resources that will support both doctors and their patients. this includes the invaluable input from patient advocacy groups. the step between presenting quality assurance of rare disease (rd) centers of excellence (coe) through designation, accreditation, monitoring and constant improvement provides a means to ensure high quality, centralization of resources and expertise, and cost-efficiency. eucerd recommendations for quality criteria of coe, issued in , are still highly relevant [ ] . in the state of art resource, almost all european union (eu) member states (ms) claim, that their coes conform to eucerd recommendations [ ] . however, national quality assurance processes differ significantly: some ms apply robust procedures, while in other ms, many of them -but not exclusively -are eu- ms, processes of quality assurance are less developed. under the subsidiarity principle embedded into european treaties, the eu plays a limited role in many areas of healthcare, and coes quality assurance processes are a choice and responsibility of ms.with the establishment of erns, another layer of quality assurance has been developed by the european commission and the ms [ ] . this new quality assurance framework may be in line, or not, with national accreditation systems and involves i) assessment of coes when they apply for full membership of erns and ii) continuous monitoring afterwards [ ] . in every ern, members have to be "equal partners in the game" and share the same goals, rights and obligations. while the ern logo should eventually be a quality mark of the highest standards, strong links of ern members to national systems, including many more and less specialized healthcare providers, are essential to ensure proper care pathways for rd patients. importantly, erns themselves and patients/non-governmental organizations provide us with additional means of "informal quality assurance". many erns are implementing their own monitoring processes through the creation of registries to collect health outcomes that allow peer-benchmarking. meanwhile, patients provide their strong voice through european patient advocacy groups (epags) and help to signpost "the best" coes through information sharing. in both these processes, the power of open, transparent information on performance may finally lead to improved transparency and accountability at a national level and, presumably, may have an impact on the composition of erns in the future. in order to improve clinical research, patient preferences and outcome measures relevant to patients should become the core of drug development and be implemented from the earliest stage of drug development. from 'bedside to bench' instead of from 'bench to bedside' . at all levels the reuse of data could and should be enhanced. patient derived or provided data are not owned by those who collected them, and their reuse should be primarily controlled by the donors of these data. researchers and health professionals are custodians (gdpr). to enable the optimal reuse of real world data, the data needs to be findable, accessible, interoperable and reusable (fair) by medical professionals, patients and in particular also by machines. for this reason the world duchenne organization published a duchenne fair data declaration [ ] . reuse of placebo data and use of natural history data could speed up research especially in the field of rare diseases at this moment, in line with gdpr, patients are in a good position to decide about the reuse of their own data and should not only have access to these data but preferable also be in charge of their own data. background: drug repurposing for rare disease has brought more costeffective and timely treatment options to patients compared to traditional orphan drug development, however this approach focuses purely on medical interventions and requires extensive clinical trials prior to approval. in the case of refractory epilepsy, practical solutions are also required to better manage daily life. here we present an example of technology repurposing as a practical aid to managing absence epilepsy. methodology: existing research tells us that seizure control is not the only consideration of quality of life in children with epilepsy and that mental health and caregiver/peer support are of utmost importance. we explored the needs of stakeholders and determined that there was a delicate balance between the individual (and those that care for them) and those that have the power to change their lives. results: across all stakeholders there was a shared common need to obtain objective data on absence monitoring to relieve the burden on families/carers to retain manual seizure diaries whilst providing accurate and timely data to medical teams, researchers and social care. epihunter is an absence seizure tracking software using repurposed technology: a headset from wellness/leisure to collect electroencephalographic (eeg) data and an ai algorithm to detect and record absence seizures on a mobile phone application in real-time. both eeg and video recording of the seizure are automatically captured. this low cost, easy key: cord- -p bnvu authors: thiberville, simon-djamel; schilling, stefan; de iaco, giuseppina; fusco, francesco maria; thomson, gail; maltezou, helen c; gottschalk, rene; brodt, reinhard h; bannister, barbara; puro, vincenzo; ippolito, giuseppe; brouqui, philippe title: diagnostic issues and capabilities in isolation facilities in european countries: data from euronhid surveys date: - - journal: bmc res notes doi: . / - - - sha: doc_id: cord_uid: p bnvu background: highly infectious diseases (hids) are defined as being transmissible from person to person, causing life-threatening illnesses and presenting a serious public health hazard. the sampling, handling and transport of specimens from patients with hids present specific bio-safety concerns. findings: the european network for hid project aimed to record, in a cross-sectional study, the infection control capabilities of referral centers for hids across europe and assesses the level of achievement to previously published guidelines. in this paper, we report the current diagnostic capabilities and bio-safety measures applied to diagnostic procedures in these referral centers. overall, isolation facilities in european countries were evaluated. although % of these referral centers are located near a biosafety level laboratory, % and % of them still performed their microbiological and routine diagnostic analyses, respectively, without bio-safety measures. conclusions: the discrepancies among the referral centers surveyed between the level of practices and the european network of infectious diseases (eunid) recommendations have multiple reasons of which the interest of the individuals in charge and the investment they put in preparedness to emerging outbreaks. despite the fact that the less prepared centers can improve by just updating their practice and policies any support to help them to achieve an acceptable level of biosecurity is welcome. findings background a highly infectious disease (hid), such as severe acute respiratory syndrome (sars) [ ] , lassa fever [ ] , or other hemorrhagic fever [ , ] , is defined as transmissible from person to person, causes a life-threatening illness, presents a serious hazard in healthcare settings and in the community and requires specific control measures [ ] . suspected hid patients should be managed in specialized isolation facilities, such as "high-level isolation units" [ ] . to prepare the hospital management of hids for possible future outbreaks, the european union (eu) recently funded the european network of infectious diseases (eunid) [ ] . following eunid, the european network for highly infectious diseases (euronhid), a new eu-funded project, performed a cross-sectional study analysis of european isolation facilities. the specific mission of euronhid is to prepare and support isolation facilities to provide appropriate infection control measures and strategies for health care worker (hcw) safety during care to patients with suspected and confirmed hids. the appropriate management of hid cases requires high-level diagnostic capabilities. in , the eunid has reached a consensus on recommended biosafety procedures for the entire diagnostic process, from specimen sampling to the transport of laboratories [ ] . the aim of this paper is to report the current inventory of the diagnostic capabilities and infection control procedures for the appropriate and safe handling of specimens in isolation facilities in european countries who participated to the crosssectional study of the euronhid. at the beginning of the project, national health authorities in all of the european countries were asked to suggest a physician with expertise in hid management as a project partner and to identify all the isolation facilities as referral centers for hid in his country. to survey only isolation facilities identified by national health authorities for the referral and management of hids, we requested official documents in which these hospitals are clearly indicated. the data were collected during on-site visits using checklists specifically developed during the first year of the project [ ] . the checklists were drafted by the steering committee members (including partners from france, germany, greece and the united-kingdom) and then discussed with and approved by all of the partners. the coordination team (based at the national institute for infectious diseases in rome, italy) considered the strength scores of these assessments to be indispensable. on-site visits were performed by the project coordinator assisted, when feasible, by the project partner of the explored country from february-november . the objective of the project was to assess the level of achievement of each surveyed facility to previously published guidelines [ , ] . with this aim, a standard evaluation form was developed. in this form, all of the data were summarized in main topics, and for each topic, an evaluation score was assigned that represented in the general lab, without closed-type automatized analyzers / ( ) / ( ) *the corresponding assessment of the evaluation scores of the three topics are summarized below: topic : management of group agent specimens (a: the unit is located in the same hospital/city as a bsl- lab and has protocols for the safe and appropriate handling of group agent specimens; b: the unit is not located in the same hospital/city as a bsl- lab but has protocols for the safe and appropriate handling of group agent specimens to another city/country; c: the unit is not located in the same hospital/city as a bsl- lab and does not have adequate protocols for the safe and appropriate handling of group agent specimens to another city/country). topic : management of group agent specimens (a: the unit is located in the same hospital/city as a bsl- lab and has adequate protocols for the safe and appropriate handling of group agent specimens; b: the unit is not located in the same hospital/city as a bsl- lab but has adequate protocols for the safe and appropriate handling of group agent specimens to another city; c: the unit is not located in the same hospital/city as a bsl- lab and does not have adequate protocols for the safe and appropriate handling of group agent specimens to another city). the level of achievement with respect to a "standard" based on the available evidence, literature data and expert consensus [ , ] . the checklists and the evaluation form are available on the website www.eunid.eu after free registration. the participant selection process led to the inclusion of isolation facilities identified for the referral and management of hids in countries table . throughout europe, ( %) of the surveyed isolation facilities have a biosafety level (bsl- ) laboratory (lab) [ ] in their country, of which are located in the same hospital or city as the isolation facility to avoid or limit the distance of the transportation of hid diagnostic specimens. among those isolation facilities not located in the same city as the bsl- lab, ( %) have written protocols for the appropriate handling and transportation of the specimens to a bsl- lab, and ( %) have no specific protocols. among the isolation facilities surveyed, ( %) and ( %) have a bsl- lab [ ] for virological and bacteriological diagnosis in their country, respectively. in total, ( %) of the isolation facilities are located in the same hospital or city as the bsl- lab, and all of the isolation facilities have an adequate protocol for the safe and appropriate handling of group agents. sixteen ( %) isolation facilities have access to adequate capabilities for other routine diagnostic tests: (i) optimal use of bed-side testing inside the isolation area, (ii) use of the central hospital lab after the inactivation of samples, and (iii) use of the bsl- lab [ ] . however, ( %) of the isolation facilities perform routine analysis (such as biochemistry and hematology) in the central hospital lab without inactivation but using special measures of biosecurity and biosafety, such as closed-type auto-analyzers. the remaining ( %) facilities perform other diagnostic tests in the central laboratory without any special measures of biosecurity and biosafety. microbiological and routine diagnostic tests are performed directly inside the isolation area in ( %) and ( %) of the surveyed facilities, respectively. microbiological testing in the majority ( ; %) of isolation facilities and routine testing in a small proportion ( ; %) of the facilities are carried out in a bsl- lab. for ( %) and ( %) of the centers, the samples of microbiological and routine testing are sent to the central laboratory, which performs the analysis in a closedtype automatic analyzer. finally, microbiological and routine tests are performed in the central laboratory without using closed-type auto-analyzers in ( %) and ( %) of the surveyed facilities, respectively. although most of the isolation facilities surveyed have appropriate diagnostic capabilities and infection control procedures for the safe handling of specimens, % and % performed their routine and microbiological diagnostic tests in the central laboratory without any measures of biosecurity and biosafety as recommended by the eunid [ ] . the delay between data collection, and publication, can be considered as one of the limit of this paper. the discrepancies among the referral centers surveyed between the level of practices and the eunid recommendations have multiple reasons. one main explanation is the interest of the individuals in charge and the investment they put in preparedness to emerging outbreaks. obviously, these centers might benefit from larger funding from their national institution, or if they better allocated their internal resources. despite the fact that the less prepared centers can improve by just updating their practice and policies any support to help them to achieve an acceptable level of biosecurity is welcome. identification of a novel coronavirus in patients with severe acute respiratory syndrome a fatal case of lassa fever in london bichat guidelines for the clinical management of haemorrhagic fever viruses and bioterrorism-related haemorrhagic fever viruses response to imported case of marburg hemorrhagic fever, the netherland framework for the design and operation of high-level isolation units: consensus of the european network of infectious diseases infection control in the management of highly pathogenic infectious diseases: consensus of the european network of infectious disease euronhid checklists for the assessment of high-level isolation units and referral centres for highly infectious diseases: results from the pilot phase of a european survey biosafety in microbiological and biomedical laboratories (bmbl) diagnostic issues and capabilities in isolation facilities in european countries: data from euronhid surveys the authors would like to thank the entire euronhid working group for the fruitful discussions and input provided: norbert vetter (austria), mira kojouharova abbreviations hid, highly infectious disease; eu, european union; eunid, european network of infectious diseases; euronhid, european network for highly infectious diseases; hcw, health care worker; bsl- , biosafety level ; bsl- , biosafety level . the authors declare that they have no competing interests.authors' contributions sdt and pb wrote the manuscript, ss, gdi, gt, hcm, rg, hrb, bb, vp, and pb substantial contributed to design the study, participated in the acquisition and interpretation of data, and gave the final approval of the version to be published; gi substantial contributed to design the study, and gave the final approval of the version to be published. all authors read and approved the final manuscript. key: cord- -due tloa authors: nan title: ecr , part a date: - - journal: insights imaging doi: . /s - - - sha: doc_id: cord_uid: due tloa nan percutaneous needle biopsy has been a mainstay of oncologic diagnosis for almost three decades, since the advent of ultrasound and ct. the basic principles of fine-needle aspiration and core needle biopsy can be applied to almost any site in the body, with subtle differences in technique depending on the organ being investigated and the imaging modality utilised. while excisional biopsy is still appropriate in certain cases, percutaneous needle biopsy has become the standard of care in the diagnosis of most tumors throughout the body and is also used to diagnose noncancerous conditions, such as infection. percutaneous biopsy is also beneficial in the staging of patients with cancer, particularly when another treatment method may be more appropriate than surgical resection. the advantages of percutaneous biopsy over surgical excisional biopsy include time and cost savings and reduction in morbidity. the aim of this course is to discuss the practical aspects of biopsy, needle selection, and guidance techniques and to show how to approach difficult lesions and avoid complications. imaging after treatment of breast cancer is for confirmation of lesion removal, identification of postprocedural fluid collections, detection of residual or recurrent cancer and screening for metachronous cancers. posttherapy changes -which include fluid collections, edema, skin thickening, architectural distortion, scarring and calcifications -are mainly due to surgery, axillary dissection and radiotherapy. the greatest treatment-related changes occur - months after therapy, and mammographic stability is achieved after two to three years. for mammography, pre-and all posttherapeutic images have to be compared. ultrasound is the method of choice for evaluation of fluid collections. mri is for problem solving (i.e. differentiation between scar and relapse) and should not be performed prior to months after therapy to avoid false-positive diagnoses. dystrophic calcifications may develop in areas of fat necrosis mimicking malignancy. fat necrosis predominantly occurs at the treated site; however, it can develop anywhere in the ipsilateral breast. its appearance may be indistinguishable from cancer at all imaging modalities. to differentiate between fat necrosis and other common post-treatment changes from relapse, it is important to know the timeline when these changes occur and schedule follow up imaging accordingly. mammography serves as the basis for postoperative surveillance. ultrasound is helpful in the early postoperative phase, whereas mri is the method of choice, especially for differentiation of scar and relapse in the later postoperative phases. breast radiologists need to be familiar with post-treatment imaging findings in patients with breast cancer, but often patients are also imaged after a diagnosis of benign entities which are treated surgically (fibroadenomas, radial scars, papillary lesions) or after a percutaneous diagnosis of a high-risk lesion which has undergone a surgical biopsy to avoid underestimation (atypical ductal hyperplasia, lobular neoplasia or flat epithelial atypia). these procedures will leave an imprint on the breast which can be a cause of concern due to the surgical scar. furthermore, imaging findings after plastic surgery for mastopexy, reduction mastoplasties, implants or auxiliary techniques (lipofilling, hyaluronic acid) are becoming frequent in our daily practice and have their peculiarities that can overshadow breast cancers. interventional percutaneous procedures can also be a cause of tissue distortion. there are advanced systems for biopsy (bles) and for percutaneous removal of benign lesions (fibroadenomas, papillomas), which use large-gauge needles and have to be accounted for due to their trace in the breast tissue. endoscopy is currently considered the reference standard for the evaluation of colonic disease activity in patients with inflammatory bowel disease (ibd). however, it only allows evaluation of the mucosal surface and is not always complete. it cannot, therefore, help to estimate the depth of involvement of transmural inflammation and extraluminal complications, both characteristics of ibd. an evolving role for cross-sectional imaging in the evaluation of patients with ibd is increasingly recognised, especially in the setting of crohn's disease (cd) since the cross-sectional imaging has been demonstrated to have a high diagnostic accuracy not only for assessing the presence and extension of luminal disease, but also for evaluating the cd-related acute or chronic complications. available evidence suggests that ultrasound, computed tomography and magnetic resonance have similarly high diagnostic accuracy in the detection of disease activity, location, severity and complications: in particular, the penetrating and stricturing lesions which are characteristic of cd. thus, the choice of the technique for assessing cd may be influenced by local availability or expertise. in the case of ulcerative colitis, cross-sectional imaging, although less evaluated, may also be helpful in certain circumstances. there is evidence indicating that cross-sectional imaging is an alternative problem-solving tool to endoscopy whenever tissue sampling is not required and can provide valuable guidance for performing medical and surgical treatment with maximised efficacy and safety. overall, findings from crosssectional imaging accurately reflect disease activity and provide reliable information for decision-making and patient care optimisation. learning objectives: . to learn about optimised examination protocols for ulcerative colitis and colonic crohn's disease in the acute, subacute and chronic disease setting. . to become familiar with the criteria for the assessment of disease activity through ct, mri and ultrasound. . to learn about an integrated approach to the use of cross-sectional imaging in colonic inflammatory bowel disease. liver transplantation is the accepted treatment of patients with irreversible liver cell failure and some metabolic disorders and in a selected group of patients with hepatocellular carcinomas. over the last decade, major transplant centres have reported improving survival rates, though during this period they have developed more complex surgical techniques, including split-liver, auxiliary and live-related transplantation, and have treated more marginal higher risk patients. this successful outcome has been dependent on appropriate recipient selection, robust surgical technique, improvements in immunosuppression and intensive care management and the prompt recognition and treatment of complications. diagnostic and interventional radiology have been core specialties in achieving the goals of improved graft and patient survival. improvement in surgical techniques has decreased the more common vascular and biliary complications, but the newer techniques present differing diagnostic and interventional challenges, particularly in paediatric recipients. developments in mr and mdct allow many of these vascular, biliary and infective complications to be diagnosed non-invasively. vascular techniques of angioplasty and stent placement may reverse the sequel of graft ischaemia or portal hypertension. mrc allows the diagnosis of biliary strictures that may be treated by dilatation or stent placement. imaging is also important in the diagnosis of recurrent disease and the acquired diseases of prolonged immunosuppression including atypical infections and the post-transplant lymphoproliferative disorders (ptld). this interactive session will present these appearances by case example and provide guidance of the appropriate diagnostic and treatment paradigm. learning objectives: . to understand the common imaging findings after liver transplantation. . to recognise significant complications following liver transplantation. a- : b. imaging of treated liver tumours i. bargellini; pisa/it (irenebargellini@hotmail.com) imaging findings after systemic and loco-regional treatments vary greatly depending on baseline features of the treated tumour and treatment modality. conventional uni-dimensional and bi-dimensional criteria (such as recist . and who criteria) have been extensively validated in metastatic lesions treated with conventional chemotherapy. however, their prognostic value is limited in patients treated with new molecular targeted therapies and after locoregional treatments. in the setting of hepatocellular carcinoma (hcc), specific response criteria (such as easl and mrecist) have been proposed that take into account variation in the size of the viable tumour. these criteria have been extensively validated, although their interpretation could be troublesome after some specific treatments, such as molecular targeted agents and y radioembolisation. on the other hand, there is no consensus regarding tumour response criteria for metastatic lesions after loco-regional treatments. the role of morphological criteria and several imaging biomarkers (such as those provided by diffusion and perfusion imaging, fdg activity, dual source ct) is under investigation, being able to provide additional information on tumour activity and biology. while new drugs with different mechanisms of action and new treatments are becoming available, the work of radiologists is changing and there is increasing evidence that tailored radiological response criteria are required for these new targeted and tailored treatments. learning objectives: . to understand the common imaging findings after chemotherapy for liver tumours. . to recognise common imaging findings after radiofrequency ablation of liver tumours. . to be aware of the common imaging findings following transarterial treatment of liver tumours. vessel size is practical, since it is related to the pathological changes as well as to the clinical and radiological presentation. the two main large vessel vasculitides are giant cell arteritis and takayasu arteritis. behcet disease may combine large and small vessel vasculitis. imaging plays an important role in primary vasculitis. chest radiographs are not especially useful in large vessel vasculitis. contrast ct and mri further detect and especially help in the characterisation of large vessel vasculitis. vessel wall changes are well detected with both techniques. contrast enhancement, distribution of vessel involvement and morphological vascular changes should be considered. today, pet/ct is the recommended imaging technique in the assessment of vessel wall inflammatory changes and in the evaluation of treatment response. although the clinical scenario differs usually between takayasu and giant cell arteritis, the radiologist should combine the imaging findings with clinical and laboratory data to suspect a specific vasculitis. therefore, this presentation will concentrate on the basic signs and associated findings in large vessel vasculitis, pathologic correlation, imaging protocols and the differential diagnosis. the term vasculitis refers to a variety of clinico-pathological entities. the most widely used chapel-hill classification divides the vasculitis syndromes into three groups based on the size of vessels primarily involved. small vessel vasculitis such as anca-associated granulomatous vasculitis (wegener's disease), churg-strauss granulomatosis and microscopic polyangitis are most often associated with pulmonary abnormalities. the spectrum of hrct findings is quite variable and differs by entity. the hrct findings will be discussed together with clinical and laboratory findings to be integrated into a multidisciplinary diagnostic approach. the typical findings in wegener's granulomatosis include solitary or multiple, often cavitary nodules or masses, or focal or diffuse consolidations, churg strauss is dominated by interlobular thickening or transient multifocal and nonsegmental consolidations frequently in subpleural distribution. all types of pulmonary vasculitis may present with focal or diffuse pulmonary haemorrhage that produces pulmonary ground glass or consolidations in various distributions. a number of collagen vascular diseases (e.g., lupus erythematosus) or other granulomatous diseases (e.g. sarcoidosis) may affect the small pulmonary vessels causing haemorrhage or pulmonary hypertension. per disease entity, the course will review typical and more rare hrct features and key features that allow for diagnosis and classification, discuss side-to-side overlapping of morphologic features important for differential diagnosis and illustrate the findings of other diseases that represent the most challenging differential diagnosis, e.g. oedema, infection or malignant diseases. learning objectives: . to learn when hrct is of value in investigating pulmonary vasculitis. . to appreciate the different appearances of pulmonary vasculitis on hrct. a- : d. inflammation and remodeling a.a. bankier; boston, ma/us (abankier@bidmc.harvard.edu) the presentation will lay out the pathological and pathophysiological basis for the complex processes of inflammation and remodelling. the implications of these processes for imaging will be discussed and illustrated by selected pathologies. finally, the presentation will indicate how the imaging reflections of these processes could be used in the future as imaging biomarkers for the diagnosis, follow-up, and outcome evaluation of disease. at (marcus.hacker@meduniwien.ac.at) appropriate diagnosis and therapy of coronary artery disease (cad) frequently require information about both the morphological and functional status of the coronary artery tree. thus, combined imaging consisting of invasive coronary angiography (ica) plus spect myocardial perfusion imaging (mpi) is practiced in clinical routine diagnostic of patients with stable angina since many years and can therefore be accepted as the reference standard in the diagnosis of hemodynamically relevant coronary artery stenoses. both morphological and functional information are mandatory for the decision of performing an interventional therapy or initiating/maintaining medical treatment in numerous symptomatic patients. the hemodynamically relevance of coronary artery lesions is a major condition to decide whether an interventional therapy should be performed or not. a non-invasive concept providing both information could provide accurate allocation of perfusion defects to their determining coronary lesion and specific morphological and functional classification of patients with coronary artery disease. in symptomatic patients, a normal stress mpi confers a very low short-term risk for cardiac death and/or acute myocardial infarction. however, a normal mpi does not exclude the presence of underlying coronary atherosclerosis, which may be extensive although not yet flow-limiting. in this regard, ct will unmask a sizeable subgroup of patients with coronary atherosclerosis who should receive more intensive antiatherosclerotic intervention than would have been indicated by mpi results alone. knowledge regarding the presence and extent of subclinical coronary atherosclerosis in patients who do not have ischemia by mpi can be of importance in patient management. learning objectives: . to appreciate the scope of information a spect/ct cardiac study can deliver. . to become familiar with protocols of spect/ct studies. . to learn a structured approach to performing and reporting a spect/ct study. a- : c. mr/pet: do we really need it? h.h. quick; erlangen/ de (harald.quick@imp.uni-erlangen.de) following pet/ct and spect/ct, mr/pet hybrid imaging is the most recent addition to the palette of hybrid imaging modalities. mr/pet synergistically combines the excellent soft tissue contrast and detailed image resolution of mr with metabolic information provided by pet. integrated mr/pet systems furthermore offer the ability to acquire hybrid imaging data simultaneously. this can be applied to mr-based motion correction of pet data. these features open up several cardiac applications, e.g. evaluation of cardiac function and viability, diagnosis of cardiac inflammatory diseases and tumorous diseases. to fully assess the diagnostic potential of mr/pet, however, several technical challenges have to be considered: attenuation correction (ac) of the patient tissues in mr/pet has to be based on mr-images and is currently hampered by a limited number of tissue classes and undercorrection of bone tissue. cardiac radiofrequency coils and ecg gating equipment are currently not considered in ac. consequently, quantification of pet data therefore might be biased. the clinical workflow is rather complex and needs to be tailored to cardiac examinations. few research groups currently explore this new hybrid imaging modality in selected cardiac applications. cardiac mr/pet: do we really need it? considering the sparse clinical experience that is available today, it is quite early to answer this question yet. however, once the remaining technical hurdles are overcome and the diagnostic potential can be fully exploited clinically, the answer is most likely positive. the majority of malignant bone tumours can be detected on plain radiography. the age of the patient, location of the tumour in a bone and history of a preexisting bone abnormality should be included in determining the likely diagnosis. careful analysis of the pattern of bone destruction, periosteal reaction and matrix mineralisation allow for characterisation of most cases of osteosarcoma, ewing's sarcoma, chondrosarcoma and adamantinoma. mri is the best imaging technique for staging by displaying . the extent of bone marrow involvement, including epiphyseal infiltration and skip lesions, . the presence and extent of extraosseous soft tissue mass, . involvement of the neurovascular bundle, muscle compartments and adjacent joint. this allows to find the best biopsy approach and establish the feasibility of limb salvage as opposed to amputation. longitudinal non-contrast t sequences are the most accurate for determining intraosseous extent. axial fat-suppressed pd or t sequences optimally demonstrate soft tissue extension, and dynamic contrastenhanced mri is useful for differentiation of extraosseous tumour from edema and for assessment of tumour response to chemotherapy. chest ct is the most sensitive modality for detection of pulmonary metastases, and technetium skeletal scintigraphy is still frequently used for detection of osseous metastases. the most valuable, readily available and easy-to-use techniques to assess response to radiation therapy and chemotherapy are dynamic contrast-enhanced mr imaging, diffusion mri and colour-doppler ultrasound. to evaluate local control of disease and for detection of local recurrence, mri is usually the best imaging technique. plain radiography may detect complications of prosthesis. learning objectives: . to consolidate knowledge on malignant primary bone tumors on plain radiography, ct and mri. . to understand the principles of tumour staging and monitoring chemotherapy. . to become familiar with imaging findings following chemotherapy and surgery. oncologic approach to malignant primary bone tumours c. dhooge; ghent/be (catharina.dhooge@ugent.be) the most common bone sarcomas are osteosarcoma (os) and ewing sarcoma. os occurs primarily in long bones, ewing sarcoma occurs also in the pelvis, spine or chest wall. localised disease confers a % cure rate, and initially metastatic disease %. a multidisciplinary approach which includes neo-adjuvant systemic chemotherapy and local control measures for primary and metastatic sites followed by adjuvant chemotherapy has become the standard of care. complete surgical excision is mandatory in os; in ewing, radiation can also be used. standard chemotherapy for os is based on a combination of cisplatin, doxorubicin and high-dose methotrexate. ifosfamide and etoposide will be further considered (euramos). systemic therapy for ewing includes vincristine, doxoribicin, cyclofosfamide, ifosfamide and etoposide. high-dose chemotherapy with autologous stem cell reinfusion is evaluated in intermediate prognosis and metastatic disease (euro-ewing). among the different prognostic factors, response to neo-adjuvant chemotherapy by measuring chemotherapy-induced necrosis is a powerful indicator of outcome in bone sarcomas. patients who achieve a good histological response to pre-operative chemotherapy, defined as < % viable tumour at the time of tumour resection, have a better survival than those who show poor response (>= % viable tumour). five-year survival for good responders is o- % compared to - % for poor responders. however, the outcome for malignant bone tumours has improved little in the last years. large international studies exploring new drugs such as biologic agents (interferon) or mechanisms conferring drug resistance (topoisomerase inhibitors) in randomised controlled trials will hopefully lead to therapeutic innovation. learning objectives: . to become familiar with the principles of chemotherapy in malignant primary bone tumours. . to understand what the oncologist expects from the radiologist. . to recognise the value and limits of chemotherapy in malignant primary bone tumours. thursday a- : surgical approach of malignant primary bone tumours g.m.l. sys; ghent/be (gwen.sys@ugent.be) in primary malignant bone tumours, three surgical procedures are necessary: biopsy, resection and reconstruction. whether a biopsy is performed in an open or a percutaneous way is a matter of preference, but in each case it should provide sufficient and representative material for pathological investigation without compromising the following treatment. the area of interest and the trajectory should be determined preoperatively in a multidisciplinary meeting. as the biopsy tract has to be removed during the resection surgery, it should be close to the planned approach for the subsequent resection. if a biopsy is performed improperly, the diagnosis may be wrong or the following resection may be impaired because of extensive contamination of compartments, resulting in a severe functional deficit. surgery planning is based on clear imaging and a multidisciplinary discussion of tumour extension, aiming for a wide resection of the tumour with clear margins. correct perioperative measurements are based on predefined fixed bony landmarks that should preferably be visible on the same image as the tumour. an amputation is performed if a surgical limb-salvage procedure is expected to result in a non-functional limb. nowadays, several reconstruction techniques for bones (bone grafts or prostheses), tendons (artificial or human ligaments) and skin (skin flaps or grafts) are available. invasion of the soft tissues such as the neurovascular structures, muscles and skin, will determine which reconstruction technique (s) are necessary to restore the patient's function. each reconstruction technique bears inherent complications requiring a regular follow-up. we outline the current opportunities and threats in diagnostic radiology: traditional diagnostic radiology has been rapidly replaced by clinical radiology and the role of the radiologist is changing from image interpreter to clinical manager of imaging data. to care for the patient`s medical problem and not only his images is important. this contains great opportunities for radiology to develop and for the radiologist to become a central player in patient management. however, this requires not only clinical knowledge and involvement, but also visibility and sometimes even / availability. the need for " only" image reporting is declining as clinical subspecialisation easily brings image interpretation into the domain of non-radiologists. to understand the importance of identity, visibility is very important. visibility can be achieved by being part of mdt s and also in many other ways. clinical radiology is an idea which, after all these years, has not been very well embedded in current radiology and radiology training. to understand the importance of certified training, maybe the most important action we have to undertake now is to capitalise on our radiological expertise. image interpretation in a clinical context can only be done or franchised (to non-radiologist) under conditions of certification and accreditation by radiology. we should never let the primate of education and certification slip to non-radiologists. radiology certification should be a quality standard, recognised by international and national medical bodies, such as medical chambers and specialist societies. the latter should have high priority. session objectives: . to learn about the current opportunities and threats in diagnostic radiology. . to understand the importance of identity. . to acknowledge the importance of certified training. where are the turf battles in diagnostic imaging? g.m. bongartz; basle/ch (georg.bongartz@usb.ch) turf battles in radiology are a foreseeable development for most imaging techniques which evolve push-button methods and readily interpretable imaging results. we as radiologists must learn to accept competition. turfs should rather be seen as challenge than as threat. predominantly, nonirradiating techniques like ultrasound and mri have become progressively easier to apply (us) or to understand (mri), where the final image in some standardised areas (eg joints) can readily be interpreted by medical specialists in this field. but also x-ray diagnostics in dental medicine or orthopedic imaging is largely applied outside the radiologic department. ct today seems still relatively excluded because of its complexity. but with increasing resolution, optimal d reconstruction and fast accessibility, it is only a question of time when the added value of the radiologist will be under question. radiology has changed. we must become experts with respect to patho-anatomy and radiological differential diagnosis. this requires profound education, both initial training and ongoing education. radiology must be advertised internally and externally. in radiological publications, outcome studies are still underrepresented. demonstration of our dedicated skills, fostering our special expertise and offering this as support to our medical colleagues will preserve radiology for the future. cooperation with partners seems the only way out of a turf battle -we need to search for alliances and integrate our partners to create a win-win situation. a- : beating threats in europe with radiological training a.k. dixon; cambridge/uk (akd @radiol.cam.ac.uk) by introducing radiology to medical students, it is hoped that they will begin to appreciate how difficult it is! likewise, when teaching residents, they must come to understand the basic anatomy and principles better than their clinical colleagues; this is very much the tenet of subspecialty training. they should also appreciate that patient care is optimal when there is close collaboration between clinicians and radiologists. passing of an examination (national or edir) offers some proof that a radiologist has attained a certain standard, but the principle of testing oneself formally or informally at regular intervals again provides a measure of continuing competence. such certification can be of value in the case of litigation. radiology is one of the fastest growing specialties and the techniques that we learn during training are quickly outdated; hence, the need for life-long learning and effective continuing professional development. learning objectives: . to understand how to teach radiology to undergraduate medical students. . to become familiar with the principles of self-assessment. . to appreciate the necessity for life-long learning. history of ultrasound in radiology: lessons learned l.e. derchi; genoa/ it (derchi@unige.it) the relationship between radiology and us has never been an easy love story, and still isn't. in the early s this new imaging field was not readily covered by radiologists and many other colleagues established us sections within their departments. then, important fields of us (cardiology, obstetrics, gynecology) became almost exclusive domain of non-radiologists. in the s, the increasing success of us initiated turf battles with other clinicians. nowadays, clinical specialists perform > % of us examinations wordwide. in europe, most in-hospital us examinations is still performed in radiology, but this is not the case in all countries. a strong practice in us is quite important since this is often the first approach to patients. if initial us referrals are kept, radiologists will continue to guide decisions in subsequent imaging workup. furthermore, us keeps close to patients; in europe the examination is usually done directly by radiologists or, if by sonographers, the radiologist usually checks the case with limited directed us imaging. this clearly shows our role as physicians. to maintain a key role in us radiologists need to: ) have the best specialists in us; attention must be given to us in residency programs. ) invest in us technology (as examples: ceus, elastography, d/ d). ) invest in us research. ) have high visibility in the us community, keeping close contact with all clinical colleagues. ) present higher visibility of us within radiology, with leaders supporting us within the radiological community and each radiologist promoting us in his/her environment. learning objectives: . to understand why ultrasound has moved out of the hands of radiology in some subspecialties. . to learn what this means for daily radiology practice and the quality of work. . to know about the threats and how we should deal with them. thursday a- : interdisciplinary cooperation without losing identity m.f. reiser; munich/de recently, major shifts in the paradigms of patient care took place: interdisciplinary counseling and including the patient in the decision making process for diagnostic and therapeutic measures. the new concept is that the patient no longer comes to the specialist, but the specialists come jointly to the patient. this is reinforced by the competition among health care providers, political decision makers and the public. in many hospitals this general trend has resulted in the establishment of various centers and boards such as the breast cancer centers, vascular centers and tumor boards for several cancer entities to name only few. at the university hospital of munich centers and boards have been implemented up to now. in almost all of them the participation of radiologists is required. this results in a major challenge in terms of personal resources. together with the regular clinical-radiological rounds the time necessary for these activities equals full-time radiologist posts. the participation of radiologists has many beneficial aspects: acknowledgement of radiology as an important clinical discipline and of the radiologist as a clinical consultant; participation in and influence on the creations of sops and (internal) guidelines; opportunity to advocate appropriate radiological methods for diagnosis and interventions; increase of knowledge in clinical medicine and new concepts of therapy; close personal links with our clinical partners. in order to prevent radiologists from losing their identity as radiologists it is mandatory that they remain firmly integrated in the radiology department and to strengthen the position of radiology as the central institution for providing cost-effective imaging and interventional services. moreover it must be taken care to offer attractive career options within the field of radiology. at (michael.fuchsjaeger@medunigraz.at) ultrasound (us) is the adjunct method of choice to mammography in breast imaging. over the past two decades, us technology has undergone profound advancements and refinements. us has, therefore, become an assessment tool with a defined field of indications as well as a unique set of diagnostic descriptors for breast lesion differentiation. the acrin study has demonstrated the special benefit of us in patients at risk with dense breast tissue. the bi-rads lexicon for ultrasound, in its second edition, expanded the role of breast us. further enhancement of image quality as well as the recent advent of automated breast us has even fueled scientific discussions on the potential role of us for breast cancer screening. automated breast us, which is based on computed generation of a d imaging data set obtained from many parallel d images, offers a different approach with a variety of benefits. images are obtained by the sonographer in standardised fashion, whole breast data sets can be reviewed at any time after the examination, reducing operator dependence, and image fusion with, i.e. mri is easily possible. the basic physical background, the significance as well as important aspects of practical use of handheld and automated breast us will be explained and illustrated by the respective imaging examples. emphasis will be laid on strengths and potential weaknesses of both us technologies with regard to breast imaging. the terms "complicated cyst" and "complex-cystic lesion" are based on breast ultrasound (us) terminology. the simple cyst is a fluid-filled, clearly defined, anechoic lesion without any suspicion of a solid intracystic mass. if a simple cyst is complicated by echogenic fluid, it is called complicated cyst. this echogenic fluid can be caused, e. g., by cell debris. thin echogenic septa are common in complicated cysts. vascularisation can never be found in these echogenic components. a complicated cyst has no solid component and no thickened wall or thickened septa. the expected probability of malignancy is %. the us criteria of a complex-cystic lesion following berg et al. categorise it into types: type , thick outer wall and thick internal septa (alone or in combination of both); type , one or more intracystic masses; type , mass of mixed cystic and solid components (at least % cystic); type , predominantly solid with eccentric cystic foci (at least % solid). complex means there is a suspicion of a tumour inside a cyst or a solid tumour with cystic components. a complicated cyst will usually be followed by us after months. a complexcystic lesion needs a correlation with mammography. us-guided large core needle biopsy in combination with a us and mammography visible marker placement is useful in type and in type complex-cystic lesions. in type and type complex-cystic lesions, a preoperative hook wire placement and excision surgery is the typical procedure. learning objectives: . to learn about the us appearance of complicated cysts and complex-cystic lesions. . to consolidate knowledge on differential diagnosis for these respective lesions. . to understand the diagnostic algorithm for a work-up of these lesions. renal colic is the most frequent non-obstetric cause for abdominal pain and subsequent hospitalisation during pregnancy. intervention is necessary in patients who do not respond to conservative treatment. ultrasound (us) is widely used as the first-line diagnostic test in pregnant women with nephrolithiasis, despite that it is highly nonspecific and may be unable to differentiate between ureteral obstruction secondary to calculi and physiologic hydronephrosis. magnetic resonance imaging (mri) should be considered as a second-line test, when us fails to establish a diagnosis and there are continued symptoms despite conservative management. moreover, mri is able to differentiate physiologic from pathologic dilatation. in fact in the cases of obstruction secondary to calculi, there is renal enlargement and perinephric oedema, not seen with physiological dilatation. in the latter, there is smooth tapering of the middle third of the ureter because of the mass effect between the uterus and adjacent retroperitoneal musculature. when the stone is lodged in the lower ureter, a standing column of dilated ureter is seen below this physiological constriction. mri is also helpful in demonstrating complications such as pyelonephritis. in the unresolved cases, computed tomography remains a reliable technique for depicting obstructing urinary tract calculi in pregnant women, but it involves ionising radiation. nephrolithiasis during pregnancy requires a collaboration between urologists, obstetricians, and radiologists. learning objectives: . to list the us, mr and ct imaging findings of urolithiasis and urinary tract infections. . to discuss the role and the appropriate uses of us, mr and ct for imaging these suspected conditions in pregnancy. . to discuss how imaging can influence the management of nephrolithiasis during pregnancy. oncology t.f. hany; zurich/ch (thomas.hany@gmail.com) in malignant tumours, pet/ct imaging using -fluoro-deoxyglucose (fdg) is widely used nowadays. fdg-uptake is an unspecific process and results in physiological as well as pathological malignant and non-malignant tissue uptake. to properly understand and interpret pet/ct imaging, knowledge of the mechanism and pathways of fdg in different tissue types like brain and the gastrointestinal tract is key. intrinsic as well as extrinsic factors influence uptake pattern not related to oncological disease like uptake in brown fat in patients exposed to cold ambient temperature and psychological stress or increased muscular uptake in nonfasting patient. typical treatment-related changes occur during and after treatment and must be recognised correctly so as not to overstage patient disease. a systematic analysis and knowledge of all these factors facilitate and improve the reading of oncological clinical cases. learning objectives: . to understand pattern of physiological fdg-uptake. . to learn about the pattern of non-pathological uptake in several tissue types. . to understand the influence of chemotherapy and other agents on fdguptake in the body. radiologists mostly prefer to use the comparative technique while evaluating radiological images, especially when the anatomy is identical for both sides. comparative analysis mostly works if the anatomic details are not complex or the right resembles the left. however, the complex anatomy and relatively high incidence of individual variations in head and neck may hinder this way of interpretation. variable pneumatisation of paranasal sinuses and temporal bone may cause difficulty. vascular system, especially venous structures may be sources for pseudolesions. for example, asymmetrically enlarged jugular vein or venous plexuses at any location may be misinterpreted as mass of any origin. veins may also become problematic on mr imaging due to entry slice phenomenon, in-plane flow, and flow turbulence effects and can have variable enhancement. normal anatomical structures such as facial nerve may enhance and can be mistaken to be pathologic on mri. technical issues can be considered as a group that may cause pseudolesions. images of the improperly positioned patient may become problematic while evaluating tiny structures. pet and pet-ct carry their own risks for pitfalls. normal structures may confound interpretation and result in false-positive findings. the situation may become much more complicated when there is asymmetry. similarly, atrophy and unilateral absence of a structure may be misread as a mass on the contralateral side. because of the complex anatomy and relatively high incidence of individual variations, head and neck imaging demands much more attention and careful analysis. the appropriate imaging technique and detailed knowledge of anatomy are essential to recognise pseudolesions. as a general rule, detailed knowledge of the patient's clinical history is essential before any imaging study is performed, to evaluate the appropriateness of the indication, tailor the acquisition protocol and correctly interpret the study. in head and neck imaging, this rule particularly fits in the emergency setting. in certain circumstances, the swelling and enhancement of soft tissues produced by infectious lesions may mimic a neoplasm. even more so, in patients already treated with surgery or chemoradiation. in these patients, the challenge is double: to identify any abnormality in the new anatomy produced by treatment and to discriminate between inflammation, complication and relapse. these can be very difficult without knowledge, for example, of the reconstructive procedure after surgery. previous images are extremely useful, not only because they help detect abnormalities, but also because through indirect information on the growth rate of the lesion they are crucial for correct interpretation of findings. tablets in radiology represent a novelty. since the introduction of the tablets on the market, the radiological field has been probably the first medical discipline to discover the many advantages of these devices. in fact, many applications for image management have been made available on the app stores (apple and android), and allow radiologists and non-radiologists to handle dicom images on the tablet, as part of the patient's record. however, the emerging applications are driving the process from the simple dicom image viewing to the full integration of the tablet with the pacs, allowing the handling of a patient's full record and presumably the possibility to report. in view of this rapid technological development, again radiology falls in the middle of a storm and is asked to find a solution to problem: are tablets suitable to read and report dicom images? and if so, which kinds of images (ct, mri, x-ray, etc) .? how can we manage the portability of patients' data (security issues, data loss, etc).? what will be the impact on teleradiology? these issues will be addressed by the panel of experts who will speak in the refresher course. session objectives: . to appreciate the current state of tablet technology and its practical use in radiology. . to understand the pros and cons of the use of tablets. . to learn about specific critical areas of utilisation (dicom images reading and teleradiology). a. tablet-computers: a technical overview j. fernandez-bayo; sabadell/es (jfernandezb@cspt.es) since their introduction in , tablet pcs have evolved extensively. they have become very popular, filling the gap between laptop computers and smart mobile phones. we will review technical aspects like the processors, storage space and memory, size and weight, connectivity and networking, software, autonomy, and battery life in different devices. special focus will be on the displays and different possible uses in radiology. for diagnostic purposes, different technical aspects must be taken into consideration, like the display size, resolution, pixel size (pixel pitch or pixels per inch), luminance, and contrast. displays in radiology normally have sizes between " and " and resolutions up to mpx, with luminance that ranges from cd/m to cd/m . by contrast, tablets have screen sizes between " and " and resolutions up to mpx, with luminance that ranges from cd/m to cd/m . due to their screen size, tablets score better in pixel pitch, with around ppi versus ppi in medical displays. the two systems have similar contrast ratios. other technical aspects of display that we should take into consideration are the number of distinguishable grays that can be represented, which in tablets correlates with the number of colours that can be displayed (color gamut) and colour accuracy (delta-e). additionally, we should take into consideration that displays should be calibrated for medical diagnostic purposes and tools to calibrate tablets have recently become available. learning objectives: . to learn about the pc evolution: from desktops, to laptops and tablets. . to appreciate the versatility features of a tablet. . to understand the hardware features with a specific focus on display. . to understand the hardware features with a specific focus on networking. a- : b. reading dicom images on the tablet o. ratib; geneva/ch touch screen tablets are becoming widely available, providing convenient mobile solutions for physicians and health-care providers. this is particularly attractive in medicine to "nomad" physicians, who need to be able to access relevant patient data and images anywhere-anytime in their daily practice where they are rarely at a single location. while they may not always be adequate for routine diagnostic tasks, they provide a convenient mobile solution for on-call and remote consultations. there are different types of software architecture that can be implemented for such tasks. two different major designs are: ( ) online web-based applications where the device serves as a "thin-client" to display images rendered and manipulated on a remote computer and ( ) local applications that reside on the mobile device and can run independently after images have been downloaded on the device. the first solution requires the user to be constantly connected to the network, while the second solution can continue to function after being disconnected from the network. most pacs vendors are starting to provide web access to their imaging solutions that can be accessed from mobile devices. web access can however be slow and dependent on reliable access to wireless network. we chose to develop a stand-alone companion application to our open source imaging platform osirix. with the increasing capacity and computing power of mobile devices, users will soon be able to perform most of the processing and image manipulation functions that are today only feasible on desktop or laptop computers. learning objectives: . to learn which dicom readers are available for tablets. . to appreciate the different approaches to dicom reading (local vs remote) and the pacs/tablets integration. . to understand the pros and cons of dicom image reading with the tablet in regards to image quality and displays. the optimisation of clinical breast cancer care comprises the prevention of over-diagnosis, reduction of over-treatment, and avoiding unsuccessful treatments. these goals are targeted in the vph-prism project by seven european and two us partner institutions. from the prospectively collected data, we will derive optimised imaging protocols that comprehensively take a woman's history and risk factors into account. the tight integration of radiological and histopathological images enables joint assessment of quantitative tissue parameters from microscopic and macroscopic imaging. more informed therapy decisions can be taken by such enhanced multidisciplinary data, backed by a powerful case database. the reduction of overtreatment, and highly individualised diagnosis and therapy decisions are closely related. we ultimately aim at deriving predictive parameters from the multi-modal and multi-disciplinary database to select the treatment option with the best chance of enduring recurrence-free survival. prospectively collected data, personal risk factors, and corresponding imaging data fed into the database from three large cohort studies are expected to provide insights into the individual preconditions and factors affecting disease progression. to optimise therapy, the decision between therapy options is key. this will be addressed by a decision support system using the project database. for surgical excision planning, vph-prism will aim at providing better estimates of tumour size, and with visual planning aids supporting radiologists and surgeons. for chemotherapy, quantitative lesion parameters, tracked over time during therapy, may help to detect success or failure of the treatment early on, such that a chemotherapy regime can for a non-responder be switched to a different regime in due time. quantitative imaging biomarkers in dementia; the fp vph-dare@it project w.j. niessen; rotterdam/nl (w.niessen@erasmusmc.nl) the number of individuals suffering from dementia today is roughly million. due to ageing societies, this number is predicted to increase to million by . worldwide annual costs are estimated to be € billion. in , the who declared dementia a global health priority, highlighting the urgent need for improvements in this area. vph-dare@it's aim is to enable more objective, earlier, predictive and individualised diagnoses and prognoses of dementias to cope with the challenge of an ageing european society. the expected impact of vph-dare@it will influence the scientific, clinical and industrial communities across europe and internationally to improve health care of dementia patients. in this presentation we will introduce the vph-dare@it project. we will then focus on the development, validation and implementation of quantitative imaging biomarkers for the early detection and differential diagnosis, which will take place as part of the project. author disclosure: w.j. niessen: advisory board; part-time detached to quantib bv as scientific director. shareholder; co-founder and shareholder of quantib bv. horizon : improving diagnosis and medical interventions and support to medical imaging j.-l. sanne; brussels/be (jean-luc.sanne@ec.europa.eu) research and innovation contribute to increasing europe's competitiveness. at the same time, research and innovation help make people's lives better by improving things like healthcare. in january the european union launches horizon , the biggest eu research and innovation programme ever, with a budget of € billion (current prices) over seven years ( ) ( ) ( ) ( ) ( ) ( ) ( ) . eu support for research and innovation helps drive international projects across the european union and beyond, and promotes the progress of knowledge and technology. one of the three key pillars of horizon will be tackling societal challenges (priority ) such "health, demographic change and well-being". one feature is the development of new and more effective diagnostics. relevant tools and technology innovations will be supported in view to improve disease outcomes through earlier, more accurate diagnosis and by allowing for more patient-adapted treatment. support will be provided through the work programme - of societal challenge "health, demographic change and wellbeing". additional opportunities are offered in the work programme - of priority (industrial leadership) "leit -information and communication technologies". practically, all imaging modalities have been used in space research since a long time ago. first of all, radiology is targeted in research activities, because by definition all cosmonauts and astronauts are healthy persons. the biggest volume of examinations had been performed on crew members for studies of organs and functions of the human body before and after long-term flights. health effects of zero gravity were of special interest. for example, multiple studies with involvement of both russian and international specialists were done on bone mineralisation (bone densitometry), muscle volume (mri) and metabolism (mr-spectroscopy) both in astronauts and healthy control subjects. first, ultrasound (us) examinations were conducted in space onboard russian orbital stations in . complex us studies were done by the author himself during his flight as a crew member of "salut- " orbital station in . the author had found some complex changes in blood and fluid dynamics under weightlessness. teleradiology was used for data transfer to medical specialists on the earth. today, we are continuing to use radiology for research on astronauts. for example, applications of sophisticated brain mri and fmri methods are of special interest for studies of subtle brain changes in space travellers. among our targets are flights to the moon, mars and large asteroids. to be better prepared for that task, we need better diagnostic tools. new versions of mobile us systems, miniature x-ray and ct machines suitable for use inside spaceships are under development. there are several mri pitfalls that should be recognised when imaging the female pelvis. mri appearances of uterus and ovaries are dependent on the phase of menstrual cycle/use of exogenous hormone therapy. normal postsurgical and post-radiation appearances of the pelvis can sometimes mimic tumour recurrence. it is important to become familiar with these appearances to avoid potential pitfalls. one very common pitfall is differentiation of transient myometrial contraction from adenomyosis. interrogation of all imaging planes over the duration of the entire mri examination can be useful to distinguish between the two, although myometrial contractions can last up to min. the choice of the correct imaging plane is crucial for the precise classification of uterine anomalies (coronal oblique) and accurate evaluation of parametrial invasion (axial oblique) in patients with cervical cancer. both dynamic contrast-enhanced mri and diffusion-weighted mri improve the accuracy of mri in the evaluation of malignant pelvic conditions. however, certain pitfalls related to each technique should be recognised to avoid misinterpretation. it is crucial to be familiar with the anatomy of the uterovesical (uv) ligament, as it is often the site of pelvic lymphoma (such as bladder or cervix lymphoma). however, some benign conditions such as endometriosis can involve the uv fold and invade both bladder and uterine wall. certain mri features can be helpful in making the correct diagnosis. learning objectives: . to become familiar with normal variations in mri appearances of female pelvis resulting from physiological conditions (e.g. different phases of menstrual cycle) and treatments (including exogenous hormone therapy, surgery and radiation) potentially mimicking disease. . to consolidate knowledge on the role of the correct mr imaging plane in avoiding potential mis-classification of uterine anomalies and parametrial invasion in patients with cervical cancer. . to learn about certain pitfalls related to dynamic contrast-enhanced mri and diffusion-weighted mri. a- : b. pitfalls in ultrasound k. kinkel; chêne-bougeries/ch (karen.kinkel-trugli@wanadoo.fr) pitfalls of sonographic findings in the pelvis can be related to technical issues, interpretation errors or to the patient's specific condition or pathology. common problems consist of insufficient bladder filling, misinterpretation of posterior enhancement or shadowing according to the anatomical structure and pathology of a size that goes beyond the field of view of the probe. organspecific problems will be illustrated in interactive questions, particularly for the uterus and the ovaries. identification of diseases can be difficult in many areas of abdominal imaging and therefore misses are easily made, also by the more experienced radiologist. it is difficult in the mesentery and peritoneum, covering a large area but being a very thin structure where abnormalities are easily missed. therefore, knowledge of the peritoneal anatomy and pathophysiology of peritoneal diseases is not widespread and differential diagnosis can be difficult. hence, special attention should be paid to prevent misses. another difficult area is diagnosing occult gastrointestinal bleeding. here, the choice of the proper imaging method is crucial as well as understanding the pros and cons of the method. the technique used should be optimised, as otherwise sometimes subtle signs of bleeding are missed. the presence of bowel dilatation itself is often correctly diagnosed, but the extent of the obstruction, the cause of the obstruction and especially complications (ischaemia) might be misdiagnosed. ct is the preferred technique for evaluation of bowel dilatation. care should be taken to scrutinise the examination for principal findings in patients with bowel obstruction. a. mesentery and peritoneum d. akata; ankara/tr (dakata@hacettepe.edu.tr) imaging findings of neoplastic or inflammatory diseases within the peritoneal cavity and the mesentery sometimes overlap and cause difficulties in interpretation. even disease processes in the peritoneum, mesentery or omentum may not be recognised on radiological examination, causing major difficulties in the management. peritoneal anatomy and physiopathology of peritoneal diseases must be well understood for better evaluation. ct is the best modality to assess the whole cavity. mr is equally sensitive with better contrast resolution; however, both modalities have advantages and limitations. ultrasound has a complementary role in evaluating the peritoneal fluid content. the presence of lacelike mobile thin septa is highly significant for tuberculosis. for better diagnosing the pathology, systematic approach is needed such as assessing the presence or absence of fluid in the peritoneal cavity, its character and location, accompanying soft tissue densities and their location. some inflammatory or infectious causes, such as acute pancreatitis and tuberculosis, involve typically parietal peritoneum and certain peritoneal reflections. peritoneal carcinomatosis involves typically both visceral and parietal peritoneum as well as subdiaphragmatic space. contrast enhancement patterns of the peritoneal membranes and the mesentery also have a complementary role in differentiating a variety of diseases. learning objectives: . to learn about the imaging characteristics of peritoneal and mesenteric masses and their differentials. . to appreciate the potential and limitations of imaging techniques in the detection of such lesions. . to understand the common pitfalls in diagnosis. a- : a. filippone; chieti/ it (a.filippone@rad.unich.it) patients with persistent, recurrent, or intermittent bleeding from the gastrointestinal tract for which no definite cause has been identified by initial oesophagogastroduodenoscopy, colonoscopy, or conventional radiologic evaluation are considered to have an occult gastrointestinal bleeding (ogib). the management of ogib is clinically challenging, since the causes of such a bleeding frequently arise in the small bowel (sb), until now considered as the ''dark continent''. conventional barium contrast studies allow only a limited evaluation of sb, whereas angiographic diagnosis is strictly related to active bleeding. the introduction of capsule endoscopy (ce) as well as of crosssectional imaging dedicated to the sb visualisation, such as multidetector-row computed tomography enteroclysis (cte) and magnetic resonance enteroclysis (mre), represents significant technological advances that have overcome the limitations of older diagnostic tests. although ce is recommended as a first-line investigation in ogib patients, cte or mre are alternative diagnostic tools when ce is contraindicated due to suspected/known obstruction or stricture. moreover, in patients in whom a tumour is suspected, cte or mre may be the preferred initial test. therefore, radiologists have to be familiar with cte and mre techniques, in terms of bowel distension, scanning parameter selection, contrast administration as well as with specific imaging findings. similarly, they have to be aware of the potential pitfalls such as suboptimal bowel distension, artefacts due to peristalsis or breathing, intraluminal food debris and previous surgery. learning objectives: . to understand the causes of gi bleeding and underlying pathophysiology. . to appreciate the strengths and limitations of the imaging techniques used in diagnosis. . to learn about common pitfalls in diagnosis. dilatation. presently, ct takes a major place in this setting with a reduced role for plain films. sonography is an alternative method when ct is not recommended. the questions of mondor in were pointed before the advent of cross-sectional imaging; these are still valid and have to be answered: ) is this a true occlusion (how to differentiate it from adynamic ileus)? ) does the obstruction concern the small bowel or the colon (impacting on the decision to operate or not)? ) what is the cause of the obstruction? ) are there signs of bowel ischaemia? the key points to avoid missing imaging diagnoses in bowel dilatation are to make the distinction between true mechanical obstruction and adynamic ileus (an early sign of mesenteric infarct) and to detect signs of strangulation leading to ischaemia (such imaging findings are present despite normal blood tests). in this setting, ct is the preferred technique; if it is not available, plain films and sonography can help in an optimal medical decision. the national lung screening trial (nlst) found a % reduction in lung cancer mortality with three annual screens using helical ct relative to chest xray. the us preventive services task force has issued draft recommendations to provide annual ct screening to high risk individuals defined by age and smoking criteria. questions remain regarding the implementation of ct screening: the determination of risk to identify those who should be screened, the frequency and duration of screening, definitions of screen positivity based on detected nodule characteristics, diagnostic algorithms for the follow-up of positive screens, overall societal costs of screening, and methods to adequately diffuse this technology across all socioeconomic groups at risk. answers to these questions are being addressed by several groups through ongoing research and secondary analyses of trial data. several models exist to identify individuals at highest risk of lung cancer (and lung cancer mortality), which may be enhanced if validated molecular markers are included. ongoing analyses of nodule features and likelihood of lung cancer will better inform screening interpretation guidelines and diagnostic algorithms. estimates from the nlst suggest that screening as performed in the nlst will be cost effective: the base case estimates $ , per quality adjusted life year gained (qaly). subset analyses suggest that screening is much more cost-effective in women than men, in higher versus lower risk groups, and in current versus prior smokers. while several variables influence cost-effectiveness, major drivers of cost are the cost of ct screening exams as well as the number of follow-up ct scans per positive screen ( . in nlst). while the national lung screening trial (nlst) had shown a significantly reduced lung cancer-specific mortality and all-cause mortality, the current results from the various european trials do not show such positive results. the dutch-belgian nelson trial, the german lusi trial and the british ukls have not yet published their results. so far, more cancers were detected in the screening arm of published danish and italian trials, but the positive effect on cancer mortality could not yet be shown. the italian mild trial even showed higher mortality in the yearly screening arm, and no difference between yearly screening and the control arm. this presentation discusses the implications of these findings and the consequences on implementation on screening in clinical practice. while limited in number of participants, european trials used advanced approaches that are most likely to be used if screening will ever be implemented. this presentation will discuss issues such as growth assessment and volumetry as critical factors for nodule assessment, handling of sub-solid lesions, computer aided detection and evaluation, and use of nonradiologists for reading and standardised follow-up. the goal is to provide an insight into how the european trials have affected our idea on practical implementation of lung cancer screening. the discussion will address the following issues: . are there sufficient data to support the implementation of individual lung cancer screening in clinical practice? . if yes, how to move from efficacy to effectiveness: what are the most optimised risk profiles for screening? what is the minimum level of infrastructure support and organisation required? how should subjects be informed about screening? what are the optimal follow-up duration and screening interval? what are the technical and positive screen management requirements? . what about the alternatives of waiting for additional data from ongoing trials or the validation of new biomarkers of lung cancer, permitting better selection of screened population before implementing individual lung cancer screening? attendees' opinions will be collected interactively. the speakers will be asked to comment and debate. treatment of prostate cancer depends strongly on the stage of the disease at the time of detection. however, treatment is controversial, even in prostate cancers found through early detection. this presentation will be based on a careful recent literature review. early detection of prostate cancer by use of psa testing leads to a significant reduction of at least % of disease-related mortality in screened men. this is however at the price of - % of overdiagnosis, usually resulting in overtreatment. diagnostic developments which reduce overdiagnosis and the proper identification of cases which may not require treatment are central issues of current research and will be addressed by referring to available evidence. active observation of potentially nonaggressive cancers is an option in the management of this disease until, preferably, the diagnosis of such cases can be avoided all together. radical prostatectomy and radiotherapy are the options for treating those cancers which are judged to be aggressive. treatment recommendations for prostate cancer continue to evolve and are affected by technological advances in surgery, new discoveries in tumour biology, and the development of predictive and prognostic biomarkers. with the increased complexity of treatment decision-making, the role of mri is also evolving. the more individualised and targeted the treatment approach, the greater is the role of imaging in treatment selection, planning and follow-up. mri is especially crucial for planning technologically sophisticated treatment approaches such as robotic surgery, igrt, or focal therapy and in assessing patients' eligibility for active surveillance. though needs for uniform interpretation and standardised reporting remain unresolved, the value of mri in pretreatment staging of prostate cancer, particularly for evaluating extracapsular extension and seminal vesicle invasion, has been documented. not only the tumour stage, but also its size, volume and grade (aggressiveness) are important factors that influence treatment selection. therefore, the introduction of functional mr is essential. when added to t weighted mri, diffusion-weighted mri, dynamic-contrast-enhanced mri, and mr spectroscopy, in addition to improving tumour detection, can provide an indication of tumour aggressiveness. mri performance on all sequences is dependent on lesion size/volume and grade, and this should be considered when applying mri results to treatment selection and planning. while it has been shown that multiparametric mri is generally more accurate and informative than anatomic mri alone, evidence-based guidelines specifying which combinations of sequences are essential for specific indications in patients with prostate cancer are yet to be developed and validated in welldesigned studies with robust outcome measures. in major trauma, it is essential to immediately recognise life-threatening conditions and to initiate early treatment. the majority of these patients succumb at the site of the injury to severe injuries of the central nervous system, heart and great vessels. however, there is another peak of early deaths within the first four hours after the injury. in the majority, these patients are at risk due to haemorrhage which is basically controllable by early minimally invasive treatment. consequently, this means that the vast majority of internal injuries can be survived as long as bleeding control is achieved within the first four hours after an injury. minimally invasive interventional techniques are already in use at a very early stage of the clinical decisionmaking process. particularly, we have witnessed a major paradigm shift in the treatment of blunt aortic injuries. endovascular repair has replaced open repair in many trauma centres. in appropriately selected patients, this adoption of endovascular stent grafts has resulted in a reduction in perioperative mortality, stroke and paraplegia, as compared to open repair. this entire session will provide knowledge about indications, requirements, standard procedures, and outcome of vascular emergencies, pardon me, urgencies. a. aortic emergencies m. cejna; feldkirch/ at (manfred.cejna@lkhf.at) aortic emergencies can be classified into several categories. "acute aortic syndrome": acute aortic syndrome is an entity of changes of the aortic wall. these include aortic dissection, intramural thrombus, and penetrating atherosclerotic aortic ulcer. besides coronary malperfusion and aortic valve insufficiency (type a dissection), emergency presentation is often due to visceral/limb ischaemia. sequelae of aortic syndromes are pseudoaneurysm formation and potentially aortic rupture. endovascular treatment has been established as the treatment option in at least type b dissections and penetrating aortic ulcers. aortic aneurysm rupture: with increasing diameter, aneurysms in the thoracic, thoraco-abdominal and abdominal aorta are prone to rupture. signs of impending aortic rupture are pain and imaging signs are sometimes subtle just like blurred contours of the aorta and (blister like) vascular wall deformation. endovascular treatment is an alternative to vascular surgery. aortic trauma: in severe thoracic trauma (mostly deceleration trauma), pseudoaneurysm formation and aortic rupture occur at the level of the aortic isthmus. endovascular treatment is considered the treatment of choice. inflammation, mycotic aneurysms and aorto-enteric fistulae: with acute and chronic inflammation, aneurysm and pseudoaneurysm formation and fistulae to enteric structures can occur. endovascular treatment is more often considered as bridging to vascular surgery instead of being a definitive cure. iatrogenic aortic injuries: besides accidental aortic injury during operations or interventional procedures, transaortic valve replacement and placement of aortic balloon pumps can result in iatrogenic trauma. endovascular treatment may be a minimally invasive approach. the focus will be on systematic presentation of aortic emergencies and the role of endovascular treatment. there are a number of vascular visceral emergencies that may be traumatic or nontraumatic in origin. the degree of urgency or potential for urgency varies with each condition and the subsequent management and intervention techniques will vary accordingly. an understanding of the natural history, pathophysiology, diagnostic tests and interventional techniques for these conditions will aid the diagnostic and interventional radiologist in identifying the pathology and knowing when to refer or initiate definitive treatment. i talk about embryonal development and anatomical differentiation of cardiovascular system, with special attention to the peripheral vascular system. the goal of this part of presentation would be to understand the anatomical structure and architecture of arteries and capillaries. we will discuss the casual classification of different kinds of peripheral arterial emegencies; e.g. congenital, posttrauma, arteriosclerotic and iatrogenic. the audience will learn about different therapy approaches, interventional and non-interventional, with accent on pad (classification, pathophysiology, risk factors, statistical facts and therapy). at the end, we will review different studies about combination therapy (conventional and interventional) with different possibilities and outcomes. modern cancer research and increasing therapy are mechanism based with the development of disease-modifying therapies that target the hallmarks of cancer. modern imaging tools enable the visualisation and quantitative assessment of the expression of molecular targets, of their interaction with potential ligands, as well as of the functional consequences of interactions at a molecular, cellular, metabolic, physiological, and morphological levels in a temporo-spatially resolved manner. the ability to gather such information from the intact organism renders imaging highly attractive for biomedical research and drug development. the determination of cell proliferation with radiolabelled thymidine is a wellestablished method in various life science branches where [ h]thymidine has been used for more than years and still represents the gold standard for the assessment of tumour cell proliferation. since non-invasive determination of this parameter is required in clinical studies, considerable efforts have been made in radiopharmaceutical research. the synthesis of a large series of proliferation markers resulted therefrom mostly focussing on pyrimidine nucleosides. the labels used were gamma as well as positron emitting radionuclides. here, [ f]flt ( '-deoxy- '-fluorothymidine) is examined and discussed with respect to its preclinical and clinical application. since apoptosis is an important mechanism of cell death in tumours responding to treatment, the non-invasive assessment of apoptosis with tracers for the detection of phosphytidyl-serine presentation and/or caspase activation could be used as surrogate marker for therapeutic efficacy. several approaches have been followed during the last years; their potential and limitations will be presented. angiogenesis is one of the hallmarks of cancer that is considerably easy to characterise with many different imaging modalities and methods. it is not only crucial for invasive and metastatic tumour growth, but is also a prerequisite for the accumulation of anticancer drugs and highly impacts the success of radiotherapy. this talk summarises our experiences in microstructural, functional, and molecular imaging of tumour angiogenesis. the microarchitecture of vessels can longitudinally be studied using high-resolution ( )ct. dce mri and hf-us are introduced as favourable tools to characterise perfusion, vessel permeability and vessel maturation during antiangiogenic treatments. using targeted probes, it is shown that "fluorescence molecular tomography (fmt)", mri and us are capable of estimating the expression of angiogenic marker molecules on tumour vessels and of matrix-associated enzymes in the interstitial space during vascular remodelling. additionally, examples are also given for epr-based nano-sized theranostics and it is shown how us can aid in better accumulating them at the target site by inducing vascular permeation. many of these novel imaging concepts and tools can relatively easily be translated into clinics. thus, one can expect them to play a major role in the clinical management of tumour treatments soon. mammography technique from the point of view of radiographic compression and positioning is no different from film screening and radiologists ignore this at our peril. we will discuss the danger areas at the edges and the back of the breast and help you to find lesions only seen on one view. with digital capture the technologist can view images immediately, but the temptation is for them to repeat exposures to obtain radiographic perfection on otherwise clinically acceptable images. however, the task of assessing quality cannot be left to them, as viewing conditions on the 'lower resolution' acquisition monitor in a bright x-ray room is not enough. there is now evidence from population screening programmes that poor image quality can reduce cancer detection from between % in france to % in ontario. 'quality' can be lost at keys points in the imaging chain: the x-ray beam and dose, the detector and finally display. in flanders, poor detector performance was compensated for by increasing the x-ray dose by % and by meticulous quality control. postprocessing is less well understood, but reader studies now show different algorithms will affect performance particularly for the detection and classification of micro-calcification. finally, it does not matter how good all the equipment is if we try and read the images with the lights on full. mammography is the most important breast imaging technique allowing the visualisation of masses and micro-calcifications. however, in conventional mammography the three-dimensional breast tissue is reduced to a twodimensional image. therefore, small lesions may be undetectable due to superimposed glandular tissue. digital breast tomosynthesis (dbt) has emerged as a new imaging modality to overcome this limitation. several lowdose mammographic projections are performed over a limited angle of up to degree using a standard mammography system. the average glandular dose of a tomosynthesis scan is somewhat higher compared to one-view mammography, but lower compared to a standard mammography in two projections. based on the low-dose projections, a stack of cross-sectional images covering the entire breast is reconstructed with an interslice distance of about mm. several studies have shown that tomosynthesis improves both tumour detection and the characterisation of focal masses. due to the crosssectional nature of dbt, the techniques allow on the one hand the reliable differentiation of true focal masses from summation artefacts. on the other hand, tomosynthesis improves the detailed analysis of the lesions' border to differentiate benign lesions and carcinomas. additionally, contrast-enhanced spectral tomosynthesis may allow analysing signal-intensity time curves. in conclusion, dbt adds important information to standard two-projection mammography and can replace spot compression and rolled views as problem solver for difficult cases. however, the impact of tomosynthesis on breast cancer screening has to be investigated in larger clinical trials. reading screening mammograms is not the same as reading diagnostic mammograms. the vast majority of women attending a screening programme are usually asymptomatic and do not have abnormalities associated with a malignancy. the task of a mammogram reader is to scan a mammogram and perceive any possible abnormalities that could be associated with a malignancy. in other words, we are talking about perception and not diagnosis of a finding. examples will be shown about this crucial step. in modern population-based screening programmes, we use computerised reading and recording systems. since the work flow is rather quick, we have to ensure basic steps such as identity control and see that both ris and pacs are synchronised so that decisions of screening readings are recorded in both the systems of one and the same patient. in computerised screening programmes, we use standard reports informing a woman that no signs of malignancy have been detected on her screening mammograms. in case of findings that need further assessment, there are several ways to deal with this situation. the most common way is via a phone call, but this can also be done with a letter. a screening report should be short and concise and should only inform whether we see signs of a possible abnormality that can be associated with a malignancy. extensive reports describing the contents of a breast or findings of no clinical relevance should not be part of a screening report. emergency radiology services are very much on the rise in the last decade. medical emergencies and trauma are of enormous importance and a leading cause of death in all age groups. the use of radiological imaging in emergency departments showed an exponential increase since the s. the annual growth rate in ct is - % per year depending on the institution. even for advanced level ° medical centres running an own emergency radiology unit, it is a challenge to integrate advanced radiology services in an interdisciplinary team treating patients with acute traumatic and non-traumatic emergencies. general principles: radiology and radiological imaging procedures should be integrated in an interdisciplinary team. imaging should be guided by interdisciplinary clinical algorithms or guidelines (e.g. nexus or canadian cspine rule). many clinical diagnoses relay today on a early and thorough initial radiological diagnostic workup -mostly based on mdct. emergency radiology (er) is still a relatively young subspecialty in european radiology -it deserves specific training. the lecture will cover: the development of emergency radiology and its future perspectives; the use of conventional radiography, ultrasound and mdct; logistics and management of the patient; clinical guidelines and mdct in the primary patient survey (e.g. using atls for trauma) and advanced scanning protocols for mdct. infants and children with abdominal emergencies present special diagnostic challenges because of similarities in clinical findings between various paediatric conditions. imaging is frequently a critical part of the diagnostic process, but the choice of the best imaging modality can be controversial. patient age and safety issues such as radiation exposure must be taken into consideration when planning imaging. because certain pathologies are more likely to occur in specific age groups, patient age is a primary determinant of the imaging approach. this presentation will illustrate the differences in pathology between several paediatric age groups and discuss effective imaging algorithms for congenital and developmental, inflammatory, and obstructive conditions. the optimal use of radiographs, ultrasound, ct, and mri will be discussed. pitfalls that should be avoided in ultrasound and ct will be emphasised. in diagnostic imaging, we tend to focus on radiation protection, justification and optimisation, as the primary opportunity for radiographers and radiologists to ensure the safety of patients, both adults and children; however, this ignores some other important issues. this session aims to raise awareness amongst medical imaging professionals of some key considerations in relation to a number of child protection issues. when imaging a child in case of suspected non-accidental injury, every detail of that examination from conversations with the child and their parents or guardians, technical aspects of the examination, observations during the examination and the language used following the examination need to be carefully considered. another area of medical imaging requiring attention is how we communicate any radiation risks associated with examinations or procedures with children and their parents. there is evidence that while consent processes for invasive procedures are usually in place, these often lack focus on the risks associated with a high exposure to ionising radiation. these two medico-legal areas, suspected non-accidental injury and risk communication, are commonly encountered by radiographers and radiologists and as such it is important to make sure that we operate at the highest possible professional standards and in keeping with both regulations and best practice including the production of witness statements/contemporaneous notes. on rare occasions, we may find ourselves involved in legal proceedings as a witness or expert witness and it is thus important that we are all aware of the key considerations in this regard. session objectives: . to appreciate the need for radiographers to focus on more than just the imaging procedure in paediatric examinations. . to understand the extremely vulnerable nature of paediatric patients and the need for all health professionals to pay particular attention to this. a. professional responsibilities: an international perspective m. davis; dublin/ie (michaela.davis@ucd.ie) radiographers play a vital role in imaging children with suspected nonaccidental injuries. this session will also explore the wider role that radiographers have in child protection. current legislation and its implications will be explored from a variety of countries including those outside europe. in addition, case studies will be presented focussing upon child protection issues that have arisen in the x-ray department during imaging procedures. several of the barriers to radiographers becoming involved in child protection will be explored. radiographers and their contribution to child protection will be discussed and practical examples given. learning objectives: . to become familiar with current legislation and guidelines which have particular relevance to radiographers from an international perspective. . to appreciate best practice in child protection and the need for all radiographers to be aware of this. . to learn about child protection case reports involving imaging, radiologists and radiographers. a- : b. risk communication in paediatric examinations j.l. portelli; msida/ mt (jonathan.portelli@um.edu.mt) medical imaging plays a vital role in the diagnosis and treatment of numerous medical conditions for millions of patients worldwide. however since most medical imaging examinations utilise ionising radiation, the associated risks of radiation exposure also need to be acknowledged, especially since high doses of radiation are known to cause adverse biological effects. medical imaging professionals should have a good understanding of the benefits and risks associated with the imaging examinations they perform, so as to ensure a high benefit-risk ratio for all patients undergoing diagnostic imaging procedures, and to be able to appropriately communicate such information to patients, their families and other healthcare professionals. this may be particularly more important when imaging paediatric patients, who are relatively more susceptible to the effects of radiation and receive a higher effective dose per unit of radiation when compared with adults undergoing the same medical imaging examination. indeed, following worldwide media attention about radiation incidents and their adverse effects, some parents/legal guardians may be concerned and reluctant to pursue important medical imaging examinations requested for their child, as they may have a misconception of the risks involved. in this regard, this lecture will seek to enhance awareness about the radiation doses associated with common paediatric imaging examinations; highlight the importance of medical imaging professionals having an appropriate understanding of benefits and risks associated with these examinations; and encourage such professionals to improve patient care by making use of this knowledge to have better discussions with referring clinicians, patients and their families. learning objectives: . to appreciate the frequency and radiation dose associated with certain paediatric imaging examinations. . to become familiar with the current status quo in the communication of radiation dose-related risks. . to understand best practice and potential considerations in providing accurate information to referring clinicians, patients and their families. c. what to do if you find yourself being called to give evidence m.d. viner; london/uk radiology is a powerful tool in the investigation of cases of suspected nonaccidental injury and suspicious death in children. the evidence obtained can be pivotal in bringing successful prosecutions against suspects and protecting the child and others from further harm. it is essential that evidence is obtained, recorded and presented in accordance with applicable rules of evidence in order to be admissible in court. it is thus important that those undertaking such examinations are appropriately trained, conversant with standards of evidence and maintain continuity of evidence throughout the investigation. this presentation will review rules of evidence for cases of suspected child abuse and their application to imaging examinations. it will examine the role of the radiographer and the importance of training and familiarity with legislation and guidelines that underpin forensic practice, highlighting the pitfalls that lawyers may exploit if procedures have not been followed. drawing on examples, it will discuss best practice from the radiographer's perspective, detailing the importance of continuity of evidence and maintenance of contemporaneous notes and witness statements. it will address key considerations for radiographers and others who provide evidence or act as an expert witness. imaging professionals play a pivotal role in the investigation of suspected child abuse. it is essential that this evidence is obtained, recorded and presented with regard to the applicable standards of evidence. for those called to give evidence, the importance of appropriate imaging protocols and procedures, detailed documentation and thorough preparation cannot be understated. learning objectives: . to understand the importance of contemporaneous notes and witness statements if involved in any forensic or suspected non-accidental injury examination. . to become familiar with the importance of continuity of evidence in forensic examinations. . to appreciate the key considerations if you are asked to provide evidence or act as an expert witness in court. brain tumours are the second most common neoplasms in children after leukaemia. as "paediatric brain is not just a small brain", paediatric brain tumours are not the same as tumours in adults. mri is a method of choice in their diagnosis in accordance with the principle alara in the paediatric population. by the age of approximately years, supratentorial tumours dominate; later ( - years of age), the majority of children show infratentorial neoplasms which are pilocytic astrocytomas, most often found in the hemispheres, and medulloblastomas or ependymomas -in the fourth ventricle. after the age of , both groups are diagnosed with equal frequency. cerebral hemispheric tumours include astrocytoma which is the most frequent, teratoma in neonates, and in older children atypical teratoid/rhabdoid tumour, ependymoma, pnet, ganglioglioma and dnet. in the midline, we deal with chiasmatic or hypothalamic glioma, craniopharyngioma and germ cell tumours of the pineal region. extraparenchymal tumours are uncommon in the paediatric population. apart from the location, imaging pattern in various sequences, and contrast enhancement of the tumour, advanced mri techniques are helpful in establishing the diagnosis. dwi helps to differentiate between tumours of high and low cellularity showing low adc values in case of medulloblastoma and high in astrocytoma. mrs, pwi and dti also add important information to the diagnosis. mri of the spinal canal is obligatory in case of these primary brain neoplasms that have a high propensity to metastasise to other parts of the central nervous system such as medulloblastomas, ependymomas, high-grade gliomas, cerebral neuroblastomas, and pineal region neoplasms. learning objectives: . to learn about the difference between paediatric and adult brain tumours. . to understand the imaging strategy for the paediatric population. . to become familiar with the most common paediatric brain tumours. pancreatic endocrine tumours (p-nets) include both pancreatic neuroendocrine tumours associated with a functional syndrome (functional p-nets) and those associated with no distinct clinical syndrome (non-functional p-nets). non-functional p-nets may show immunohistochemical positivity for hormones which may be produced, but not secreted, which are clinically inert and whose serum concentrations are insufficient to induce symptoms. among functional p-net, the most common are gastrinomas and insulinomas, whereas those secreting vasoactive intestinal peptide, glucagon, somatostatin and other hormones are considered together as a group called rare functional p-nets. non-functional p-nets are classified, according to the who classification, in well-differentiated neuroendocrine tumours and poorly differentiated neuroendocrine carcinomas (necs) of small or large cell type. well-differentiated neuroendocrine tumours are then divided according to a grading scheme based on mitotic count or ki index in nets-g (with a mitotic count < per high-power fields (hpf) and/or % ki index), and nets-g (with a mitotic count - per hpf and/or - % ki index). all poorly differentiated neuroendocrine carcinomas are graded g (with a mitotic count > per hpf and/or > % ki index). most pancreatic nonfunctional p-nets are well-differentiated tumours, whereas poorly differentiated neuroendocrine carcinomas are uncommon. pancreatic endocrine tumours (p-net) are divided into functional p-nets and non-functional p-nets. diagnostic imaging of these tumours includes detection, characterisation and staging. there are significant differences when dealing with functional p-nets and non-functional p-nets. in fact, in the first case the main aim of diagnostic imaging modalities is the detection of the tumour, while for non-functional p-nets the main aim is the characterisation and the differential diagnosis with other tumours of the pancreas, mainly with pancreatic adenocarcinoma. staging is also important for both tumours, but mainly for non-functional p-nets. a large number of imaging modalities are available, both "morphological" and "functional". we can arbitrarily divide the imaging modalities for the diagnosis of p-nets into three levels: the first level includes us (with contrast-enhanced us), mdct, mri and nuclear medicine techniques; the second, endoscopic us, angiography and venous sampling; the third, intra-operative us. functional p-nets are represented mainly by insulinomas and gastrinomas which are small in size in most cases. their detection is still difficult, but significant improvements have been made with us, mdct and mri, so that these tumours are detected in most cases if appropriate imaging techniques are performed. however, still, no single modality is % effective. non-functional p-nets can be differentiated from pancreatic adenocarcinoma, since they are hypervascular and usually large in size. since they are potentially malignant, they require accurate staging by imaging modalities, both morphological (mdct, mri) and functional (pet/ct) to plan surgery and chemotherapy. there is worldwide consensus about surgical treatment of resectable p-nets in the following clinical situations: proven malignancy, whenever it is possible to remove the primary tumour and at least % of liver metastases (debulking); p-nets associated with a clinical syndrome caused by hormonal hypersecretion (insulinomas and gastrinomas mainly); non-functional p-nets with a size larger than cm. vice versa, there are different expert opinions about the surgical treatment of functional and non-functional p-nets in men syndromes and in sporadic non-functional p-nets less than cm in size. in men all insulinomas should be resected, while for gastrinomas the extent of resection is patient tailored, deserving standard whipple for selected cases. in the most common sporadic small incidentalomas, where long-term prognosis is favourable, surgical treatment has to be weighted with co-morbidities and morbidities of the patient and the procedure planned. these tumours have a risk of node metastases at the time of diagnosis in less than % of patients. whenever possible, mini-invasive approach with pancreas-sparing operations should be recommended. in elderly patients, a first period of follow-up should be preferable and surgery should be offered only if the size of the tumour demonstrates increasing trends. the "acute aortic syndrome" includes three closely related emergency entities of the thoracic aorta: aortic dissection, intramural hematoma and penetrating atherosclerotic ulcer. as these conditions present with similar symptoms, cross-sectional aortic imaging is essential for diagnosis, triage and treatment planning. initial imaging features of these pathologies including typical early complications as well as their differential diagnosis will be reviewed. as nowadays several of these patients will undergo early endovascular thoracic aortic repair, an update on its indication, the different technical options and their clinical results will follow. finally, the focus will be on repair-related late complications and progression of aortic disease. imaging follow-up protocols, features of chronic complications and their interventional repair will be discussed. a. pre-therapeutic radiological evaluation j. ferda; plzen/cz (ferda@fnplzen.cz) the acute thoracic aorta syndrome covers patients suffering from acute chest pain and those injured during high-energy trauma. despite the similar symptoms, the aortic pathologies are heterogeneous, including acute aortic injury, intramural haematoma, aortic dissection, and ruptured thoracic aortic aneurysm and mycotic aneurysms. the diagnostic approaches could include the whole pallete of the imaging modalities: chest x-ray, transesopheageal echocardiography (tee), multidetector ct angiography (mdcta), mri and pet/ct. because of some disadvantages of tee, like problems with intubation in injured patients or problems of mri with seriously ill patient monitoring, mdcta has become the most used modality due to its ability of discrimination between aortic and non-aortic pathologies, and also due to its high sensitivity, specificity, availability and especially superior geometrical resolution and potential of d imaging. the proper pre-therapeutic mdcta protocol is using sub-millimeter isotropic resolution covering the entire aorta with pelvic arteries. when the pathology of thoracic aorta is found during the triple rule-out triage protocol, additional mdcta of the rest of the aorta and pelvic arteries has to be completed. ecg triggering could be an important advantage in the reduction of pulsation-derived artefact. the volume of applied contrast material used should be as small as possible to plan applications during endovascular treatment. d reconstruction of the thoracic aorta with its side branches and advanced vessel analysis have a crucial role in the measurement of the aortic diameter, diameter profile and lengths in the selection of proper stent graft type and size. they can be accommodated even in patients with small vessels and tortuous anatomy. calcified arteries can be no longer considered absolute exclusion criteria. some devices are designed to reduce the incidence of endoleaks, improving sealing capabilities by the use of a polymer to better adjust to the variable morphology of the proximal neck or to fill the aneurysmatic sac completely. more importantly, the introduction of fenestrated devices has resolved the problem of facing difficult anatomies when the aneurismatic pathology involves renal or visceral arteries. obviously, the use of these new devices requires a learning curve. beyond an increase of cost, they permit treating those patients who in the past were considered unfit candidates for evar. the evar new frontier requires performing the procedure in an outpatient fashion. a multicentre study was conducted on patients who were discharged / hours after uncomplicated stent-graft implantation. cases ( %) with access vessel complications required additional procedures and patients were hospitalised overnight. the -day readmission rate was % for access vessel stenosis or false aneurysm. from the time of the first evar procedure, significant improvements have been achieved but there is still room for further progress. the standard imaging modality after endovascular treatment for thoracic aortic pathology (tevr) is ct angiography (cta). cta enables the radiologist to assess for the patency of the endografts, the presence of any complications after tevr, and the development of any new pathology related not only to the aorta, but elsewhere above and below the diaphragm. the role of mr is reduced compared with cta, particularly in the presence of stainless steel endografts. however, good images can be obtained by mri and mra if nitinol endografts have been inserted. mr is a useful modality for follow-up in young patients who require lifelong follow-up and, therefore, limited exposure to ionising radiation. ultrasound has no practical role in the follow-up after tevr, and although plain thoracic radiographs are routinely performed by some centres post-tevr, in practice almost equivalent information can be obtained by ct. the main complications after tevr are endoleaks, type a dissection, paraplegia and stroke. the classification of endoleaks is similar to the classification for evar. type and type endoleaks require treatment by additional endograft coverage with or without supraaortic or visceral artery bypass. other treatment options include the use of branched, fenestrated endografts or the use of chimneys and snorkels. type endoleaks are less common than post-evar. similar to the evar population, intervention is mandated for an enlarging aortic diameter and involves embolisation in most cases. interventional oncology is a discipline that uses imaging-guided procedures in the treatment of cancer patients. when the malignant disease in not curable, interventional techniques such as biliary and oesophageal stenting can provide useful palliation and improve the patient's quality of life. in patients with potentially curable tumours, thermal ablation, cryotherapy and radioembolisation provide useful alternatives to radiotherapy and surgery in carefully selected patients. interventional radiologists who treat patients with cancer should practise as clinicians, participating in multidisciplinary meetings where treatment choices can be discussed with specialists from other oncological disciplines. they should involve themselves in all stages of clinical practice, including pre-treatment consultations, ward rounds and patient follow-up. radiation oncology and interventional oncology have much in common, including the potential for curing cancer using minimally invasive techniques. interventional oncologists can usefully adopt many of the principles and methods of practice in radiation oncology, including quality assurance and the collection of robust outcome measures. an alliance between these disciplines would be beneficial to clinical practice and research in both disciplines. session objectives: . to understand the contribution of interventional radiology to oncology. . to appreciate the areas of overlap between radiation oncology and interventional oncology. . to understand the need for a clinical pattern of practice in interventional radiology in oncology. the current scope and future challenges j.i. bilbao; pamplona/es (jibilbao@unav.es) together with chemotherapy, surgery and radiotherapy, interventional radiology (ir) is the fourth pillar in oncology. transcutaneous and endovascular procedures, guided by imaging, constitute a fundamental knowledge essential for the multidisciplinary management of oncologic patients. the range is wide and covers most of the needs in patient´s care. in short: obtaining samples for making an accurate diagnosis and also to know more about its specific biology (biomarkers); ir covers any possibility needed for vascular access; percutaneous needle ablation is the best alternative in selected patients; endovascular embolisation with "vehiculisation" of therapies is an outstanding method for selective treatment, and sometimes precise ablation, of different tumors; ir is a unique way to offer palliation in a wide range of tumoral complications, such as embolisation for bleeding, stenting for vein obstructions or drainage of fluid collections. there is an obvious need of continuous technical refinement. image-guided therapy is a continuous process that implies many disciplines and that includes every step, from staging through guidance of the procedure to the evaluation of the outcomes, both immediately, during and in the follow-up. future developments in ir may be focused towards the personalisation of therapies to each patient and to a better understanding of the biological mechanisms of tumour response or progression and their image expression. finally, radiologists must be actively involved in the whole process of clinical trials, from the creation of new lines to the performance of therapy until the evaluation and diffusion of the data. radiation treatment has a fundamental role in the multidisciplinary management of people with cancer. the evidence base for radiation therapy is robust and approximately % of people diagnosed with cancer will benefit from radiation treatment. most of this benefit comes from an increase in cure. the technical quality of delivery of radiation treatment is of great importance in avoiding a geographic miss and also in reducing the amount of normal tissue being irradiated unnecessarily. as well as the excellent technical delivery of any treatment offered, of paramount importance is the intelligent integration of care with attention to evidence base choices for both local and systemic treatment. multidisciplinary teams are in the best position to integrate surgery, radiation treatment, interventional oncology and systemic therapy in a planned and deliberate fashion. best practice: how to organise an interventional oncology unit a. gangi, j. garnon, g. tsoumakidou; strasbourg/fr (gangi@unistra.fr) interventional oncology is a fast growing field. interventional oncology is a clinical and technical speciality. solid clinical knowledge associated with the technical skills is mandatory for a successful practice. the technical skills should be associated to the clinical part. the knowledge of all treatment modalities and the participation of a multidisciplinary team is vital for the best practice. every single patient should be seen by the interventional oncologist and followed-up. the interventional oncologist is part of a multidisciplinary team, taking care of the patient. the team includes oncologists, radiotherapists, pain specialists, psychologists, surgeons, and anesthesiologists. the interventional oncology unit should include specialised nurses, technician, anesthesiologist, anesthesiologist nurses, and the interventional radiologist. the team should be able to take care of the patient's follow-up and eventual complications. to summarise, high-quality interventional oncology services are essential for safe and effective patient care. the technical skills of the interventional oncologist should be associated with an excellent clinical knowledge of the disease. a multidisciplinary team work is mandatory for a successful interventional team. harmonisation of training and appropriate credentialing is required for clinical services of the best quality. interventional radiologists continuously improving their skills and knowledge, patient access to safe and effective minimally invasive treatment options is increasing. whereas cardiac ct and mri are routine examinations for specialised radiologists, it has been necessary to wait for the introduction of fast scanning techniques to modify the behavior of nonspecialised radiologists regarding the interpretation of cardiac cavities. the objective of this session is twofold. first, cardiac radiologists will underline the role of these imaging modalities in the assessment of congenital heart disease but also in ischaemic and nonischaemic cardiomyopathies. in the second part of this session, chest radiologists will focus on the diagnostic information accessible at the level of the right ventricle, known to be the target of numerous respiratory disorders. two practical situations will illustrate this approach: pulmonary thromboembolic and chronic respiratory diseases. accurate assessment of the right ventricle (rv) is crucial for the management of congenital heart disease. echocardiography can provide some information, but as the rv sits directly behind the sternum and as many patients have undergone multiple operations, visualisation can be difficult. furthermore, for long-term follow-up and treatment planning, accurate measurement of ventricular size and mass, and valvular flow (tricuspid and pulmonary), and assessment of anatomy in d and d are vital. this presentation will outline the role of cross-sectional mri and ct for assessing the rv, and describing the indications, imaging protocols, technical pitfalls and relevant information for treatment planning. right heart failure in ischaemic and non-ischaemic cardiomyopathies m. grothoff; leipzig/de (matthias. grothoff@herzzentrum-leipzig.de) the role of the right ventricle (rv) has been underestimated for many years. today, we know that the rv is of importance for keeping up a sufficient circulation and that right heart failure in both ischemic and non-ischemic cardiomyopathies can result in hemodynamic compromise. rv ischemia has a higher in-hospital morbidity and mortality but post-infarction chronic right heart failure is a rare finding. as the rv has a different physiology from the left ventricle (lv), also the pathophysiology of ischemia is different here. using our standard cmr tools t weighted imaging for detection of edema and late gadolinium enhancement imaging for detection of necrosis/scar we can find that the rate of myocardial salvage is much higher in the rv than in the lv and that rv failure in the acute phase is basically caused by reversible ischemic effects. in non-ischemic cardiomyopathies like hcm and dcm changes in the rv myocardium mostly resemble changes of the lv. in arrhythmogenic right ventricular cardiomyopathy/dysplasia (arvc/d) however, there are specific findings in the rv and the role of imaging in diagnosing arvc has been strengthened by the revised task force criteria published in . cine imaging should be performed carefully for detection of rv akinesia, dyskinesia or small aneurysms as well as the planimetry of the rv. findings like the detection of fibrosis in lge imaging and the detection of fat in t imaging can provide additional information, but are not diagnostic for arvc. learning objectives: . to learn about the different ct and mri techniques for evaluating right heart morphology and function. . to understand the prognostic impact of right heart involvement in ischaemic heart disease. . to learn about the basic morphological and functional features of different non-ischaemic cardiomyopathies. right heart and chronic respiratory diseases: can ct be used as a onestop-shop? m. remy-jardin; lille/fr (martine.remy@chru-lille.fr) over the last decade, ct technology has evolved towards fast scanning capabilities and high temporal resolution, enabling improved evaluation of the heart and surrounding structures in the course of routine thoracic ct imaging. this has progressively modified the radiologist's implication in the interpretation of chest ct examinations. firstly, it is more and more common to integrate the physiological interactions between heart and lung when analysing the ct features depicted on a given examination. secondly, it is now possible to integrate cardiac functional information into a diagnostic ct examination of the chest, providing prognostic information in the management of patients with a wide variety of chronic respiratory disorders. the purpose of this presentation is to provide non-specialised radiologists with some practical guidelines in the choice of the scanning protocols and range of functional information that can be deducted from chest ct examinations. right ventricular dysfunction and levels of pressure within the pulmonary circulation are the key points of such approaches. learning objectives: . to appreciate the clinical impact of a cardiothoracic evaluation from the same ct examination. . to learn about the scanning protocols enabling such a combined analysis. . to understand its compatibility with other non-invasive modalities. rv and prognosis in pulmonary thromboembolic disease n.j. screaton; cambridge/uk (nicholas.screaton@papworth.nhs.uk) right ventricular failure is the most common cause of early death following acute pulmonary embolism and in patients with chronic thromboembolic pulmonary hypertension. echocardiography is sensitive in diagnosing and quantifying rv dysfunction. ct is the reference standard for the diagnosis of acute pe diagnosis and has the potential to provide comprehensive evaluation of the pulmonary arteries, heart and lung parenchyma. ct signs of right ventricular dysfunction which have shown predictive value for adverse outcome include: flattening of the inter-ventricular septum or bowing towards the left ventricle, reflux of contrast medium into the ivc, relative rv/lv diameter ratios on transverse and -chamber reconstructions, and ventricular volumetry as assessed both on routine non-gated ctpa and ecg-gated ct. in patients with cteph, mri permits imaging of the pulmonary vasculature and functional evaluation of the right ventricle and pulmonary circulation, enabling both morphological assessment and prognostic evaluation. learning objectives: . to learn about clinical and imaging factors associated with prognosis of thromboembolic disease. . to understand the benefits and weaknesses of different imaging modalities in predicting prognosis in thromboembolism. . to discuss the merits of comprehensive imaging evaluation in the routine diagnostic workup of suspected thromboembolic disease. multimodality imaging including ultrasound has within recent years been introduced in several anatomical regions. pet, ct and mri datasets can after an initial co-registration be fused and shown either side by side or using overlay on screen while doing real-time ultrasound; thus, the pet, ct or mri dataset is reformatted in a projection to fit the real-time ultrasound images. several methods of aligning the images are available and have been shown to have a high accuracy. image fusion involving ultrasound has been tested in phantoms, animals and in patient studies, mainly in abdominal imaging. by combining several imaging modalities, it is possible to benefit from the strengths of each modality. it is useful for characterisation of and intervention on liver lesions, abscesses containing air and lesions in areas with poor overview by ultrasound. also, the method is useful for characterisation and biopsy of pet-positive lesions, especially in patients with a history of cancer. the background of the method and clinical examples, mainly from abdominal applications, will be covered. also, perspectives and future research topics will be highlighted. contrast-enhanced ultrasound (ceus) has proved to improve the detection and characterisation of pathologies compared to conventional ultrasound, ct and mri in a number of indications. ultrasound contrast agents (uca), which are purely intravascular, do not show any interstitial diffusion or glomerular filtration like iodinated complexes or gadolinium chelates. dynamic contrastenhanced ultrasound (dce-us) allows to display the enhancement of a lesion with a high frame rate after bolus or infusion administration of uca, and to compare enhancement profiles between normal and abnormal tissue. quantification of dce-us is useful to quantify tumour enhancement and to limit intra-and interobserver variability. mathematical models with several perfusion parameters can be used. dce-us has shown to be of interest after antiangiogenic therapies as it allows an earlier evaluation of tumour response than usually done with ct and mri, which remain mainly based on the recist criteria. the common perception of d ultrasound (us) is of surface-rendered images, especially of the foetus. the same approach, enhanced by new transducer technology and software, can be used to take ultrasonography to another level in general body imaging, providing different image presentations and working practices similar to those of ct and mr. the technology involved in a us volume acquisition was represented initially by mechanical d transducers, and more recently by purely electronic, matrix probes. this latest technology allows bi-plane real-time acquisitions, fast switch to volume acquisition up to degrees, and d acquisitions with acceptable frame rates. protocols have been described for major abdominal and retroperitoneal organs, with fast optimisation of acquisition settings. the technology supports contrastenhanced ultrasound (ceus). after volume acquisition, interpretation for volume us datasets requires a post-treatment phase, including multi-planar reconstruction, multislice imaging, and volumetric analysis. virtual cystoscopy and volume ceus quantification are new options. the implementation of this new, efficient us modality in a radiology department supposes deep changes in the daily practice with delayed post-treatment reporting times. it should represent a more effective way for comparison between different examinations and a better communication tool with clinicians. the proven and potential benefits, in terms of clinical accuracy, training, workflow and overall efficiency, will be discussed. the purpose of this lecture is to emphasise some pitfalls in liver imaging. morphologic changes in the liver are usually attributed to chronic liver disease where liver cirrhosis represents the most important cause. however, noncirrhotic diseases may also induce atrophy-hypertrophic changes of the liver. the most common mechanisms are related to venous obstruction (either portal or hepatic venous) and biliary obstruction. multidetector ct and mr imaging are essential to highlight these abnormalities. when dealing with liver tumours, the most important question that has to be solved is tumour characterisation. yet, it is often difficult to assess whether a large tumour is intra-or extrahepatic. imaging findings that might be helpful will be shown. last, some liver lesions can mimic liver tumors. vascular disorders and focal fatty changes or focal fatty sparing are the most common causes. some other conditions can be also mimickers and such cases will be shown. the technique and the anatomy of the bile duct and pancreatic duct will be described. the anatomical variants of the biliary duct and the pancreatic duct system will be analysed and their possible role in generating diagnostic imaging pitfalls described. strategies to avoid pitfalls in diagnostic imaging of the bile duct and the pancreatic duct system will be illustrated, considering the possible source of pitfalls. diagnostic imaging findings of different diseases involving the biliary ducts and pancreatic duct system will be illustrated, as well as the diagnostic imaging criteria useful for the differential diagnosis. over the last two years, the number of ct and mri units has increased by %, and modern pet centers and leading centers for radiotherapy have been built. we can identify an approximately % increase in the number of radiological research projects--mainly ultrasound, ct, and mri investigations. during the last ten years the total number of vascular interventions and mr studies has increased by ten times, four times for ct, and the annual growth of us and x-ray investigations is about %. unfortunately, these technical innovations are not always accompanied with pacs and ris installations; this is a reason why all advantages of digital equipment are not achieved. we see rapid development of private medicine; new private hospitals and ambulances have grown by more than % and investment companies are looking for new projects in this field. however, the professional education of radiologists is not enough to work with new diagnostic equipment. this is a reason why in we will start a pilot project to create a new system of continuing medical education in clinical medicine, including diagnostic radiology. we hope, this pilot project will increase the efficiency of clinical practise. differential diagnosis (dd) including tumour and non-tumour brain pathologies is an integral part of neuroradiological diagnostics and in some cases is a challenge to experienced neuroradiologists. more than patients with different cns pathologies have been investigated in the department of neuroradiology, burdenko neurosurgical institute since . in the vast majority of the cases, there were tumours. we used deconvolution method for bv, bf, mtt and ps maps reconstruction on commercially available software (perfusion . . , . protocols, adw, ge). ct perfusion has demonstrated high level of information in determining the degree of malignancy in glial brain tumours. a strong correlation between bv and ps and tumour degree has been demonstrated. the use of ct-pwi makes it possible to successfully carry out dd between radiation necrosis and tumour progression. some histological types of brain lesions are characterised by unique haemodynamic properties which enable using this method as non-invasive "biopsy". ct-pwi has been especially useful in dd of the skull base region, bone and extracranial pathologies. the absence of artefacts created by the skull base bones makes this method especially valuable as compared to the mri t *-and asl-perfusions. ct-pwi makes it possible to successfully differentiate tumours and a number of non-tumour cns lesions including demyelinating diseases, infectious and non-infectious granulomas, etc. ct-pwi is a powerful method for differential diagnosis of brain pathology. t-pwi can provide unique functional information regarding tumour pathophysiology and haemodynamics, which are not available with routine ct and mri. learning objectives: . to become familiar with the technique of brain perfusion ct. . to understand the added diagnostic value of perfusion ct. . to learn about input of ct perfusion for assessment of treatment. interlude: development and use of web-based teleradiology in russia o.s. pianykh; newton highlands, ma/us (opiany@gmail.com) the largest territory in the world, russia, presents a perfect teleradiology use case of extremely unevenly distributed health care, concentrated in moscow and spread thin outside over an enormous area and sparse population. this unevenness deeply affects the quality and availability of radiological services, and cries out for efficient and practical teleradiology solutions. however, the same unevenness in medical expertise, financing and it infrastructure frequently acts against the advancement of teleradiology projects, blocking their natural progress. as a result, technical, financial, educational and clinical matters, interleaved into the nascent russian teleradiology ventures, require serious and predictable solutions, based on analysis and adaptation of already developed, internationally proven strategies. currently, russian teleradiology simply takes advantage of whatever is available to make its progress better, but needs to develop itself up to the tasks it faces. tuberculosis in russia: a challenge for a national radiological service i.e. tyurin; moscow/ ru (igortyurin@gmail.com) approximately, . billion people worldwide are infected with mycobacterium tuberculosis, of which million are active cases. the reasons for resurgence of tb infection include the hiv epidemic, a rise in reactivation disease in the elderly, a growing migrant population and spread of drug-resistant strains. the incidence of atypical tb infection is also on the rise, of which mycobacterium avium intracellulare is the most important human pathogen. the pathologic form of the pulmonary infection depends on the sensitivity of the infected host and is classified as primary or postprimary. primary tb pattern represents infection resulting from recent contact with the pathogen. postprimary tb pattern results from reactivation of a dormant focus within the lungs. thoracic tuberculosis produces a broad spectrum of radiographic abnormalities. the radiological patterns had been described as parenchymal, airway, vascular, mediastinal, pleural, and chest wall lesions. common causes of a missed diagnosis of thoracic tuberculosis are failure to recognise hilar and mediastinal lymphadenopathy as a manifestation of primary disease in adults, overlooking of minimal productive lesions or reporting them as inactive, and failure to recognise that an upper lobe or superior segment of lower lobe mass might be tuberculosis. in aids patients, the imaging features depend on the degree of immune suppression. a pattern of postprimary tb is also usually seen among patients with decreased immunity due to alcoholism, renal failure, diabetes mellitus, ageing, malignancy, and renal and cardiac transplantation. learning objectives: . to understand the scope of the problem with tuberculosis screening in the russian federation. . to learn about the changing imaging patterns of thoracic tuberculosis. . to appreciate the current role of imaging in the differential diagnosis and follow-up of this pathology. interlude: mystery of denisov's cave and paleoradiology m. mednikova; moscow/ ru (medma_pa@mail.ru) this study focuses on the anatomical identification of fossil bones from altai highland in southern siberia, their radiological description, and parallels among other hominin fossils. methods describe the small and isolated tubular bones. we used microct versaxrm- by xradia inc. and digital microfocus x-ray. denisova cave is the best studied paleolithic cave in north asia. archaeological data suggest that about thousand years before the present, a group of humans migrated to southern siberia from the west. their descendants had lived in the altai mountains in relative isolation for thousand years. the taxonomic status of the early inhabitants of southern siberia was clarified thanks to palaeogenetic studies. it was concluded that the altai hominin was genetically twice further from modern humans than were neanderthals. phylogenetic analysis suggests that this clade diverged from that ancestral to humans and neanderthals very early -about billion years ago. descendants of this clade apparently survived in the altai refugium until the period - thousand years ago. due to structural patterns of preserved pedal and manual phalanxes, humans from denisova cave could be expected to be the bearer of "archaic" traits. the extraordinarily thick walls of tubular bones in adults reflect a high level of occupational stress. radiological volumetric microscopy of the denisovan girl phalanx gave evidence for the histological estimation of the developmental age. radiological methods seem to be the perspective approach in the study of rare and fragmentary fossil human remains, differentiating between archaic and modern humans. advances in imaging of pancreatic masses g.g. karmazanovsky; moscow/ru (karmazanovsky@ixv.ru) modern diagnostic radiology allows not only to detect pancreatic tumours, but also to carry out their differential diagnosis with high accuracy. this is especially important because many benign tumours of the pancreas do not require immediate surgery. at the same time, many benign tumours may become malignant. the differential diagnosis of the pancreatic cystic lesions using endoscopic ultrasound and magnetic resonance pancreaticocholangiography has now become particularly accurate. contrast enhancement allows detecting signs of tumor invasion into the main blood vessels and adjacent organs. modern developments of radiology help to use extended operations for malignant tumours and sparing surgery with minimal removal of intact functioning pancreatic tissue in benign tumours. learning objectives: . to become familiar with major types of pancreatic masses and their radiological manifestations. . to understand approaches to the selection of diagnostic pathways in pancreatic tumours. . to become acquainted with new developments in diagnostic imaging of pancreatic tumours. interlude: an artistic view of ct a.l. yudin; moscow/ ru (prof_yudin@mail.ru) radiology is a specialty based largely on figurative perception of diagnostic imaging. the radiological picture of many symptoms reminds one of objects and environmental phenomena (for example, vanishing lung, draped aorta, geographic liver, porcelain gallbladder, dirty fat and others). training of creative mentation promotes lasting assimilation of educational material at the associative level. metaphorical thinking skills can also be improved by searching for amusing fragments in radiological images similar to the objects in everyday life; animals, cartoon characters, and others. the essential help in the study of the physical foundations of radiology and functional capabilities of workstations provide the execution and study of x-ray films, ct or mri of fruits and vegetables, flowers, various artifacts and other objects in the environment. eventually, it is just beautiful. ultrasound of the breast is an established imaging technique with several advantages. it has been used to differentiate solid from cystic breast lesions for a long time. it has also been increasingly used to visualise small lesions in mammographically dense breasts and is the initial imaging method of the breast in young women. the features of the lesions that need to be analysed are: shape, relation to anteroposterior to laterolateral diameter, margins, internal structure and absorption of sound (distal acoustic phenomena). color doppler enables the thorough assessment of flow within the breast lesions. elastography is established as a useful method to differentiate breast lesions on the basis of their stiffness, which can be quantified. the accurate staging of axillary disease is an important aspect in the management of patients with breast cancer. it is recognised, however, that underlying malignancy can be found in lymph nodes that appear morphologically normal. a variety of morphological features that may be seen in pathological nodes have been described (cortical thickness of > . mm and absence of a fatty hilum). the automated breast ultrasound technique has been known for a long time, but with recent improvements now may play an important role. the automated breast volume scanner (avbs) employs frequencies of - mhz. avbs has various major advantages, such as being independent of the experience of the operator as well as supplying consistent and reproducible results. additionally, for the first time it has become possible to obtain coronal images of the complete breast, which greatly assists in the planning of surgical interventions. to study the technical basics in breast ultrasound (us), a breast scanning phantom helps to understand different us settings of greyscale imaging. typically, breast us work has a linear array transducer with a minimum field of view of . cm and a high frequency of minimum mhz. a broadband transducer (e.g. - mhz) with different bands is preferred. in the central axillary area, a lower frequency band offers a deeper penetration depth. high frequencies improve the tissue contrast and optimise the axial and lateral resolution. the transmitter focus is adapted to the region of interest. highly sensitive color doppler detects slow flow rates ( cm/s and less). for frequencies and power settings used in diagnostic greyscale and color doppler studies, no safety limitations have to be considered. breast us is mainly used as a second-line imaging tool to mammography (mg). a correlation between both imaging modalities is crucial. skin markers and additional mgs may help to solve the correlation problem. in bi-rads or us lesions with unclear correlation in mg, a marker placement after us-guided large-core needle biopsy helps to clear the problem. the pathomorphological growth pattern of a lesion is the basis to understand different imaging aspects with different modalities. second-look us after mri is an important correlation of lesions, which were first visible with mri. understanding the complementary imaging criteria will help to reduce the number of false-positive results. a practical imaging approach to brain and neurovascular trauma will be reviewed, with an emphasis on understanding the correlation between pathophysiology and imaging signs. guidelines on when to order acute neurovascular studies will be presented. special technical considerations to help optimise ct and mr imaging protocols for suspected brain injury, cns haemorrhage, and arterial dissection will be discussed. neurological emergencies are often associated with high morbidity and mortality, and thus require prompt diagnostic and therapeutic action. nontraumatic emergencies may however have a subacute onset, and radiological signs may be subtle, which can lead to delay in diagnosis and treatment. since clinical features are often nonspecific, the radiologist may be the first to point the clinician in the direction of the correct diagnosis. it is therefore of great importance that the radiologist is aware of and familiar with the various imaging findings, on both computed tomography (ct) and magnetic resonance imaging (mri), of non-traumatic neurological emergencies. these include vascular, infectious and inflammatory diseases. commonly encountered emergencies are ischaemic and haemorrhage stroke, venous thrombosis, arterial dissection, abscess, acute disseminated encephalomyelitis (adem), and encephalitis. radiological findings in rarer diseases may mimic those in the more commonly occurring diseases, but need to be correctly interpreted as therapeutic strategies and prognosis may be entirely different. such entities include for instance posterior reversible encephalopathy syndrome (pres), reversible cerebral vasoconstriction syndrome, susac's syndrome, and status epilepticus. furthermore, initial findings of (impending) complications of brain disease, such as hydrocephalus and herniation of brain structures, may be subtle, while early recognition allows for prompt and adequate intervention. finally, diagnostic and therapeutic interventions performed in an emergency setting may interfere with the diagnosis and interpretation of clinical and imaging findings. associated limitations and pitfalls therefore need to be recognised to avoid false-negative or false-positive diagnosis, respectively. learning objectives: . to learn about the modalities (ct/mri) and protocols for non-traumatic neurological emergencies. . to learn how to diagnose the main non-traumatic neurological vascular and non-vascular emergencies. . to become aware of the pitfalls and limitations of clinical presentation and imaging findings in non-traumatic neurological emergencies. the current century has brought with it new challenges in radiation protection, many of which follow the development of mdct which greatly enhanced diagnostic capabilities and substantially reduced scanning time to just a single breath-hold for chest ct, making ct scanning both patient-friendly and the physician's preferred tool. ct has also been added to nuclear imaging systems to create hybrid scanners (pet/ct; spect/ct image gently years later: lessons learned in radiation protection for children. the alliance for radiation safety in pediatric imaging was founded in to promote radiation protection for children worldwide through awareness, education and advocacy. comprising over medical professional societies and agencies, the alliance seeks to change practice locally for improved paediatric patient safety in medical imaging. since its inception, the alliance has had campaigns in ct, diagnostic and interventional radiology, nuclear medicine, digital radiography and an outreach campaign to parents and educational summits. yet, there is still work to be done. there is a need for parents and patients to receive information about their imaging test prior to the performance of imaging procedures, particularly those involving ionising radiation. the paediatric medical imaging community should work tirelessly with the manufacturers of imaging equipment to tailor equipment for children to optimise the study and ensure that the users of the equipment are knowledgeable in its safe use in children. the routine display of size-specific dose estimate on ct scanners will better estimate patient dose for quality improvement within facilities and in dose registries. finally, there is a need for diagnostic reference levels in paediatric radiology. this presentation will update the audience in progress to date regarding these challenges. the image wisely programme continues to move forward, guided largely by its steering committee, composed of two representatives each from acr and rsna and one representative each from aapm and asrt. the image wisely website (www.imagewisely.org) has educational content for imaging professionals (radiologists, medical physicists, and technologists) and referring physicians. a patient tab features links to radiologyinfo.org and the image gently website. a tab for "my equipment" links visitors to radiation dose-related 'microsites' developed by ge, hitachi, philips, siemens and toshiba. as of august , , , individuals, facilities and associations have taken the image wisely pledge. to attract traffic to its website, image wisely has recently instituted 'news' and 'top reads' sections on the splash page highlighting recent articles and announcement, highlighting breaking news coverage and journal publications on relevant topics. in , content on nuclear medicine procedures was finalised, completing two of the three targeted foci of the initial action plan_ct and nuclear medicine. in october , a launch meeting will initiate the process of developing content for the third target area, fluoroscopy. in and , image wisely has concentrated on efforts to engage referring physician communities, with particular interest in paediatricians and emergency medicine physicians. progress has been slow, but has recently gained traction with emergency medicine in the form of radiologist participation in a symposium, diagnostic imaging in the emergency department: a research agenda to optimise utilisation, organised by the society of academic emergency medicine. learning objectives: . to learn about the image wisely campaign for radiation protection of adults. . to understand the lessons learnt while communicating messages. lessons from a national approach to patient safety in radiation protection p. cavanagh; taunton/uk (petecavanagh@gmail.com) in considering a european campaign on radiation awareness, it is useful to gain information from similar campaigns in health care. there have been a number of national patient safety campaigns based on the institute for healthcare improvement (ihi) , lives campaign initiated in the usa in . following the success of this, a similar campaign was launched in england. part of this process included an analysis of the success of the programme and the key interventions and drivers that were considered essential at both the national and local level. these included strong leadership, . bone (pseudo)-tumours: the majority of bone tumours can be detected on plain radiography. the age of the patient, location of the tumour in a bone and history of pre-existing bone abnormality should be included in determining the likely diagnosis. careful analysis of pattern of bone destruction, periosteal reaction and matrix mineralisation allow for characterisation of most bone tumours and differentiation from pseudotumours. ct may be useful for osteoid osteoma and mri is the best imaging technique for further diagnosis and staging by displaying tumour composition and extent of bone marrow involvement, including skip lesions, presence and extent of extraosseous soft tissue mass, and involvement of neurovascular bundle, muscle compartments and adjacent joint. this allows to find the best biopsy and surgical approach. to evaluate local control of disease and for detection of local recurrence, mri is usually the best imaging technique. . soft tissue (pseudo)-tumours: the majority of soft tissue tumours can be detected on ultrasound. except for cysts and subcutaneous lipoma, mr will be the next imaging technique for diagnosis, local staging, and eventually to find the best biopsy and surgical approach. the age of the patient, location of the tumour and careful analysis of the signal characteristics on mri allow for characterisation of many soft tissue tumours and differentiation from pseudotumours. plain radiography and ct may be useful for detection of calcifications and myositis ossificans. to evaluate local control of disease and for detection of local recurrence, ultrasound and mri are the best imaging techniques. in this honorary lecture, i will review a number of personal impressions that may contribute to the successful performance of research projects in cardiovascular radiology. also, issues and obstacles in performing successful research in the field of radiology will be considered. key factors for success include guidance by inspiring mentor, genuine interest and motivation, some creativity and serendipity. radiology has developed tremendously with many innovative modalities over the last years, providing ample opportunity to participate in clinical research. potential issues that should be addressed are subspecialisation, turf battles, competition from other clinicians/ imaging specialists and adequate training in radiology research. the field of imaging research includes traditionally more clinically oriented research by focussing on developing and testing of new modalities and applications, now becoming more basic by including molecular imaging, population imaging, outcome studies and alike. silicone-filled implants (single or double lumen) require a dedicated sequence where silicone exhibits a high signal intensity, whereas signals from fatty and fibro-glandular tissues are cancelled. diagnosis of intracapsular rupture relies on the detection of a linguine sign (direct sign) or multiple "key-hole" and/or " teardrop" images (indirect signs). extracapsular rupture is defined as the presence of silicone outside the capsule (breast, soft tissues, lymph nodes). breast reconstruction after mastectomy may involve autologous tissue flaps (skin, fatty tissue, muscle and fascia). the transferred muscle is inserted in front of the major pectoralis muscle and enhances after contrast medium injection. the most common benign findings seen in such reconstructed breasts and also after a breast-conserving treatment are oedema, seroma, haematoma, fat necrosis and fibrosis. diffuse thickening of the skin and trabeculae is usually observed the first year after completion of radiation therapy. small focal areas (less than mm) of non-mass-like enhancement and thin linear or rim enhancement at the lumpectomy site can all be expected findings. these benign findings decrease progressively, but residual enhancement may be observed in % of women at or more years. in patients who were switched from tamoxifen to aromatase inhibitors, a stronger or re-appearance of a background enhancement may be observed. before classifying such findings as benign or probably benign, clinical examination and standard imaging data must be taken into account. highly suspicious lesions in such patients are similar to bi-rads lesions in the nontreated breast. prevention strategies for women at high lifetime risk of breast cancer are usually adapted to the individual woman's risk of breast cancer. women at high risk (brca mutation carriers or women with a % likelihood of brca-heterzygosity) are offered primary prevention, which includes chemoprevention by tamoxifen or surgery (risk-reducing mastectomy and/or salpingooopherectomy, where the latter also helps reduce the risk of subsequent breast cancer by %). if women opt for secondary prevention (active surveillance/screening), this has to start at at the latest. mri is considered a compulsory component of all programs worldwide. due to the very limited added cancer yield of mammography in this subset of women, and because of the still unsettled issue of possibly increased radiosensitivity of brca-mutation carriers, mammographic screening is not recommended until the age of . there is broad agreement that (at least) an annual screening is important; especially for brca mutation carriers, screening every months may be more appropriate. we recommend interleaved screening, with annual mri and annual ultrasound (± mammography) phase-shifted by months. there are important differences between brca -and brca -associated cancers in terms of age distribution and incidence rates, histologic features, but also specific imaging features which must be known by radiologists. mri screening protocols must be adjusted to the specific needs of this (usually very young) screening population. this presentation will review the current evidence with respect to screening of women at high familial risk of breast cancer and will provide information on all aspects mentioned above. chest trauma is directly responsible for % of all trauma deaths and is a major contributor in another % of all trauma mortality. blunt trauma, accounting for % of chest injuries, is the third most common site of injury in polytrauma patients. plain radiographs still have a role in recognition of some acute thoracic pathology that requires immediate further management, either diagnostically and/or therapeutically, such as tension pneumothorax, major transdiaphragmatic herniation, large hemothorax or obvious mediastinal hematoma. mdct of the chest is now typically included in a whole body scan with iv contrast to facilitate rapid diagnosis on polytrauma cases using less radiation than selected segmental scans. mdct is the well-proven diagnostic gold standard for chest injury evaluation. the major advantages of mdct over other modalities include identification of active bleeding, direct signs of trachea or esophageal injury, direct evidence of major arterial vascular injury, such as pseudoanurysms, pneumo and hemopericardium, location and extent of lung contusion and laceration, and assessment for thoracic spine, shoulder girdle and rib fractures. diaphragm injuries are well depicted by mdct, especially on the left by identifying both the torn diaphragm edges, herniation and constriction of abdominal contents at the level of the torn diaphragm (collar sign), and direct contact of herniated structures with the posterior chest wall (dependent viscera). tracheal injuries are suggested by diffuse and progressive pneumomediastinum, dilated tracheostomy cuff, ectopic endotracheal tube, and direct connection of mediastinal air with the trachea lumen. ct-angiography eliminates the majority of indications for diagnostic catheter angiography. pulmonary symptoms such as chest pain, shortness of breath or wheezing are common non-traumatic symptoms prompting er visits. because clinical symptoms are very non-specific, imaging plays a major role in differentiating life-threatening from less severe diseases and forming a diagnosis. the chest radiograph remains the first imaging despite its limited sensitivity for certain diseases and being prone to inter-observer variability. comprehensive cardiothoracic ct examinations using most modern ct equipment are well evaluated in their diagnostic accuracy to determine the presence of vascular life-threatening events such as aortic dissection, acute coronary disease or pulmonary embolism. protocols and literature evidence will be discussed. the main focus of the course, however, will lie on the analysis of pulmonary ct findings and its contribution to the differential diagnosis of pulmonary emergencies (e.g. pneumonia, oedema, pneumothorax, exacerbation of fibrotic or obstructive lung diseases) and how to further integrate imaging findings, laboratory findings, patient history and clinical information to tackle the differential diagnosis. imaging findings will be discussed by clinical case studies, key findings and also overlapping morphological features of other differential diagnosis will be discussed side by side, illustrating the options and also limitations of imaging findings. mentor was first introduced in greek mythology, and in its modern usage first recorded by a french writer, françois fénelon, in . since then, this concept has been widely used in the development of mentoring, mentorship and more recently of mentees, with an overarching principle based on accompanying, sowing, catalysing, showing and harvesting. this concept remains today as a fundamental one in all teaching activities and implies generally a close contact between the mentor and the mentee. however, the development of internet with concomitant e-learning capabilities, which are hugely increasing, raises the very challenging necessity to re-think this concept and to adapt it to this new era. the esr is very concerned by this endeavour in the context of the striking development of esor, on one hand, and with the launching of its e-library, on the other. n. gourtsoyiannis; athens/gr (gournick@gmail.com) the presentation "esor in action" gives an overview on all esor activities in and . a detailed report on the esor courses and teaching programmes in is presented, containing statistics about the number of course participants and of scholarship and fellowship grants. additionally, all educational activities in , including dates, venues, topics and local organisers, are announced. evidence-based radiology: the basics evidence-based medicine (ebm) was derived from clinical epidemiology during the last century. it "is the conscientious, explicit, and judicious use of current best evidence in making decision about the care of individual patients" (dave l. sackett, ) . this means integrating ( ) individual clinical expertise with ( ) the best available external evidence from systematic research and ( ) patient's values and preferences. when considering evidence-based radiology (ebr), the needed expertise is not only clinical, but also technical, while ( ) radioprotection issues assume a relevant value according to the "as low as reasonably achievable" (alara) principle. ebm (and ebr) can be practised according to the "top-down" model, when using guidelines issued by governmental, professional, or academic bodies, or the "bottom-up" model, when local physicians ( ) formulate an answerable question, ( ) search for the best evidence; ( ) appraise critically; ( ) apply findings to practice; and ( ) evaluate the performance. a hierarchy of radiological studies has been established according to the investigated matter: . technical performance; . diagnostic performance; . diagnostic impact; . therapeutic impact; . patient outcome; . societal impact. different degrees of recommendations are based on different levels of evidence, with experts' opinion as the lowest level and meta-analyses of high-quality homogeneous studies and multicenter studies being the best level of evidence. to practise ebr, radiologists should be familiar at least with basic statistics and epidemiology as well as of methods for study design, thus recognising the principal types of bias which can limit the value of published studies. learning objectives: . to understand the historical development and general principles of evidence-based medicine (ebm) with its top-down and bottom-up approaches. . to learn how to apply ebm principles to radiology (evidence-based radiology, ebr) and to illustrate the main reasons for the relative delay in developing ebr, including the challenges of fast technological innovation in medical imaging. . to learn about the relationship between levels of evidence and the strength of recommendations for diagnostic imaging and interventional radiology. . to learn how radiologists could play a stronger role in building the evidence in favour of diagnostic imaging and interventional radiology, taking the patient's interest as the primary aim of clinical practice and science. education in research: action plan j. hodler; zurich/ch a survey regarding education in research has been commissioned by the esr and published in insights imaging ( ) : - . the results indicate that education in research, associated with career models, needs promotion. the research committee of the esr is working on possible future steps to promote research through education. several problems will have to be addressed, most notably inhomogeneity within europe, motivation issues, illdefined career paths as well as funding issues. several stakeholders will play an important role, including scientific societies, although this is the core business of academic radiology chairmen. most young professionals starting a career in academic institutions are eager to expand their professional and personal aptitudes. an effective way to support young professionals in this regard is by mentoring. mentorship is a relationship where the mentor supports a junior faculty member (mentee) in personal and professional development. the mentee benefits from early determination and initiation of career-relevant steps, greater productivity in research and publishing activity and developing techniques in networking and collaboration. moreover, the mentee gains perspectives on how his/her discipline operates academically, socially and politically, improves self-efficacy and deals more confidently with challenging intellectual work. the mentor, a knowledgeable, experienced and regarded person, benefits from fresh ideas, energy and curiosity of mentees, as well as through acceptance as a mentor an enhanced status in the department. it is a trusted relationship of dialogue, learning and challenges, based on mutual interests. however, the mentor is not automatically the best friend or the sole exclusive advisor of the mentee. more and more academic medical institutions offer facilitated mentoring programmes, which help the mentoring pairs to create an effective mentoring relationship to accomplish the desired skills. evaluation of faculty mentoring programmes in academic medical institutions demonstrates the importance of mentoring in research and academic development as well as in patient care. especially in radiology where over many years there have been challenges in recruiting and retaining talented young professionals, mentoring is considered as an essential way for preparing the next generation of scientific and intellectual leaders. pitfall: "a hidden or unexpected danger or difficulty". imaging methods can provide an extraordinary amount of useful data to specialists treating head and neck (cancer) patients. it is crucial that these data are used to full advantage of individual patients. the most important factor in this process is mutual cooperation between the physicians in charge of patient care and the diagnostic imaging specialist. pitfalls in the head and neck may present in various ways: normal variants may look like disease, incidental findings are frequently encountered and suboptimal technique may obscure important findings. moreover, many pitfalls are directly related to technical errors. the presentation aims to familiarise general radiologists, who have an interest in head and neck imaging, with common pitfalls encountered on ct and mr studies focussing on the neck. in an interactive setting, examples from daily practice will be discussed. imaging of the skull base and maxillofacial skeleton requires a meticulous imaging technique and a good knowledge of normal anatomy and possible anatomical variants. asymmetry in the pneumatisation of the paranasal sinuses, skull base or temporal bone is a common reason for misinterpretation. for example, hypoplasia of the maxillary sinus may be misinterpreted on conventional radiography as maxillary sinusitis or orbital blowout fracture, depending on the context; asymmetric pneumatisation of the petrous apex or mastoid bone may mimic, respectively, a cholesterol granuloma and fluid effusion in the non-pneumatised side on mri. vascular variants may also cause interpretation problems. for example, turbulent flow in a large jugular bulb may mimic a jugular foramen tumour on mri. variants in the vascular plexus surrounding the trigeminal or facial nerve may occur, and cause asymmetric findings on mri, possibly mimicking neuritis or perineural tumour spread. incomplete maturation or arrested development of skull base structures may also cause confusion. examples are the cochlear cleft, not to be confused with otosclerosis, or arrested pneumatisation of the sphenoid sinus, possibly mimicking a tumoural lesion in the central skull base. to avoid problems, one should keep in mind the existence of such variants, while correlating the imaging findings with the clinical problem; in some cases, an additional imaging study may be needed to exclude a pathological process more confidently. this short -min introduction by the session chairman will give an overview of the current state of cardiac computed tomography (ct) for diagnosis and prognosis. in addition, the introduction will provide the basis for the subsequent dedicated lectures covering low radiation dose, diagnostic accuracy, and incidental findings in cardiac ct. what are the protocols with the lowest radiation dose in clinical practice? j.-f. paul; le plessis robinson/fr (pauljf@ccml.fr) the radiation dose delivered for coronary ct angiography using retrospective gating is high, because only a small part of the total radiation delivered is used for the reconstruction of the image in the retrospective mode. on average, % of the radiation burden is used to reconstruct one phase of the cardiac cycle. ecg-triggered tube current modulation allows reducing the nominal dose by up to % in the systolic phase. another approach for reducing radiation exposure is to use prospective, sequential acquisition, with this approach being associated with % dose reduction compared to the spiral retrospective acquisition. exposure time may be also shortened by an increase of pitch, in particular on the dual-source ct. however, applicability to each dose-sparing technique depends on the heart rate and its regularity. dose-sparing strategies should be modified in case of high or irregular heart rate and radiation dose reduction may be less effective. for coronary ct angiography, individually weight-adapted protocols have been successfully applied, by adjusting mas to patient's weight. more recently, settings of kv and even kv have been successfully used for cardiac studies, especially in slim patients or children. reduction of exposure time and individual adaptation have an additive effect on dose reduction: it has thus been possible to scan down under msv in selected, slim patients, using this combined approach. in many cases, using the latest technology, radiation dose may be lower than the mean radiation dose associated with conventional angiography. learning objectives: . to understand various protocols to lower radiation dose in cardiac ct with regard to the clinical situation and the risk of image quality impairment. . to learn how to tailor the radiation dose level to each patient in terms of morphology. . to become familiar with modulating exposure windows in regards to a patient's ecg. how accurate and prognostically valid is coronary ct angiography? f. cademartiri, e. maffei; monastier di treviso/it (filippocademartiri@gmail.com) the role of cardiac computed tomography in cardiovascular diseases is becoming increasingly important. cct is a robust and reliable investigation for the detection and exclusion of significant coronary artery disease. in addition, cct provides reliable noninvasive information concerning coronary plaque burden (severity, distribution, type), morphology and function of left and right ventricle, valves, aorta, and so forth. in the past years, the prognostic role of cct has been established. the prognostic information that can be extrapolated from cct is very diverse and most of it is still under investigation. there are converging evidences that cct can stratify cardiovascular risk better than conventional clinical methods, especially in symptomatic patients. for asymptomatic individuals, the role is still controversial since coronary calcium score alone has a very important role already. several aspects of cct information seem to have importance for prognostic stratification: plaque presence, obstruction, plaque type and distribution. more recent studies have started showing the relationship between cct and pharmacological treatment. this is a very new and appealing topic, since the evidence is towards the fact that the most effective pharmacological treatment (e.g. statin) should be restricted to patients with at least some nonobstructive cad as detected by cct. this might become a revolutionary concept in cardiovascular medicine with several implications. learning objectives: . to learn about and integrate the prognostic information from coronary ct angiography into conventional methods. . to learn about coronary ct angiography and conventional methods. . to appreciate the prognostic impact of coronary ct angiography and conventional methods. incidental findings in cardiac ct: how to report and proceed s. leschka; st. gallen/ch (sebastian.leschka@kssg.ch) rapid advances in computed tomographic (ct) technology have facilitated the widespread use of ct for cardiac imaging worldwide, and can be considered nowadays an essential part of the clinical workup of patients with suspected coronary artery disease. albeit that the performed ct study is focussed on the cardiac structures and the coronary arteries, adjacent noncardiac structures are in the field of view whenever a patient undergoes cardiac ct to quantify coronary calcium or to perform a noninvasive coronary angiogram. in addition, the location of the heart near other anatomic regions, including the lungs, mediastinum, upper abdomen, and bones, necessitate that these structures are included when acquiring coronary ct angiographic images. several publications have demonstrated a high frequency of incidental findings on coronary ct angiography. the availability of this information is part of an ongoing clinical debate. while some radiologists believe that every image should be interpreted completely, others believe that examining noncardiac structures reveal too many incidental findings of uncertain clinical significance, which may harm because additional testing increases risk, cost, and patient anxiety. the aim of this lecture is to separate solid facts from opinions and beliefs. the diagnosis of pulmonary embolism (pe) is usually established by a combination of clinical assessment, d-dimer testing, and imaging with either pulmonary ventilation-perfusion (v/q) scintigraphy or pulmonary multidetector ct angiography (ctpa). v/q scintigraphy is a functional imaging method using isotopes for the visualisation of ventilation and perfusion distribution in the lungs. in recent years, the imaging techniques for diagnosing pe have improved. many nuclear medicine centres have adopted the single photon emission computed tomography (spect) technique as opposed to the planar technique when diagnosing pe. the introduction of -dimensional v/q spect technology instead of -dimensional planar v/q scintigraphy has resulted in fewer indeterminate results and a higher diagnostic value. the latest improvement is the addition and combination of a low-dose ct without contrast to the v/p spect (spect/ct) technique. the spect/ct in combination has been compared to ctpa, planar scintigraphy and spect alone, and one study has shown that spect/ct in combination had the highest diagnostic accuracy for pe diagnosis. spect in combination with low-dose ct without contrast enhancement is superior, especially for small subsegmental emboli; however, consensus is lacking regarding the clinical impact and treatment. spect and spect/ct may provide alternative diagnosis if pe is refuted and are feasible in almost all patients, because there are no definitive contraindications. in the present lecture, spect and spect in combination with low-dose ct are discussed in the context of diagnosing pe. over the last few years, emergency radiology (er) has been a subspeciality with growing interest. because of rapid developments in technology, ct has become the most useful tool in evaluating trauma and other emergency patients. in this lecture, the basics of planning and organising an er department are presented and discussed. besides optimising technical equipment and protocols for imaging, different logistic concepts have to be considered in planning and organising er departments. first of all, logistic concepts have to be considered for obtaining an optimal workflow: the radiology department has to be in close proximity to the emergency department and the admitting area, in particular. the whole workflow must be optimised for speed and accuracy. this also mandates having dedicated and standardised examination and viewing protocols for ct. in contrast to the usa where dedicated emergency radiology departments are well established, nonspecialised radiologists still frequently do the reading of emergency radiology cases in european countries. the radiologic staff involved has to be trained for interpretation of trauma and other emergent cases. this does not only account for residents, but also for consultants and attending radiologists. since a large number of cases will arrive during after hours and on weekends, staffing has to be adjusted to this fact, which includes attending radiologists to be available during these hours on call or, preferably, on-site. this lecture will give an overview of logistic concepts and organisation of an emergency department and will also discuss critical issues in polytrauma imaging. at least during the radiological training in hospitals, every radiologist will personally be confronted with the treatment of polytraumatised patients. it is a fact that standardised whole body mdct (wb-ct) as an independent predictor is able to save around % of patient lives. thus, wb-ct is the major diagnostic tool and should be performed as fast as possible. some institutions bypass the emergency room and directly start patient stabilisation in the ct cabinet. however, radiographs and focused ultrasound remain important in general settings as long as they are performed in a way which does not delay ct diagnostics. after scanning, specially trained radiologists support the whole interdisciplinary team with correct, appropriate and prioritised diagnoses. besides optimisation of parameters like logistics, patient positioning preferably 'feet first', reading and communication, 'choosing the right protocol' is a crucial factor for ideal radiological patient care. four types of protocols should be differentiated: there is a wide agreement that for patients with polytrauma, integrating whole body computed tomography (ct) scan into early trauma care significantly increases the probability of survival. therefore, ct is today considered the most important imaging technique in the diagnostic workup of polytrauma patients. the downside of whole body ct represents the relatively high radiation exposure. to overcome this handicap, the following measures can be taken: . positioning of the arms during whole body ct above the head results in reduction of radiation dose; . adaption of scan parameters, especially reduction of tube voltage to kv reduces radiation dose; . installation of recent technical improvements like iterative reconstruction algorithm reduces noise in the image, allowing a downward adjustment in radiation dose to obtain standard diagnostic quality images; . installation of dose control software enables optimisation and improved dose management over time. another disadvantage of high-resolution whole body ct is the amount of acquired data. to optimise the workflow in the evaluation of a couple of thousand submillimetre images it is necessary to connect the mdct scanner with a dedicated workstation for volume image reading, which offers the possibility of quick multiplanar online evaluation. in this introduction i will define the connectome (a comprehensive map of neural connections in the brain) and related concepts such as scaling, structural and functional connectivity as well as the integration-segregation paradigm. the development of mri-based techniques such as white matter tractography and segmentation of white and gray matter has played a crucial role in the emergence of connectomics by providing tools to map in vivo the entire human structural connectivity at a macroscopic scale. i will explain these methods and how from mr imaging a human connectome can be mapped and represented as a network (set of nodes and edges). finally, i will also review the validation work related to those techniques and mention the foreseen technical advancements in the field. session objectives: . to learn what the human connectome is and how it is affected by brain disease. . to understand how the human connectome can be imaged and characterised with mri. . to become familiar with the network formalism and its interpretation. . to understand the relationship between brain function and underlying structural connectivity. . to understand the conceptual ideas behind connectomics. . to learn about the general workflow, from diffusion imaging to mapping a structural brain network. . to become familiar with the robustness of the technique by reviewing validation studies and getting a glimpse of the technical challenges. the economics of brain networks e. bullmore; cambridge/ uk (etb @cam.ac.uk) we review the methods and recent results of network analysis of human neuroimaging data. human functional mri and structural mri data can be analysed using mathematical tools drawn from graph theory to quantify the complex (non-random) topological properties of brain networks. these results can be substantiated by meta-analysis of prior neuroimaging results and analysis of non-human nervous systems. brain networks consistently express complex topological features, such as small-worldness, hubs, modules and rich clubs. some of these features entail disproportionate biological cost, but may be "worth it" by supporting integrative information processing and adaptive behaviours. highly connected hub nodes are high cost/high value network components that likely also represent special points of vulnerability for diverse brain disorders. these economical principles of brain network organisation are expressed at the microscopic scale, e.g., in the nervous system of the nematode worm c. elegans, and may therefore provide a new axis for translation between macro and micro systems neuroscience. neuroimaging can be used to elucidate the economical principles of human brain network organisation in health and disease. connectomics in brain pathology m.p. van den heuvel; utrecht/nl (m.p.vandenheuvel@umcutrecht.nl) healthy brain function depends on efficient functional communication within a complex network of structural neural connections, a network known as the connectome. conversely, damage to the brain's network, disrupting local neuronal processes and/or global communication between remote functional systems may lead to brain dysfunction. in the last few years, emerging evidence from a wide variety of studies suggests that connectome abnormalities may indeed play an important role in the aetiology of several brain disorders. in my talk, i will discuss the results of recent studies suggesting an important role for affected connectome organisation in a number of neurological and psychiatric disorders. in particular, i will highlight the findings of affected functional and structural brain network in neurodegenerative disorders such as alzheimer's and als, as well as discuss how the application of network science and connectomics may aid our understanding of the biological basis of psychiatric disorders such as autism and schizophrenia. learning objectives: . to understand the role of connectome architecture in (cognitive) brain function and dysfunction using diffusion mri/functional mri. . to become familiar with connectomics as a tool for examining disease pathology in a wide range of neurological and psychiatric brain disorders. linking structure and function: the role of modeling in understanding the pathophysiology the interplay of the brain's intrinsic activity and the external world has seen a revival in the last decade, especially in neuroimaging. a long-held assumption in many of these studies has been that ongoing brain activity is sufficiently random that it averages out in statistical analysis. hence, imaging studies are termed 'activation' paradigms, where experimental manipulation results in the activation of cerebral circuits that are necessary for performing the task. nevertheless, a large amount of recent literature reports a body of observations that there are consistently distributed patterns of activity during rest. this fact has led to the suggestion that it might be possible to characterise network dynamics without needing an explicit task to drive brain activity. indeed, numerous neuroimaging experiments have evidenced the solid existence of spontaneous long-range correlations, i.e. functional connectivity (fc), by fmri, meg, and eeg techniques. the functional connectivity is defined as the statistical dependence between remote neurophysiological dynamics. the emergence of resting functional connectivity is intrinsically linked with the underlying anatomical connections between those areas, i.e. the structural connectivity. whole brain modeling can indeed establish a direct link between structure and function. furthermore, the modeling can shed light on the origin of pathological functional disfunction by making the link with the underlying anatomy explicit. we will show how, structural dti tractography, functional imaging and modelling studies can be combined for reaching this goal. traumas to the paediatric pelvis and hip include traumatic dislocations of the hip, fractures of the femoral neck, fractures of the pelvic ring, acetabular fractures and apophyseal avulsion fractures. traumatic dislocations of the hip, fractures of the femoral neck, fractures of the pelvic ring and acetabular fractures are rare in children (less than % of paediatric fractures), as compared to adults. these fractures are commonly the result of high-energy trauma. imaging is based on plain radiographs, but ct and mri are very useful to precisely assess bone (ct) and cartilage and soft tissue (mri) lesions. imaging enables accurate diagnosis, appropriate treatment and detection of potential complications (femoral head osteonecrosis, premature physeal closure …). in contrast with the previous injuries, apophyseal avulsion fractures of the hip and pelvis are common in children and adolescents, usually associated with athletic activities. in most cases, these fractures are of good prognosis and can be treated conservatively when minimally displaced. plain radiographs confirm avulsion injuries to ossified apophyses, but mri and ultrasound are the modalities of choice to demonstrate injuries to nonossified apophyses and to assess apophyseal displacement. the elbow is a very common site for fractures in a child. the challenge for the radiologist is to differentiate normal variants of growth from possible injury and this is usually achieved by having good-quality radiographs and an understanding of normal growth. it is important to recognise those fractures which require surgical intra-operative treatment. in a small number of cases when assessing for vascular integrity, intra-articular extent and injury to cartilaginous structures, ultrasound and mr imaging have a valuable role. this lecture will give an overview of the radiographic appearance of fractures, highlight the features which require orthopaedic intervention and illustrate the use of additional imaging modalities. learning objectives: . to become familiar with the types of injuries seen in the paediatric elbow. . to understand the strengths and weaknesses of different imaging modalities. a- : the diagnosis of c-spine injury is more complex in children than in adults. early diagnosis is crucial since delayed diagnosis results in high morbidity and mortality. leading mechanisms of pediatric c-spine trauma are motor vehicle accidents, sports and pedestrian injuries. due to their anatomy, children are prone to different types and locations of injuries. children < yeas of age are more likely to injure the spinal cord itself and are prone to dislocations and high (c -c ) bony injuries. children > years of age more often sustain c-spine fractures. in children under the age of years, radiographs are rarely helpful. ap and lateral radiographs are helpful in children aged between and years. in children aged > years, additional lateral and odontoid views are obtained. anatomical variants such as pseudosubluxation of c -c , widening of the atlantodental interval and ossification centres may appear to be of concern on imaging, but are normal. abnormal radiographic findings require additional imaging to differentiate them further with ct on the area of concern. mri is mandatory if signs of atlantorotary subluxation and spinal cord injury without radiologic abnormality (sciwora) are present. mri identifies injuries to the spinal cord that are not apparent with other modalities, and should be used when a child presents with a neurologic deficit but normal radiographs on ct scan. nowadays, ultrasound (us) has become the first choice for performing most breast biopsies. the main advantages of this technique are non-ionising radiation, full control of the needle position in real time and widespread use of us equipments at all centres. us allows access to difficult places (such as the axilla or near the nipple), multiple lesions can be safely biopsied in one single session, the breast is not compressed, there is excellent comfort for patients and radiologists, local anaesthesia and haematoma do not hide the lesion and it is a cost-effective technique. however, the main limitation is that the lesion must be visible on us. us-guided fine-needle cytology was widely used in the past, but today its use has decreased. us-guided core-needle biopsy has proven to be a reliable technique to perform a biopsy for breast lesions, showing a sensitivity value of about %. furthermore, it can be safely used for performing biopsies of axillary lymph nodes. vacuum-assisted devices can be used not only for diagnostic purposes, but also for therapeutic ones, because small palpable benign lesions (such as fibroadenomas), papillomas and radial scars can be completely removed. stereotactic breast biopsy is the performance of tissue sampling under mammographic guidance. as with any imaging technique, some lesions are only seen with mammography. when these lesions are suspicious (e.g. birads or higher), there are no imaging techniques that can downgrade this suspiciousness directly, although for birads lesions follow-up might be a viable alternative. in mammography, most lesions that are only visible on the mammogram consist of clusters of microcalcifications. these harbour a likelihood of around % being malignant and should thus be classified as birads . this implies that tissue sampling is mandatory. different from ultrasound-guided biopsy, it is not possible to perform a real-time biopsy under mammographic guidance. rather, stereotactic biopsy uses triangulation to assess the depth of a lesion within the breast, while the lesion location in the xand y-plane is assessed on a scout view. the needle is positioned using the coordinates thus obtained. since there is no real-time feedback of the accuracy of needle positioning, it is necessary to obtain substantially more tissue than under ultrasound guidance. therefore, vacuum-assisted systems are essential. in case of calcified lesions, the biopsy result is controlled by x-ray. calcifications should be present in the sample. biopsy complications that occur under stereotactic guidance are usually limited. most common is the formation of large haematomas. scarring may also occur. breast magnetic resonance (mr) imaging is the most sensitive modality available to evaluate the breast for cancer. it can detect lesions that are occult at mammography and ultrasound. it has a limited sensitivity and the positive predictive value ranges between and %. histopathological assessment of mr-detected lesions is therefore mandatory. some of these lesions will be visible at second-look ultrasound and can be biopsied sonographically. a significant number however will only be discernable on mr, and mr-guided biopsy is therefore the sampling technique of choice. mr biopsy has become an essential component of any breast imaging practice. several mr-biopsy techniques are available including the grid-localising, pillar and post and the freehand techniques. the technique for preparing and performing a mr-guided vacuum-assisted biopsy using the grid-localising technique is reviewed. potential complications, limitations of mr breast biopsy and actions to prevent failure are discussed. imaging-histologic correlation is essential to ensure accurate sampling. clip placement and follow-up imaging should be performed to ensure sampling, including ultrasound correlates. finally, new developments in mr-guided therapeutic interventions are discussed. abdominal injuries require a timely and reliable diagnosis to prevent potentially lethal outcomes. the armoury of clinical tools (physical examination, lab tests) does not fulfil these criteria, since they are either not fast or not reliable. imaging diagnostic modalities help the clinician to acquire the necessary amount of information to initiate focused and effective treatment. however, the selection of the appropriate imaging algorithm, modality and technique, as well as the precise detection and interpretation of essential imaging findings are frequently challenging, especially because the circumstances, under which these examinations are performed (open wounds, bandages, non-removable life-supporting equipment, lack of patient cooperation, etc)., are frequently less than optimal. knowledge of critical imaging signs, symptoms and the role they play in the evaluation of the patient's condition, as well as fast decision-making and ability to closely cooperate with the clinicians are skills of key importance for radiologist members of the trauma team. this presentation will describe the technique and important findings for ct stone studies as well as the accuracy of this study. an explanation of when iv contrast should be given to augment a ct stone study and findings of gu and non-gu diseases that can mimic symptoms of stone disease will be provided. examples and descriptions of acute mesenteric vascular abnormalities, bowel obstruction and infectious conditions will be included. ways to differentiate and categorise emphysematous infections for proper treatment will be described. lower tract emergencies will also be illustrated and described. a systematic approach to evaluating abdominal cts will be described to avoid mistakes. using cases and an audience response system, this segment of the course will go over the optimal imaging approach for patients presenting with acute abdominal pain and abdominal injuries. ct findings will be emphasised. key imaging findings of traumatic and nontraumatic causes of acute abdominal pain including gastrointestinal tract and urinary tract pathology will be explained. a systematic approach for the imaging evaluation of patients with abdominal emergencies will be illustrated and explained including proper scan protocols and analysis of imaging findings. imaging diagnosis of blunt and penetrating abdominal injuries, urinary tract obstruction, infection, bowel obstruction, and ischemia will be emphasised. transcatheter renal denervation represents a novel therapy for treating patients with treatment-resistant hypertension, leading to higher risk of major cardiovascular events. an overview of sympathetic nervous system anatomy, physiology and physiopathology will be followed by a description of the technical aspects of renal denervation. first, results in patients with therapyresistant hypertension look very promising. however, these data need to be confirmed. importantly, long-term efficacy and safety need to be assessed. as a consequence, the treatment cannot be considered, at the moment, standard therapy. future studies need to address other disease conditions characterised by sympathetic hyperactivity, including heart failure, chronic kidney failure and others. it is a general opinion that in the near future the treatment will also be applied to lesser severe hypertensive patients. future studies should also include formal cost-effectiveness analyses. renal denervation can be achieved by a number of different techniques that include radiofrequency ablation, intraluminal and high-intensity focussed ultrasound ablation, chemical ablation and radiation ablation. radiofrequency and ultrasound ablation are currently the most commonly used techniques. most of the evidence to date is derived from studies and trials based on devices that use radiofrequency ablation technology. there are a number of currently ce-marked devices available and each device differs in the catheter design, generator design and energy delivery. these device types include single-point ablation, multi-point ablation, balloon-mounted systems and irrigated technology. the indications for renal artery denervation have been formulated by various international and european societies based on the various clinical trials and studies. these indications include patients with true resistant hypertension, age - years, egfr > ml/min/ . m (mdrd formula), single renal arteries with diameter > - mm with a landing zone of - mm depending on the device, no significant renal artery stenosis > %, no renal artery aneurysm, no previous renal artery angioplasty or stenting and the absence of significant valvular heart disease where lowering blood pressure will be dangerous. renal denervation of the sympatic nerves around the renal arteries is a new catheter technique to treat patients with resistant hypertension. resistant hypertension is seen in about %- % of all patients with hypertension. the mechanism of sympatic nerve driven resistant hypertension will be explained. the available techniques for rdn will be discussed, focusing on the pros and cons of each technique. currently, rf ablation, hifu and cryo-ablation are the techniques under investigation. the technique and physics of renal denervation with radiofrequency ablation will be explained in more detail, focusing on available devices in the market. patient selection and nonresponders will be discussed. the current evidence from the available trials will be reviewed. the future of rdn and the role of radiology and the radiologist will be illuminated. the adrenal glands are composite endocrine organs consisting of the steroid hormone-producing cortex and the catecholamine-synthesising medulla. the increased use of imaging modalities has demonstrated the presence of varying sized mass lesions in up to % of individuals subjected to ct studies for reasons unrelated to adrenal dysfunction. most of these incidentally discovered lesions are non-functioning benign lesions of cortical origin. however, incidentalomas may also represent functioning lesions and malignant masses. clinical diagnostic and biochemical evaluation is used to further subdivide functional and non-functional adrenal lesions. f-dopa has been found to be of high sensitivity and specificity in pet imaging of pheochromocytoma. ct and mr imaging are first choice in characterisation of adrenal lesions. techniques of dual energy ct and histogram analysis may offer additional information. pet-ct has been shown to contribute to the diagnostic power, especially in oncologic patients. knowledge of the physiologic appearance of adrenal glands in -fdg pet is necessary to correctly identify pathologic processes. fdg-pet also has the ability to detect metastatic lesions in non-enlarged adrenal glands. in addition, fdg-pet has the advantage of simultaneously detecting metastases at other sites. the role of mr-dwi and mr-spectroscopy in characterising adrenal masses has to be defined by further studies. differentiating benign from malignant adrenal masses using non-invasive imaging methods can reduce the need for percutaneous adrenal biopsy in patients with underlying malignant disease and the follow-up imaging of incidentally detected adrenal adenomas. renal cell carcinoma (rcc) is the most common malignant tumour of the kidney. as response rates to radiation and nonantiangiogenic chemotherapy are low, surgical excision, i.e. radical nephrectomy has been the treatment of choice. however novel treatment options have emerged, so that imaging of rcc is of increasing interest. nephron sparing partial nephrectomy has become an accepted operative option and antiangiogenic agents such as inhibitors of tyrosine kinase (tk) and mammalian target of rapamycin (mtor) have been approved for treatment of advanced rcc. thus, it has become important to perform correct local staging and identify patients suitable for partial nephrectomy using standardised scoring systems such as renal or padua. furthermore, optimal imaging strategies for monitoring of advanced and metastatic rcc are discussed, as current antiangiogenic therapy evaluation in clinical routine is based only on morphological imaging information, but changes in tumour size may lag behind functional changes. finally, common side effects, i.e. pneumonitis, of vascular disruptive agents are addressed. characterisation of renal masses can be performed using three categories with respect to the lesion size and gross architecture: the indeterminate very small masses, the cystic, and solid renal masses. lesions with diameters below mm are usually difficult to classify due to partial volume effect that prevents accurate ct attenuation measurement. in the general population, these lesions are likely to be microcysts and do not require further workup. better characterisation remains needed in selected patient populations such as patients with hereditary renal tumour disease and previous history of renal carcinoma. in this case, mri combining t w, diffusion-weighted imaging and dynamic contrast enhanced sequences or contrast-enhanced us may help differentiate very small cysts from solid neoplasms. cystic renal mass characterisation still relies on bosniak's classification with categories: benign (i) and minimally complicated (ii) cysts, indeterminate cystic lesions (iif and iii) and malignant cystic masses (iv). some cystic masses remain unclassified at ct because of atypical attenuation characteristics or enhancement properties. us, contrast-enhanced us, and mri are now playing a key role by providing additional diagnostic information that help distinguish between atypical fluid filled masses and atypical solid neoplasms, especially poorly enhancing solid papillary rcc. the characterisation of small solid renal tumours starts at ct with the identification of macroscopic fat, a typical feature of angiomyolipoma. in the case of non-fatty indeterminate renal neoploasms, percutaneous-guided biopsy can be performed when accurate characterisation is needed before surgery or when renal metastases or lymphoma are suspected. drug development being a complex and costly process, there is an increasing need for imaging biomarkers to take go/no go decisions in the early clinical phases. the recist criteria based on tumour size measurements at ct are currently used for this purpose. however, additional functional and molecular biomarkers have been developed to assess the early biological effect of drugs on tumours. development of imaging biomarkers is a structured process in which new biomarkers are discovered, validated and qualified against biological processes and clinical end points. the validation not only concerns the determination of the sensitivity and specificity, but also the measurement of reproducibility. reproducibility assessments, standardisation of the acquisition and data analysis methods and quality control are crucial when imaging biomarkers are used in multi-centre trials. functional and molecular parameters obtained at perfusion imaging, diffusion-weighted mr imaging and pet are being developed and validated. the perspectives (earlier assessment of response to treatment) and limitations (limited validation and standardisation) of these imaging biomarkers in cancer drug development will be presented. more recently, pharmacodynamic imaging biomarkers such as fdg-suv and k trans have been introduced. drug developers are reluctant to use more exploratory unvalidated imaging biomarkers (i.e. cannot distinguish a true negative from a false negative). the extensive literature on biomarker validation mostly refers to biochemical biomarkers extracted as analytes from biospecimens and is unhelpful to radiologists. unlike biospecimen biomarkers, the quality and validity of imaging measurements as biomarkers depend crucially on the use of a diagnostic imaging device, in the presence of the patient, in a manner for which the device (a) was not designed, (b) has not received regulatory approval and (c) may be unfamiliar to the user in the trial site. technical validation and biological validation are orthogonal activities. ''technical validity'' is confidence that the imaging biomarker can be measured reliably anywhere in the world. ''biological validity'' is confidence that the biomarker correctly reports some underlying biology that is important to the patient's future clinical outcome. since the physician always has access to the patient's clinical status and history, the biomarker is only useful if it provides a better forecast than clinical data alone. (if the forecast is near perfect, the biomarker might be a surrogate end point). technical and biological validations are massive undertakings best achieved by consortia, and in particular publicprivate partnerships, of which the innovative medicines initiative in europe and the biomarkers the standard imaging assessment of tumour response relies on size measurements, which, with predominantly cytostatic targeted agents, may not reflect the drug effect. functional imaging biomarkers have the potential to quantify the biological characteristics of tumours and measure on-target and off-target effects that indicate early likelihood of response to a specific therapy, which can then be used to guide the optimal biological dose and drug schedule. serial, non-invasive assessments of whole tumour are possible. this is particularly important in the context of inter and intra-patient tumour heterogeneity, as different parts of the tumour and primary vs metastatic lesions may be biologically different and these characteristics may change with treatment. however, functional imaging end points suffer from variability, which can be very significant in a multicentre setting. strict quality assurance and quality control measures need to be implemented at the start of a trial and the variability across centres documented. data acquisition protocols need to take account of equipment variations. data analysis methodology needs standardisation of software, central review and preferably double reading of scans. automation may not always prove the most robust and reliable option. this presentation will focus on the factors that are crucial in determining the compatibility of data in multicentre trials with functional imaging end points. learning objectives: . to learn about the potential role of quantitative imaging in processes related to tumour growth such as cell metabolism, cell death, and vascular function in the assessment of tumour response. . to become familiar with the issues of accuracy, reproducibility and standardisation for using functional imaging biomarkers in drug development. quantitative nuclear medicine in drug development w. weber; new york, ny/us nuclear medicine techniques can detect and quantify very low concentrations of radiolabelled pharmaceuticals in the human body. this allows investigators to use nuclear imaging for various purposes during drug development. nuclear imaging can visualise drug targets that are only present in nanomolar concentrations and can thus identify patients most likely for therapies directed against these targets. a recent example is folate receptor spect imaging for the selection of patients for treatment with a folate-targeted drug conjugate. in addition, nuclear imaging and specifically pet can measure the concentration of pharmaceuticals within tumour and normal organs over time. nuclear imaging can also be used to monitor target inhibition, for example the blocking of oestrogen receptors by anti-oestrogens. finally, nuclear imaging can assess tumour response to therapy by measuring changes in tumour metabolism or proliferation, e.g. with fdg-and flt pet/ct. this allows an earlier and more sensitive detection of tumour response than morphologic techniques. since pet is a whole body imaging technology, pet imaging can also be used to study the heterogeneity of target expression, tissue pharmacokinetics, target inhibition and response. pet imaging is now widely available in many countries and has become internationally standardised. it is therefore a robust clinical technique that will increasingly be used during drug development. there are about . patients per year in europe who lose their leg. about % are due to diabetic ischaemic and ulcerative leg problems. in diabetic patients the prevalence of foot ulcerations is about % and the risk for developing foot ulcerations is increased four times. diabetic foot problems may be due to neuropathy and malperfusion or a combination of both. the neuropathic ulcer and the neuroischaemic ulcer are usually at the plantar pedis, have a punched-out appearance and are painless. bone deformities (charcot foot) may be associated. the arterial ulcer is usually at the toe, forefoot and ankle, with pale, cold skin and it may be painful. patients need a multimodality approach involving a diabetologist, vascular specialist (angiologist, vascular surgeon, interventional radiologist) and podiatrist. pain control, antibiotic and antithrombotic treatment and the treatment of cardiovascular risk factors and other co-morbid disease have to be done first. in case of ischaemic pain and ulceration imaging such as mra and cta is the next step. patients should be referred to a team of vascular specialist early in the course of their disease to plan for revascularisation options. revascularisation is the optimal treatment for patients with an ischaemic and neuroischaemic diabetic foot. a. pathophysiology of the diabetic foot v. bérczi; budapest/hu (berczi@hotmail.com) the incidence of diabetes mellitus, associated with both predisposing genetic and environmental factors, is increasing globally. several major clinical trials have proved that complications may occur many years following proper glycaemic control. besides peripheral arterial disease, sensory and motor neuropathy along with an altered response to infection is of crucial importance. recent studies have showed that microvascular occlusive arterial disease is not a major factor affecting the diabetic foot; infrapopliteal macrovascular disease and microvascular dysfunction (e.g. arteriovenous shunting, precapillary sphincter malfunction, capillary leakage, venous pooling) are major components of impaired perfusion of diabetic foot. there are no randomised controlled trials analysing the major outcome following endovacular or open bypass surgery. the major outcomes, however, were similar in the case series: -year limb salvage showed a median of % (interquartile range - %) and % (interquartile range . - . %) following open surgery and endovascular treatment, respectively. limb salvage rate, however, was considerably higher with either type of revascularisation compared to medical therapy. negative pressure wound therapy, hyperbaric oxygen therapy, effective off-loading or nonweight-bearing therapies (total contact casts rather than removable devices) have also shown promising results in recent publications. endovascular arterial revascularisation is today a solid option in the management of cli with low complication rates and limb salvage rates comparable with surgery. the restoration of adequate blood flow to the foot is crucial to facilitate wound healing, provide pain relief, and avoid whatever amputations. the angiosome concept was first introduced in by taylor and further developed by attinger for planning treatment of ischaemic lesions of the foot. they divided the foot into six distinct angiosomes, arising from tibial and peroneal arteries. planning the procedure on the basis of this concept will yield the best local results of wound healing, compared with the indirect intervention. a proper pre-procedure assessment through colour doppler us and dsa of lower limbs is mandatory for guiding the procedure through the vessels of the foot. several studies have evaluated the efficacy of pta in the btk and the reliability of the angiosome model, approaching % with a limb salvage rate of up to % at months. on the basis of these data, we can conclude that pta in diabetic patients with btk disease is a safe and effective technique. the first endovascular treatment option is related to the angiosome model, but when not feasible the indirect technique is also a valid and similarly effective procedure. to face technical failures, up to % in crural chronic total occlusion a decade ago, different approaches and dedicated devices and technologies have been developed in the last few years. furthermore, percutaneous revascularisations are gaining more interest, particularly in patients with critical limb ischaemia not only as first-line treatment, but also as the only possible treatment for complex lesions and high-risk patients. special techniques for crural endovascular revascularisations are by design techniques rarely used in routine practice, but could offer wide possibilities for the interventional radiologist to solve challenging situations and manage complex lesions. these techniques include options for arterial access (trans-popliteal, trans-tibial, trans-pedal), approaches for crossing chronic total occlusion (assisted endoluminal, lambda technique, re-entry technologies) and uncommon routes for angioplasty/recanalisation (trans-metatarsal loop technique, trans-collateral techniques). combined antegrade and retrograde approaches have also been developed as well as related methods for successful re-entry. deciding when the patient/lesion is a potential candidate for special techniques is also a crucial issue. the interventional radiologist should be able to convert at any time if necessary the strategy of treatment to another one. he should also be able to select the most appropriate technique for the patient in planning revascularisation. continuous medical education and training is however mandatory and for some techniques the learning curve is relatively long. pathology of the eye and orbit is rare in the radiologist's practice. however, differential diagnosis is not too difficult if the compartment model is applied. in the orbit, different anatomical structures like the optic nerve as part of the cns, muscles for moving the globe, vascular structures and glandular tissue are present in a very small space. each anatomical structure is found in a special compartment; each compartment may give rise to a different group of pathologies, and only to these pathologies: glioma in the optic nerve, rhabdomyosarcoma in the eye muscles, varix in the venous vessels (intraconal compartment) and pleomorphic adenoma in the lachrymal gland. understanding the compartments in the orbit is therefore the key to differentiating different pathological entities. this presentation explains the orbital anatomy, how the compartments are differentiated, and what changes in anatomy treatment may induce. orbital congenital lesions are uncommon. they can be diagnosed prenatally, at birth or later during childhood. several orbital components can be involved. we will focus on congenital globe lesions (such as staphyloma, coloboma, persistent hyperplastic primary vitreous, coats disease), the developmental cysts such as epidermoids and dermoids, and vascular malformations such as lymphangioma and vascular tumours, e.g. capillary haemangioma. orbital inflammatory and infectious lesions are on the other hand common. orbital pseudotumour can involve any area of orbit, being one of the great mimickers in the orbit. if located in the orbital apex and/or cavernous sinus, it will be called tolosa-hunt syndrome. other inflammatory processes in the orbit are sarcoidosis and wegener and sjogren syndrome. among infections the most common is the orbital cellulitis, mostly secondary to a sinusitis and frequent in the urgency setting. the role of the radiologist is to assess whether it is preseptal or already postseptal or complicated by a subperiosteal phlegmon or abscess. also of importance is to know and evaluate the possible intracranial complications. finally, we will focus on inflammatory and infectious lesions of some specific regions such as the globe and lachrymal gland and give some clues for their differentiation. as more than pathologies can be seen in the orbit, a systematic approach is very important to come to the right diagnosis. the main and most helpful criteria of differential diagnosis of any orbital pathology is the definition of the affected orbital compartment, as some tumours may only or preferentially involve specific orbital structures. the criteria of the most frequent masses of the globe, malignant melanoma and retinoblastoma are presented as well as those of cavernoma and lymphoma, the main representatives of intraconal tumours. there are numerous extraconal neoplasms, only few arising from the nasal sinuses, and only a little number of tumours of the optic nerve. the presentation will include the most frequent as well as rare, but important tumours. the routine practice of oncologic imaging requires standardisation, which means that we need to harmonise technical protocols and agree on the meaning of selected words for the radiological report. the words "response, "progression" and "stable disease" are precisely defined according to internationally accepted thresholds and criteria. although the rules are quite simple and rather easy to apply, they are very efficient in the classification of the response to treatment, and therefore for the medical decisions. however, the role of the radiologist is not limited to measurements and calculation. the detection of new lesions may be challenging and requires experience. the differential between cancer progression and complications of the treatment might be very difficult and requires an adequate communication with the referring clinician. overall, most of the decisions taken by the clinician will be related to imaging results, stressing the importance of adequate protocols and reports. in solid as well as non-solid tumours, pet/ct imaging using -fluorodeoxyglucose (fdg) has demonstrated the ability to a) correctly stage disease, b) demonstrate therapy response and c) predict therapy outcome. fdg uptake can be measured objectively; however several factors in the standardisation processes of tracer application, image acquisition and post-processing are needed for reproducibility. the term standard uptake value (suv) measurement is used for compensating the influence of injected dose, decay time and body mass and represents fdg uptake in any selected pixel of the image. for therapy assessment, drop in fdg uptake represents tumour cell kill, notably a negative pet scan does not exclude viable tumour cells but overall has a better outcome. pet response criteria in solid tumours (percist . ) have been introduced to refine previously established pet response criteria by eortc. major changes concern the use of lean body mass-based suv (sul), sulpeak measurement in a fixed roi, use of only a single target lesion and normalization to liver uptake. metric measurements in ct component of the pet/ct as an intrinsic asset like in recist . have not yet been introduced, but might be crucial in the future. the proposed percist . criteria are not yet standard, since several limitations hamper its general use but may improve metabolic tumour response assessment. malignant gliomas (who grade iii and iv) are the most common primary tumours of the brain. according to consensus guidelines, the standard of care of these tumours includes maximal safe surgical resection followed by combined treatment with chemoradiotherapy. radiological assessment is critical in the follow-up and should be performed at four different times: . within hours after surgery, an early post-operative mri must be done to evaluate residual tumour and to be used as baseline for follow-up. . two to six weeks after completing radiotherapy, a new mri examination is recommended to evaluate the response to treatment. four possibilities are envisaged according to the rano criteria: complete response, partial response, stable disease or progressive disease. the evaluation of this first mri examination after rt is challenging and the pseudoprogression phenomenon can appear. advanced mr techniques may be of help in this respect. . additional follow-up should then be performed by mri every to months to rule out clinically silent progression. . in any case, an mri examination must be completed when there is a clinical suspicion of progressive disease. several options for treatment can be offered when progressive disease is detected, including antiangiogenic drugs. evaluation of response to these drugs is challenging too, and the pseudoresponse phenomenon can then appear. due its complexity, it is recommended that the management of malignant gliomas be performed in the context of multidisciplinary teams and that the radiologists are strongly involved in these teams. we are facing complex times with no parallel in human history. the worldwide economic crisis, in combination with a non-regulated process of globalisation, is pressuring countries to change their social and political model. these changes are affecting academic institutions and the health-care sector. higher education was always more internationally open than most sectors because of its immersion in knowledge, which was never worried about country juridical boundaries. it is well recognised that it was the knowledge created by universities research that contributed to the development of societies. we, as european citizens, must be able to transmit the message that having more knowledge and being capable of understanding better the world we live in will make us stronger, more successful and more competitive. european academic institutions must be prepared for this shift of paradigm and understand that they are no longer educating only for their country but for the globalised world. it is also important to bear in mind that developed health-care systems are suffering several pressures and a call for a new inter-professional collaboration concept is needed, based on each one's roles and responsibilities, towards a better and more efficient health-care delivery for the patients. in this session, the invited speakers will give us an overview about the challenges that academic institutions will face in the near future and the way they must adapt to become more attractive and at the same time guarantee that radiographers and radiologists are educated and trained according to the highest professional standards. session objectives: . to appreciate the challenges that academic institutions must face in order to educate graduates with the ability to work internationally and to actively contribute to the healthcare sector of the future. . to explore innovative approaches in education and assessment. . to highlight the importance of inter-professional collaboration in order to ensure that graduates are ready to meet the demands of the modern healthcare sector. challenges in undergraduate education l.j.o.c. lanca; lisbon/pt (luis.lanca@estesl.ipl.pt) radiographers operate a wide range of sophisticated equipment and perform a range of techniques in several radiological procedures. they are responsible for the radiation delivered to the patient while assuring safety and image quality at an acceptable level for an accurate diagnosis. education and training in radiography, in line with the constant technological developments, is a requirement to be fulfilled by higher education institutions (hei). this is of major importance in undergraduate education where radiographers learn to play an effective role as healthcare providers in planning, preparing and performing accurate and safe radiological procedures within the diagnostic or therapeutic field. the european qualifications framework (eqf) for radiography, in terms of the development of knowledge, skills and competences, provides a european standard for the development of radiography learning outcomes at eqf level . this constitutes an opportunity to develop, tune, advance and promote standards of radiographic practice, education and research throughout europe. the eqf provides guidance to increase the educational and professional recognition of radiography in europe. at a european level, the cooperation between hei in undergraduate radiography education would be an opportunity to provide discussions of mutual concern, explore new perspectives and views of different cultures, and also share learning and teaching methods. international cooperation is an opportunity to improve and provide the harmonisation of education regarding a professional group that plays an effective role in healthcare in their field of competence and expertise. there is a vast amount of evidence available from the published literature that higher education institutions continue to explore innovation in both teaching and assessment practices. this is especially true of health professional programmes as evidenced by the abundance of literature relating to medicine, nursing, radiography and other allied health professions. such higher education, health professional programmes would appear to be more innovative than many other disciplines often as a direct attempt to mirror the dynamic clinical environment in which most graduates will work. there are, however, challenges associated with any such educational innovations and novel approaches to delivery and assessment. these specific challenges must be carefully considered in parallel with the larger challenges facing higher education institutions, both nationally and internationally, along with the professions which they contribute to. through a case study approach, supported by published peer-reviewed literature, institutional reports and research, this presentation will consider international best practice in education along with the value of collaborative, inter-professional approaches to teaching and assessment. haemodialysis accesses have become the most commonly performed type of vascular surgical procedure. however, only % of all haemodialysis accesses remain patent at years. autogenous fistulas have a higher rate of primary failure compared to prosthetic grafts ( % vs. %), but the long-term patency of fistulas is superior to that of grafts. autogenous fistulas fail after a median of to years, whereas prosthetic arteriovenous grafts fail after a median lifetime of only to months. the pathogenic stenoses causing access failure occur in various locations, but the most common site in prosthetic arteriovenous grafts is at the anastomosis between the graft and outflow vein, as identified in % ( , ) to % of cases, or in the outflow vein itself in autogenous arteriovenous fistulas. clinical assessment alone will detect a large number of failing fistulas. by assessment of the thrill, pulse character, palpation of the fistula itself, excessive bleeding and difficulty needling a fistula may all indicate a failing fistula. if the venous pressures during haemodialysis exceed mm hg, fistula flow falls to< ml/min, graft flow decreases to< ml/min, or access blood flow falls by more than %. a fistulogram is recommended if monitoring or surveillance suggests that thrombosis is either imminent or has already occurred. duplex can be used to assess the fistula, but cannot visualise the central veins. mr and ct venography has also been used in many patients successfully. treatment of symptomatic stenosis of vascular access is essential as it can lead to thrombosis and loss of vascular access. but treatment can lead to restenosis. the treatment of stenosis remains a challenge which requires a good knowledge of different materials (guidewire, catheters, balloons and stents). though the pathophysiological mechanism of stenosis is different for native fistulas, grafts or restenosis, the treatment technique remains the same. the basic principle is the use of high inflation pressure balloon angioplasty. whatever the material used, the key is to have no residual stenosis greater than %. so far, no study has demonstrated the superiority of the "new" tools (cutting balloon, drug-eluting balloon, cryotherapy) regarding restenosis. except perhaps for anastomotic grafts stenosis, stents should be reserved to treat complications or failure of balloon angioplasty. the stents indications are residual elastic stenosis, wall obstructive damage, acute rupture during angioplasty, aneurysm or restenosis. complications of stenosis treatment are rare, but can lead to the loss of vascular access. the main complications are thrombosis and acute rupture. as with any treatment, there have been contraindications: infection, distal ischaemia, high flow, newly created or surgically revised access (< weeks). when dealing with a vascular stenosis access, never compromise on the future, but think about the surgical alternative. so, the management of stenosis vascular access must also be multidisciplinary. percutaneous treatment of a thrombosed dialysis access can be extremely challenging. the major concern does not stem from the fact that the procedure can be complexly long, but arises rather from the management of its complications, which can be serious particularly when the access is branched onto the brachial artery. access infection is an absolute contraindication. temporary contraindications include fluid overload and severe hyperkalaemia. percutaneous thrombectomy of grafts is very well standardised and predictable: size of mm, well palpable wall which is easy to needle, small average clot burden, and an underlining stenosis almost always found at the venous anastomosis. thrombectomy of avfs is subject to variations in approach and technical difficulties depending on their anatomical particularities. all techniques employed follow rules: first, the removal of thrombi and, second, dilation of the stenosis responsible for the thrombosis. heparin and antibiotics must be injected. the basic principle is to place in the arterialized vein or graft introducer sheaths in opposite directions to work on both the venous outflow and arterial inflow. thrombus lysis or removal can be achieved by mechanical, pharmacological and pharmaco-mechanical methods. all techniques may work in grafts given that the modest amount of thrombus ( . ml on average) can be simply pushed into the lungs. prosthetic grafts are thus easier to declot than native fistulas, but they are much more prone to early rethrombosis. thrombi located at the arteriovenous anastomosis form a firm and rubbery plug, frequently and notoriously resistant to thrombolysis. drug-induced lung disease is an increasingly common cause of morbidity and mortality. the diagnosis is based on clinical history and consistent radiologic findings. lung biopsy is performed in a small percentage of cases. highresolution ct may demonstrate different parenchymal patterns including diffuse alveolar damage, acute or chronic alveolar haemorrhage, nonspecific interstitial pneumonia (nsip), hypersensitivity pneumonitis, organising pneumonia, and eosinophilic pneumonia. the imaging and histologic manifestations are often nonspecific. a systematic approach to the radiological evaluation of drug-related lung diseases is essential and includes not only chest imaging pattern recognition, but also integration of available clinical information. in this interactive session, we will present and discuss several cases illustrating thoracic changes after instrumental procedures in the thorax such as surgery, radiofrequency ablation, interventional endoscopy, and intensive care. radiologists have an important role in assessing the results of these procedures as well as in depicting the complications. however, knowledge of the normal appearance after those procedures has to be presented. the initial imaging workup of polytrauma patients remains challenging. besides hardware requirements, workflow issues continue to evolve, with the current focus on the introduction of whole body ct into the early resuscitation phase of severely injured patients as a standard and basic diagnostic imaging method. this session is aimed at a thorough discussion on the requirements for advanced imaging in the early clinical situation in emergency radiology. focus is directed on the issues of radiation dose as well as on ct and contrast media protocols. a. chest and abdomen m. scaglione; castel volturno/it (mscaglione@tiscali.it) thoraco-abdominal injuries are a significant cause of death in the polytraumatised patients. early recognition and communication of lifethreatening thoraco-abdominal injuries is the major task of the radiologists involved in the emergency room. although most of these patients reach the hospital prior to dying, lethality continues to remain high. heart, thoracic great vessels, trachea, bronchus, pleura, lung, diaphragm, abdominal/retroperitoneal, vascular and solid organ injuries are potential cause of death. any appropriate surgical/interventional management approach must be carried out "around the clock", before thoraco-abdominal injuries reach the level of clinical evidence. on the other hand, non-operative management has actually become the standard of care for the most serious thoraco-abdominal injuries. these goals become feasible if a correct contrast-enhanced mdct diagnosis, in a dedicated facility in which the trauma team works effectively h a day, days a week, is performed. thus, in this lecture, the most serious thoraco-abdominal injuries will be illustrated, with special emphasis on vascular/injuries as well as the value of post-processing techniques, protocols, pitfalls, tips and tricks. furthermore, the importance of a rational and integrated imaging approach will be pointed out and, finally, the role of the radiologist in the emergency room will be emphasised. spinal and musculoskeletal trauma account for significant morbidity in severely traumatised patients. traumatic injuries to the spine encompass a variety of frequently occurring disorders and primarily result in stable injury. although rare, unstable disorders comprise injuries to the bone, the discs, and the ligaments. in the majority of cases, spinal cord injuries result in devastating medical and social consequences. severe musculoskeletal disorders usually are the result of high-impact accidents, such as motor vehicle accidents and falls from a height. for instance, the injured pelvic ring causes one of the most life-threatening conditions that the trauma team must handle. pelvic injuries are often associated with severe arterial, venous, and/or osseous bleeding. radiography is no longer recommended as the primary screening modality in spinal and pelvic trauma for adults. despite the radiation dose burden of ct, patients with a high risk of spine injury receive mdct imaging, as it is the method of choice. mr imaging is indicated primarily when patients present with myelopathy, and to search for spinal cord pathology. interventional radiology plays a major role in the therapy of complex traumatic pelvic disorders. extremity injuries in patients after polytrauma can be complex and are initially often difficult to be fully diagnosed. emergency radiology diagnosis is today mostly based on a standardised whole body ct (wbct), which can be extended with cta and adapted to cover extremity injuries. extremity injuries comprise: fractures of ( ) long bones, ( ) articular joints, ( ) complex fractures of hands and feet, ( ) vascular, ( ) soft tissue, ( ) nerve and plexus injuries and ( ) amputations. imaging protocol: *mdct is indicated in all major and complex bony fractures and is carried out early or integrated with wbct. cta using mips and mprs enables a thorough workup. * the role of us and cr is limited. * the role of mr and mra (in stable patients only) is to evaluate unstable articular injuries, injuries of tendons or major ligaments and nerve and plexus injuries). * the role of dsa is mostly for intervention. clinical findings and findings from wbct determine how to proceed, "first things are done first" in a priority-oriented clinical algorithm. treatment of extremity injuries must therefore be priority oriented and carefully planned in the context of possible concurrent injuries and a possible risk of multi-organ failure (mof). systemic drugs can be used for the treatment of cns and non-cns diseases. both classes can affect the brain inadvertently. common general drugs that affect the brain in a bystander manner are steroids (~ % brain volume reduction), recreational drugs (alcohol, cocaine, heroin, xtc), metronidazole and anti-epileptic drugs (corpus callosum demyelination) and immunosuppressants like cyclosporine/tacrolimus (pres/rpls) and methotrexate. mechanisms of action include neurovascular compromise, fluid/metabolite shifts and toxic effects to myelin of other tissue components. among cns-targeted drugs, especially immunomodulating agents may cause specific side effects. cytokine-release syndromes may occur with broad-acting agents such as general t cell antibodies. specific side effects may occur in multiple sclerosis, where treatment with natalizmumab may cause reactivation of jc virus leading to progressive multifocal leukencephalopathy (pml). upon withdrawal of therapy, this may then evolve into an immune reconstitution inflammatory syndrome (iris). in alzheimer disease, novel antibodies or vaccinations against amyloid may cause amyloid related imaging abnormalities (aria), which may present with microhaemorrhage on t * images, or with edema and effusion on flair. the objective of this session will be to review the diagnostic value of conventional sequences as well as the use of contrast in the monitoring of brain tumours, with special attention to gliomas. also, we will address the advantages and limitations of advanced techniques: perfusion, diffusion, and spectroscopy. immediately after surgery, the main objective of neuroimaging is the detection of the remaining tumour. it may also be necessary to rule out complications such as haemorrhage, ischaemia or infection. the fundamental technique is mri with contrast in the first hours and also diffusion sequences. in late follow-up, the goal is to differentiate the changes secondary to treatment of those related to tumour progression or recurrence. in these cases, conventional sequences present important constraints and are useful studies of diffusion, perfusion and spectroscopy. during follow-up after chemoradiotherapy, the objective is to assess tumour response. the response according to the new rano criteria will be revised. the combination of chemotherapy and radiotherapy, as well as the use of anti-angiogenic drugs causes changes that complicate the assessment of the response to treatment, with cases of pseudoprogression and pseudoresponse. in these cases, perfusion techniques, diffusion and spectroscopy can provide relevant information, although it is necessary to standardise the quantification to make its wide use possible. the speakers in this course will update the audience on contrast media safety such as steps to be taken before contrast administration and present the newest safety guidelines. the first speaker will cover new concepts of non-renal reactions to contrast media explaining which hypersensitivity reactions are allergic and non-allergic. the audience may learn assessing symptoms according to the ring and messmer classification and understand the importance of tryptase sampling and skin testing in the follow-up. the second speaker will address nsf. the presentation will review the pathophysiology, risk factors, recent recommendations and prevention of nsf. patients with gfr less than ml/min/ . m have increased risk of developing nsf. lowstability gadolinium contrast media show the strongest association with nsf. following existing guidelines on the use of gadolinium contrast agents minimises the risk of nsf. potential long-term harm from gadolinium accumulation in the body and legal issues are discussed. the last speakers will cover contrast medium-induced nephropathy with more recently published guidelines related to that issue. the presentation will include the definition of cin and the choice of contrast medium and prophylactic measures. recent changes in esur guidelines will be explained. the risk of cin is considered significantly lower following iv. cm administration and patients referred for enhanced ct are genuinely at risk if they have an egfr < ml/min/ . m . volume expansion with isotonic saline or sodium bicarbonate may be used for preventing cin in at-risk patients. acute immediate hypersensitivity reactions occur within the hour following the administration of contrast media. they can be seen with iodinated and gadolinium-based contrast agents. over the last ten years, new concepts have emerged in the way of understanding, managing and exploring hypersensitivity reactions. the clinical appearance is best classified by the ring and messmer scoring, from grade (cutaneous and subcutaneous signs) to grade (cardiovascular arrest). the mechanisms involve either true ige-mediated hypersensitivity or non-allergic hypersensitivity. the differential diagnosis in favour of allergy is made on a triad: clinical signs (the more severe, the more are the chances to be allergic), elevated tryptase levels in the plasma (indicating mastocyte triggering) and positive skin tests performed one month after the reaction. these new concepts induce important consequences in managing hypersensitivity reactions: be prepared to treat the patient adequately, be prepared to draw blood after the reaction to dose tryptase levels, send the patient to a dedicated allergologist, and forget about the preventive role of premedication against severe reactions. the authorities have introduced several restrictions on the use of high-risk agents, which will be followed by every physician in the eu. the agents are contraindicated in ) patients with severely reduced renal function including dialysis, ) acute renal insufficiency, ) neonates and ) pregnant women. they may only be used with caution in patients with moderately reduced renal function and children less than year old. there must be at least days between injections in those patients. renal function must always be determined by laboratory methods before use of high-risk agents. women should stop lactation for hours. these agents should never be given at doses higher than . mmol/kg per examination in any patient. for the intermediate and low-risk agents, the restrictions are significantly less; they should only be used with caution in patients with severely reduced renal function including dialysis. if a physician does not follow these rules, he or she will have legal problems as they have been introduced into the spc by the authorities. the contrast media safety committee (cmsc) of the esur has updated its guidelines on contrast medium-induced nephropathy (cin). the acr recently updated its guidelines as well. new guidelines were produced by nephrological societies. the topics reviewed include the definition of cin, the choice of contrast medium, and the prophylactic measures used to reduce the incidence of cin. the cmsc considered it appropriate to keep the definition of cin that was agreed in . however, nephrologists have recently agreed on a new definition. in the previous guideline, a number of risk factors were listed (raised s-creatinine levels, particularly secondary to diabetic nephropathy, dehydration, congestive heart failure, age over years, concurrent administration of nephrotoxic drugs). the significance of these risk factors has been confirmed and new risk factors were added. the cmsc agreed that the risk of cin was significantly lower following intravenous cm administration and concluded that patients referred for enhanced ct were genuinely at risk if they had an egfr < ml/min/ . m . the previous cmsc guideline suggested the use of low or iso-osmolar cm in patients with risk factors for cin and the cmsc considered that this previous guideline should not be changed. the cmsc considered that there was enough evidence to recommend that either volume expansion with isotonic saline or sodium bicarbonate may be used for preventing cin in at-risk patients, while the efficacy of nac and other drugs in reducing the incidence of cin remained unproven. guidelines produced by other societies provide very similar suggestions, thus further validating these recommendations. jia is the most common rheumatic entity in childhood and includes a subset of childhood arthritis, all of which are characterised by chronic synovitis with a potential risk of progressive joint destruction. radiological investigations in jia should ideally be able to determine the presence and degree of active inflammation, precursors of bony destructions and established erosions. however, there are many pitfalls in the interpretation of joint pathology in children. ultrasonography is often the initial tool in the assessment of arthritis and can depict joint fluid and synovitis. erosions and cartilage destruction of small joints may also be seen. the major problems are standardising the imaging technique and the lack of normal standards of anatomy in us in children. radiographs can show bone erosions and may depict cartilage loss indirectly through joint space narrowing, but are insensitive to inflammation and early joint destruction. mri is the only imaging modality that can assess all relevant anatomical structures in joint inflammation and is sensitive to early inflammation and destruction. however, large variations in the amount of joint fluid, bone marrow oedema-like lesions and changes resembling erosions are seen in children and also in healthy individuals. the differentiation between true pathology and normal findings on mri in children remains a challenge, particularly in early disease. in this lecture, the role of radiographs, ultrasound and mri and the typical radiological findings in joint pathology in jia will be presented. current knowledge on validity and reliability of the different imaging techniques in jia will be discussed. brain mri plays an important role in those criteria, as it can demonstrate the classical dissemination in time and space and helps earlier diagnosis, which is of major importance since the present recommendation in children is to start immunomodulating treatments as soon as diagnosis is established. among paediatric ms, % begin before the age of years and % before the age of years, frequently with an adem presentation in young children as initial manifestation of ms. however, only % of the adem indicate ms onset and the most predictive factors should be known, i.e. periventricular, deep white matter, corpus callosum high t signal lesions and black holes on t sequences. mri evaluation is also instrumental in differential diagnoses such as nonrelapsing adem, vasculitis, immunogenetic diseases and occasionally leukodystrophies. finally, brain mri is useful to evaluate the risk of more severe ms. the obesity epidemic represents one of the most significant european and public health challenges in the st century with prevalence of the disease having tripled in many countries during the past years. this is resulting in an ever increasing cost to healthcare systems including hospital and in particular radiology services which face unique challenges when imaging this group of patients. obesity significantly increases the patient's risk of various comorbid diseases including the incidence of cancer, diabetes, and cardiovascular and liver pathology with a multidisciplinary team approach mandatory for optimal patient care. this multidisciplinary symposium will review the implications of the epidemic with cutting edge, in-depth lectures presented by european experts addressing the epidemiology, role of imaging in the bariatric surgical patient as well as the importance of abnormal fat deposition in the liver. session objectives: . to learn about the impact of the obesity epidemic on european healthcare. . to appreciate the value of imaging techniques in the management of the post-operative bariatric patient. . to understand the role of radiology in fatty liver disease and the importance of imaging during subsequent patient surveillance. obesity: causes and consequences to the patient r. batterham; london/uk (r.batterham@ucl.ac.uk) obesity is one of the greatest st century public health challenges. its prevalence has tripled in many european union (eu) countries since the s. currently, % of eu adult population is overweight and % obese, and the numbers of those affected continue to rise. overweight and obesity are risk factors for numerous health problems, including hypertension, diabetes, cardiovascular diseases, respiratory problems, musculoskeletal diseases and some forms of cancer. mortality also increases sharply once the overweight threshold is crossed. because obesity is associated with higher risks of chronic illnesses, it is linked to significant additional health care costs and is already responsible for - % of health costs. changes in our environment are the main driver for this increase in overweight/obese. however, a person's genetic make-up can either increase or decrease their chances of becoming overweight. the gastrointestinal tract is the body's largest endocrine organ producing hormones that regulate bodyweight. dietary modifications, such as caloric restriction, are the first-line obesity treatments. however, dieting produces only moderate weight-loss with poor weight-loss maintenance. compensatory gut hormone changes induced by dieting are thought to contribute to the failure of dieting. in contrast, bariatric surgery is an efficacious treatment modality for obesity, producing durable weight-loss, amelioration of obesity-associated co-morbidities and reduced mortality. consequently, the number of bariatric procedures undertaken within europe has doubled in the last years with , procedures undertaken in . there is increasing evidence that surgically-induced alterations in circulating gut hormones mediate the weight-loss and metabolic beneficial effects of bariatric surgery. learning objectives: . to understand the epidemiology of obesity and its impact on european healthcare provision. . to appreciate the aetiology of obesity and the scientific rational for surgical treatment. . to learn about the effects of obesity on health. imaging of modern surgical procedures and their complications m. rengo; latina/ it (marco.rengo@gmail.com) we will illustrate the common bariatric procedures, in particular their normal appearance on different diagnostic technique as well as their early and late complications. we will illustrate the correlation between conventional barium studies and advanced imaging with mdct and mr. we will explain how to optimise mdct and mr acquisition protocol according to the clinical indication. we will explain what to evaluate before redo surgery, in particular quantitative and functional analysis. we will illustrate the role of interventional radiology in the management of early postoperative complications, in particular in the management of patients subjected to gastric banding. non-alcoholic fatty liver disease comprises a variety of pathological disorders ranging from simple steatosis to steatohepatitis. this condition is common in the western population and is typically associated with obesity and the metabolic syndrome. its incidence increases dramatically. diagnosis of fatty liver and distinction between simple steatosis and steatohepatitis are keys because the latter can lead to extensive fibrosis and cirrhosis with an increased risk of hepatocellular carcinoma (hcc). imaging plays a crucial role in diagnosing fatty liver. the two most important imaging modalities are ultrasound and mr imaging. ultrasound can exclude major steatosis but lacks accuracy to precisely quantify fat, while mr is the most accurate for quantification. the technique of reference is mr spectroscopy, but sophisticated sequences based on chemical shift principle have been shown as accurate as mr spectroscopy. unfortunately, imaging has still limitations to assess the presence of fibrosis and inflammation which are associated with steatohepatitis and functional tools could be of interest. today, imaging is combined with clinical and biological biomarkers to evaluate the risk of steatohepatitis. due to the increased risk of hcc and cirrhosis-related complications in patients with steatohepatitis, patients at risk should be enrolled in the surveillance programme. learning objectives: . to understand the pathophysiology of fatty liver disease and its link to cirrhosis. . to become familiar with the role of imaging in the detection and quantification of fat in the liver. . to learn about the importance of imaging in the surveillance of patients with fatty liver disease. the concept of breast cancer units originates from the need for making available to all women in europe high-quality breast services where breast disease could be looked after by specialists working as teams. such teams have to provide all the services related to breast cancer, including genetics and prevention, treatment of the primary tumour, care of advanced disease, palliation and follow-up of previously treated women. the breast unit is made up of a group of dedicated breast cancer specialists including a radiologist, radiographer, surgeon, reconstructive surgeon, pathologist, medical oncologist, radiation oncologist and breast care nurse. the specialists involved in breast cancer units have access to all the facilities required for high-quality care and spend most of their working time dealing with breast cancer. in our hospital, we routinely work as a team and to make it possible we organise weekly meetings involving specialists from different disciplines to evaluate and plan patient care at any step of the diagnostic and therapeutic process. each year, we take care of about . newly diagnosed patients with primary breast cancer and of . patients in follow-up. about are patients who undergo rt. after a short introduction from our fellows from pathology, surgery, oncology and radiation therapy, who will discuss their role in the breast unit, we will try to give a practical demonstration of the everyday work of breast units with particular emphasis on the role of the radiologist. the european society of urogenital radiology (esur) published, in , the new clinical guidelines for evaluation of the multi-parametric mri of the prostate. this structured reporting/scoring system (pi-rads) is based on literary evidence and consensus of experts' opinions. the pi-rads scoring system is similar to that already employed by breast imaging (bi-rads) and reflects the probability of a prostatic lesion to be significant. each lesion can be scored in all sequences used in the multi-parametric prostate mri protocols (t -and diffusion weighted imaging, dynamic contrast enhanced imaging and mr spectroscopy) based on defined mri criteria, which are specific for each sequence. based on the scoring in the particular sequences a final pi-rads score for each lesion can be assessed. for each lesion a five-grade scoring systems was created where, for example, the score means that clinically significant disease is highly unlikely to be present and score clinically significant disease is highly likely to be present. this lecture elucidates the principles of this scoring system and its impact on the target definition for the invasive diagnostics and therapy. prostate cancer screening with psa and extended systematic biopsy protocols have led to the over-detection and over-treatment of small and well differentiated cancers, considered clinically insignificant. these cancers cover up to % of the overall prevalence and may not be of any threat. multiparametric (mp)-mri has shown recently its value in the detection, localisation and characterisation of prostatic tumour foci larger than . cm, and may be of value to address the issue of over-detection and over-treatment. an mri-targeted biopsy strategy alone, without any additional systematic biopsies, has been suggested to decrease the detection rate of insignificant tumours while increasing that of potentially aggressive tumours. in patients with no evidence of lesion on mp-mri, the biopsy may probably be deferred. in patients with localised prostate cancer, mp-mri may also become a cornerstone in the selection, guidance and surveillance of patients managed with focal therapy. the rational for this new modality of treatment is to decrease over-treatment by destroying exclusively the index lesion detected on mp-mri. the precision of such treatments may be increased with mritransrectal ultrasound image fusion, allowing for real time navigation during the procedure. the accuracy of multiparametric mri has greatly improved the ability of localising tumour foci of prostate cancer. this property can be used to perform a trus-mr image registration, a new technological advance, which allows for an overlay of an mri onto a trus image to target a prostate biopsy towards a suspicious area. three types of registrations have been developed: cognitivebased, sensor-based and organ-based registration. cognitive registration consists of aiming at a suspicious area during biopsy with the knowledge of the lesion location identified on multiparametric mri. sensor-based registration consists of tracking in real time the trus probe with a magnetic device, achieving a global positioning system which overlays in real time prostate image on both modalities. its main limitation is that it does not take into account prostate and patient motion during biopsy two systems (artemis and uronav) have been developed to partially circumvent this drawback. organbased registration (koelis) does not aim at tracking the trus probe, but the prostate itself to compute in a d acquisition the trus prostate shape, allowing for a registration with the corresponding d mri shape. this system is not limited by prostate/patient motion and allows for a deformation of the organ during registration. the pros and cons of each technique and the rationale for a targeted-biopsy only policy are discussed. radiotherapy of the prostate typically takes the form of either external beam radiotherapy or alternatively, brachytherapy using radioactive seed implants. in both cases, the use of ultrasound has been proven to be highly useful and this presentation will give an current state of the art overview of ultrasound in external beam and radioactive seed radiotherapy. in the case of external beam radiotherapy, accurate localisation of the prostate is essential to ensure adequate target coverage with minimal damage to normal tissue. dedicated ultrasound scanners are able to provide sub millimeter localisation of the prostate superior to the imaging traditionally obtained using ct scanners. this positional information can be obtained daily. in addition, ultrasound allows the imaging of patients with metal hip implants which are normally difficult to image using ct. apart from the benefits of the technique, some pitfalls will also be highlighted. in the case of prostate brachytherapy, the use of ultrasound in the -d volume imaging of the prostate both prior and during treatment will be discussed. in both cases, some time will be spend discussing the quality assurance requirements for ultrasound imaging systems. image-guided radiation therapy (igrt) with onboard kilovoltage cone-beam ct (cbct) allows image guidance during radiotherapy treatments for patient setup and dose replanning. all these items will be discussed in connection with other topics: organ dose and image quality. it is compulsory to convert cbctimage's pixels from arbitrary grey scale to hounsfield unit (hu). this conversion was obtained using a catphan phantom. the same phantom is used for image quality evaluation; standard ctdi head and body phantoms and a farmer chamber were used to measure the cbdi to estimate organ and effective doses by monte carlo software of different protocols of acquisition. ctdosimetry software (ver. . . -impact) and pcxmc . rotation software were used. to verify the dose replanning techniques by cbct, horizontal and vertical dose profiles were compared with the same obtained from ct. patient replanning was verified using cbct vs ct in terms of conformity index. image quality parameters of cbct (in comparison with ct images) are fine for spatial resolution, but are less useful for low contrast. cbdi of . mgy/ mas was measured; msv effective dose, mgy prostate dose and mgy bladder dose were evaluated and discussed during the presentation. replanning on cylindrical phantom shows a mean percentage difference for each profile % and a variation on d mean in all inserts < %. the mean percentage difference between parameters characterising ct and cbct-based plan values is less than %. the replanning showed a substantial agreement with doses evaluated on the reference ct-image; patient dose must be evaluated for all radiation sources. cranial nerves i-vi have an anteroposterior course and are best examined in the coronal plane. cranial nerves vii-xii run in an anterolateral direction and are therefore best examined in the axial plane. these two planes also allow left-right comparison which makes lesion detection easier. lesions involving the cranial nerve nuclei can be detected on axial proton-density/t /multi-echo-ge images and diffusion-weighted images can exclude acute infarction. the cisternal segment of the cranial nerves is best detected on heavily t -weighted images (drive/fiesta/ d-tse/ciss). at -tesla, d-sequences (e.g. b-ffe) can be used and it is often possible to cover the cisternal segments of all cranial nerves. in the cavernous sinus, the cranial nerves can only be evaluated on coronal gadolinium-enhanced high-resolution t -weighted images and similar axial images are needed to evaluate the nerves in the jugular foramen and hypoglossal canal. axial and coronal gd-enhanced highresolution t -weighted with or without fat suppression are used to image the extracranial course of the cranial nerves. today, the tse-dixon sequences provide non-fatsat and fatsat images simultaneously, making cranial nerve imaging faster and easier. time-of-flight images can be used to study neurovascular conflicts, although these can also be detected on submillimetric gd-enhanced t images. of course, it is important to know the major anatomy of the cranial nerves and the most frequently occurring lesions. the abovementioned imaging techniques, the most important anatomy and the most frequently occurring lesions will be demonstrated and discussed in this lecture. mri study of the lower neck space includes the study of anatomy and pathologies of the thoracic outlet, or cervicothoracobrachial junction, extending from the cervical spine and the mediastinum to the lower border of the pectoralis minor muscle, the brachial plexus and the supraaortic vessels (carotid, subclavian arteries and veins). for this, use of a dedicated head and neck coil is fundamental to avoid frequent artefacts arising from air and the passage between different surfaces. only in case of thoracic outlet evaluation, a dynamic angio-mri study should be performed and this means the use of a surface coil. mri protocol of the lower neck should include t -and t weighted sequences. both fat-saturated proton density and t with stir sequences can be used to overcome the inhomogeneity of the magnetic field, especially with large fov. sand bags can be placed on either side of the neck and suprascapular region of the patient to improve image quality. also, flow saturation bands can be utilised to limit blood flow artefacts. also, volumetric sequences are in common use to obtain a quick examination. breath or cardiac gating can be helpful. abnormal findings of brachial plexus consist of nerve signal abnormalities with mild or marked hyperintensity on t -weighted, being aware of magic angle effects and swelling. in case of thoracic outlet syndrome, mri protocol should define the compression of brachial plexus components arteries and or veins, both in indifferent and dynamic mri acquisition. learning objectives: . to learn how to overcome difficulties in performing a lower neck study. . to understand how to avoid the most common pitfalls. . to become familiar with differential diagnosis. c. ct and mri of temporal bone: user's guide f. veillon; strasbourg/fr ct and mr imaging of the temporal bone must be performed with a precise technique. in ct it is important to locate the box of the study above and not in the orbit to avoid the lenses. irradiation is divided by compared with a study through the orbit. the axial sections must be parallel to the lateral semicircular canal the coronal, sagittal sections are completed by oblique coronal views through the long process of the incus and the head of the malleus to get the ossicular v. double oblique sections through the componants of the v permit a view of the stapes and also the malleus and incus in d (mip : mm). mri must be performed parallel to the roof of the orbit permiting very good axial sections in the plane of the lateral semicircular canal (t and t ). the internal auditory meatus and the inner ear must be analysed with mm axial sections (se) after intravenous gadolinium injection completed by a high resolution t (gradient echo or se) . - . mm, depending on the machine ( t, , t). the middle ear in chronic otitis media must be studied with t , diffusion and high resolution t . there is no need of contrast medium injection apart from the complication: fistula of the lateral semicircular canal, thrombosis of the sigmoid sinus. the different pathologies are then discussed: external auditory meatus and middle ear pathologies: ct first. mri is useful for postoperative cholesteatomas. inner ear malformations, otosclerosis, trauma: ct. labyrinthitis, inner ear hemorragia, schwannomas, internal auditory canal content: mri. revascularisation time windows for patients with acute ischemic stroke are generally restricted up to . hours in the anterior and up to hours is the posterior perfusion area. later treatment attempts require more accurate prediction of risk and benefit, as safety and efficacy at these time strata are less well. thus, rapid and effective imaging is important for decision-making concerning intrarterial catheter based recanalisation and/or thrombolytic therapy. advanced imaging techniques identify irreversible infarction as well as tissue at risk. diffusion-weighted mri detects ischemia within minutes of onset, whereas perfusion-weighted mri and ct perfusion studies disclose the ischemic penumbra. combined, they provide information on mismatched tissue, i.e. potentially salvageable brain. in addition, non-catheter angiographic techniques like ct or mr angiography are a useful adjunct to localise arterial occlusion. as an attempt to a reliable emergency examination, the following protocol has been proven to be robust: for the anterior perfusion area, a non contrast ct may exclude cerebral bleeding and is followed by ct angiography (including supraaortic and intracranial vasculature). if technical available, ct perfusion should be performed in addition. in ischaemic strokes of the vertrebrobasilar region, cta is essential to exclude basilary stenoses or thrombotic occlusion. if the stroke onset remains unclear or might extend the above mentioned time window, mr stroke imaging (i.e. diffusion, flair and perfusion sequences) is suggested as mr offers a higher sensitivity. anyway, the best method for each emergency stroke imaging center is depending on clinical availability h/ days, technical equipment and -finally -individual experience of the emergency team. some recent publications have questioned interventional treatment of stroke as an alternative to iv thrombolysis only. these papers -albeit published in highranking sources -usually do not describe the modern concepts of interventional stroke treatment. interventional radiologists, therefore, are convinced by their practical experience that the modern concept of combining iv thrombolysis with mechanical thrombectomy by stent retrievers offers a benefit to a subgroup of patients with severe stroke. this includes anatomical level of occlusion -basilar artery or single vertebral artery, proximal carotid occlusion with distal tandem occlusion, carotid t obstruction and m occlusion. m occlusions are debatable. besides location, the clinical status of the patient before stroke and time of onset, absence of early ct signs of stroke or bleeding and clinical contraindications to iv thrombolysis such as recent surgery influence the decision-making. more difficult than the description of technical success is prediction of clinical success. there are a couple of scores such as thrive available that may be used for outcome forecasting. the amount of collateral flow is frequently used as a decision tool, but is not always easy to quantify. mismatch scores and penumbra have been questioned recently. in conclusion, anatomical factors are pretty easy to identify to indicate treatment, but the clinical and functional setting still lacks quick and reliable parameters that allow a clear decision-making, particulary in borderline cases. the refresher course addresses the current state-of the-art use of different mechanical revascularisation strategies, devices and potential complications. in addition, multimodal imaging applications with a focus on patient selection for endovascular recanalisation therapies, as well as new techniques to guide endovascular therapy within the angio suite are presented. finally, some organisational aspects important for providing an interdisciplinary interventional stroke service are discussed. the ultimate goal of an acute endovascular stroke intervention is neurological recovery or improvement. recanalisation of an arterial occlusion is key in achieving this goal. clinical data suggest that endovascular stroke treatment results in higher recanalisation rates and may provide superior clinical outcomes when compared with intravenous thrombolytic therapy only. however, these higher recanalisation rates are far away in being paralleled by equally higher rates of favourable outcomes in recanalised patients. thus, patient selection remains crucial. besides the careful neurological assessment, brain imaging is here of major importance. the case presentations illustrate that imaging may help in patient selection for subsequent thrombolytic/endovascular therapies by differentiation of patients who may profit from intravenous or interventional therapy in an even extended time window from those who do not. there are a number of key areas supported by evidence-based medicine necessary for a high-level interventional stroke service. as a precondition, a neurointerventional stroke service has to be organised within a multidisciplinary acute stroke team. inside the hospital, it is all about streamlined pathways. any possible delays should be minimised at every step. sacroiliac joint pain may arise from a number of conditions including inflammatory arthritis, degeneration, fractures and tumours. studies suggest a prevalence of % to % pain arising from the joint in patients with positive clinical signs. temporary effect is provided by a mixture of local anaesthetics with steroids with a response varying between % and % in reported series. dual blocks using agents of differing duration are considered more precise, but are less often used in practice. imaging including mr and scintigraphy are of limited predictive value. injections may be into the synovial joint, around the joint or adjacent to the nerve innervation of the joint. there is evidence that para-articular sources of pain are common and injection outside the joint may be more effective. sl-joint injections are performed through a dorsal approach guided by ultrasound, fluoroscopy or low-dose ct. shortacting agents may have lasting benefit, but radiofrequency ablation has been employed in an attempt to obtain long-term response. the evidence for lasting therapeutic response to intra-articular or periarticular injection of steroids and conventional radiofrequency neurotomy is weak. there is fair evidence of longterm response to cooled radiofrequency neurotomy. facet joints account for - % of all low back pain. they are affected by osteoarthritis, joint space narrowing, intra-articular vacuum phenomenon/fluid, osteophytes, synovial cyst and ligament hypertrophy. conservative therapy is initially proposed. percutaneous facet joint steroid infiltrations are minimally invasive procedures involving injection of corticosteroid with or without local anaesthetic inside the joint. they also can provide diagnostic verification of a certain facet joint acting as the pain source. the injectate usually contains a long-acting corticosteroid mixed with a local anaesthetic. sodium hyaluronate solutions or ozone were tested; however more and extensive studies are necessary. other options are either percutaneous ablation or surgical arthrodesis. fluoroscopy, computed tomography or magnetic resonance can be used for guidance. fluoroscopy has the advantage of real-time imaging. cone-beam ct can also be used. computed tomography provides better anatomy information, but has increased radiation dose for the patient. magnetic resonance has higher cost and longer duration. a recent study concluded that it is twice the cost of ct-guided infiltrations. mri can be used in combination with focal ultrasound for ablation, a technique which is still under investigation. success depends upon patient selection ( - % immediate and - % long-term relief). the level of evidence is moderate for lumbar spine concerning short-and long-term improvement. however, the most recent guidelines released (american society of pain physicians) state that it is the oldest and most commonly used technique. the success rates with its safety profile and least invasiveness seem to make it an attractive therapy. although promising for early assessment of response to treatment, these newer functional biomarkers need extensive validation and standardisation for their wide clinical use. validation includes the assessment of reproducibility and accuracy, whereas standardisation concerns image acquisition and postprocessing. the added value of the more complex functional biomarkers relative to the viability parameters should also be shown. viability and functional imaging biomarkers are evolving and emerging parameters for the early assessment of response to treatment. mri biomarkers must be able to show how tumours respond to specific treatment. they need to allow assessment of the effectiveness of new treatment more rapidly than classical clinical end points. these biomarkers must be easy to obtain to facilitate a large spread of the technique. they have to be reproducible. the longest diameter of the tumour remains the easiest biomarker that can be obtained from any kind of morphologic acquisition with no need of post-processing. additional information about the tissular organisation and cellularity can be now easily obtained using modern scanners through diffusion-weighted sequences. the ease with which those sequences are obtained for a while masked the necessity to perform a more complex postprocessing than the one initially done to get reliable biomarkers. there are numerous mri biomarkers of microcirculation, reflecting o consumption, blood volume, blood flow, vessel permeability and extravascular volume. to get them, we need more sophisticated acquisitions and image processing that take into account the t of the tissue, arterial input function, respiratory motion, etc. most of these new mri biomarkers are now used in research and in phase i studies, but have not been validated in more advanced clinical trials or in clinical practice. to use them widely and reliably, we need to perfectly understand the consequences of the choices we make during the acquisition and post-processing of these biomarkers. new targeted treatments in cancer can be effective without significantly reducing tumour size. there are already a large number of targeted treatments that are licensed to treat a range of cancers. for some of these cancers, there is currently no reliable method to tell whether the drug is effective and new response, predictive and prognostic biomarkers are required. functional imaging techniques such as diffusion-weighted mri, dynamic contrastenhanced mri and fdg-pet imaging are being developed or applied as response biomarkers. however, for these to be useful in a wider multi-centre setting, the measurements need to be precise, repeatable and reproducible. we discuss these properties in the context of emerging imaging response biomarkers. learning objectives: . to understand imaging biomarker precision (repeatability and reproducibility) and accuracy and how it is evaluated. . to learn how to interpret biomarker precision and accuracy in the context of the biomarker's intended use. coronary artery disease (cad) and its related cardiac disorders are still the number one cause of death in the usa and the western world. up to date, single photon computed tomography (spect) using traditional radiotracers like thallium- or tc- m sestamibi is the most utilised imaging technique for the assessment of myocardial perfusion. however, over the past decade, there has been a growing interest in cardiac imaging with positron emission tomography (pet) and, indeed, a paradigm shift has been witnessed in the use of myocardial perfusion imaging (mpi) with pet taking advantage of the superior imaging properties of pet over spect. therefore, pet mpi is now being increasingly used for routine clinical evaluation of patients with known or suspected cad. furthermore, it is being used not only at large academic institutions, but also at community hospitals and even in private practice. several factors contribute to this shift in the use of pet mpi, including the growing availability of combined pet and computed tomography (ct) systems, mainly driven by oncological applications, radiotracer, like rubidium- or fflurpiridaz, which can be used in clinical routine, changes in reimbursement, and the increasing clinical evidence supporting the value of pet/ct mpi. the lecture "pet for evaluation of perfusion, absolute myocardial blood flow and coronary flow reserve" will cover several aspects of the growing field of pet mpi. besides the visualisation of coronary morphology, computed tomography (ct) has shown feasibility to also assess myocardial perfusion. currently, there are two different approaches to ct-based myocardial perfusion imaging: singleshot and dynamic, sequential acquisitions over a predefined scan time. the presentation will cover basic concepts of both approaches and highlight protocol details and findings in these stress acquisitions. also, emerging scientific results with respect to diagnostic accuracy, the detection of hemodynamically relevant coronary stenosis and prognostic implications will be discussed. over the past few years, cardiovascular magnetic resonance imaging (cmr) has been increasingly established as an important method in the diagnosis of cardiovascular disease. many studies have shown the equality or even superiority of cmr compared to other imaging modalities (e.g. nuclear medicine and echocardiography). cmr offers important advantages like the absence of ionising radiation, high spatial resolution, and the combination of perfusion imaging with tissue characterisation. the main clinical applications in the assessment of coronary artery disease (cad) include ventricular function, myocardial viability and perfusion. in clinical routine, myocardial perfusion is determined by contrast-enhanced first-pass perfusion techniques during pharmacological stress using coronary vasodilators (e.g. adenosine) or ßadrenergic agents (e.g. dobutamine). non-invasive characterisation of myocardial microcirculation is thought to reflect myocardial tissue supply much better than mere luminographic detection and quantification of epicardial coronary stenosis, and has been shown to be useful for planning of revascularisation procedures and cardiac risk stratification. in several studies on the prognostic value of cmr in cad assessment, normal stress perfusion cmr was highly predictive for a good prognosis, thus able to identify patients in whom invasive angiography can be deferred safely. the purpose of this lecture is to demonstrate how it is possible to sensitise the mr signal to water molecules diffusion in tissue and how to use calculated indices to reflect structural integrity. the concept of diffusion-weighted imaging will be introduced with particular emphasis on the pulsed gradient spin echo (pgse) sequence. methods to calculate the apparent diffusion coefficient (adc) will be described and the concept of the diffusion tensor (dt) will be explained. image processing of diffusion data includes steps like eddy current distortion correction, model fitting and potentially registration of maps to a reference space. indices that are reproducible and rotationally invariant will be described, such as fractional anisotropy (fa), mean diffusivity (md), axial diffusivity (ad) and radial diffusivity (rd). examples of when to use the adc maps or the dt indices will be given in relation to pathologies such as stroke, multiple sclerosis and neurodegenerative diseases. strategies for result presentation such as region of interest approach, histogram analysis and tractbased methods will be shown. limitations and advantages of diffusion imaging methods such as dt do not preclude the use of this technique in research and clinical radiology for investigating structural changes in disease. diffusion imaging methods have a lot of potential to be used in clinical practice. however, although diffusion-weighted imaging has been increasingly applied, clinical use of diffusion tensor imaging is limited to date. this is in part due to the lack of standardisation and the need for more complex analysis tools to evaluate the data. during this lecture, an overview will be provided of some analysis methods for diffusion-weighted and tensor imaging. i will discuss the different parameters that can be obtained as well as some advantages and limitations of the different analysis methods. it is highly recommended to include diffusion-weighted imaging (dwi) in the routine protocols for mri of the brain. dwi as well as automatically calculated apparent diffusion coefficient (adc) map is used for evaluation. the adc values of gray and white matter are identical in the adult brain. restricted diffusion (high signal on dwi and low signal on adc) is seen not only in acute stroke, but can also be present in some brain tumours, the necrotic centre of abscesses, some acute mr plaques, some contusions, encephalitis, creutzfeldt-jakob disease, metabolic diseases, etc. this finding is thus not specific, but is very helpful combined with clinical information and the signal pattern on other sequences. acute stroke has restricted diffusion during the first - days and the diffusion gradually increases to become very high in a chronic infarct. the diffusion changes in ischemic brain and spinal cord tissue are usually irreversible, but may be reversible. brain tumours with high cellularity, such as lymphoma, glioblastoma multiforme, medulloblastoma and metastases from small cell lung carcinoma, usually have restricted diffusion. the surrounding vasogenic oedema has increased diffusion. in the central necrosis of a pyogenic abscess the diffusion is restricted, while it is increased in the necrotic centre of a malignant brain tumour. reversible diffusion restriction in the cortex, the hippocampi and thalami can be seen in patients with status epilepticus. diffusion tensor imaging has so far had limited clinical use, but may be useful, e.g. for the preoperative evaluation of brain tumours. learning objectives: . to understand the differential diagnostic possibilities of high signal intensity lesions on diffusion weighted images (dwi) of the brain and spinal cord. . to become familiar with the appearance of acute, subacute and chronic stroke on dwi. . to learn about the appearance of cerebral tumours, infection/inflammation, neurodegenerative diseases and traumatic lesions on dwi. . to understand the present use of diffusion tensor imaging (dti) and diffusion tensor tractography (dtt) in clinical neuroradiology. the signal intensity in diffusion-weighted imaging (dwi) reflects the cell density in the tissue. dwi, including apparent diffusion coefficient (adc) maps, can therefore be used to differentiate highly cellular from acellular regions of tumours, distinguish cystic from solid lesions, and monitor change in tumour cellularity over time, reflecting response to therapy. in general, tumours have high signal intensities in dwi, but low corresponding adc values compared to normal/benign/reactive tissues. dwi has a wide range of clinical applications, which includes cancer imaging, imaging of infections and inflammations, evaluation of trauma and visualisation of peripheral nerves. of these, the most promising application appears to be in oncological imaging. there is, e.g. now evidence to recommend inclusion of dwi in mri protocols for lesion detection and characterisation in the liver. the advent of whole body (wb) mri including wb-dwi (from root of the neck to groin) has introduced tumour imaging with a systemic approach compared to established multi-modal diagnostic algorithms. it has been found valuable for staging, therapy evaluation and surveillance of tumours, especially in children. however, dwi is generally recommended to be incorporated into oncologic mri protocols of both wb-mri and of selected organs, because it provides additional valuable information to the conventional mr sequences. dwi of the body is frequently prone to imaging artefacts, which can obscure or mimic lesions. to minimise misinterpretations, analysis of the raw b-value images directly in conjunction with adc maps and conventional coregistered sequences is recommended. spinal surgery is most frequently performed to decompress (disc herniation, stenosis, malignant infiltration), fuse and stabilise (particularly following trauma or infiltrative destructive processes) and correct deformity. often, there may be a combination of these procedures at one operation. surgical instrumentation or bone graft is sometimes employed. patients may present themselves with symptoms early or late following the procedure. this interactive session seeks to address the variety of surgical procedures undertaken and subsequently imaged post-operatively because of symptoms. the session aims to help one to understand and become familiar with the expected post-operative imaging appearances related to the surgical procedure, learn about abnormal pathological features as a cause of symptoms in the acute and more chronic situation and explore the diagnosis and differential diagnosis. this may include post-operative fibrosis versus recurrent disc herniation versus post-operative infection. failure of fusion due to failure of instrumentation or inadequate take of bone graft can give rise to pseudoarthrosis. recurrent stenotic symptoms may relate to an inadequate decompression, recurrent disc herniation, postoperative haematoma, extension of a malignant process or ischaemic damage. joint replacement surgery for the treatment of arthritis most often offers the patient excellent results. however, there are potential complications that a radiologist should know about. it is essential to understand the importance of pre-and postoperative imaging for evaluating patients. most commonly, standard radiographs are used to assess the patients after joint replacement. however, ct, nuclear medicine methods as well as mr imaging play an increasing role in such patients. the radiologist should be aware of the most common type of prosthesis and the most common complications after joint replacement. these complications include postoperative prosthesis loosening, prosthesis fractures, periprosthetic fractures, postoperative infection, rotation failure of the prosthesis, and soft tissue abnormalities such as surrounding tendon tears. this interactive session seeks to address the variety of joint replacements which are undertaken and subsequently imaged postoperatively as a result of symptoms. the session aims to help one to understand and become familiar with the expected postoperative imaging appearances related to joint replacement, learn about abnormal pathological features as a cause of symptoms and explore the diagnosis and differential diagnosis. learning objectives: . to learn about changes related to surgery. . to understand changes related to non-surgical treatments. serbia is a beautiful country where east and west merged and often collided for centuries, and where nowadays eastern cultural heritage meets and mixes with the western heritage in a lovely and unprecedented way, with a lot of charm and with a variety of extraordinarily natural beauty and plenty of both traditional and gastronomical wonders to explore. it extends from the edges of the pannonian flatland in the north, over the danube and sava rivers to the gradually growing wonderful mountains in the south. already in , the first x-ray unit was installed, and only twenty years later x-ray units are present in most state-owned and private hospitals in serbia. nowadays, slightly more than half of the thousand radiologists in serbia perform a broad variety of diagnostic imaging and interventional techniques in different radiological units, including numerous ct, mri, and few pet/ct units distributed in five university centers, a number of public clinical centers and hospitals, and also in the private sector. several centers are fully equipped with pacs and ris systems, and are capable of performing teleradiology services. there are still significant challenges in serbian radiology in the field of education and research, planning and implementation of national radiology networks, improvement of standard clinical practice and financing the new equipment. yet since radiologists' enthusiasm overwhelms most of the difficulties, radiology in serbia provides nowadays a broad variety of modern and competent diagnostic and interventional procedures, improving the management and treatment of our patients. radiology in serbia, since its beginning, has been inextricably linked to european radiology. there are written documents testifying to the fact that the first x-ray appliances in serbia had emerged and had been used for medical purposes only two years after wilhelm conrad röntgen discovered x-rays in . the real beginnings of diagnostic radiology in serbia can be linked to the establishment of the general state hospital in , where the first x-ray cabinet was installed. later discoveries in the field of radiological techniques, materials and contrast agents were also immediately accepted by the serbian radiology community. the first cerebral angiography was performed in , which may be considered as the beginning of neuroradiology in serbia. the second half of the twentieth century is characterised by the introduction and wide application of ultrasound, ct and mr diagnostics and outbreak of vascular and nonvascular interventional radiology procedures in the areas of neuroradiology, uroradiology, and cardiovascular and gastrointestinal radiology. for years, serbian interventional radiology has successfully kept pace with achievements of the most relevant centres worldwide. nowadays, serbian radiologists monitor and enforce the most complex modern procedures in different areas of radiology. the rapid development of science and information technology has enabled the daily monitoring of all modern developments, methods and approaches and their increasingly faster introduction into practice. non-ischaemic cardiomyopathies (nicm) refer to myocardial diseases caused by mechanical and/or electrical disorder in the absence of significant coronary artery disease, valvular heart disease, hypertension, or congenital heart disease. nicm often present with genetic mutations, but no clinically apparent disease. in some cases, the presence of fibrotic tissue could cause adverse events and, therefore, the use of late enhancement (le) post-gadolinium technique is necessary in evaluating these patients. le suggestive of fibrotic tissue could be found in up to % of patients with nicm. in the majority of cases (up to %), midwall/subepicardial enhancement could be found; patchy enhancement, often at septal junctions, has been registered in % of patients, while the least common pattern of le was subendocardial ( %). in ischaemic cardiomyopathy (icm), subendocardial or transmural is a typical pattern of hyperenhancement and is related to coronary artery distribution. furthermore, le has the potential to provide important information for risk stratification in clinical practice. in nicm, the extent of fibrosis has been shown to be independently associated with increased rates of future adverse events including all-cause mortality, while in icm myocardium with le in more than % of wall thickness is unlikely to recover contractile function following coronary revascularisation. as many nicm disorders are associated with the presence of scar tissue, le in nicm is nonspecific and should be correlated with clinical presentation and history, which is critical in the evaluation of these patients. interlude: hop-on, hop-off sightseeing tour of serbia s. stojanovic; novi sad/ rs (tupsons@gmail.com) in the interlude between scientific lectures, you are most welcome to hop on a sightseeing tour around serbia during which we shall stop at a few stations where the natural and cultural heritage of our country will be shown. nota bene: some of the sights will be visible to everyone and some only to radiologists. ten thousand years of human effort and five billion years of european soil refinement will blink in front of your eyes through the window we have made with red churches' walls, azure skies and a silver sparkle of our spirit. all the time, a bit of grayscale shadow will supplement the picture as we in radiology are so used to. fragments of past and present will make a mosaic, the glance of which will hopefully light up your day. when you hop off our tour, we believe, some pictures will become part of your emotional memory and maybe will make you wish to see them in vivo. the role of prenatal mri in foetal central nervous system abnormalities: a case-based pictorial review k. koprivsek; sremska kamenica/rs (katarina.koprivsek@gmail.com) mri provides a unique opportunity for studying in vivo central nervous system development. its higher contrast resolution compared with prenatal sonography allows better visualisation of sonographically occult normal cns structures as well as structural abnormalities. in clinical praxis, prenatal mri has become a mandatory tool in evaluating: a) foetuses that have abnormalities suspected on the basis of foetal sonography; b) foetuses with increased risk for brain abnormalities even in the setting of a normal prenatal sonogram (family history of cns anomalies or a genetic/metabolic disorder); c) foetuses potentially at risk due to maternal illness; d) foetuses with congenital malformation, which are candidates for prenatal or neonatal surgical treatment; e) foetuses that could not be evaluated by sonography due to oligohydroamnion, maternal obesity, difficult position of the foetal head, or us reverberation on foetal calvaria in advanced gestational age. we will present a selection of foetal cns abnormalities detected in our institution during the last years, including ventriculomegaly, abnormalities of the posterior fossa, agenesis of the corpus callosum, cerebral cortex developmental abnormalities, spine developmental abnormalities and a variety of encephaloclastic lesions. in all previously listed cases, prenatal mri could provide either crucial or additional information, which can lead to the accurate diagnosis of different cns developmental abnormalities (cortical, commissural and posterior fossa malformation), specific disorder or syndrome, further enabling adequate pregnancy management and parents' counselling. this lecture will cover the standard mammographic views required, the reporting systems for mammograms together with birads and other european systems, breast density, feature analysis, information the clinician requires in a cancer case and how to approach a b lesion. acr practice guidelines ( ) recommend obtaining previous mammograms as you can dismiss an abnormality that is unchanged for years or that has been previously worked up and are able to detect subtle new lesions more easily, although this has not been confirmed in the screening situation. in a structured report the following information is recommended: indication, breast density, description of lesion, size, location, comparison to previous findings, overall assessment, management recommendations and a score to indicate the level of suspicion. examples of different common conditions seen on mammography will be shown including soft tissue masses, microcalcification, architectural distortion and asymmetry. breast ultrasound is one of the main imaging modalities in breast radiology, it allows us to characterise lesions and also guide interventional procedures. the bi-rads categorisation of ultrasonographic findings facilitates the diagnostic approach and also helps the radiologist to use a common language, understood not only by the rest of the radiological community, but also by other breast cancer professionals. ultrasound can be a diagnostic procedure on its own, but is mainly a modality that characterises the findings of other modalities (mammography and mri) and, as such, correlation between all these techniques is the mainstay of everyday clinical practice. this act of correlating and integrating the information of all modalities is what makes a breast radiologist a key actor in the diagnosis, staging and follow-up of breast cancer and other benign or high-risk entities. the final product of this integration will be the radiological report, the means by which we convey all the information we have gathered through all the procedures to our clinical colleagues. this report shall also follow some composition rules to be clear and concise. as with any other modalities, breast magnetic resonance imaging (mri) examinations should be reported in a structured way following the guidelines of the respective national, continental or international societies. adequate nonclinical information (i.e. patient name, date and type of examination, etc). is indispensable. any breast imaging report has to follow a stringent structure including indication, clinical history, clinical findings, brief description of technology used, assessment of parenchymal density, detailed description of significant findings, comparison with previous imaging studies and final assessment according to bi-rads. indications and contraindications of breast mri will be discussed. significant findings at breast mri include foci (small contrast-enhancing spot, nos, < mm), mass lesions, "non-mass-like enhancement" (no mass lesion, partly diffuse regional contrast enhancement of various size) and associated findings. furthermore, as breast mri is a functional study, the different appearance patterns of kinetic contrast enhancement will be presented. the current status and the appropriate use of the bi-rads mri lexicon will be discussed. any breast mri report should not only follow the guidelines, but also follow a red thread, be consistent, express confidence and be comprehensible to clinicians. the overall final bi-rads assessment is based on the most worrisome finding, taking into account both breasts and all imaging methods (mammography, ultrasound, mri) evaluated. furthermore, adequate communication of the result, as well as do's and don'ts of the report wording will be discussed. the prostate is a small glandular organ, whose main function is to secrete seminal fluid. in young men it comprises of mainly the peripheral zone ( %) and a smaller central gland. however, the central gland enlarges with age to form the larger part. the main diseases of the prostate are inflammation, hyperplasia, and cancer (the most important). approximately % of cancers arise in the peripheral zone. the main applications of mri are for cancer detection, staging, and recurrence. mri is increasingly used for radiotherapy planning owing to its good soft-tissue contrast when compared with ct scanning. endo-rectal rf receiver coils, usually inflated with air or perfluorocarbon but sometimes of a rigid design, improve signal detection of the prostate. phased-array surface coils may also be used or (preferably) in combination. t -weighted imaging helps to distinguish haemorrhage caused by biopsy. on t -weighted images the peripheral zone appears brighter, owing to a larger proportion of prostatic ducts, while tumours generally appear darker. more sophisticated mr methods are increasingly used to help diagnosis. in diffusion-weighted images (dwi) tumours appear relatively bright, but dark in the corresponding calculated map of apparent diffusion coefficient (adc), owing to the more cellularly-dense nature of tumours, and hence, reduced diffusion. in dynamic contrast-enhanced mri (dce-mri) tumours appear bright owing to increased vasculature. magnetic resonance spectroscopic imaging (mrsi) of normal prostate is dominated by high levels of citrate from the prostatic ducts; in cancer this is reduced while choline is increased. prostatectomy samples allow one to verify image contrast against histology. recent studies on prostate cancer have provided a number of relevant targets for imaging and treatment purposes. today, imaging plays an important role in different aspects of the disease, but its role should grow in answering clinical questions at various phases of the disease. relevant targets for imaging include metabolites (including glucose, acetate, choline, and amino acids), antigens (including prostate-specific membrane antigen), receptors (grp receptor, cxcr , androgen receptor), proliferation, angiogenesis, and hypoxia. in this presentation, a range of radiotracers for potential use in the imaging of prostate cancer will be discussed, as well as options to tailor nuclear imaging tracers to the various phases of the heterogeneous disease. the fusion of morphologic (ct and mri) and functional (pet, spect) imaging modalities (pet, spect) has become widely available and part of the standard diagnostic workup in cancer patients. for many years, neurosurgeons have already relied on multimodal imaging data during brain tumour surgery to identify hyper-or hypometabolic tissue. the availability of pet and spect data during percutaneous ablation procedures may also be helpful to facilitate treatment planning, probe placement, probe re-positioning, and early detection of residual disease following percutaneous image-guided ablation of cancer. however, up to now conventional us-and ct-guided thermal ablation procedures rely on morphologic information only. intraoperative implementation of multimodal data requires the application of frame-based or frameless stereotactic techniques, both relying on the establishment of an accurate relationship between the patient's preoperative image space and the intraoperative physical space. in image-guided neurosurgery, ct/mrt/spect/pet data are registered to the patient using rigid body transformation. due to respiratory motion and different patient positioning during image acquisition and the actual intervention, soft tissue deformations in the liver occur. thus, the images and the surgical presentation do not match between preoperative imaging and intervention, leading to an extensive degradation of spatial guidance. this presentation illustrates solutions for these fundamental problems that occur during d-navigated liver interventions, including patient immobilisation and repositioning, respiratory triggering and intraoperative imaging. r. bale: equipment support recipient; isys has supported the section of microinvasive therapy with testing equipment. patent holder; rb is a (co-) inventor of the atlas aiming device and the bodyfix immobilisation device and a (co-)shareholder in its financial returns. the goal of ultrasound-triggered, image-guided drug delivery is to increase the therapeutic index and decrease adverse effects of drugs. the bio-effects of focused ultrasound can lead to local tissue heating, cavitation, and radiation force, which can be used for ) local drug release from nanocarriers circulating in the blood, ) increased extravasation and cellular uptake of drugs and/or carriers, and ) enhanced diffusivity of drugs. thermo-sensitive liposomes have been suggested for local drug release in combination with local hyperthermia more than years ago. microbubbles may be designed to enhance cavitation effects. real-time imaging methods, such as magnetic resonance, as well as optical and ultrasound imaging have led to novel insights and methods for ultrasound-triggered drug delivery. image guidance of ultrasound can be used for: ) target identification and characterisation; ) spatio-temporal guidance of actions to release or activate the drugs and/or permeabilise membranes; ) evaluation of biodistribution, pkpd; ) physiological read-outs to evaluate the therapeutic efficacy. liposomes may carry both hydrophilic and hydrophobic drugs in their aqueous interior and lipid bilayer membrane, respectively. the circulation half-life may be increased by incorporating polyethylene glycol (peg)-lipids in the bilayer. recent publications have shown that ultrasound-triggered delivery is feasible. realtime imaging methods, such as magnetic resonance, optical and ultrasound imaging may lead to novel insights and methods for ultrasound-triggered drug delivery. up to now, the success of surgical tumour resection has always been limited by the surgeon´s vision. the human eye is not an accurate detector of small cancer cell clusters and it cannot accurately differentiate cancerous from healthy tissues due to lack of "visible" contrast. by combining a fluorescent probe targeting the folate receptor and a multi-spectral real-time fluorescence camera to observe the operating field, we found that, using fluorescence molecular imaging, -fold more lesions can be identified compared to conventional surgery. these first in-human results point to molecular-based clinical decision-making in surgical and endoscopic procedures as a paradigm shift over decision-making based on human vision. the talk describes current progress with instruments, methods and applications in the field of intraoperative imaging. pre-clinical and clinical results are presented and the advantages and limitations of the method as well as future directions will be discussed. the emerging field of multispectral optoacoustic tomography (msot) is also outlined. mysteries of the human brain unveiled: imaging of white matter microstructure and neuroplasticity p.m. parizel; antwerp/be (paul.parizel@uantwerpen.be) the human brain remains one of nature's great mysteries, and is considered science's final frontier. this greyish lump of tissue with a weight of . kg contains some billion neurons, each of which are connected to thousands of other nerve cells in an intricate network of white matter fibers. in the previous century, the brain was perceived as a fixed three-dimensional landscape, with brain functions confined to certain locations (e.g. motor and sensory cortex, visual cortex, auditory cortex). this concept of neurolocalisationism was in line with the philosophy that there should be "a place for everything, and everything should be in its place". advanced quantitative mri techniques for acquisition and data analyis (fmri, dwi, dti, vbm), help to unravel white matter connectivity of the human brain, and are able to show that the brain can modify its structure and function in response to changing circumstances (such as learning, memory, hormones). this process, which is known as neuroplasticity, occurs at different levels and different time scales. for example, in response to injury, changes may occur at cellular level as well as on a larger scale with cortical remapping. some processes may take months or years (e.g. physical therapy and training) whereas some forms of neuroplasticity happen within hours or days (e.g. changes in brain volume and connectivity during the female menstrual cycle, which have been linked to behavioral changes). in summary, quantitative mri techniques help to unveil the mysteries of the human brain and have opened exciting new fields of active research such as white matter connectivity and neuroplasticity. there are a host of inflammatory and infective insults that can manifest focally or diffusely within the musculoskeletal system. the appearances of the underlying pathological processes in both the soft tissues and skeleton cover a very wide imaging spectrum. the appearances vary, depending on the timing and degree of inflammatory insult and the host response in the involved tissues. the approach of this lecture will cover the imaging manifestations using all modalities covering radiography, ultrasound, ct, scintigraphy and magnetic resonance imaging. the basic knowledge that is required will be displayed in major musculoskeletal categories covering disorders involving the soft tissues, joints, bones and entheses. the imaging manifestations will also be linked with the evolution of the pathological processes covering acute, sub-acute and chronic stages of the inflammatory/infective disorders. by the end of the session the audience should have a clear understanding in making best use of the imaging modalities in the correct diagnosis of a wide variety of inflammatory and infective conditions that can affect the musculoskeletal system. arterial pta and stenting is firmly established as primary treatment in the management of athero-occlusive disease, particularly of peripheral limb arterial disease (pad). patients are selected on the basis of symptoms such as short distance claudication or critical limb ischaemia for lower limb disease or appropriate ischaemic symptoms in other territories in the renal and mesenteric territories and head and neck vessels. commonly used grading systems such as the fontaine or rutherford scores can be useful in patient selection. where treatment is required, noninvasive imaging is utilised to look for the site, severity and extent of arterial disease. most centers use duplex imaging as the primary imaging modality; mra or cta are often used with a higher sensitivity and specificity; and invasive angiography is reserved for problem solving or immediately prior to intervention. the tascii classification has been devised to select patients for either endovascular or surgical intervention. however, most centres will adopt an endovascular approach first. all patients should be on good medical therapy, i.e. aspirin and statins, with good blood pressure and diabetic control where appropriate. when carrying out interventions, an access sheath is used, peri-procedural heparin administered and lesions crossed using selective catheters and commonly hydrophilic guidewires. subintimal crossing of occlusions can be much simpler than trying to cross lesions luminally. balloon angioplasty and or stenting may be applied differently depending on the site and extent of the lesions. the evidence for drug-eluting technologies has also been growing with improved patency rates compared to standard balloon angioplasty. during the efrs meets russia session, radiographers have the opportunity to get information about the role and education of radiographers in the different european countries. the radiographer is a health-care team member who interacts with other professionals in the primary and secondary health-care environment to provide an optimum diagnostic or therapeutic outcome. radiographer education therefore requires that the curriculum covers a wide range of scientific, medical, pathological, sociological, ethical and technical subjects together with the development of appropriate clinical skills. the curriculum should also include the development of research and audit skills to ensure the constant improvement of service quality for the benefit of service users. the session will include a panel discussion about the role of the radiographer in image acquisition and processing. the tasks of the assistant to the radiologist are: filling of the protocol of research, survey of the patient, instructing the patient according to safety measures, positioning the patient, placement of coils of strengthening, beginning of magnetic and resonant research -the research is approved by the doctor, introduction of contrast substance (in the presence of the certificate of the procedural sister) and supervision over the patient during mr research. furthermore, the other tasks are briefing on patient safety, excluding introduction of the patient to metal objects on mr tomography, explaining the procedure of the study, informing about the duration of the study, wearing noise-cancelling headphones, warning about possible vibration and loud sounds, and providing a globular button to call the nurse. today coronary ct-angiography has become a well-established and proven diagnostic modality. quality of coronary and cardiac cta datasets has tremendous significance for correct diagnosis. a radiographer has a pivotal role in performance of coronary cta. coronary cta is one of the most challenging examinations for radiographers because even a small mistake at any stage of examination may ruin the whole study. before the scanning procedure, the radiographer performs a check for possible contraindications to contrast media injection (history of adverse reactions to contrast media and impairment of renal function are the most important ones). the radiographer checks the patient's heart rhythm and takes part in administration of betablockers for control of the heart rate. a detailed knowledge of different scanning modalities of ct-machine is a must. radiation exposure to the patient from coronary cta examinations has been one of the major public concerns. this is why the radiographer should be aware of different approaches to the performance of low-dose cta (prospective gating, tube current modulation, high-pitch scanning) according to patient body composition. the radiographer should have knowledge of when and how to use radiation dose control in an appropriate way. another big issue is injection protocols. selection of optimal bolus timing and iodine load results in highquality cta images. the radiographer has to know the basics of ct image reconstruction and interpretation in order to perform different types of reconstructions from the raw data according to the radiologist's requests and be able to recognise image artifacts. clinical examples of the radiographer's role for performance of coronary cta will be given. every country is famous for its architecture, history, literature, music, museums, etc. however, there are some distinctive features for russia. "russia is a riddle wrapped in a mystery inside an enigma." these words by the famous british statesman winston churchill aptly throws light on the pulsating art and culture of russia. the different aspects of russian art and culture find its best expression in the rich heritage. just feeling the nature and boundless expanses of the country can understand the mysterious russian soul, works of art and real masterpieces. cardiac mdct in children with congenital heart diseases v. bereznitskiy, k. serkova; moscow/ ru (vsber@mail.ru) in ct studies of children with congenital heart diseases, it is mandatory to keep the x-ray dose, amount of contrast agent and study time as low and as short as possible and simultaneously get the best diagnostic image quality. how can we reduce the x-ray dose? by minimising the volume coverage, reducing the voltage, minimising the effect of "overranging", use of x-ray protective equipment, use of tube current modulation technique, use of iterative reconstruction technique. how can we reduce the amount of contrast agent by volume? by covering of only necessary anatomic volume, minimising scanning time, use of adequate speed injection in ml/sec, which depends on catheter position and size, blood flow and anatomic structures. how can we reduce the amount of iodine concentration in the contrast agent? by lowering the voltage we may reduce the iodine concentration and get higher absorption ct values (hu). how can we reduce the time of cardiac ct studies? by preparing the study protocol ahead, preparing all necessary equipment in the ct room, warming up of the contrast agent before the study. what is the radiographer's role in cardiac ct studies? to take care along with the responsible radiologist the above-mentioned needs, to take care of adequate room and instrument temperature, to take care that only absolutely necessary people with adequate x-ray protection are in the study room, for performing the necessary image post-processing, archiving and distribution of studies. the purpose is to evaluate the quality of imaging in forced position of patients with anesthetic support. the role of laboratory technician in ct post processing. we analysed images of unconscious patients on artificial lung ventilation on -and -slice computer homographs ge and siemens and monitored them through the infusion. during investigation the patients were in forced positions, as a result they got artifacts from upper extremities on the abdominal cavity. we analysed patients -hands under the butt, patients -hands on the chest (abdomen), patients -arms along the body. according to the images the smallest number of artifacts was identified when the patient was on the table with hands under the butt. different variations of the patient's hands position and the problem of choice of this position will be offered for researching. the reconstruction the laboratory assistant can execute and also the results of tests will be presented. by order of the doctor the laboratory technician builds d and d reconstructions of fractured ribs, pelvis, facial skeleton, sagittal and coronary reconstructions of spine in case of its fracture. gastrointestinal stromal tumours (gist) are a rare disease that metastasises in up to % of patients with subsequent median progression-free survival (pfs) of around months. tumours are characterised by activating mutations in the kit or the pdgfra gene and treatment is mainly based on tyrosine kinase inhibitors designed to block mutated receptors. however, drug resistance is often based on mutations changing the conformity of the receptor, leaving little effective therapeutic options. to date, second line chemotherapy offers a median pfs of - months and external beam radiotherapy is limited by organs at risk close to the tumour. alternative approaches like endoradiotherapy or minimally-invasive ablation techniques are effective for local control but are inconsistently used and are not tailored to the individual patient's type of disease. to address these issues, the mitigate project proposes a closed-loop personalised treatment concept combining endoscopic-assisted tissue sampling, inline biotechnology and targeted molecular pet imaging probe development combined with minimally-invasive treatment monitored by new mr imaging techniques. a consistent value chain across european research institutes and smes will be established for mass spectrometry of tumours, linkage of radiochemical molecular imaging probes, design of new animal models and targeted therapeutic radiopharmaceuticals. this closed-loop platform will minimise fragmentation of treatment approaches by a coherent molecular-based multimodality concept, thus providing new treatment options. the mitigate platform can be expanded to further patient cohorts with oligometastatic diseases such as other sarcomas or renal cell carcinoma. gastrointestinal stromal tumours (gist) are characterised by highly specific genetic mutations. several specific targets are overexpressed in a majority of the tumours. on the other hand, secondary resistancies limit the usability of highly specific chemotherapeutics such as imatinib. instead of applying cascades of alternative tki inhibitors in order to find an efficient one, quantitative molecular imaging and targeted endoradiotherapy could be considered. for the detection of gist by imaging, ffluorodeoxyglucose (fdg)-pet is widely accepted to visualise the tumour burden. however, as fdg gives no molecular information of potential targets for treatment, new radiopharmaceuticals have to be developed to visualise and quantify other target-structures expressed by gist that would be able to be addressed. this molecular information can be used both for designing a conventional treatment or an advanced strategy: i.e. targeted endoradiopharmaceuticals. compared to conventional chemotherapy this method has the great advantage that not only tumour cells expressing the targeting vector are affected but also tumour cells within the range of the radiation of the decaying radiopharmaceutical. this method is used with great success in the case of neuroendocrine tumours using labeled octreotate derivatives (a small cyclic peptide binding to the somatostatin-receptor) and non-hodgkin's lymphoma applying y- labeled zevalin (a monoclonal antibody binding to the cd antigen). the success of these methods relies on the crossfire-effect, meaning that a certain margin of tissue surrounding the cell targeted by the specifically accumulating endoradiopharmaceuticals is destroyed as well, depending on the penetration range of the emitted radiation. thus, the development of new radiopharmaceuticals for the specific diagnosis and treatment of gist would strongly improve the outcome of the tumour patient. stereotactic radiofrequency ablation of liver tumours: does improved targeting increase tumour response? r. bale, g. widmann, p. schullian, m. haidu, w.r. jaschke; innsbruck/at (werner.jaschke@i-med.ac.at) the purpose is to describe stereotactic radiofrequency ablation (srfa) and to review its inital results in the treatment of patients with primary and secondary liver tumours. one retrospective review includes consecutive patients with srfa sessions for the treatment of hepatocellular carcinomas (hcc) and metastases ( ). in two other studies the outcome after srfa of intrahepatic cholangiocellular carcinomas (icc) ( ) and colorectal liver metastases (crlm) ( ) was evaluated. the overall technique effectiveness was . % with a local recurrence at year of . %. technique effectiveness was not significantly different for lesions< cm ( . %) and - cm ( %). after treatment of inoperable consecutive patients with iccs a median overall survival of months was achieved. a median os of months and os rates of %, % and % at , and years in patients with resectable crlm were achieved. tumour size did not affect os and dfs. rfa probes can be precisely planned, positioned and distributed by means of stereotaxy. especially in large liver tumours the local recurrence rate can be greatly reduced by precise stereotactic placement of multiple radiofrequency probes. these improved local control rates result in better long-term survival rates. author disclosure: r. bale: shareholder; coshareholder atlas aiming device. other; coinventor atlas aiming device. functional and molecular imaging in tumour therapy response assessment s. aime; turin/ it (silvio.aime@unito.it) in the mitigate project further development of currently used mri and ct techniques will be tackled in order to obtain functional and metabolic information which are mandatory to detect early response to the targeted radiopharmaceutical approach. new gd-based agents are under scrutiny in order to improve dce-mri procedures and attain more precise information on the tumour microvascularity level as well on the vessel permeability and the assessment of extracellular ph and mmp activity. the x-nuclei mri ( na sodium) is a non-invasive imaging technique, which enables the measurement of the tissue sodium concentration (tsc) in humans. the direct link of the tsc with the tissue integrity and viability provides a promising approach for monitoring tumour tissue, and could also help to assess cytoxicity and cell death by means of breakdown of the sodium-potassium-pump. however, na mri is challenging because of the low in vivo abundance, the fast transversal relaxation rate and the low gyromagnetic ratio of the na ions in human tissue. new mr measuring sequences for x-nuclei mri will be developed to establish a standardised tsc measurement method within the eu for better comparability. furthermore, the x-nuclei mr data will be reconstructed, scaled and fused with further imaging modalities. lung cancer is the leading cause of death related to cancer. most patients are inoperable as they present with advanced stage disease or even a localised tumour associated with poor general condition, limited cardiopulmonary function or a too high surgical risk. according to the stage of the disease, chemotherapy, radiotherapy and percutaneous ablation therapies are the current therapeutic options for inoperable patients. it is important that radiologists are familiar with the various response and complication imaging patterns related to each of those treatments. the timeline modifications after radiation therapy and percutaneous ablation and the diagnostic management of drug-induced lung disease will be reviewed. this presentation will summarise the current evidence and how to detect early recurrences after those treatments. learning objectives: . to learn about changes after radiotherapy. . to learn about sequela after percutaneous ablation of lung tumours. . to understand changes after chemotherapy of lung cancer. cardiovascular medicine owes much of its spectacular development to the parallel evolution of imaging techniques in the last thirty years. the complex contemporary techniques in interventional cardiology, electrophysiology, and cardiac surgery require advanced imaging modalities. in current times, a wide spectrum of imaging techniques -such as fluoroscopy, mri, non-contact mapping, echo techniques -is performed in organised cardiovascular labs while computed tomography and nuclear techniques are provided by the collaborative departments of radiology and nuclear medicine. it is indeed the collaborative spirit among specialists that bring about the results patients desire. cardiac magnetic resonance (cmr) is a very versatile imaging technique for the assessment of patients with left ventricular dysfunction and has become a central method for characterising the etiology of the dysfunction. indeed, wall thinning less than mm and aneurysmal motion are morphological signs of transmural necrosis. furthermore, a low dosage of dobutamine ( - microg/min) is used to detect functional reserve in the case of viable tissue. a further option offered by cmr is late contrast enhancement (le) imaging, which allows identifying the presence of fibrotic tissue as well as necrotic infarcted myocardium. the sensitivity and specificity of late hyperintensity is, respectively, . and . . similar results can be obtained with mtctetrofosmina g_spect. there is a relation between transmural extension of hyperenhancement and the recovery of contractile function after coronary revascularisation. the chance of recovery decreases progressively with the transmural extension of late hyperintensity, since the average value of the latter is ± % in the segments that recover and ± % in the segments that do not recover contractile function. cmr was also shown to be able to predict response to cardiac resynchronisation therapy differentiating between nonischemic and ischaemic cardiomyopathy le-cmr may allow to precisely delineate the presence and the location of fibrosis, avoiding the placement left ventricular leads in necrotic areas and in particular when the scar is located in the posterolateral area. at the beginning of the presentation, a short overview will be given on the relevance and prevalence of valvular heart diseases with special focus on europe and the data collected in the euroheart survey. also, the aetiologies of the various valve diseases will be briefly mentioned. in the second part, special attention will be given to aortic stenosis (as) and mitral regurgitation (mr), as these two entities are responsible for up to % of all valvular heart diseases. echocardiography is well accepted as the backbone examination of any valvular heart disease evaluation. echocardiographic criteria for severe as and mr are reviewed as well as their importance for the indication of valve repair or valve replacement. in the third part, we will focus on newer "cross-sectional" imaging technologies such as cardiac mr and ct. in particular, the strengths of cardiac mr in the assessment of regurgitant lesions will be demonstrated. in addition, the value of cardiac mr for the pre-interventional evaluation of congenital heart diseases will be discussed. similarly, the crucial role of cardiac ct in the workup for transaortic valve implantation (tavi) will be illustrated. finally, the yield of these novel techniques in the context of combined valvular and ischaemic heart disease will shortly be reviewed. learning objectives: . to better understand the role of cardiac mri in valvular heart disease. . to appreciate the added value of mri vs other imaging modailities. cardiac resynchronisation therapy (crt) and transcatheter valve heart (thv) interventions have been important therapeutic breakthroughs for heart failure patients and patients with valvular heart disease and contraindications for surgery. patient selection is crucial to optimise procedural outcomes and therapeutic efficacy. multimodality imaging plays a central role in patient selection and procedural strategy planning. assessment of left ventricular dyssynchrony, cardiac venous anatomy and extent and location of myocardial scar are key to identify heart failure patients who will respond to crt. stepwise use of non-invasive imaging modalities, including echocardiography, nuclear imaging, magnetic resonance imaging and multidetector row computed tomography (mdct), has been common practice to select patients for crt. recent technological advances have allowed integration or fusion of imaging modalities to create -dimensional models that provide relevant information. multimodality imaging is also crucial to optimise the results of thv interventions. detailed anatomical and geometrical evaluation of the mitral valve with current -dimensional imaging techniques has permitted better selection of patient candidates for transcatheter edge-to-edge mitral valve repair and accurate procedural guidance. for transcatheter aortic valve implantation, accurate measurement of the aortic annulus with -dimensional echocardiography or mdct allows us accurate selection of the prosthesis size to minimise the risk of complications, and evaluation of the procedural access (transarterial or transapical) with mdct permits accurate procedural strategy planning. the learning objectives of this session include: providing an overview of current imaging modalities to select patients for crt and thv interventions and how to use them in clinical practice. the inner layer of the anal canal consists of squamous and columnar epithelium with the transitional zone at the dentate line. the muscular component of the anal sphincter consists of an inner layer of circular smooth muscle (internal sphincter), extending downwards from the rectum, and an outer striated muscular layer extending downwards from the levator ani muscle, comprising the puborectalis muscle and the external sphincter. between these layers is the fat-containing intersphincteric space, including the continuation of smooth-muscle fibers of the longitudinal muscle of the rectum. outside the anal sphincter is the fat-containing ischioanal space. magnetic resonance imaging (mri) and endoscopic ultrasonography have become the mainstay for preoperative imaging of perianal fistulas. for complex tracts, mri seems preferable. mri can be performed using an endoluminal coil or a phased-array surface coil. a state-of-the-art imaging protocol should include t tse sequences in three orthogonal planes, with the axial and coronal sequences angulated at the anal canal. addition of a fat-saturated t tse sequence is recommended for optimal conspicuity of inflammatory changes and post-contrast t -weighted imaging can also be helpful. intersphincteric infection is the principal feature of perianal fistulas; this is generally not found in other conditions. veins can be mistaken for fistulas, but in contrast to fistulas, veins usually are thin-walled, tortuous, symmetric structures. a pilonidal sinus may resemble a fistula, but absence of extension to the intersphincteric space helps one to discriminate between them. haemorrhoids and anal tags may resemble small submucosal fluid collections, but are easily diagnosed at clinical examination. this lecture will describe the pathogenesis of fistula-in-ano, with a focus on cryptoglandular disease, so that the radiologist can understand how the various classifications of fistulas arise. the park's classification for fistula-inano will be described with reference to cryptoglandular disease and other aetiologies. the surgical questions that need to be answered by imaging will be presented, followed by a description of what the radiologists need to include in their report for it to be clinically useful. the role of anal ultrasound and mri for preoperative fistula imaging will be described, with explanation as to why the latter is fundamental to modern management of the disease. perianal fistulising disease develops in approximately half of the adult and paediatric patients with crohn's disease (cd), with a relevant impact on their quality of life. in patients with cd, particularly those with severe fistulising ileal or colo-rectal disease, complex ramified anal fistulas associated with abscesses (parks' classification type - or s.james hospital classification type - ) are more frequently observed than in other patients. perianal disease thus represents a severe complication of cd, which may change disease management, by requiring specific pharmacological and/or surgical treatments. a non-responsive disease eventually may lead to total proctectomy with ileostomy. mri represents the gold standard diagnostic modality, because it provides a comprehensive staging of both enteric and perianal disease, with high accuracy and without invasiveness. moreover, mri is the ideal tool for monitoring disease response to therapy, being able to detect the inflammatory activity of perianal fistulas with high accuracy. so far, several clinical indexes have been proposed to measure fistula's activity, including the perianal disease activity index (pdai). similarly, several mri classifications of fistula's activity, based either on t -weighted or t -weighted contrast-enhanced imaging, have been proposed, although a widely accepted imaging severity score is not available yet. several cases of complex perianal fistulising cd of increasing severity will be shown, with anatomical and clinical correlations, staging and activity description, particularly in patient follow-up and treatment monitoring. finally, differential diagnoses with other benign conditions which may affect the anal canal, including inflammatory infective diseases, hydradenitis and pilonidal disease will be discussed. ground glass opacity (ggo) is characterised on hrct by the presence of a hazy increase in lung opacity that does not cause obscuration of underlying bronchial and vascular margins. although a very common finding, it also constitutes a very nonspecific term since it can be seen in a variety of different intraalveolar and interstitial processes with different histology including inflammatory, infectious and neoplastic diseases that have a common physiologic mechanism: partial displacement of air. ground glass opacity may even be seen in normal processes such as poor ventilation in dependant lung areas and in expiration. moreover, ggo can represent either an ongoing, active and potentially treatable disease or an irreversible process. to interpret correctly this highly nonspecific but very significant finding, it is crucial to attempt to further classify the different large main entities in which this radiological finding appears. are there specific radiological and hrct findings that can help us differentiate ggo in autoimmune inflammatory conditions from infectious and neoplastic processes? are there associated findings other than ggo, such as nodules, reticulation or focal disease, and distribution of findings that can narrow the differential diagnosis? systematic evaluation of ggo and associated findings as well as integration with clinical information (acute, subacute or chronic symptoms) is essential in defining ggo subtypes to improve the radiological diagnosis. radiologists who regularly review high resolution ct (hrct) should be aware of the range of patterns and, more importantly, their potential pathological meaning. a pattern of ground-glass opacification is one of the more common hrct findings but, to the unwary, its interpretation can be problematic. an important underlying principle is that a ground-glass pattern may be caused by any process -physiological or pathological -which partially displaces air. physiological (i.e. non-disease-related) ground-glass opacification is perhaps most commonly seen in subjects who, for whatever reason (e.g. breathlessness, obesity), are unable to maintain or achieve a satisfactory inspiratory effort during scanning. a generally increased lung density (in contrast to adults) is also a feature in infants and young children simply because there are fewer alveoli in the developing lung. finally, it is worth noting that intravenous contrast administration (presumably because of a relative but temporary increase in capillary blood volume causing partial displacement of air) can unpredictably increase lung density. disease processes which lead to partial filling of the air spaces, thickening of the interstitium, partial collapsing of alveoli, and/or increased capillary blood volume will also manifest as a pattern of ground-glass opacification. in clinical practice, the recognised causes of ground-glass opacities on hrct include pulmonary oedema (cardiogenic or otherwise), infections (e.g. pneumocystis jiroveci pneumonia) and some of the idiopathic interstitial pneumonias (e.g. non-specific interstitial pneumonia and respiratory bronchiolitis associated interstitial lung disease). the presentation will review and revise the causes of physiological and disease-related ground-glass opacification on hrct. ground-glass opacity (ggo) is defined as increased attenuation of the lung parenchyma without obscuration of the pulmonary vascular markings on ct images. ggo may be the result of a variety of interstitial and alveolar infectious and noninfectious inflammatory diseases. as an imaging finding alone, ggo does not usually allow a specific diagnosis. ggo in inflammatory disorders is often present in the company of other interstitial or alveolar findings. however, the number of diseases that cause diffuse isolated ggo or ggo as the predominant finding is relatively small and can be prioritised with clinical information. the most common cause of diffuse isolated ggo in immunocompromised hosts are a variety of diffuse, opportunistic pneumonias, e.g. pneumocystis jiroveci pneumonia (pcp), cytomegalovirus pneumonia (cmv) or herpes simplex pneumonia (hsv), which constitute the first differential. chronic onset disorders in immunocompetent patients include cellular nonspecific interstitial pneumonia (nsip), subacute hypersensitivity pneumonitis (hp), organising pneumonia, air-space sarcoid, and drug-induced lung disease. in these disorders, ancillary findings such as an associated reticular pattern with traction bronchiectasis/bronchiolectasis (nsip), mediastinal lymphadenopathy (sarcoidosis), superimposed nodularity or cysts or may help to refine the diagnosis. in patients with collagen vascular disorders, e.g. scleroderma, ggo secondary to pulmonary involvement needs to be differentiated from drug-induced lung disease. this refresher course will put ggo in the context of outpatients versus inpatients, the acuity of clinical symptoms, e.g. fever, cough and dyspnoea, signs of massive systemic inflammation, and the clinical situation such as inhalational history, pneumotoxic drug administration, immunocompromise, or bone marrow suppression. in this course, we will review the dysplastic and neoplastic conditions that are associated with persistent ggo in the lung parenchyma. we will separate these conditions into those that are responsible for localised diseases such as single nodular ggo and those responsible for more extended diseases. nodular ggo can be separated into pure or mixed or part-solid nodules. pathology-radiology correlations show that nodular ggo are related to atypical adenomatous hyperplasia (aah), adenocarcinoma (adc) in situ (ais), minimally invasive adc (mia), and invasive adc according to the new iaslc/ats/ers lung adenocarcinoma classification. differential diagnosis includes exceptional metastases of angiosarcoma and melanoma as well as infection, inflammation and localised fibrosis. diffuse ggo related to neoplastic conditions is rare and may be due to lepidic adc (former advanced adc with bac component), diffuse large b-cell non-hodgkin's lymphoma, intravascular lymphomatosis (ivl) or mucosa-associated lymphoid tissue (malt) lymphoma. these neoplastic diseases should be differentiated from infectious and inflammatory causes of diffuse ggo. we will review the value of different morphological ct criteria to differentiate benign from malignant localised ggo, such as the size, the morphology (round, oval, flat), presence of mixed vs. pure ggo, and the multiplicity of nodular shadows. discussion will also include the changes that may occur within the nodule as well as the mean doubling time. the role of pet ct and transthoracic biopsy will be discussed. finally, we will present the current recommendations regarding the management of nodular ggo. it is the purpose of this lecture to briefly discuss ankle distortion-related lesions with emphasis on failed radiographic diagnosis, discuss their significance and management implications and define the diagnostic pathways in the acute and subacute stage. significant lesions are those with no return to sports activities after months; these have our major interest. in the acute setting of ankle distortion, the ottawa clinical decision rules are accepted to decide whether or not radiographic evaluation is needed; about % of radiographic examinations can be avoided. additional evaluation is restricted to patients with residual pain during reevaluation one week after trauma. assessment of fractures with the lauge hansen classification has a major advantage, as it defines the stability at the talocrural and distal tibiofibular joint. only in restricted cases, additional examinations may be needed to rule out instability. although radiographs easily detect displaced fractures, occult fractures may occur; also, grade ii-iii ligament lesions and peroneal tendon dislocation may go undetected. additional imaging evaluation should focus on the detection of significant lesions. significant lesions are talar fractures and complete calcaneofibular ligament tears. the latter is explained by the instability at the level of the subtalar joint with associated sinus tarsus syndrome. in case of residual local tenderness posterior and inferior to the lateral malleolus, the fibulocalcanear ligament and retinaculum peroneorum lesions are best evaluated with (dynamic) ultrasound. mri or ct is used to detect occult fractures in case of residual talocrural joint effusion with anterior talar tenderness during endorotation and plantar flexion. the ankle and foot can be affected by inflammation from a number of diseases. the main focus of this lecture will be inflammation secondary to infection and systemic inflammatory disorders. due to its function in locomotion and weightbearing, the foot is specifically susceptible to infections secondary to penetrative trauma including foreign bodies or to abnormal repetitive pressure. in addition, both seropositive and seronegative arthropathies can affect the foot. other unusual causes of inflammation include chronic recurrent multifocal osteomyelitis, osteoarthritis, mechanical disorders and sensory loss. this lecture will deal with the various patterns of inflammation in the context of pathogenesis and discuss the imaging features that aid in diagnosis. the importance of the clinical context in diagnosis will also be discussed. characterising tumours before deciding on the most appropriate treatment is a general rule in oncology. cystic tumours are now adequately graded based on bosniak's classification. however, this pre-therapeutic characterisation is not currently systematic in case of solid renal tumours, as it remains difficult in clinical practice due to the high prevalence of renal cell carcinomas (rcc) and the lack of reliable imaging criteria for recognition of benign versus malignant tumours. as a result, approximately % to % of surgically excised renal masses are actually benign. to avoid these unnecessary ablations, pretherapeutic image-guided biopsy has been proposed, but its use is still debated as it is invasive and time consuming. new validated imaging criteria, able to accurately differentiate the most common tumour types but also indolent vs more aggressive malignant lesions, would be useful to reduce the number of unnecessary biopsies or to consider the most appropriate treatment for a tumour or active surveillance. among these, mr imaging plays a major role based on chemical shift gradient echo (gre) sequences, signal intensity on t -weighted images, dynamic contrast-enhanced sequences, diffusionweighted sequences and late contrast-enhanced images. multiparametric mr imaging is now performed in clinical practice in most primary care centres. therefore, using different combinations of two or several parameters, it is now possible to distinguish certain renal tumours. a larger validation of all these combinations is still necessary to define those having a clinical significance for routine practice. targeted therapies such as vegf mab bevacizumab, vegf tyrosine kinase inhibitors (sunitinib, pazopanib, axitinib and sorafenib) and mtor inhibitors (temsirolimus and everolimus) have been approved and included in european guidelines. pre-therapeutic imaging prerequisites are mainly based on the natural history of mrcc. this justifies imaging through a thoracoabdominal ct scan. outside clinical symptoms, there is no need for bone imaging or brain ct scan; the latter nevertheless could be done because of influencing the therapeutic strategy. pet scan should not be done due to low sensitivity. the evolution of metastatic sites by comparison from previous imaging is useful to evaluate the natural history. outside clinical trials or research imput the strategy is to use the drug in an optimal way to increase pfs. so far, monitoring has to inform about change in recist criteria for cr, pr, sd or pd, and if pd is a real pd such as recist pd or a slowing pd. in conclusion, standard ct scan and recist are adapted to real life. nevertheless, more information on the mode of action of targeted drugs should be interesting for knowledge and research purpose, without any impact on the management of patients. additional imaging could be done with additional criteria than recist on ct scan, by vascular functional imaging and pet scan. breast mri is recognised as a useful tool for a number of clinical indications, but remains relatively challenging. recognising the importance of an evidencebased approach to the development of protocols will influence the further integration of this technique into everyday clinical practice in the management of breast cancer patients. there remain concerns regarding the potential of over-diagnosis from breast mri. the aim of this integrated rc is to inform the clinical practice with regard to the establishment of breast mri in specific clinical and imaging scenarios, namely, using mri for surveillance in high-risk patients, monitoring response to neo-adjuvant chemotherapy and developing protocols for the indeterminate imaging scenario of non-mass-like enhancement. the establishment of protocols to maximise the specificity and sensitivity of the technique of breast mri in such indeterminate clinical and imaging scenarios will be discussed. non-mass like enhancement is a frequent finding in breast mri. it relates to the fact that enhancement occurs in the normal-appearing fibroglandular tissue that surpasses that of the other parts of the parenchyma. there is no associated space-occupying lesion. in pre-contrast non-fat-suppressed or fatsuppressed t -and t -weighted images, there is usually no correlate observable. the differential diagnosis of non-mass-like enhancement is between benign nonspecific fibrocystic disease/adenosis, hormonal stimulation, and subclinical mastitis on one hand, vs dcis or (less likely) diffusely infiltrating (usually lobular) cancer on the other. key components of differential diagnosis is configuration of the enhancement (does it follow the orientation of the milk duct or not?) and symmetry (symmetric or asymmetric). less important criteria are internal enhancement (internal architecture) and enhancement kinetics. it is important to realise that enhancement kinetics can only be used to corroborate the suspicion of dcis -but they cannot be used to alleviate the indication to biopsy a finding which, based on configuration and asymmetry, is suspicious. management depends on the different constellation of clinical, mammographic, and mri findings. it usually includes short-term follow-up ( months) and, if stable/persistent, mr guided vacuum biopsy. assessment of the change in tumour burden is an important feature of the clinical evaluation of cancer therapeutics. tumour shrinkage and time to the development of disease progression are important end points in cancer clinical trials. however, these end points are useful only if based on widely accepted and readily applied standard criteria. criteria, known as recist, were published and are updated on a regular base. the revised recist includes a new imaging appendix and underlines the importance of moving from anatomic unidimensional assessment to either volumetric anatomical assessment or functional assessment with pet or mri. the goal of the lecture is to discuss the value of the different mr imaging techniques (e.g. mr spectroscopy, diffusion-weighted imaging, angiogenesis mapping, pet-mri), which evaluate response to treatment in breast cancer. participants will learn about particular imaging challenges of assessing response to neo-adjuvant chemotherapy. in addition, they will learn to understand clinical aspects of neo-adjuvant chemotherapy based on recist. currently, the esr has been using social media in many successful ways to improve the contacts among members and to boost our fast development as one of the most advanced scientific communities in the world. our session "the role of social media in radiology" presents the view of experts, acknowledged leaders of international it projects. it seems to be important to discuss the major implications of social media for radiology, the leading clinical discipline in information technology. davide caramella will clarify the relationship of social media to professional societies and especially to the esr, including their basic features. the "social media generation" has developed from schoolkids to students and, recently, to teachers as well. alexander sachs represents the young professional expert of elearning in radiology and will reflect the current user behaviour. osman ratib will share his view on how social networks enhance the communication between radiologists, nonradiologists and patients. critical aspects of the pros and cons of large communication scenarios are followed by some ideas of future developments. jürgen brandstätter will indicate the patient´s view. most patients see social network communications with their radiologist as useful for, e.g. appointment management or other non-sensitive communication. besides communicating personal health data, a wide area of potential useful other possibilities are at hand, like anonymised health data for reviewing purposes, second opinion or education of patients. the fast development of social media may induce hopes and fears; the four speakers of this session will present them in a panel discussion. the widespread use of social networks in all professional activities has attracted a great interest in the recent years. in medical applications, the main concerns include the possible consequences in terms of patients' confidentiality protection, perceived role of the physician, patient-physician relationship, and caregivers' distraction during professional activities. there are no doubts, however, that social networks are beneficial in enhancing the information flow between scientific societies and their professional members, without any interference at the point of care. social networks have proved to be extremely effective in updating members about relevant news concerning their areas of interests and in allowing them to be informed about the educational activities promoted by their scientific societies. this presentation focuses on the social network activities promoted by the esr via facebook, youtube, twitter and other blogs. impact of social media on the training of radiologists a. sachs; vienna/ at (alexander.sachs@meduniwien.ac.at) in modern times, social media have gained more and more important value in exchanging first experiences in radiology from resident to resident. social networks such as facebook, twitter, google plus and others are used to start discussion groups about cases and share the growing knowledge with different educational concepts in each hospital. growing e-learning modalities as moocs, applications for smartphones or tabs and web-based case series or learning platforms support the training of radiologist in social media. modern elearning platforms use social media as their first choice of sharing knowledge around the whole world about certain illnesses, cases or different decisions of therapy. though sharing information seems to be easier and shows several advantages than in former times, social media can also highlight new problems. this lecture focuses on user behaviour to improve learning, the pedagogic evidence for large group learning with networks and the importance of using networks to manage time. social media networks in communication between radiologists, nonradiologists and patients o. ratib; geneva/ch while social networks have gained unprecedented popularity in our daily environment, they have only limited utilisation in medical applications. the paradigm of communication within a given community for exchange and sharing of imaging data would certainly have its place in a medical environment where there is an increasing need for multidisciplinary collaborative work. the required security and confidentiality in management of sensitive patient data have often precluded the usage of existing public social networks. professional proprietary systems have emerged in a more restricted way with much less flexibility leading to limited use. the exchange of medical images adds a level of complexity due both to the size of the data and the need for appropriate browsing and visualisation tools. traditional pacs and teleradiology system offer such features, but in a very limited way, far from the desired convenience of social networking between groups of users. a prototype of such a system aimed essentially at the community of users that need to exchange anonymised medical images and data for academic and research purposes will be presented and discussed. the project nicknamed dicom sandbox is being tested and evaluated by some users in europe under the coordination of the esr subcommittee of ehealth and informatics. developed with open source software components, it allows using existing cloud-based storage services with additional layers of secure dicom file management and visualisation. it allows the community of users to share dicom data with all social networking and notification features beyond simple peer-to-peer exchange. interactions between patients and radiologists through social media j. brandstätter; wiener neudorf/ at (j.brandstaetter@codewerk.at) social networks have become integral parts of most citizen's use of the internet. especially, the younger generation uses social networks extensively; however the way those tools are used should be chosen carefully. it is one thing to use a social network for private interests, but a different one to use it as transport media for sensible data like personal health-care data. the sensitiveness of citizens regarding privacy has been increased recently by different events of misuse or fraud, especially accelerated by the recent privacy affairs caused by governmental agencies. though most patients see social network communications with their radiologist useful for, e.g. appointment management or other non-sensitive communication, personal medical reports or images shall not be shared across this media. besides communicating personal health data, a wide area of potential useful other possibilities are at hand (e.g. anonymised health data for reviewing purposes, second opinion, education of patients, …). future developments shall make use of social media where useful, but always with deep respect to the patient's privacy rights. apart from the historical, but ever present clinical expectation from the neuroradiologist is the answer to the most important question if the patient has a brain tumour or not. nowadays, due to the outstanding technological advances in imaging techniques during the last decades, the stakes are growing much higher and it is the neuroradiologist on whom lies the initial responsibility of answering a large number of questions that are of utmost importance for further brain tumour patient care and treatment. having the opportunity to provide the usual and generally basic information regarding tumour morphology and exact location and to narrow the differential diagnostic possibilities, neuroradiologic expertise now opens a wide spectrum of techniques, especially those derived from mri, such as mr spectroscopy, mr perfusion, dwi, dti, swi, etc. these provide us the unique possibility of not only obtaining and comparing images, but also of exploring multiparametric brain maps, allowing us to gain insight into the internal architecture of the tumour and the functional and molecular features of the tumorous tissue. this consequently defines the actual stand of neuroradiology, indicating the uprising clinical urge not only to provide the answers regarding the tumour type, grade and possibilities of the most appropriate and patient-individualised therapy modality, whether surgical or nonsurgical, but also to accurately assess the post-treatment tumour behaviour and status, by differentiating the true tumour progression from pseudoprogression, represented by inflammatory responses to the radio-and/or chemotherapeutical treatment, and to monitor other neurooncological response assessment criteria. a. differential diagnosis and pseudo-tumoural lesions h.r. jäger; london/uk (r.jager@ucl.ac.uk) conventional contrast-enhanced mr imaging gives an indication about the likely tumour type and histological grade, but advanced mr imaging methods, including perfusion-weighted (pwi) and diffusion-weighted (dwi) imaging, mr spectroscopy (mrs) and pet imaging provide additional information which helps to refine the diagnosis of intrinsic brain tumours. contrast enhancement can be a feature of low-grade (pilocytic astrocytoma and ganglioneuromas) as well as high-grade gliomas (who grade iii anaplastic gliomas and who grade iv glioblastomas). infiltrative low-grade (who grade ii) astrocytomas do not usually enhance, whereas who grade ii oligodendrogliomas can enhance. higher-grade glial tumours (who grade iii and iv) tend to have an elevated relative cerebral blood volume (rcbv) on pwi, a decreased adc on dwi and increased choline turnover on mrs and pet imaging. a raised rcbv can, however, also be seen in low-grade oligodendrogliomas, particularly in those with a p/ q chromosomal translocation, which is associated with a better response to chemotherapy. pwi and dwi are useful in the differentiation of glial neoplasms from cerebral lymphoma, which has an increasing incidence in the immuno-competent population. dwi and pwi are also very useful in the differentiation of enhancing pseudo-tumoural lesion such as cerebral abscesses, tumefactive demyelination, pseudoprogression and radiation necrosis. perfusion imaging plays an important role in the characterisation and management of brain tumours. low-grade brain tumours may transform into high-grade gliomas at some point, which is a transformation that is highly variable and difficult to predict in an individual patient. conventional contrastenhanced computed tomography (ct) and magnetic resonance (mr) imaging are found to be insufficient to depict this transformation and angiogenesis and therefore to detect malignancy. however, perfusion imaging provides additional information over conventional imaging in terms of tumour physiology and haemodynamics, providing important biomarkers for malignancy and prognosis. perfusion ct and mr perfusion techniques such as arteriel spin labelling and dynamic susceptibility-weighted perfusion imaging are more practical and widely used compared to nuclear medicine methods. these perfusion techniques should be considered and used in routine clinical workup of brain tumours to asses grading and prognosis. maximal safe resection, radiation therapy and temozolomide chemotherapy are the current standard of care for newly diagnosed high-grade gliomas. still, hgg have a poor survival rate. a contributing factor to the poor survival is the inability of currently available imaging techniques to accurately delineate the tumour, with the result that targeted focal treatment my not be effective. conventional imaging is not able to give an early assessment of the effectiveness of radiation and/or chemotherapy. in addition, conventional imaging has difficulties in differentiating pseudo-progression, which is a common phenomenon in conventional chemo-radiation therapy, from true progression. early identification of patients who suffer from tumour recurrence can be of great advantage: it provides the opportunity to adjust individuals more rapidly, sparing patients unnecessary morbidity and delay in the initiation of other, maybe more effective, treatment. in recent years, different functional imaging approaches such as perfusion mri, diffusion-weighted imaging and spectroscopy, have been complementarily used for imaging evaluation of treatment response. in this lecture, different advanced mr and ct imaging methods used to support differentiation between pseudo-progression and true tumour progression to assess treatment response will be discussed. in addition, a novel recently published promising technique, the parametric response mapping (prm), a novel voxel-wise analytical method of monitoring physiological and environmental changes in a tumour volume during treatment will be presented and compared to the traditional methods used. finally, the aim of the lecture is to consolidate the present knowledge and novel ideas in brain tumour imaging for assessment of pseudo-progression versus true tumour progression. learning objectives: . to understand the challenges and limitations of routine mri in monitoring brain tumour treatment. . to become familiar with the role of advanced imaging biomarkers for early assessment of treatment response. . to learn how to integrate routine and advance mri into clinical practice after tumour therapy. the purpose of this lecture is to describe the role of functional radiological imaging modality-ct and mr perfusion in the evaluation of head and neck tumors. conventional contrast-enhanced ct and/or mr imaging are the current standard techniques for the diagnosis and treatment evaluation of the head and neck tumors. however, this method is limited in its ability to depict the angiogenesis which is a hall-mark of tumor growth. perfusion imaging provides a rapid evaluation of tissue perfusion and can be easily implemented in every head and neck ct or mr protocol. the determination of tissue perfusion is based on examining the relationships between the arterial, tissue and potentially the venous enhancement after the introduction of a bolus of contrast material. the quantification of the perfusion values helps to outline the malignant tissue as well as to differentiate recurrent disease from nonspecific post-therapeutic changes and can be used as a therapeutic monitoring tool during and after tumor therapy. in recent years diffusion-weighted mri (dwi) gained increasing importance and also acceptance in head and neck radiology. although this functional technique is challenging due to air tissue interfaces, its ability to improve the diagnostic accuracy is widely accepted. dwi can be applied to detect lesions not only in adults, but also in children. it is helpful in lesion characterisation and has a great potential to differentiate recurrence from posttherapeutic changes. initial promising results to improve lymph node staging however are difficult to reproduce. image interpretation is performed qualitatively based on visual assessment of the high b-value images (b= - sec/mm ) and the corresponding apparent diffusion coefficient (adc) map. quantitative image analysis is based on the measurement of the underlying adc value. solid tumors including recurrences are bright on the high b-value images and dark on the corresponding adc map, whereas posttherapeutic changes are bight on both the high b-value images as well as the adc map. cystic and necrotic lesions are typically dark on the high b-value image and bright on the adc map. furthermore, malignant lesions tend to have a lower adc compared to benign and cystic lesions provided that no necrosis is present. therefore, image interpretation always has to be performed together with morphological images to avoid misinterpretation of functional images. pet/ct has established itself as a robust, rapid and reliable technique in head and neck oncology. it is mainly used to stage nodal disease, to detect distant metastases and synchronous tumours, to identify unknown primary tumours in patients with metastatic neck nodes, to assess treatment response and prognosis after chemoradiotherapy and for radiotherapy planning. mri with diffusion-weighted imaging (dwi) is routinely used for the assessment of submucosal tumour spread, to stage nodal disease, to monitor treatment response and to detect recurrent disease. the information provided by pet/ct and mri is often complementary and the recent implementation of hybrid mr/pet systems in clinical settings holds promise because it can combine morphologic, functional and molecular information. this lecture focuses on clinical applications of pet/ct and mr/pet in the head and neck. current evidence about the combined use of pet/ct and mri with dwi is discussed. the principles of mr/pet data fusion are summarised as well as current knowledge regarding the diagnostic performance of mr/pet in the head and neck. typical radiologic findings of tumour manifestations are reviewed with particular emphasis on the early detection of lesions, their appearance on different imaging modalities and the added value of hybrid imaging techniques. the variable appearance of functional phenomena mimicking disease as well as the potential pitfalls of image interpretation and how to avoid them are equally addressed. major emphasis will be put on how to report the findings in a comprehensive way. multi-energy and functional imaging ct has reached a robustness allowing the use of this new technology in clinical routine for a variety of different clinical questions. this presentation will discuss the basic principles and the strengths and limitations of the techniques. implementations of multi-energy methods for material characterisation and of ct methods for functional imaging will be discussed. contrary to normal single-energy ct systems, multi-energy ct scanners allow simultaneous scanning at two peak x-ray energies. when the attenuation is measured at two energies, their values are not exactly proportional to each other, which open new diagnostic possibilities. measurements at two spectra can be achieved using multiple kvp and/or filtration or with detectors with energy discrimination. these methods have different pros and cons such as sensitivity to subject motion and dose efficiency. to be able to acquire functional data such as perfusion, images are acquired dynamically following the injection of a contrast agent and physiological models are used to convert the measured contrast agent concentration to perfusion estimates. methods that acquire multiple images have the potential to increase the radiation dose to the patient, so ct protocols need to be optimised. in computed tomography, d images are reconstructed from a number of x-ray radiographs that were acquired at different angles. tomographic reconstruction relates to the generation of a d image based on these acquired radiographs. most tomographic reconstruction methods are based on a technique called filtered back projection (fbp) in which a sharpened version of the radiographs are back projected to generate the d image. while it has been the standard reconstruction method, mostly due to its speed and good quality if many radiographs are available, iterative reconstruction methods are emerging. the reason for that is that fbp suffers from important disadvantages. in this talk, the basic concepts of iterative reconstruction are explained and its virtues are detailed. the flexibility of iterative reconstruction methods is demonstrated for the field of x-ray computed tomography. finally an outlook is given with respect to future research on tomographic reconstruction. interventional radiology (ir) has been increasingly applied in the management of obstetric and gynaecological haemorrhage. national reviews of maternal deaths from post-partum haemorrhage have recognised that maternal deaths may be prevented by ir and that all obstetric units should have protocols and arrangements in place to ensure appropriate and timely referral to ir. the important role that irs play in the management of fibroids has also highlighted how these techniques can be applied to other gynaecological conditions which may result in haemorrhage. this session will start by describing which gynaecological disorders may result in haemorrhage and the technical aspects of treatment followed by the published evidence for embolisation. the session will then focus on obstetric haemorrhage, with an emphasis on prophylaxis and how women deemed at high risk for haemorrhage might be managed safely. the techniques used for prophylaxis of haemorrhage will be described in detail and the published data presented. the final presentation will concentrate on the technical aspects of embolisation when haemorrhage has occurred to ensure the best results by knowing the relevant anatomy and appropriate technique of embolisation. the published evidence of the results of embolisation in pph will be presented. these presentations will be followed by a panel discussion on how irs can reduce the radiation dose whilst simultaneously ensuring successful outcomes for their therapies, an important issue in a young and fertile group of women. gynaecologic massive bleeding related to benign and malignant causes is under the scope of all radiologists because of being diagnostic and also because of its implication in interventional radiology procedures. the causes are uterine fibroids (uf) and post-partum hemorrhage (pph). however, oncologic disease, arterio-venous abnormalities, or iatrogenic injuries are less commonly known to be eligible for embolisation procedures. usually, surgical homeostasis attempts or radical hysterectomy is preferred to embolisation procedures. despite the good clinical outcome for hysterectomy and the hazardous success rate of conservative treatment, both are unsatisfactory to patients. pelvic artery embolisation (pae) could dramatically change the patients' clinical issue. the purposes of this lecture are to: revise the pelvic arterial anatomy (possible arterial anastomosis or variation); outline the aetiology of non-obstetric massive menometrorrhagia; expose the typical clinical presenting features; describe the imaging appearances of the etiologies; and discuss patient section, procedure technique, pitfalls and complications. obstetric haemorrhage occurring mainly in the postpartum period is a significant cause of maternal morbidity and death. blood loss of over and ml after vaginal delivery or caesarean section, respectively, complicates % of deliveries. major blood loss of more than ml occurs following less than % of all deliveries. the causes include uterine atony, retained placenta products, placental abnormalities, uterine rupture, lower genital tract laceration, cervical ectopic pregnancy and coagulopathies. generally, treatment is based on administration of uterotonic drugs, vaginal packing, surgical ligation of uterine arteries and even hysterectomy is sometimes needed. transcatheter embolisation of uterine, pudendal, vaginal arteries and ovaries is considered to be superior and should be the first-line treatment for intractable obstetric haemorrhage where interventional radiology is available. angiography reveals extravasation of contrast agent in more than % of these patients. the most frequent extravasation is from vaginal artery, and then from pudendal and uterine arteries. depending on the extravasation location and haemodynamic stability of a patient, f catheters or coaxial microcatheter technique is utilised for catheterisation. gelatin foam, microparticles acrylic glue and microcoils are the most frequent embolic agents. the embolisation procedure usually does not require general anaesthesia and can be repeated if bleeding continues. primary success rate in bleeding control is reported from to %. the most commonly reported long-term side effects after embolisation were transient buttock numbness and urinary frequency. the transcatheter pelvic artery embolisation for obstetric intractable bleeding is a fertility-preserving alternative to hysterectomy. medical imaging has nowadays integrated the diagnostic armamentarium of anosmic patients regarding not only qualitative assessment of the olfactory tract, but also quantitative evaluation of olfactory bulb volumes which are known to closely correlate to the olfactory function. many clinical studies on various pathological conditions have evidenced the value of such measurements in the workup of olfactory dysfunction for both aetiologic and prognostic purposes. imaging workup also plays a role in the medico-legal evaluation of post-traumatic anosmia together with electrophysiological and clinical olfactory tests. technical improvements in fibre tracking (ft) using diffusion-tensor imaging (dti) and appropriate designs of olfactory stimulation at bold-based functional mri (fmri) are expected to allow insights into the neurophysiological processes and circuitry of olfaction in the very near future. imaging workup of the anosmic patients will be the cornerstone of this lecture. the relevance of the different imaging techniques will be detailed. beyond the workup of anosmia, a comprehensive overview of the most common lesions of the olfactory tract seen in clinical practice will be given. additional review of lesions observed at the anterior cranial fossa in the near vicinity of the olfactory tract will be done, which comprehensively includes all commonly observed developmental, traumatic, inflammatory and neoplastic conditions not arising from the olfactory tract. their potential impact on the olfactory function or on other sensory/neurological functions will be underlined. clues to clinical/radiological differential diagnoses of the most common conditions and radiological features of the anterior cranial fossa will be highlighted. the central skull base, the floor of the middle cranial fossa, has a complex anatomy and is pierced by a variety of foramina and canals providing crossroads for spread of disease between the extra-cranial head and neck and the intracranial compartment. disease spread into the intracranial compartment indicates a dismal prognosis, as often it becomes inoperable due to involvement of vital structures including cranial nerves, carotid artery and/or brain parenchyma. imaging plays a pivotal role in the assessment of these structures which are not amenable to clinical inspection and can dramatically change the patient's management. in this complex anatomical area, ct and mr have a complementary role in providing a comprehensive roadmap for treatment planning. of major importance is the pterygopalatine fossa, the cavernous sinuses and orbital apex. the former, lying between the pterygoid plates and the maxillary sinus, is a major crossroad between the intra-and extracranial compartments and should be carefully inspected. pathologies affecting the central skull base are varied and may originate from the skull base proper, from the middle cranial fossa or from the extracranial head and neck. by looking at the centre of the lesion, its tissue characteristics and pattern of growth, it is often possible to provide a useful differential diagnosis and, most importantly, to map the lesion's extent. here, we present a radiologist-friendly approach to central skull base pathology highlighting the most important features in the differential diagnosis and treatment planning. . the jugular foramen: the jugular foramen is an opening in the skull base. the radiologic evaluation requires high-quality imaging with ct and mr. angiography is reserved for preoperative embolisation. it is important to recognise the "pseudo lesions". the most common tumour of the jugular foramen is the paraganglioma. the second is the schwannoma of the lower cranial nerve, and the jugular foramen meningioma is the third most common. the differential diagnosis shall be discussed. . the cerebellopontine angle (cpa) and the internal auditory canal (iac): the iac is a bony conduit for several nerves and a vessel, the neurovascular bundle. the cpa is a cistern of the peripheral cerebral spinal fluid, and several anatomical structures pass through this. it is also the place of some frequent disease processes. most of the lesions in the cpa are benign tumours with order of frequency: vestibular schwannoma and meningioma. the third most common lesion is a benign cystic lesion: the epidermoid cyst. in the iac, you can encounter the same benign tumoural lesion as in the cpa, but also inflammatory lesions, viral lesions and malignant lesions. why is it important in your imaging protocol of the iac and cpa region to use gadolinium? the differential diagnosis of different lesions shall be discussed. learning objectives: . to learn about the imaging techniques and anatomy in the posterior cranial fossa. . to become familiar with the imaging findings of common posterior cranial fossa pathologies. . to learn how to differentiate between the lesions in the posterior cranial fossa. health care is characterised by several specific features making it different from traditional markets. two of the key features are: the impossibility of patients to make choices, due to asymmetry of knowledge between them and their health-care providers; the fact that patients create an agency relationship with health-care providers, delegating to them the decision about the care delivered. due to these characteristics and to the fact that patients are fragile when they need care, there is a natural tendency for patients to rely on healthcare professionals. it is in this complex socio-technical environment that radiographers must be aware of their roles and responsibilities and ensure professional autonomy and accountability; demonstrate an ethical and knowledgeable understanding of the profession; apply professional practice in securing, maintaining or improving health and well-being; develop knowledge, skills and competences that underpin their education and training, contributing to the wellbeing of the patient; understand that professional advancement arises out of evidence-based practice and is acquired through focused research. this demands for a focus on patient care and safety, based on high professional standards. ebp is the best way to achieve this desideratum, as it combines the best research evidence with clinical knowledge and expertise, addressing the patient in a holistic way. since the radiographer is the ultimate interface between patient and technology, it is crucial to create a real teamwork concept, using guidelines for the roles and responsibilities of each team member as the pathway to minimise practice error, maximising patient and staff safety. the use of ionising radiation in medical exposures is well documented across countries. recent studies indicate that a substantial fraction ( to %) of these procedures is unnecessary when taking into consideration patient clinical indication. defensive medicine, economic interest, poor patient clinical information, patient pressure and the lack of knowledge about other alternative procedures, taking into account the benefits and risks, are the principal causes of unnecessary radiological examinations. patients must be referred to a medical imaging department for an examination that should be justified according to clinically appropriate image criteria and informed of the potential benefits and risks. following justification, the optimisation of practice is essential to ensure the minimal exposure of radiation dose to the patient whilst diagnostic efficacy is maintained. the principal aim of optimisation is getting a narrower dose distribution, with lower mean and th percentile values of the diagnostic reference level. optimisation processes need to consider several factors including the type of examinations, body region, clinical information, available equipment technology and image processing tools. to implement this process, a multidisciplinary teamwork is essential where radiographers, being the pivot between patient and technology, have an essential role. to understand and apply optimisation procedures, radiographers should be aware of the international guidelines, standards and directives. all medical imaging procedures must be audited to analyse if the workflow processes (referral, justification, optimisation and radiation protection principles) are according to the recommendations to create a radiation protection safety culture. although we are approaching the year , a lot of clinical radiography is not based on best evidence. lot of clinical decision-making is based on, for example, traditional practices that we cannot argue as being the best possible ones. in this presentation, barriers to using research and other sources of evidence and facilitators of evidence-based (eb) radiography are described. the role radiographers have in developing eb guidelines and procedures may vary according to educational level and work description. developing eb guidelines is not the work of every radiographer, but obeying them is. the speaker gives some suggestions on what to do in a situation if there are no eb guidelines and procedures in ones' country or institution and no group that develops them. with the framework of "the new work order" and increasing implementation of new work processes, professionals are faced with new modes of collaboration requiring communicative skills as well as multiprofessional competencies between all the stakeholders across the hospital. some evidence about the consequences of ineffective communication in the field of radiography is presented. in addition to benefits associated with the procedure of clinical audit, uniform good quality protocols for examinations and uniform national/international criteria help to perform comparisons between radiology units. clinical audit also works as an educational tool and as a feedback tool for the staff on their performance. it also points out the need for corrective actions. initiatives from professional organisations are probably the most powerful engines for the construction of europe, and in many cases open the road for legal and political advances. many medical specialties felt that there was a need for the harmonisation and standardisation of medical training over europe. the esr has shared this opinion and has elaborated a unanimously appreciated training curriculum, which is constantly updated. establishing the ebr (european board of radiology), with the mission to organise a european diploma was the logical next step. today, examinations have already been organised and more that candidates from european countries and abroad have applied. the trainee who passes the examination proves his professional skills and has reached the objectives assigned in the curriculum. in this session, we will explain how esr organises the diploma. we will review, capitalising on our current experience, the main lessons learnt from the past editions, the keys for success and reasons for failure, as seen by the examiners. we will also learn about the "diploma adventure", from the point of view of a successful candidate. national radiology training programmes in europe differ to a large extent, regarding length of training, content, and the presence or absence of a national board examination for certification. thus, for cross-border radiologic activities, the level of training and expertise applying to radiologists is difficult to assess under these circumstances. there is an increasing number of medical subspecialties, which organise european diplomas. more than subspecialties (e.g. anaesthesiology, ophthalmology, urology, etc). organise european examinations, which are well received among the respective communities. growing numbers of residents in training successfully stand this test of knowledge and practical skills. recognition of european diplomas by national authorities is an issue, which will increase the level of acceptance. the present status of the recognition process in european countries will be summarised. case material for the diploma will test knowledge in general radiology at a standard to be expected at the end of training as outlined in the esr curriculum. all ten esr-recognised subspecialty areas of the curriculum will be tested. the written exam comprises two computer-based sections, a multiple response section, and a short case section. there will be multiple response questions. these questions comprise both multiple options format, a single stem with answer options, and short imaging studies where candidates are asked to either annotate the image from a drop-down menu, or select the radiological features displayed or differential diagnosis from the options given. a special scoring system has been developed to prevent guessing. the short case section comprises or case-based questions. you may practice using the self-assessment tests on the esr website which follow the same format as the written examination. for the oral cases, you must have a systematic method and perform under pressure, which takes practice. you may rehearse with experienced radiologists who feedback on your performance. you can break a radiology report into areas, observations, interpretation and conclusion. conclusions should take into account the clinical information. candidates often forget this under pressure, so ask for it to be repeated. your differential diagnosis should be logical and comprehensive. the commonest causes of an abnormality is a good topic to revise. suggest if further imaging tests are indicated and give an indication of urgency. each case is marked separately. as a well-known statement in the academic world of education, "assessment drives learning". with the help of carefully prepared and performed examinations, it is possible to emphasise and instill the training curriculum. due to different reasons, training of radiology in european countries is highly varied. like in our experience in turkey, a central examination system, accompanied by a neatly prepared training charter and curriculum, is a good starting point to solve the problem. we believe that a common european assessment system like edir will contribute a great deal to the harmonisation of radiological training and establishment of quality standard in every country. the need for harmonising the training is not unique to radiology. this is why in more than medical disciplines, european diploma examinations are now being organised. furthermore, in some disciplines like urology, annual and online "in-service assessment" of residents and practising urologists allows the participants to evaluate their current knowledge base against the current european standards, and also provides the trainers and directors knowledge about the teaching and learning process in education centres. although there are huge diversities among different european centres, the urgent need of harmonising radiology training and setting a standard of quality are felt by all radiologists in europe. establishing and institutionalising a common assessment system in european radiology appears to be the shortest way to reach this goal. this is why we should support "european diploma in radiology" by any means and work hard to promote its recognition throughout the continent. the majority of emergency ct studies reported by junior radiologists or general radiologists out-of-hours comprise brain scans. brain ct studies are often performed to detect acute life-threatening abnormalities, such as stroke, intraparenchymal or subarachnoid haemorrhage, cerebral edema, etc. misrecognition of these often subtle, but life-threatening abnormalities can lead to inappropriate patient management and worsen patient outcome. errors in interpretation can generally be categorised as either perceptual or cognitive in nature. perceptual errors are those in which the radiologist does not see the abnormality, resulting in a false-negative interpretation (e.g. basilar artery thrombosis, deep cerebral venous thrombosis, pres,…). cognitive errors, on the other hand, are those in which an abnormality is identified but the meaning or significance of the abnormality is not recognised. cognitive errors can result in false-positive interpretation if, for example, a normal anatomic variant is mistaken for a pathologic condition. while the more common normal variants and artefacts often do not present a problem to experienced clinicians and neuroradiologists, less experienced individuals should beware of these diagnostic hazards. to reduce false-negative and false-positive reporting, a checklist of the review areas (blind spots) to be verified on any brain ct scan will be offered and the most frequent normal variants will be discussed. pitfalls in brain imaging comprise perceptual or cognitive errors of interpretation of imaging findings. errors of perception refer to a missing of a pathology that is present on an image, while cognitive errors consist of a wrong interpretation of a seen structure or pathology. interpretation may be false negative or false positive. in addition, a wrong conclusion may be drawn resulting from missing, incomplete or false clinical information, or as a consequence of a lack of knowledge of differential diagnoses. perception errors are frequently the consequence of incomplete examination protocols where lesions are missed (for instance, acute ischaemic changes without the use of diffusion-weighted imaging) or signals, for instance, flow-related phenomena are misinterpreted. asymmetric positioning of the patient`s head may also lead to misinterpretation of side differences, especially in the temporal lobe. another source of perception errors might be reduced attention (especially at night shifts) and have been proved to occur in regions that are "potential blind spots", as the sulci, cavernous sinus, meckel cave, dural sinuses, brainstem, and skull base foramina. perceptual errors may be avoided by following a checklist when reading images. all sequences should be used for comparison of signal intensities to distinguish artefacts from pathology. another very important fact to avoid misinterpretation lies in a profound knowledge of morphologic and metabolic age-related changes of the brain and of anatomical normal variants (such as, for instance, accentuated virchow robin spaces in certain locations). cystic liver lesions can be classified based on their nature as benign and malignant. in the benign category are, among others, developmental and infectious/inflammatory cysts, while neoplastic cyst can be subdivided into primary and secondary. developmental cysts originate from abnormal ductal plate malformation and consist of hepatic (bile duct) cyst, bile duct hamartomas, caroli's disease and polycystic liver disease. infectious/inflammatory cysts include, among others, abscesses (pyogenic and amoebic) and hydatid cysts. primitive neoplastic cystic lesions are cystoadenoma and cystoadenocarcinoma. secondary lesions can originate mostly from mucinous tumours such as colon and ovary. the role of crosssectional imaging in the detection and characterisation of these entities will be discussed, with an emphasis on the differential diagnosis with ct and mr imaging. benign hepatocellular neoplasms are being recognised with increased frequency using cross-sectional imaging. one of the main goals is to be able to make a clear-cut differential diagnosis between focal nodular hyperplasia (fnh) and hepatocellular adenoma (hca) since patient management is substantially different. despite the sound knowledge acquired in the last few years about the morphological features of fnh using various cross-sectional imaging techniques, new players in the field have arisen such as diffusionweighted mr imaging (dwi) and use of hepatobiliary contrast agents. these new biomarkers offer a different view over fnh and allow a more accurate characterisation even in more atypical cases. concerning hca, current knowledge implies that the sub-types of this neoplasm should be known since, again, patient management differs and these patients are no longer seen as compulsory surgical candidates. these sub-types will be further discussed and the role of imaging for risk stratification (haemorrhage and malignant transformation) will be addressed. further, illustrative cases of combined fnh/hca cases will be demonstrated along with the main imaging clues for a successful differential diagnosis. imaging in the early postoperative period is complex. it is essential for radiologists to know the new anatomic arrangement after surgery and to understand the range of normal postoperative appearances. this latter aspect can be particularly difficult, because the radiological findings are affected by the type of operation performed (open vs. laparoscopic) and the complexity of the surgery (such as the presence of adhesions or haemorrhage) as well as the underlying comorbidity of the patient. patients may be considered in broad categories: haemodynamically unstable patients who may be bleeding; patients with distension or unopened bowels who may have obstruction or ileus; patients with sepsis in whom to search for an intraabdominal collection or leak; and patients who do not follow an expected recovery for which the cause is unclear. ct is the primary imaging technique, with ultrasound reserved predominantly for liver and renal transplants or assessment of perihepatic collections, and fluoroscopy for routine postoperative assessment of upper and lower gi anastomoses. ct protocols should be optimised to detect particular complications with positive luminal contrast for assessment of anastomotic leaks ( %), triple phase imaging for bleeding (without positive oral contrast) and delayed phase imaging to detect ureteric injury. since each operation has a specific range of expected complications, these should be considered when deciding on the protocol and in light of the patient's clinical status. excellent communication with the surgical team is required to make an accurate diagnosis and in particular where haemostatic compound or mesh for hernia repair was used. learning objectives: . to understand the best imaging options for investigating suspected postoperative complications. . to appreciate the normal appearances of the abdomen and pelvis after surgery, in particular with regard to free fluid, haematoma and free gas, and when to suspect complications. radiologists should be aware of the newer surgical techniques and expected post-operative alterations, to yield a correct interpretation of a post-surgery imaging examination, thus being able to differentiate a normal post-operative finding from a potential complication. in this setting, it is crucial to perform a technically adequate imaging examination, so that post-operative anatomical and functional findings may be evaluated. therefore, the aim of this lecture is to give an overview of the contribution of various imaging modalities in the diagnosis of late post-operative complications following gi tract surgery. it will include a mention of the technical issues that need to be considered to achieve better diagnostic accuracy, as well as a description and illustration of the main imaging findings of late post-operative complications. those include diseaserelated (recurrence for malignancy or inflammatory disease) and procedurerelated (anastomotic strictures, internal herniation, adhesions and intussusception, among others) complications. focus will be placed on crosssectional imaging techniques, which at present constitute the workhorse for detecting and characterising late post-operative complications after gi tract surgery. in this way, imaging has an obvious and pivotal role in planning therapy, since some late complications are usually treated in a conservative way while others require a surgical approach. lung biopsy of a suspected lung cancer remains a well-established technique for histopathologic examination in patients with suspected lung cancer. complications can nevertheless occur and therefore indication should be given after carefully balancing the benefits and drawbacks for a particular patient. the method should be clearly considered complementary to alternate methods of tissue sampling, fiberoptic endoscopic, mediastinoscopy or us-guided endoscopy, surgical biopsies and resection during a multidisciplinary discussion. several steps are needed in the preparation of the biopsy procedure, including informed consent, management of anticoagulation and anti-platelet medications, imaging assessment of the lesion and its environmental lung, and planing of the needle trajectory procedure. the choice of the guiding method and the most appropriate biopsy needle is still largely dependent on local skills and habits. ct-guided percutaneous biopsies using coaxial automated core biopsy needles offer many advantages. needle manipulation can help to reach difficult targets. participation of the patient is needed in terms of respiratory manoeuvres and breathhold. complications may occur such as pneumothoraces haemorrhage and air embolism. knowledge of the respective risk factors and the potential method of prevention or treatment by operators are essential. the therapeutic response of lung cancer can be objectively evaluated on the basis of changes in the tumoural size or depiction of metastases, such proposed in the recist criteria (response evaluation criteria in solid tumours). recent advances in cancer biology have triggered development of novel targeted therapies designed to disrupt specific biologic pathways. among them, antiangiogenic drugs represent a promising strategy for non-small cell lung carcinomas (nsclc). these agents are more cytostatic rather than cytotoxic, explaining the limitations of tumour response assessment based on morphological criteria. dynamic contrast-enhanced imaging techniques have the potential to quantify tumoural angiogenesis. in particular, dynamic contrastenhanced multidetector-row ct (dce-ct) represents a promising tool to provide non-invasive and repeatable assessment of the angiogenic process within non-small cell lung cancers (nsclc), offering the possibility to generate morphological and functional information from the same examination. dce-ct allows the calculation of regional tumour blood flow, blood volume, flowextraction product, and permeability-surface area product, over the entire tumoural volume, using mathematical models and dedicated softwares. dce-ct can depict early changes in lung cancer vascularity, before tumour shrinkage, that could help predict response to antiangiogenic drugs. becoming familiar with this technique enables the radiologist to integrate it into clinical practice and to assist the thoracic oncologist in the pre-therapeutic and follow-up evaluation of nsclc patients treated by antiangiogenic drugs. this new approach should improve patient's selection for targeted therapies. a- : it is now accepted that pet-ct is of value in the staging of lung cancer and reduces the number of unnecessary thoracotomies. its role has been further clarified by recent publications on its value in assessing mediastinal nodal involvement, tumour aggressiveness using either standardised uptake value or total glycolytic volume, and response to neoadjuvant chemotherapy. more recently, mri has been investigated as a tool to detect distant disease using diffusion-weighted imaging, and there are a number of trials that have compared both pet-ct and dwi mri retrospectively and prospectively. this presentation will summarise the current available data and the likely benefits of either imaging modality neurological deficits in children are an urgent condition that depends significantly on imaging for a prompt and accurate diagnosis because of the significant overlap present in clinical history, presentation and neurological examination. different imaging modalities, such as ultrasound, computed tomography (ct) or magnetic resonance imaging (mri), are utilised dependent on the age of the patient and neurological symptoms. the purpose of the present lecture is to discuss the causes and imaging appearance of acute neurological conditions in childhood, broadly classified into stroke, infection, inflammatory demyelination, metabolic disorder, cerebral neoplasm or neurotoxicity. imaging also plays an outstanding role in the management of paediatric oncology patients presenting with acute neurological symptoms related to disorders of blood cell counts, immunosuppression, neurotoxicity of therapy, or progression of the primary malignancy. a review of the main indications to perform a neuroimaging procedure in these children will be undertaken as well as a differential diagnosis based on representative cases selected from the daily routine in a paediatric tertiary hospital. practical algorithms with the preferential use of either ct or mri will be developed for each section. ct continues to be the first imaging modality in these patients in many centres, despite the accompanying radiation, because of the easy availability and no requirement of sedation. however, mr imaging is nowadays better for imaging these children owing to no radiation, the more completely provided information and the useful advanced techniques that can be used, such as diffusion imaging (di), spectroscopy, arterial spin labelling (asl), and susceptibility weighted imaging (swi). learning objectives: . to learn about the currently limited role of ct in the non-traumatic acute setting. . to become familiar with radioprotection strategies and protocols adapted to children. . to consolidate knowledge on the role of mri as the modality of choice for acute non-traumatic neurologically ill children, with an emphasis on newer techniques. . to become familiar with imaging findings and the main differential diagnosis of acute neurological conditions in children. b. imaging of acute chest pain and distress in children c.e. de lange; oslo/no (clange@ous-hf.no) acute chest pain in children is a common complaint in the emergency department, but patients rarely present with significant distress or lifethreatening symptoms requiring immediate care or resuscitation. the most frequently reported cause is benign musculoskeletal pain followed by respiratory and gastrointestinal causes, while cardiac causes are less frequent. a thorough clinical history and careful physical examination will determine, in most cases, the patients in need of further investigation to establish a diagnosis. in this regard, radiology plays an important role, especially in the emergency setting in patients with more serious associated symptoms like acute breathing difficulties, swallowing problems, fever or sepsis. when choosing the appropriate technique for investigation, the consequences of radiation exposure in children must be considered. plain radiography and fluoroscopy still remain the most important and frequently used tools to gain information on various acute chest/pulmonary problems. ultrasonography is the first choice for diagnosis/treatment of pleural effusions. multidetector computed tomography and magnetic resonance imaging are mainly used for investigating pulmonary/mediastinal masses and congenital abnormalities of the great vessels and the lungs. this lecture will discuss the choice of imaging technique and the urgency of radiological management depending on the symptoms and age of the patient. the imaging characteristics of the different causes of acute chest pain and/or distress in children will be reviewed, represented by the more common conditions involving the chest wall, respiratory tract, oesophagus and the heart, as well as less frequent causes such as tumours, manifestations of congenital malformations and nonaccidental trauma. outcome data from trauma series have demonstrated improved patient survival by the prompt diagnosis of a range of injuries and the recognition of life-threatening sequels, principally that of active arterial haemorrhage. widely used trauma scoring systems are applicable in the paediatric population, although the mechanisms and combination of injuries may differ from those encountered in adults. a ct diagnosis of active haemorrhage demands early operative or non-operative intervention with embolisation or use of alternative endovascular techniques including deployment of vascular stents. this principle is applicable to blunt or penetrating liver, splenic, renal or pelvic trauma. it is also applicable in some cases to severe mediastinal and limb vascular injury. important caveats are firstly the use of ct in an overall trauma service and avoidance of any inappropriate irradiation. secondly, in paediatric interventional practice, due regard needs to be given to the delivery of implantable devices that may have adverse sequels in later life. nevertheless, the demand for immediacy in intervention may outweigh any such consideration. this presentation will summarise the importance of imaging and intervention in both the early and delayed complications of trauma with an emphasis on abdominal and thoracic trauma. brachial plexus originates from the ventral branches of the cervical nerve roots from c to t . it is composed of roots, trunks, divisions, cords, and branches, topographically divided into supraclavicular (roots and trunks), retroclavicular (divisions), and infraclavicular sections (cords and branches). mri has the advantages of multiplanar imaging, high tissue contrast, and relative paucity of artefacts. the t -weighted sequences display topographic anatomy including muscles, blood vessels, and nerves stressed by tissue fat planes. the nerves appear as elongated fibres that are isointense to the scalene muscle, posteriorly and superiorly to the curvilinear flow void of the subclavian artery. the fat-suppressed pd and t -weighted and stir sequences detect signal alterations of the bp. general abnormal findings include: loss of fat planes around part or all bp components, nerve signal abnormalities with mild or marked hyperintensity on t -weighted images that can be associated with swelling, focal or extensive enhancement of the nerves after gad injection on t fat sat images. differences in signal intensity should be judged by visual inspection, which is susceptible to a subjective point of view; the comparison of both sides of bp has a relevant role in judging the presence of signal alterations, especially when they are smooth and/or focal. the pathology of brachial plexus can be divided into nontraumatic and traumatic brachial plexopathies. among nontraumatic brachial plexopathies, we find radiation fibrosis, metastatic breast cancer and primary or metastatic lung cancer. other pathologies are tumours and inflammatory disease or thoracic outlet disease. the peripheral nerves of the upper limb are affected by a number of entrapment and compression neuropathies. these syndromes involve the brachial plexus as well the musculocutaneous, axillary, subscapular, ulnar, radial and median nerves. clinical examination and electrophysiological studies are traditionally the mainstay of diagnostic workup. however, ultrasonography and magnetic resonance imaging (mri) may provide key information about the exact anatomic location of the lesion or may help to narrow the differential diagnosis. in certain patients with the diagnosis of a peripheral neuropathy, imaging using either ultrasonography or mri may help establish the cause of the condition and provide information crucial for conservative management or surgical planning. in addition, imaging is particularly valuable in complex cases with discrepant nerve function test results. a variety of peripheral neuropathies can be encountered in the lower limb. most are entrapment syndromes affecting many nerves, such as the sciatic, gluteal, femoral, lateral femoral cutaneous, obturator and pudendal around the hip, the peroneal and its branches and the saphenous at the knee, the superficial peroneal at the lateral leg, the tibial with its plantar and calcaneal branches at the ankle, the deep peroneal and the interdigital nerves in the foot. although clinical examination and nerve conduction studies are the mainstay of the diagnostic workup of peripheral neuropathies, ultrasound (us) and magnetic resonance (mr) imaging may provide key information about the exact anatomic location of a lesion and the nature of the constricting finding or may help narrow the differential diagnosis. in patients with peripheral neuropathies of the lower extremity, us and mr imaging may provide critical information for planning an adequate treatment strategy. although us and mr imaging have followed parallel paths for nerve imaging with little comparison of the two modalities, us seems to have some advantages over mr imaging, including higher spatial resolution, time effectiveness, the ability to explore long nerve segments in a single study and to examine tissues in both static and dynamic states. advances in diagnosis of acute stroke have been achieved by brain imaging. ct or mri distinguish ischaemia from haemorrhage and may identify acute clot (dense artery sign) or vessel wall pathology by "black blood mri". dwi provides evidence of early infarction. ct and mra enable identification of large vessel pathology as potential source of embolic or haemodynamic ischaemia. supplementary techniques are ct or mr perfusion. on mri perfusion, a mismatch between the area of restricted diffusion and perfusion (pwi) is a signature of the penumbra. the tissue of pwi restriction outside of the dwi abnormality represents "tissue at risk". similarly on ct perfusion, the region of reduced cerebral blood volume presumably represents irreversible infarction and the area of reduced cbf and extended mtt visible beyond the cerebral blood volume abnormality represents potentially reversible ischaemia. morphologic imaging and advanced standardised perfusion/penumbral stroke protocols aim at identifying lesions that are amenable to acute stroke therapy and exclude pathologies that mimic stroke but do not represent vascular disease. since iv tpa has become an approved therapy from , the number of acute stroke patients entitled for treatment has remained limited (~ ) despite extension of the time frame from to . h. dwi-flair mismatch and an individualised assessment of the ischaemic penumbra may serve in the identification of patients within the appropriate time frame. by selection of patients with advanced brain imaging and initiation of appropriate therapy, a further increase in good outcomes and reduction of the incidence of symptomatic haemorrhage may be achieved. in recent years, the role of imaging in the patient admitted with an acute neurologic deficit has changed significantly, due to the arrival of new treatments for acute stroke, aimed at re-establishing blood flow, reducing infarct size and protecting the brain at risk. the first goal is to differentiate haemorrhagic from ischaemic stroke and to rule out other stroke mimics. this can be achieved by performing nonenhanced ct scan of the brain. the next strategic imaging objectives include: demonstration of major blood vessel occlusion (e.g. by ct angiography) and identification of potentially salvageable brain tissue, the so-called "penumbra" (e.g. by ct perfusion). ct angiography is performed during rapid intravenous bolus injection of a high-concentration iodinated contrast agent. the fov should cover not only the intracranial arterial circulation, but also the arteries of the neck, to exclude dissection. ct perfusion is also achieved by bolus injection of contrast, followed by a series of fast images or volume acquisition. ct densities change over time and reflect the iodine concentration. perfusion images are processed to generate parametric maps, which reflect: regional cerebral blood flow (rcbf), blood volume (rcbv), mean transit time (mtt), and time to peak (ttp).the combination of ct angiography and ct perfusion provides a unique insight into the pathophysiology of the cerebral circulation. these techniques are now essential tools in the management of acute stroke and in selecting those patients who are potential candidates for advanced therapies such as thrombolysis or thrombectomy. learning objectives: . to become familiar with a comprehensive imaging protocol in patients with suspected stroke. . to understand the advantages and limitations of cta and perfusion in the initial work-up of stroke patients. . to recognise the different imaging patterns in stroke and their prognostic value. a- : b. stroke mimickers and pitfalls p. vilela; lisbon/pt (ferrovilela@sapo.pt) "stroke mimic" is classically used to describe nonvascular diseases that present with acute focal neurologic deficit corresponding to a consistent vascular distribution, which may resemble or may even be indistinguishable from stoke. it is estimated that up to % of stroke-like presentations are due to mimickers. there are several neurological and psychiatric disorders that can have such a clinical presentation, such as seizures and/or postictal status, the most frequent one. the other classical clinical stroke mimickers include: multiple sclerosis, migraine (specially hemiplegic migraine), metabolic disturbances (more commonly hypoglycaemia/hyperglycaemia), intracranial tumours or infections and the conversion reactions. brain imaging plays a key role in the diagnosis and management of acute stroke by differentiating ischaemic from haemorrhagic lesions, identifying the vessel occluded and estimating the viable brain tissue that is at risk for stroke. imaging is also important to exclude some of the aforementioned stroke mimics. the more comprehensive stroke imaging protocols, with ct perfusion/angiography and/or with mri, have reduced significantly the misdiagnosis of stroke. mri-dwi is the most sensitive and specific imaging modality for depicting ischaemic stroke. however, it is also essential to be aware that there are some pathological processes that can mimic stroke in imaging studies. these include some disorders that may present themselves with lesions associated with restricted diffusion, like transitory postictal brain abnormalities, migraine and encephalitis. the author reviews the most common clinical and imaging stroke mimics and highlights the importance of brain imaging in depicting these mimickers and avoiding the potential adverse effects of stroke therapy in these patients. learning objectives: . to learn the differential diagnosis of stroke. . to understand the role of ct and mri in the work-up of stroke and related disorders. . to recognise imaging patterns that may mimic stroke clinically and radiologically. imaging plays a central role in the management of stroke, which is an important health issue according to the number of patients involved and the severity of the disease. it helps to select patients who will benefit from a revascularisation therapy (iv thrombolysis or endovascular treatment) at the acute phase of stroke. if the value of perfusion/diffusion mri mismatch in the evaluation of penumbra is still a matter of debate, other imaging factors are associated with the clinical outcome including length of clot, flair positivity, and collateral circulation. if iv thrombolysis using rtpa is still the reference treatment, endovascular treatment is emerging since several years as a feasible and efficacious alternative singularly since the appearance of stent retrievers. recent neutral randomised trials comparing iv thrombolysis and endovascular treatment have shown the critical role of appropriate imaging in the selection of patients who can benefit from acute revascularisation as well as the importance of using the most efficacious endovascular devices. prevention of stroke is partially based on the treatment of cervical and intracranial stenosis. several randomised trials comparing carotid angioplasty and carotid endarterectomy were initially negative, but a recent meta-analysis showed that below the age of years both techniques were equivalent. the place and value of endovascular treatment of intracranial arterial stenosis is still controversial after the publication of the sammpris results. the management of thyroid nodules has been continuously evolving. i hope that during this session you will understand the important differences in the upto-date national and international guidelines on thyroid nodule management and understand the role of the radiologist in the multidisciplinary thyroid meetings. the features that are suggestive of benign, indeterminate and malignant nodules will be demonstrated. advances in both radiological and cytological techniques such as us elastography and braf mutation analysis that may help triage patients with thyroid nodules will be covered. the differing biopsy techniques (fine needle aspiration, non-aspiration (fna and fnnac) and trucut biopsy) and their roles will be highlighted. there will be time for an open question and answer discussion with all the speakers before the end of the session. the management of thyroid nodules and the guidelines of the various interested parties, surgeons, endocrinologists, nuclear medicine physicians, radiologists and ultrasound specialists, vary across the world, indeed between different countries within the eu, and are constantly evolving. i will discuss how the thyroid multidisciplinary meeting (mdt) works at our institution and the role of the radiologist in it. the current differing international guidelines on the management of thyroid nodules will be mentioned, highlighting their salient points and differences. the role of the differing imaging modalities used in the management of thyroid nodules will be discussed based on differing clinical scenarios. finally, the management of the common clinical scenario of the incidental thyroid nodule will also be discussed. learning objectives: . to understand the role of the radiologist as part of a multidisciplinary team. . to learn about current guidelines on the management of the thyroid nodule. . to understand the role of various imaging modalities in the patient with thyroid nodule. thyroid disease is ubiquitous and usually benign; therefore accurate imaging of the thyroid is a major challenge for radiology. significant advances in conventional ultrasonography allow superb visualisation and anatomic characterisation of thyroid pathology and the impact of broader bandwidths, higher frequencies, spatial compound imaging and novel signal processing techniques will be discussed in this session. the key b mode features of thyroid nodules are discussed with reference to relevant clinicoradiological guidelines, specific pitfalls and limitations. a significant number of nodules remain indeterminate after conventional assessment, and imaging advances that enable more accurate, non-invasive thyroid characterisation are highly attractive, potentially reducing unnecessary biopsy and surgery for a large number of patients. ultrasound elastography offers a potentially useful adjunct to conventional sonography in thyroid disease with a growing evidence base indicating that thyroid malignancies are typically stiffer than benign lesions and that a 'soft' cutoff value with a high negative predictive value is feasible. however, elastography studies have included a variety of different techniques, patient/nodule selection and assessment methods; therefore the current role of elastography will be discussed. fine needle biopsy of the thyroid nodule f. campoy-balbontín, m.c. jurado-gomez; sevilla/es (fcampoy @gmail.com) thyroid nodules are very common. the clinical importance of thyroid nodules rests with the need to exclude thyroid cancer, which occurs in - %. ultrasound (us) has been widely attempted to differentiate benign from malignant nodules and to guide fine needle biopsy (fnb). however, considerable overlap between benign and malignant characteristics has been found. nowadays, there are a number of different guidelines for the management of thyroid nodules. at our hospital, we have adopted the guideline proposed by the society of radiologist in ultrasound ( ). the different elements necessary to perform the fnb procedure are demonstrated; the transducer probe, the needles, the slides, the syringe. the way of managing these elements is shown by figures and videos, with different skills and performances to optimise the procedure and get the best sample. uscontrol fnb can be in parallel or perpendicular, both methods are demonstrated. thyroid fnb is usually non-aspirated, but sometimes it has to be undertaken with aspiration; when and how to change from one method to another is explained. with good technique, the insufficient cytological rate can be significantly reduced, with which the need for core biopsy diminishes. although the complication rate for core biopsy is low, fnb is safer and if performed correctly has a high diagnostic yield. core biopsy should be limited to those nodules with a second insufficient fnb cytological result, with us findings suspicious for malignancy. multimodality breast imaging is emerging as radiologists have access to new technologies coinciding with the refinement of operative techniques. one of the challenges for radiologist remains the correct preoperative staging, especially the metastatic involvement of axillary lymph nodes. multimodality imaging has new accurate solutions. the imaging follow-up of patients with sophisticated oncoplastic techniques is another challenge where multimodality breast imaging is the solution. coincident with this increasing contribution to the diagnostic process is the emerging role of the breast imager as a therapeutic clinician, making use of innovative image-guided procedures. in this integrated rc, experts will present a clinical update on these newer techniques and there will be an opportunity to discuss how the evolution of such techniques is changing the nature of the modern multidisciplinary team meeting. a. conventional, functional and interventional lymph node assessment r.m. pijnappel; utrecht/nl (r.m.pijnappel@umcutrecht.nl) metastatic involvement of axillary lymph nodes has been known to be one of the most important prognostic factors for women with breast cancer. the traditional approach to staging the axilla is either lymph node sampling procedures or sentinel node biopsy. the main diagnostic features of potentially malignant involvement of the axilla are abnormal lymph node morphology (shape and cortical thickness). there has been a concerted effort in recent years to find imaging techniques that might facilitate accurate axillary staging. so far, no imaging technique alone (ct, mri, scintigraphy and ultrasound) has been capable of differentiating between positive and negative lymph nodes of the axilla in breast cancer. recent techniques like gadofosveset-enhanced magnetic resonance imaging and high spatial resolution -t mri imaging appear promising; however ultrasound-guided sampling of suspicious lymph nodes by core or fna remains the standard of care in assessing the axilla prior to treatment planning. ultrasound-guided axillary lymph node sampling is an effective method of assessing the axilla. a recent meta-analysis shows that ultrasound can be expected to achieve sensitivity approaching % for detecting abnormal nodes in those that eventually prove to be nodal metastatic disease. the false-negative rate where biopsy is performed is around %. it is impossible to detect the sentinel node by ultrasound alone. therefore a new promising technique was introduced, injecting an ultrasound-contrast agent around the nipple. using this method, it is possible to detect the sentinel node in % and therefore lower the false-negative rate of us-guided biopsy with another %. oncoplastic breast procedures were introduced to fill the gap between comprehensive oncologic surgical treatment of breast cancer and the achievement of cosmetic results that fulfill patient expectations in both, body imaging and psychological well-being. due to its complexity and relevance for the patient, the decision of performing these procedures must be made as part of the multidisciplinary approach of breast cancer treatment. as a consequence, the role of breast radiologists has expanded beyond the anatomic region of the breast and the usual imaging techniques. basic knowledge of the different oncoplastic techniques is mandatory to understand the spectrum of findings from a multi-modality approach. implants and/or autologous reconstruction techniques (pedicle, free or perforator flaps, as well as lipofilling techniques) are widely applied. the role of the radiologist in the multidisciplinary team is twofold: assessment during the planning stage, and imaging evaluation at follow-up. the assessment during the planning stage includes the determination of the local extent of the disease that makes the choice of the appropriate surgical technique possible, and the imaging study of the donor site in those cases where autologous reconstruction is elected. imaging evaluation at follow-up comprises the recognition of changes and potential pitfalls after reconstruction, the identification of short/mid/long-term reconstruction complications, and the detection of recurrent/second carcinomas. controversial aspects will be reviewed, such as the probability of recurrence after oncoplastic surgery, the need to establish multimodality follow-up protocols and the interrelations between the autologous tissues and the mastectomy bed or remaining breast. the use of image guidance for surgical planning has only recently been accepted. the use of intraoperative ultrasound in women undergoing lumpectomy is associated with a higher rate of negative margins. the combination of image guidance and radiological intervention techniques also harbours the possibility of tumour treatment without surgery. most techniques make use of the placement of needles within the tumour. treatment is performed by heating, freezing or extracting the tumour tissue. currently, radiofrequency ablation under ultrasound guidance is being furthest evaluated. however, in rfa even after optical complete tumour ablation, residual vital tumour tissue is still found in many cases. this is mainly dependent on tumour size with poorer results ( - % success rates) in tumour over cm, and better results (~ % success rate) in smaller tumours. adequate tumour size estimation is thus of vital importance. currently, since remaining vital cells cannot be excluded, the technique is limited to use in patients who cannot undergo surgery due to poor health state. larger series in which rfa is combined with radiotherapy have not been reported. it is therefore not clear whether it is important to actually excise residual vital tumour. similar restrictions currently hold for cryotherapy, laser ablation and hifu. large vacuum-assisted needles and the breast lesion excision system can be used to remove the tumour through a minimal incision in the skin. however, margin evaluation is difficult. nevertheless, these techniques are already optional for the removal of high-risk lesions and can potentially be combined with (focused) radiotherapy. the müllerian ducts are paired embryologic structures that undergo fusion and resorption between the th - th weeks of gestation to give rise to the uterus, fallopian tubes, cervix, and upper two-thirds of the vagina. non-development, defective vertical or lateral fusion, or resorption failure of the müllerian ducts can result in formation of müllerian duct anomalies (mdas), whose reported prevalence ranges between . % and %- %. the classification revised by the american society for reproductive medicine in is the most widely accepted. obstructive mda can occur in infants as palpable pelvic mass, at menarche with cyclic pelvic pain, or pelvic mass, and primary amenorrhea. complications include hematocolpos, hematometra, hematosalpinx, endometriosis, pelvic adhesions, and obstruction of urinary tract. mdas are commonly associated with renal and other anomalies; thus, identification of both kidneys is important. accurate diagnosis of an mda is essential because of the high associated risk of infertility, endometriosis and miscarriage, and since the management approach varies depending on the type of malformation. hysterosalpingography (hsg) is routinely used in evaluation of infertility. because a key component of mdas characterisation is the external uterine fundal contour, hsg is limited for this purpose. ultrasonography (us) is initially performed to evaluate uterine anatomy and to assess for associated renal abnormalities. mri is reserved for complex or indeterminate cases to characterise the uterine anomaly and evaluate the possibility of a vaginal abnormality, or after a seemingly normal us when clinical opinion ensures its use, or to make decisions on management strategies and preoperative planning. this refresher course will focus on the radiologist's approach to using pet-ct within the context of the gynaecologic oncology multidisciplinary meeting. the current indications for using fdg-pet/ct will be discussed for cervix, endometrial and ovarian cancer as well as the potential use of fdg-pet/ct in rare gynaecologic cancers. the value of adding fdg-pet/ct to the patient management pathway will be presented alongside the potential pitfalls in interpretation as well as controversies. future developments, including potential new tracers, will be mentioned. cystic lesions account for the vast majority of ovarian masses and are detected incidentally in - %. their spectrum ranges from non-neoplastic cysts to benign tumours, e.g. cystadenomas and dermoids to the rare ovarian cancer. diagnostic criteria in us, ct and mri are based on the clinical background (age, menopausal status, medical history, and tumour markers) and the morphology of the ovarian mass. typical of a benign cystic lesion is its pure cystic structure or thin septations, fatty or haemorrhagic contents, and mild enhancement of solid components in a complex solid and cystic ovarian mass. thus, patients can be categorised into different risk groups. sonography has been established as the first-line imaging modality to assess the ovaries. recently, management guidelines have been adopted for cystic ovarian lesions. in these guidelines, also cystic adnexal incidentalomas are included. mri is most useful as a complementary technique in sonographically indeterminate masses. an algorithmic approach will render a specific diagnosis in the vast majority of cases. in complex cystic lesions, integration of dwi and dynamic contrast-mri allows differentiation of rare benign complex tumours, e.g. cystadenofibroma from ovarian cancer. pitfalls of cystic ovarian tumours include cystic fibroids, peritoneal cysts, and extraperitoneal cystic tumours. compared to us, both ct and mri are superior in assessing large cystic pelvic masses. criteria to differentiate between intra-and extraperioneal origin include displacement patterns and identification of the vascular pedicle. careful analysis of imaging and clinical findings usually allows differentiation of inflammatory tumours from ovarian cancer. learning objectives: . to learn about specific imaging algorithms of ovarian cystic tumours. . to understand certain imaging features that can differentiate ovarian from non-ovarian cystic tumours in the pelvis. . to become familiar with the pitfalls in imaging of ovarian cystic tumours and the lessons to be learned from them. the diagnostic methods available in patients with conductive hearing loss are: personal and family history, otoscopy, audiology, imaging, surgical inspection and counseling. the most important reasons for imaging are: atypical history, suspicion of congenital hearing loss, suspected otoscopic image, asymmetric bone conduction thresholds, profound mixed hearing loss and suspected tympanometry. ct can be used to detect otosclerosis, tympanosclerosis, posttraumatic ossicular lesions, superior semicircular canal dehiscence and cholesteatoma. mri can be used to exclude schwannoma, congenital cholesteatoma and residual and recurrent cholesteatoma after prior surgery. this lecture will illustrate the value of ct and mri in the evaluation of cholesteatoma patients as well as prior to first stage surgery and prior to second look surgery. different subtypes of surgery will be highlighted and illustrated. the growing importance of mr imaging and diffusion weighted (dw) imaging in particular will be illustrated in the evaluation of cholesteatoma patients. the role of echo-planar and non-echo-planar dw imaging will be discussed as well as the eventual use of delayed gadolinium-enhanced t weighted imaging. mr is the modality of choice in patients with congenital or acquired sensorineural hearing loss (snhl) and the complete auditory pathway from the cochlea to the auditory cortex must be studied in these patients. the labyrinth, internal auditory canal (iac) and cerebellopontine angle (cpa) are best studied using submillimetric heavily t w images, showing the intralabyrinthine fluid and nerves. but submillimetric gd-enhanced t w images or d-flair images remain more sensitive. the auditory pathway in the brainstem is best studied with a multi-echo sequence (m-ffe/medic/merge) or t w tse sequence and the auditory cortex is best studied on t w tse images. the clinical information provided by the otologist will direct the radiologist to a certain part of the auditory pathway or will let him use certain imaging techniques. for instance, congenital malformations in the labyrinth and iac must be excluded in case of congenital snhl, and in case of mixed hearing one will start with ct to exclude otosclerosis. the most frequent pathology in the labyrinth (labyrinthitis, intralabyrinthine schwannoma, congenital malformation, post-traumatic lesions), the iac and cpa (cochleovestibular schwannomas, meningiomas, epidermoid tumours and facial nerve neuritis) and central auditory pathway (ms, infarction, tumor and trauma) will be illustrated using clinical examples and the important "otologist-radiologist" team work will be stressed. for tumours that nearly always arise from the investing mucosa, like in the larynx and hypopharynx, it is endoscopy that directly maps the superficial extent and grades the functional impairment (arytenoid and vocal cord movement, pyriform sinus distensibility). in addition to white light, the recent use of narrow band imaging endoscopic technology, enhancing fine capillaries in the mucosal surface, enables the detection of neoangiogenesis caused by early squamous cell carcinoma. conversely, submucosal spread is the domain of cross-sectional imaging techniques. therefore, it is the integration of the two classes of data, along with the information about patient conditions, which permits the choice of the most proper treatment strategy. in early and advanced glottic cancer, treatment planning is modulated by information about deep neoplastic invasion into the paraglottic space, the cartilage or the suprasubglottis subsites. this information can be derived by discriminating the tumour tissue from intra-laryngeal muscles, fat spaces and the cortical rim of ossified cartilages. though mr shows a greater contrast resolution than ct, it is hampered by a significantly longer acquisition time. this means more artefacts caused by breathing and movement. nevertheless, mr has the potential to unravel the signals from submucosal tissues, separating tumour from oedema, muscles, fat and ossified cartilages. advanced laryngeal and hypopharyngeal cancers need to preliminary assess local (nodes) and distant (metastases) spread. ct-pet is the more accurate tool. the multidisciplinary approach to the processing of the whole frame of data is essential to offer the patient the best care. traditionally, x-ray ct scanners were equipped with a single or a few detector rows only, image reconstruction was assuming parallel slices, which was a good approximation for these fan-beam systems. about a decade ago the number of slices increased to or more and the parallel slice approximation was no longer valid. the cone-beam nature of these multi-slice ct (msct) scanners had to be taken into account by specific cone-beam reconstruction algorithms. while these msct systems -today they are simultaneously acquiring up to slices -are cone-beam ct systems in a general sense, the notion of cone-beam ct (cbct) is also associated with a specific kind of conebeam ct, namely those equipped with flat detectors, i.e. the non-diagnostic or non-clinical ct systems. msct, in contrast, is a cone-beam ct system for diagnostic use, i.e. a clinical ct system. this distinction between msct and cbct is useful, as there are decisive differences in application, performance, and in image reconstruction. these differences will be reviewed in the presentation, with a particular focus on why image quality in msct is so much better than in cbct. cone-beam (cb) imaging with flat detectors is performed on a variety of different imaging systems. the most important area is cb imaging using interventional c-arm systems. the minimally invasive procedures carried out on these systems benefit from the availability of high-resolution d images for intervention planning, guidance and outcome control. interventional cb imaging was first applied in neuroradiology using rotational angiography acquisitions. today, it is used for a variety of procedures in interventional radiology, cardiology and oncology based on angiographic and soft tissue protocols. one of the inherent advantages of this approach is the direct registration of the volume images into the interventional procedure. other application areas of cb imaging using different scanning systems are onboard imaging in radiation therapy or the integration of flat detector tomography and spect. this part of the course teaches the generation of d volume information from flat detector systems and its utilisation in different clinical applications. image acquisition protocols, scan modes, and system design parameters are explained. application-specific calibration and processing steps are introduced to show how cb imaging is tailored for specific clinical applications. examples are presented for angiographic and soft tissue volumetric imaging. medical applications utilising the tomographic images for diagnosis, intervention planning, guidance, or outcome control are discussed. in the last decade, three-dimensional dentomaxillofacial imaging through conebeam ct (cbct) technology has become widely available. dental cbct (digital volumetric tomography; dvt) equipment is compact and often affordable to dentists. the driver for dental cbct was implant dentistry, but its use has spread to other areas, including paediatric applications. radiation doses are variable. some systems offer a fixed, large, field of view and others fixed exposure factors, obstructing attempts at optimisation. typically, doses are at least an order of magnitude greater than for "conventional" imaging. image quality is also variable, which means that certain equipment may be unsuitable for some clinical applications. there is often scope for lower exposure factors to be used than those recommended by manufacturers. a key aspect of using cbct relates to justification. imaging in three dimensions may be perceived by dentists as inevitably superior, a view which is often implicitly encouraged by those selling equipment. the research on diagnostic efficacy is, however, limited. in , the european commission published "radiation protection : evidence-based guidelines on cone beam ct for dental and maxillofacial radiology". this provides a comprehensive set of recommendations, including referral criteria, a quality assurance programme and optimisation strategies. priorities for future research relate to exposures to the necessary image quality requirements for specific clinical applications, along with research on the impact on clinical outcomes of cbct-based treatments. it is clear that much work remains before the place of cbct in dentistry can be established. the role of nonvascular interventional radiology of thorax for both diagnosis and treatment has significantly increased in the last years. although these procedures are technically easy with highly successful results, thoracic nonvascular interventional procedures are not unfortunately routinely performed in all of the interventional units. the most commonly performed procedures generally under ct guidance are transthoracic biopsies of the lung, mediastinal lesions and pleura. the others include percutaneous drainage of thoracic collections such as pleural effusions, empyema and mediastinal collections. image-guided tumour ablation performed by either radiofrequency ablation (rfa) or microwave (mw) as a nonvascular thoracic intervention has become a serious treatment alternative in patients with lung tumours (either non-small cell lung cancer or lung metastasis) since the year . interventional radiologists are expected to understand the clinical indications of these procedures at the first step. in addition, the techniques of the procedures and possible risk of complications in their management methods should be learnt by interventional radiologists as well. this refresher course will help us to discuss the main indications, techniques and complication management of these procedures. a. lung biopsy t. sabharwal; london/uk (tarun. sabharwal@gstt.sthames.nhs.uk) lung biopsy is most commonly now performed under ct guidance. it has a high technical success rate. common complications include pneumothorax, bleeding, sepsis, pain and failure. air embolism is a rare complication. pet scan is a useful tool to guide for appropriate target. ultrasound is useful for biopsying pleural-based lesions. drainage of thoracic fluid collections is a relative emergency. we will review the different thoracic fluid collections that can be drained by radiologists: pleural (parapneumonic, empyema, and malignant effusions), mediastinal, chest wall, pulmonary (lung abscesses and infected tumours) and pericardial effusions. we will answer the following questions about the main thoracic fluid collections: how to diagnose, when and how to drain, which guidance (ultrasound or ct-fluoroscopy), and procedures to perform before and after the drainage. the choice of imaging to detect thoracic fluid collections and guide drainage depends on the availability of the modalities, the imaging characteristics, location of the collection, and the comfort and expertise of the operator with the specific modalities. we will also review the main complications of percutaneous drainage of thoracic fluid collections (pneumothorax, pain, drain dislodgement and drain blockage) and compare these complications with those related to large-bore chest catheters inserted by thoracic surgeons. the incidence of lung cancers continues to increase and primary lung cancer remains the leading cause of cancer-related deaths in both women and men. therapy of lung tumours includes resection, radiation therapy, chemotherapy, thermal ablation or a combination of these treatment modalities. image-guided percutaneous thermal ablation therapies are minimally invasive techniques established in the local treatment of hepatic, renal, or osseous tumours. among these techniques, radiofrequency ablation (rfa) has now attained consideration for therapy of small pulmonal tumours. other ablation techniques that have been used for treatment of lung tumours include cryoablation, laser (litt) and more recently microwave ablation (mwa) and irreversible electroporation (ire). ablative techniques may produce a complete and irreversible tumour tissue destruction through application of either hot or cold thermal energy, or through permeabilisation of the cell membrane while using ire. ct is currently the imaging modality that is most widely used for planning, monitoring the ablation's course and for assessment of treatment response. only patients with stage i and stage ii lung cancer and those with small metastases are potential candidates for ablation. the medical history and physical examination of the patient as well as recent imaging with ct should be evaluated to determine the indication for thermal ablation. percutaneous lung rfa is considered as a safe procedure with an overall morbidity rate from . % to . % and mortality rate from % to . %; it requires less costs and offers faster recovery, and reduced morbidity and mortality. sorafenib, a tyrosine kinase inhibitor, has shown clinical efficacy in patients with hepatocellular carcinoma (hcc) and is the standard of care for patients with advanced-stage hcc. nowadays, many targeted therapies are evaluated in hcc either as sole treatment or in combination with other treatments such as tumour ablation, chemo-embolisation, and surgical resection. therefore, there is a need to assess the efficacy of targeted therapy in hcc. recist is the reference method to evaluate treatment efficacy in solid tumours, but does not seem appropriate in evaluating targeted therapy as objective responses are seen in very few cases in patients treated with sorafenib or sunitinib. new criteria have been proposed to evaluate the treatment efficacy of nonsurgical treatments in patients with hcc. the most common ones are the choi criteria, the easl criteria, and the modified recist criteria. all these criteria mainly focus on internal tumour changes such as appearance of necrosis or disappearance ot tumour hypervascularity. many examples will be shown during the lecture. another approach is based on functional imaging and especially perfusion-related imaging. contrast-enhanced ultrasound, ct perfusion and dynamic contrast-enhanced mr imaging have the capability to assess perfusion changes in patients under treatment. the advantages and disadvantages of these modalities will be discussed. lastly, other functional tools that are not routinely used will be presented. the evaluation of treatment efficacy is a key issue with prognostic and patient survival implications. it is crucial to have objective and reproducible criteria for specific groups of patients. the goal of ablative therapies of hcc is to induce tumoural tissue destruction. complete response (cr) after initial chemical and thermal percutaneous ablation, defined as the absence of contrast enhancement of the treated tumour at ceus, dynamic ct or dynamic mr, has been reported to correlate to long-term survival. nevertheless, the clinical effectiveness of imaging techniques to assess initial treatment success differs according to tumour size. the success rate of rf has been demonstrated to be superior to pei in hccs > cm, and depends on the ability to ablate all viable tumour tissue including an adequate tumour-free margin all around the lesion of . to cm. thus, the effectiveness of rf directly depends on the tumour location and size. rf is considered an effective treatment in lesions cm and its effectiveness is progressively reduced along with tumour size and it is not effective in lesions > cm. ceus beyond month may confirm or detect residual tumour, deserving a final ablation procedure. ct and mr are more effective in the follow-up to confirm cr and detect local recurrence, or additional hcc lesions in the liver parenchyma. the presence of transient hyperaemic inflammatory changes in the periphery of the treated area is a common finding that should be considered to avoid overestimation of the recurrence rate. one of the major characteristics of medical imaging in the twenty-first century is the dramatic influx of novel technology. this impact of new technology and techniques is experienced in all imaging modalities. the continuous development and implementation of highly sophisticated medical products and devices is key to the evolution of medical imaging leading to the improvement of patient care in terms of quality and positive outcomes. this session will analyse state-of-the-art displays (monitors, smartphones, tablets) and their applications in modern imaging. the session will provide a brief overview of displays in the current imaging chain. it will then evaluate the various specifications and parameters associated with displays. the session will evaluate current qa standards of modern displays as well as the challenges surrounding such novel technologies. image quality can be described in terms of resolution and contrast. an image displayed on a monitor consists of differences in brightness. the display monitor is at the end of the image chain in radiology, which starts at the x-ray tube. there are several factors in this chain that have influence on the contrast displayed on the monitor. diagnostic image quality can be described as observed contrast, which is not only dependent on the displayed image, but is also influenced by ambient conditions. for the primary class displays, typical characteristics of display monitors; such as resolution, homogeneity, luminance, contrast, monochrome or color, must be appropriate for the radiologist to execute a reliable diagnosis. calibration of display monitors is also important. which parameters are significant and what is the effect of variation of these parameters. there are standards like dicom gsdf and technical standard from acr-aapm-siim. both within and outside the clinical environment, the use of smartphones and tablet computers is increasing. reported applications range from teaching and education to navigation in surgical procedures. interest in the use of smartphones and tablets for viewing radiological images has been growing. however, these portable displays can differ significantly from conventional radiological displays. in order to determine where and how smartphones and tablets can be used appropriately, it is important to understand these displays and the challenges associated with them. this presentation will outline the current and potential uses of smartphones and tablet computers in radiology and compare their performance with other radiological displays and standards. the possible limitations/benefits of smartphones and tablets as display devices for radiological images will be discussed. recent research in the field will be reviewed, with emphasis on studies of diagnostic efficacy. in modern clinical environment, diagnostic monitors have replaced glowboxes and films and have become an important part of the imaging chain. the required specifications and the quality of monitors largely depend on their purpose, beginning from quite simple monitors used for the preview of radiographic images placed on modality or in control room to monitors used by high-end diagnostic workstations. decision about the kind of monitor for certain workplace is also very important from the economic standpoint as there are large differences in their prices. besides monitor specifications, also ambient conditions and ergonometry in the reading rooms are also very important issues which need to be considered to ensure optimal environment for clinical image reading. some of good and bad examples will be given in the presentation. as with other radiological equipment, also diagnostic monitors need to be controlled to ensure their optimum performance. quality control begins with monitor acceptance testing and setting up measurement parameters which are to be followed. practically all monitor vendors include some qc software within their workstations which can, together with some independent measurements, form an efficient quality control programme. in the last steps, how to deliver all information regarding image quality and possible presets on the diagnostic monitor and how not to confuse the user are important. sitting an examination is always a source of anxiety. the edir exam is no exception. this workshop is dedicated to those who want to take the edir exam, or become examiners, and would like to learn more about its format and discover some tips and tricks. the edir organisers and examiners have decided to build an interactive, friendly and enjoyable session in order to teach the essentials. some will play the role of the examiners, some the role of the candidate. in order to simulate the candidate's experience, this session will have no safety net. some senior members will expose themselves to real difficult questions in general radiology. fortunately, we hope that the attendees will be able to help the "candidates" to succeed. hopefully, you will enjoy this session and get some very useful information for your edir exam. abdominal malignancies are an important health problem in mexico. like in many other middle-income countries, there has been an epidemiological transition with an overall increase in oncologic diseases. colorectal, gastric and liver carcinomas are some of the frequent abdominal malignancies. colorectal cancer is a disease that is curable if detected early and even preventable if precursor polypoid lesions are removed. imaging plays a critical role in staging at diagnosis. additionally, virtual colonoscopy is an accepted modality in cancer screening. high-resolution magnetic resonance imaging (mri) has become a pivotal modality in the pretreatment assessment of rectal carcinoma. computed tomography (ct) is an excellent modality in the preoperative staging of gastric cancer and follow-up after treatment. positron emission tomography (pet) combined with ct (pet/ct) is particularly helpful for gastric and colonic carcinoma staging. hepatocellular carcinoma (hcc) is also common. cirrhosis related to alcohol and hepatitis c infection are by far the commonest aetiologies. ultrasound (us) in conjunction with alpha-protein is the most widely used modality for screening purposes. ct and mr are confirmatory modalities. the advent of hepatospecific gadolinium contrast agents has been very helpful in the characterisation of focal lesions in cirrhotic patients. the use of multidetector ct (mdct) in pancreatic carcinoma for the detection of vascular and adjacent organ invasion is crucial in treatment planning. over the last few decades, there has been an important improvement in imaging techniques resulting in better quality images. imaging is critical not only for the diagnosis, but also for treatment planning and follow-up. learning objectives: . to learn about the imaging of common oncologic gastrointestinal diseases. . to understand the importance of imaging in liver and colon cancer. . to learn how imaging supports oncologic institutions. activity, ultrasound is readily available and gives detailed local information but is limited by the restricted field of view, communication of results to clinicians and comparison of examinations on time. computed tomography (ct) is fast, readily available and gives a detailed, reproducible overview, but radiation exposure and contrast resolution are limitations. magnetic resonance imaging combines a good, reproducible overview with high contrast resolution, dynamic information and no radiation exposure and is therefore preferable in many situations, but the longer examination times than for ct, availability and costs are limiting factors. for the diagnosis of stenoses, fistulas and abscesses, either technique can be used, although the unrestricted view of ct and mri favour these techniques in many situations. in acute situations, us and ct are more accessible than mri. in that setting us can be considered, but when the examination is inconclusive or the patient has clear inflammatory signs, ct is preferable. endoscopy is currently considered the reference standard for the evaluation of colonic pathologies, including colitis. it allows direct visualisation of the mucosa and in obtaining tissue sample. however, it has major limitations including the invasiveness of the technique, incomplete endoscopy and risk of perforation. endoscopy cannot, therefore, help to estimate the depth of involvement of transmural inflammation and extraluminal complications. by contrast, an evolving role for cross-sectional imaging in the evaluation of patients with colitis has been increasingly recognised, especially in the setting of crohn's disease (cd), since cross-sectional imaging has been demonstrated to have a high diagnostic accuracy not only for assessing the presence and extension of luminal disease, but also for evaluating cd-related acute or chronic complications. establishing the ultimate cause of colitis may sometimes be challenging and histology cannot be conclusive. however, cross-sectional imaging may provide additional information that is useful in the workup of colitis. the use of cross-sectional imaging has been increasing in the evaluation of inflammatory bowel disease. following their presentations on imaging protocols and features of small bowel disease and colitis, each speaker will present a case to illustrate the learning points covered in their lectures. the session will be moderated by the chairman and the audience will have full opportunity to ask questions of all the panel. multimodality imaging is essential in a wide variety of oncology situations. anatomic imaging, whether using us, ct or mri, is mandatory for tumour localisation. moreover, the evaluation of treatment response mostly relies on size assessment, whether uni-or bi-dimensional. with the advent of new targeted bio-therapies, functional imaging has progressively been integrated in the imaging strategies whether for better tumour characterisation or for optimal evaluation of treatment response. hence, assessment of molecular targets by pet is supplemented by the recent developments of diffusion mri, reflecting tissue architecture and cellularity, tissue perfusion, reflecting angiogenesis, and magnetisation properties of tissues. hcc is a primary liver tumour where the use of multimodal anatomic, functional and metabolic imaging appears of particular interest. according to easl and aasld recommendations, noninvasive diagnosis of hcc can be performed using dynamic contrast enhanced cross sectional imaging. liver mri provides optimal sensitivity and specificity for the detection of as small as cm large hcc. recent reports have suggested that the prognosis of hcc lesions could be anticipated based on the combined analysis of metabolic f -fdg pet ct, as well as with diffusion weighted imaging. the objective of this lecture will be to highlight the potential of combining both metabolic and multimodal cross sectional imaging in order to improve the management of patients with hcc. the (extra)ordinary night shift at the er k. petrovic; novi sad/rs (smakap@sbb.rs) besides the accuracy, working at the er requires maximal shortening of the diagnostics time, which has a significant influence on the patient's outcome. thus, it is the radiologist's duty to be familiar with the pathophysiological mechanism of the disorders, specific protocols, possibilities and limits of certain examinations, and also to be aware of all possible pearls and pitfalls, to make the correct diagnosis. pathology encountered at the er has an extremely wide range and requires expertise in all imaging modalities from head to toe. the more the radiologist is aware of all possible situations, the more is the diagnosis accurate, and information given to the clinician is more valuable. the aim of this presentation is to point out the possibility of encountering different pathological conditions in different body regions. moreover, it is important to be reminded that in radiology there are different modalities, each of which has its own indications and limits. this lecture reviews the selected series of cases from the er department of clinical center of vojvodina, pointing out the most common possible differential diagnosis and possible diagnostic mistakes, which often occur due to the lack of experience. learning objectives: . to point out the in-depth knowledge and experience in all imaging modalities and all body regions required for a radiologist to perform a routine work at the er. . to present a series of cases that are most and less often encountered at the er, discuss the possible differential diagnosis and mistakes which could potentially be made. diagnosis and management of pancreatic cystic lesions r.j. méndez; madrid/ es (ramiro.mendez@salud.madrid.org) pancreatic cystic lesions are frequently diagnosed in patients with clinically suspected pancreatic disease, but they are also increasingly detected on imaging studies performed due to other abdominal processes. the prevalence of pancreatic cysts increases with patient's age. most symptomatic cysts will be surgically resected, whereas asymptomatic lesions should be managed depending on the likelihood of causing harm to the patient. cystic lesions can be inflammatory or neoplastic. the incidence of pseudocysts is low without a history of pancreatitis. some pancreatic cystic neoplasms are malignant, but even benign lesions can become symptomatic depending on its size and location in the pancreas. the radiologist detects most of the pancreatic cystic lesions and should also play an important role in the clinical decision-making process. cyst size, morphology, calcifications, contrast enhancement, location in the pancreas, relation with pancreatic ducts, patient's age and gender are important data to classify the lesion. a specific radiological diagnosis is not possible in every patient, but in many cases this information is enough to decide on a conservative approach with imaging follow-up. if a more aggressive lesion is a concern, then endoscopic ultrasound and fluid analysis can help to better characterise some pancreatic cystic lesions. cyst puncture is only recommended if fluid analysis results will modify the patient's management. patient's age, status and preferences should also be taken into account. when follow-up is recommended, the preferred imaging technique and interval should be indicated. the chair will set the scene for the need for effective radiation protection in clinical radiology and highlight the role of clinical audit as a tool in achieving this. clinical audit is defined as a professionally led method of improving patient care through the systematic examination of systems, processes and outcomes against chosen and agreed standards. it can readily be carried out by individuals, groups or whole departments. if properly conducted, clinical audit can be an effective way of examining what we do to provide reassurance on performance, and to improve patient care. the clinical radiology audit committee (crac) of the royal college of radiologists (rcr) co-ordinates national radiology audit activity. it promotes and facilitates audit through nominated audit leads in each hospital, who act as a link between the rcr and their department. at least one national audit is carried out annually, with data collected via electronic submission and the anonymised results presented at an annual audit forum. individual departmental results are analysed using statistical process control (spc) methodology. this enables identification of departments underperforming against the national mean, and recommending corrective action, by redesigning the process being audited, or by identifying and eliminating specific root causes locally. the committee has developed a web-based tool for facilitating local audit, "auditlive", a fully searchable collection of templates which can be downloaded and adapted. fellows are also able to submit their own templates for publication hence sharing best practice. trainees are encouraged to participate in audit through audit poster competitions at national radiology scientific meetings. our experience leads us to believe that audit succeeds when relevant, locally owned and properly structured, and multi-professional, and the rcr model encourages this. models of external audit in the netherlands s. geers-van gemeren; utrecht/nl in the netherlands clinical audit is legally obligatory for healthcare professions in order to be able to practise since . clinical audit is a tool designed to improve the quality of patient care, experience and outcome through formal review of systems, pathways and outcome of care against defined standards, and the implementation of change based on the results. the quality of the provision of care by professionals is assessed by peers. in the fields of radiology, nuclear medicine and radiotherapy, different models of clinical audit are used. for radiotherapy a multidisciplinary audit has been used since . for nuclear medicine a multidisciplinary audit has been implemented since . for radiology the clinical audit for radiologists and for radiographers are separate. implementation of the multidisciplinary audits needs requirements and adjustments of the audit system. this process is complex and needs approval of the members of all involved societies. to support the clinical audit a web based tool adas (general digital audit system) is used. the development of professional standards is a prerequisite to start clinical audit. the use of adas in multidisciplinary audits is a requirement to be able to audit different professions and focus on the content and the quality of their contribution to patient care. clinical audit is a good tool to improve the quality of patient care. important are the professional standards, the culture of learning and willing to improve by the professionals. "every defect is a treasure." learning objectives: . to learn about the different dutch models of external clinical audit. . to comprehend the importance of professional standards, the culture of learning and willingness to improve. . to become familiar with the use of a digital audit system to support multidisciplinary clinical audit. external regulatory audit in finland r. seuri; helsinki/fi (raija.seuri@hus.fi) directive / /euratom states that the clinical audit of medical imaging should be carried out in accordance with national procedures. the finnish solution has been regulatory external audits every five years by radiological professionals with special training for audits. all radiological units have so far been audited at least twice, and the third time is performed on site in the unit. although the purpose is to audit the process of radiological practises compared to "good practise", special focuses are recommended by the finnish advisory committee for clinical audit, set by the national institute for health and welfare. at the beginning focus was on the organisation and documentation of the structure and processes of imaging practise like justification, optimisation and quality control, but also encouragement and guidance to self-assessment. later special focus has been on paediatric imaging and ct, and the third cycle will target deeper to ct practises. the auditing group includes at least a radiologist and a radiographer to audit a small unit, and a medical physicist if the unit has high-dose modalities like ct. during the audit the auditors work for an independent organisation. multi-professionality gives both insight and the possibility to contact all professionals in imaging practise. the focus of clinical audit is on quality improvement; it is a way to both give and get feed-back and education. it is not control or inspection, but encouragement and guidance to self evaluation and implementation of good practises. we often think of the scientific researcher as detached, objective, and dispassionate, nobly labouring without any expectation of reward. nothing could be further from the truth. scientists are as subject to ambition, competitiveness, envy and even guile as any person. lifting the veil on scientific discovery reveals to us the human dramas that underlie not only the coveting of recognition, but also great conflicts over priority and credit. nowhere are the stakes higher than for that pinnacle of scientific honour: the nobel prize. mri has had a notably tempestuous genesis -a cogent episode that has much to teach us. it involved an explosive clash of personalities; deceit and rampant self-interest; challenges to the basic concept of scientific behavior; legal suits and validation by the united states supreme court; and declamations of denunciation in the international press. it persisted as a year battle -a prize fight, a blood feud -between two protagonists: one whose seminal contribution established the biologic basis of mri, and the other whose flash of insight served as a cornerstone of diagnostic imaging. . metabolic disorders of the skeleton involve the mineralised components of the skeleton. they affect all bone components histologically, but involvement patterns may vary depending on the age of the patient (growing versus adult skeleton) as well as the type of bone (cortical versus trabecular bone). they can be depicted on radiographs and ct images, but remain occult on mr images because the bone marrow is spared in the vast majority of these disorders. mr imaging can help in the assessment of local complication such as fractures. . metabolic disorders of the bone marrow can affect either red (anemia…) or yellow marrow as well as the balance between these marrow types in the body. medical imaging plays a limited role in the assessment of these disorders, but marrow changes associated with these metabolic conditions must be recognised to avoid confusion with neoplastic conditions. in primary and secondary malignant tumours of solid organs, in most of the cases surgical resection is considered the curative treatment. however, this is possible only in about % of the cases with tumour-dependent recurrence rates of % and even more. the main components for adjuvant, neo-adjuvant and finally palliative therapy are the permanently growing number of chemoand antibody therapies. nevertheless, in many cases these therapies provide no definitive or long-lasting success, necessitating multimodality treatment concepts. in the meantime, local ablative techniques represent the main components of these concepts. three main minimally invasive tumour-ablative techniques can be differentiated: chemo-ablation (percutaneous alcohol injection, transarterial chemo-embolisation, chemotherapy, chemo-perfusion), thermo-ablation (radiofrequency-, microwave-, laser-ablation, high-intensified focused ultrasound, cryo-ablation) and radio-ablation (radio-embolisation, interstitial brachytherapy, percutaneous stereotactic radiation). these ablation techniques differ significantly among each other with respect to their mode of action and indications; nevertheless, their benefit is high tolerance and at the same time high therapeutic efficacy, which can easily be combined with other treatment modalities. the different minimally invasive techniques will be discussed together with specific short-and long-term results and complications. missed lung lesions are one of the most frequent causes of malpractice issues. chest radiography plays an important role in the detection and management of patients with lung cancer, chronic airways disease, pneumonia and interstitial lung disease. among all diagnostic tests, chest radiography is essential for confirming or excluding the diagnosis of most chest diseases. however, numerous lesions of a wide variety of disease processes affecting the thorax may be missed on a chest radiograph. the chest radiograph will also help narrow a differential diagnosis, help to direct additional diagnostic measures, and serve during follow-up. the diagnostic usefulness of the radiograph will be maximised by the integration of the radiological findings with the clinical features of the individual patient. ct has a tremendous spatial resolution that helps detect lesions in the chest and has proven to be more sensitive and specific than chest radiographs. however, missing lesions or misinterpreting lesions in ct of the chest is not uncommon. in this session, we will provide interactive cases of chest examinations (radiographs and ct) in which lesions have been missed and or misinterpreted, with a special focus on how correlation with mdct of missed lung lesions can help improve interpretation of plain chest radiographs. this session will explore the value of imaging for facilitating precision medicine, in which molecular data (including genomics, proteomics and metabolomics) is used to classify patients into subpopulations and tailor treatments to the specific molecular characteristics of their diseases. imaging already plays a limited role in precision medicine, primarily through the use of molecular imaging techniques (e.g. pet/ct, mri/pet and hyperpolarised mri) and theranostics (the combination of an imaging agent with a therapeutic entity). in the near future, the role of imaging in precision medicine will increase at a gradually accelerating pace through the development of new molecular imaging probes and theranostic agents and through progress in radiogenomics (the correlation of radiologic findings with genomic features). this session will discuss recent findings and methodologies in radiogenomics as well as the use of nanotechnology for designing novel molecular imaging probes and theranostics. in addition, as theranostics is often considered the essence of precision medicine, the session will provide an in-depth look at current and emerging theranostic strategies and their special benefits for treatment selection, assessment of dose distribution of targeted therapies, and treatment follow-up. radiogenomics and personalised (precision) medicine g.p. krestin; rotterdam/nl (g.p.krestin@erasmusmc.nl) "precision medicine" as well as related notions, such as personalised medicine, or stratification medicine, all revolve around the idea that the consideration of individual characteristics -molecular and otherwise -can improve medical research and practice. precision medicine is a multi-faceted approach to medicine that integrates molecular and clinical research with patient data and outcomes. individual assessment of the location and extent of an alteration is and always has been the basis of medical imaging, whether the 'alteration' is a disease, a malformation, or an injury. as such, medical imaging intrinsically enables "precision medicine". the addition of genomic data in the last twenty years allows new correlations to be made between cellular genomics and tissue-scale imaging. structural and functional imaging and the automated analysis of large amounts of image data have only recently reached a stage where they can be used on a large scale and in a population setting. identifying common genetic variants that contribute to explain variance in imaging phenotypes by a systematic analysis of the genome is based on genome-wide association studies (gwas). the power of gwa analyses has been recently demonstrated with the identification of susceptibility genes involved in a range of imaging phenotypes like coronary artery calcifications, intracranial volume, or size of the hippocampus. these developments highlight new etiological pathways and are expected to improve the understanding of the molecular basis of some diseases. correlation between genotype and imaging phenotypes may be relevant for further characterising the development of disease states. use of nanotechnology, imaging and therapy t. lammers; aachen/ de (tlammers@ukaachen.de) advances in nanotechnology and chemical engineering have led to the development of a significant number of novel materials for diagnosis and therapy. many different diagnostic and therapeutic nano-and micro-materials have been designed and evaluated over the years, including, e.g. gadoliniumcontaining dendrimers, uspio nanoparticles and microbubbles for functional and molecular imaging, and drug-loaded liposomes, polymers and micelles for temporally and spatially controlled drug delivery to tumors and to sites of inflammation. in the literature, such advanced nano-and micromaterials are generally claimed to be highly useful and broadly applicable. a critical reflection on their specific capabilities, however, as well as on their pharmacokinetic properties, biodegradability and toxicity is often lacking. in the present lecture, i will briefly introduce the rationale for using diagnostic and therapeutic nanomaterials. i will highlight several clinically relevant examples in which nano-and micromaterials hold potential for improving disease diagnosis and detail how combining diagnostic and therapeutic properties within a single 'nanomedicine' formulation can be used to individualise and improve (chemo-) therapeutic treatments. learning objectives: . to learn about the design and construction of nanoparticles for use in imaging and therapy. . to appreciate both the advantages and limitations of using nanoparticles as agents for both imaging and therapy. . to learn how nanotechnology enables multimodality imaging and therapeutic agents. theranostics w. weber; new york, ny/us precision medicine requires diagnostic tests that predict the effectiveness of specific therapeutic interventions for individual patients. this close interaction between therapeutics and diagnostics is expressed in the term "theranostics". a frequently used example for a theranostic is the staining of tumours for her expression prior to therapy with her antibodies, such as trastuzumab or pertuzumab. theranostic has also been practised since many years in nuclear medicine by using radioiodine scans to select patients for treatment with iodine- . a more recent example is the selection of patient for peptide receptor-targeted radiotherapy (prrt) by somatostatin receptor imaging. imaging is highly attractive for theranostics, because it allows investigators to study the spatial and temporal heterogeneity of target expression, whereas in vitro assays generally analyse a small part of the tumour at one point in time. several molecular imaging probes, such zr-trastuzumab or folate receptor targeting ligands, are in clinical development as in vivo companion diagnostics. imaging companion diagnostics can also be used to assess tissue pharmacokinetics and thereby help to determine the optimal dose and dose schedule of therapeutic agents. they can also identify unexpected interactions between two therapeutic agents. for example, studies with c-docetaxel have shown that bevacizumab can markedly decrease the intratumoral delivery of docetaxel. finally, imaging-based companion diagnostics can assess target inhibition, as demonstrated by the inhibition of uptake of the androgen receptor ligand f-fdht in metastatic prostate cancer treated with anti-androgens. imaging-based companion diagnostics are therefore expected to play an important role in establishing precision medicine. erus technique as well as the state-of-the-art mr imaging protocols tailored according to the presenting rectal tumor site, allowing trained radiologists to obtain all necessary information for appropriate treatment decision-making, will be described. normal cross-sectional appearances as well as morphological and signal changes encountered in pelvic structures/tissues that may be involved in primary rectal cancer will be illustrated. the influence of imaging findings on initial therapeutic approach and potential limitations of different imaging techniques will be discussed. the staging and categorisation of malignant lymph nodes in patients with rectal cancer is a topical issue and has resulted in some degree of confusion. much enthusiasm has been expended in determining whether mesorectal nodes are, or are not, involved in the primary disease process, as there is a perception that nodal disease is an important determinant of local recurrence. this incorrectly perceived association of nodal status and a high risk of pelvic recurrence have propagated the concept that this should be the primary indication for neoadjuvant therapy regardless of whether tme surgery is to be performed. furthermore, misinterpretation of involved local nodes results in over-treatment of patients, if firstly involved nodes alone are considered as a poor prognostic factor, and secondly the optimal treatment of involved nodes is pelvic radiotherapy plus a non-systemically acting but radiosensitising dose of chemotherapy within chemoradiotherapy (crt) schedules. in this lecture, the evidence for risk factors associated with nodal disease and validated risk factors for local recurrence will be reviewed. the evidence for both the optimal techniques and objective criteria for assessing lymph nodes with magnetic resonance imaging (mri) will be presented. the standard treatment for advanced rectal cancer is preoperative chemoradiotherapy (crt) followed by standard resection of the rectum and mesorectum. neoadjuvant crt allows downsizing and downstaging of the tumour, leading to improved resectability and local control. while the role of mri in rectal cancer treatment is recognised and mri is recommended as part of the standard staging workup, its role for restaging after preoperative treatment is more controversial. this lecture will provide an understanding of whether and how mri can assess treatment response in rectal cancer and how it may impact treatment decision. the attendees will learn about the difficulties in image interpretation and learn about potential new imaging techniques to improve its performance. primary tb typically presents with consolidation in the middle and lower lobes, and necrotic lymphadenopathy. fibrosis, tuberculomas and calcified nodes may result as sequelae. cavitation and tree-in-bud appearance involving the apical and posterior segments of the upper lobes and the superior segments of the lower lobes are the hallmarks of reactivation tb is typically seen in immunocompetent patients. in immunocompromised patients, primary tb findings are more commonly observed, as miliary or disseminated disease or atypical manifestations in case of severe immunosuppression. pleural effusion and tracheobronchial involvement may be observed. the "classical" appearance of non-tb mycobacteria (n tm ), less common and more indolent than tb, typically affects males more than years old with pre-existing pulmonary disease or underlying immunologic disorder. despite a great overlap in the radiologic appearances of both infections, the presence of cavities on sites other than the upper lobes should suggest an n tm infection. the "nonclassical" form related to mac infection that predominantly affects elderly women with no pre-existing pulmonary disease mainly consists in mild bronchiectasis and centrilobular nodules predominantly located in the lingula and middle lobe. a high rate of lymphadenopathy and disseminated disease are seen in immunocompromised patients. the radiologist has to ensure the diagnosis of active tb and mention scarring before starting specific treatments; to suggest n tm infections, immune reconstitution inflammatory syndrome with paradoxical worsening in haart treated hiv-infected patients as multi-drug resistant tb; to appreciate the extent of disease and follow-up; to diagnose the complications; to propose mri or pet scans in some situations. learning objectives: . to appreciate the imaging features of primary and post-primary infections in tb. . to learn about the similarities and differences between tb and non-tb mycobacterial infections. . to understand the radiologist's role in diagnosis. the appropriate investigational technique, frequently targeted differential diagnosis, and the special needs of immunocompromised patients need to be understood by the referring physician as well as by the radiologist. thus, an intensive interdisciplinary co-operation on a patient basis, as well as on a department basis is essential. early detection of a focus is the major goal in febrile neutropaenic patients. as pneumonia is the most common focus, chest imaging is a special radiological task. the sensitivity of chest x-ray, especially in the supine position, is known to be low. therefore, the very sensitive thinsection multislice-ct became the gold standard in neutropenic hosts and might be cost-effective in comparison to antibiotic treatment. the infiltrate needs to be localised, so that this information can be used as guidance for invasive procedures for further microbiological workup. furthermore, the radiological characterisation of infiltrates gives a first and rapid hint to differentiate between different sorts of infectious (e.g. typical bacterial, atypical bacterial, fungal) and non-infectious aetiologies. follow-up investigations need careful interpretation according to disease, recovery, and concomitant treatment. due to a high incidence of fungal infiltrates, interpretation of the follow-up of an infiltrate must use further parameters besides the lesion size. besides the lungs, also other organs systems such as brain, liver and paranasal sinuses need attention and are to be imaged with the appropriate technique. despite advances in diagnosis and treatment, new pulmonary infections have been diagnosed. streptococcus pneumoniae remains the main aetiological agent in outpatients with community-acquired pneumonia (cap). elderly patients or those with toxic habits and various comorbidities favour the development of severe cap. in addition, the development of nucleic acid amplification techniques has emphasised the role of concomitant bacterial and viral pneumonia in the outcome of cap in elderly patients. healthcareassociated pneumonia has been recently defined as a different infectious condition by the american thoracic society/infectious diseases society of america (ats/idsa). the main concern of this new disease is the risk of having an infection due to multidrug-resistant pathogens. with the advent of haart and increased long-term survival of hiv-positive patients, the range of pulmonary manifestations has also evolved. in patients with haematological malignancies or after hsc transplant, aspergillus is a common infection. actually, aspergillus spp. isolation from lrt samples in copd may indicate an increased diagnosis possibility of ipa. new emerging viruses such as human metapneumovirus (hmpv), sars-associated coronavirus, and avian influenza caused by the h n virus have been diagnosed. in , an outbreak of a novel swine-origin influenza a (h n ) virus was reported. the clinical diagnosis of new pulmonary infections as well as the presence of concomitant bacterial and viral infections has been significantly enhanced by improved laboratory methods. a systematic approach to the radiological evaluation of lung infections is essential and includes not only chest imaging pattern recognition, but also integration of available demographic, clinical and laboratory information. the paediatric central nervous system is a complex structure undergoing rapid development. as such, there is a rapid, continuous modification of what is "normal" in relation to age and the stage of development. knowledge of the normal patterns of brain development in the clinically relevant ages from to years is necessary to interpret neuroimaging findings correctly. knowledge of embryology and normal variants is also greatly helpful. mr imaging equipment and parameters need to be adjusted and optimised for paediatric studies. pitfalls often occur from the misunderstanding of normal conditions that are perceived as abnormal based on a comparison with the appearance of the normal brain in adults. this includes, for instance, the evaluation of the brain in the first - years of life during the course of the process of myelination. a summary of the most frequent conditions that may lead to misinterpretation of findings will be provided. a wide spectrum of anomalies of thoracic and abdominal organs may be encountered on radiologic evaluation. these anatomic variants and developmental anomalies can all pose a diagnostic challenge to the radiologist. familiarity with these anomalies, the imaging techniques available for their study, and their variable imaging manifestations is necessary for differentiating them from pathology. a basic understanding of the embryologic development and normal anatomy of thoracic and abdominal organs and vessels is also essential for identifying these anomalies. this review explores the wide variability in appearance of the normal thoracic and abdominal organs during imaging, stressing a thorough understanding of normal anatomy to recognise normal variants. the skeleton of a child is a developing system with a variety of changing normal appearances. imaging studies, especially plain films, are requested for many clinical reasons, and the radiologist is in the position to determine if an image is a normal finding or we are dealing with a lesion. the way the physis and epiphysis grow, ossify, and fuse constitutes a great source of physiologically bizarre appearances, which the radiologist must be familiar with. this talk will concentrate on the plain film diagnosis of some of the most common musculoskeletal variants. other imaging modalities will also be shown when appropriate for the case. irregularities, asymmetries, partial fusions, hypo-or hyper-dense bone areas, accessory bones, prominent normal structures, external artifacts, and potential fracture lines are the most often encountered pseudolesions. a defective radiological technique may also be potentially misleading. patient age, location of the supposed "abnormality" and lack of significant local symptoms are key factors. usually plain films, correlated with regional clinical findings, are the only imaging method that is required. however, in certain doubtful situations, ultrasound, ct, mri, bone scan, or even biopsy may be needed to reach the right diagnosis. unnecessary overuse of these imaging modalities, and the subsequent family anxiety that ensues from this overuse, should be avoided with careful analysis of the x-ray and clinical findings. the purpose of this lecture is to cover the clinical presentation, underlying pathological processes and essential mri features of relatively common conditions affecting the hip. as the hip is afflicted by different conditions according to age, this is how the various pathological entities will be presented. a simple imaging algorithm is presented showing the role of mri. the role of mr arthrography in the assessment of the dysplastic hip and femoroacetabular impingement is covered. a structured approach to mri reporting is outlined. (claudia.schueller-weidekamm@meduniwien.ac.at) mri of the hand requires high spatial resolution, perfect immobility, and homogeneous fat suppression. the hand can either be placed in the superman position, which often is uncomfortable for the patients, or both hands can be placed above the abdomen next to each other while the patient is in the supine position. standard sequences are axial pd, followed by coronal t tse, and t tse fs in the coronal and sagittal planes. additional d gradient echo sequences with fat saturation are recommended to assess ligaments and cartilage, as well as the triangular fibrocartilage complex (tfcc). in certain cases, mr arthrography with thin-slice t fs is helpful for further evaluation of the wrist ligaments. in inflammatory diseases, such as rheumatoid arthritis, static or dynamic t fs sequences are useful to assess tenosynovitis and synovitis. the most common injuries and inflammation of the hand and wrist are discussed with special emphasis on key findings for an accurate diagnosis. the awareness of advantages and drawbacks of other imaging modalities, such as conventional radiographs and ct, should be strengthened. the clinical impact of the radiological report is discussed to strengthen the importance of proper terminology for the description of pathological findings. the structure of the report should be clear and concise, and should allow an interaction and broad communication with clinicians. gastro-enteropancreatic neuroendocrine tumours (gep-net) are a heterogeneous group of cancers that differ in their biology and clinical presentation. diagnosis of these tumours has been improved by advances in pathology and classification and by the combined use of structural imaging and functional imaging modalities. multimodal imaging is increasingly recognised both in detecting and staging disease and also in characterising biological patterns of lesions that may be relevant to the selection and delivery of therapy. in this course, the complex nature of gep-net and the intrinsic uses and limitations of each diagnostic imaging modality will be underlined. insights to hybrid structural and molecular imaging techniques will be provided and discussed. a. tumour biology, pathogenesis and classification b. wiedenmann; berlin/ de (bertram.wiedenmann@charite.de) neuroendocrine tumour cells are characterised by the coexpression of neuronal and epithelial proteins and cellular organelles such as synaptic vesicles containing synaptophysin of neurons and intermediate filaments/cytokeratins of the epithelial cells. based on the presence of secretory vesicles and the continous, uncontrolled vesicular release of biogenic amines, neuropeptides and hormones, patients suffer in half of the cases of so called functional symptoms and syndromes. examples are the carcinoid syndrome (excessive release of serotonine) or the zollinger-ellison syndrome (excessive release of gastrin). activation or inhibition of certain g-protein coupled receptors (e.g. somatostatin receptors) or channel proteins (r-type calcium channels) can lead to the control of the hypersecretion or so called functionality of the affected patients. medical interference with signal transduction pathways involving tyrosine kinase receptors as such mtor and channel proteins can lead to an inhibition of cellular and tumour growth. these observations have led to the establishment of new therapies, especially for pancreatic nets using especially mtor and tyrosine kinase inhibitors. whereas the tumorigenesis is unknown in the case of sporadic nets, hereditary nets appear to develop via a menin mutation through the intermediate stage of hyplasia before they develop the full metastatic potential. based on the above given tumour biological and histopathological findings, a rather robust classification for nets has been developed by the european neuroendocrine tumor society (enets) using a tnm-classification together with a gradingsystem. furthermore, the formerly used terms such as apudoma, neurocrestoma, carcinoid, etc. have been largely replaced by the meanwhile generally accepted term neuroendocrine tumour. this presentation will focus on the pivotal role of nuclear medicine in the diagnosis and treatment of neuroendocrine tumours (nets). the metabolic and molecular imaging capabilities of both positron emission tomography (pet) and single photon emission tomography (spect) have made a great impact on the clinical management of patients with these tumours. a poster child of molecular imaging in oncology is scintigraphy of the somatostatin receptor (ssr). in-dtpa-octreotide has a substantial value in detection, diagnosis and staging of net and more particularly in gastroenteropancreatic nets. newer ssr-binding radiopharmaceuticals have been developed for pet, mainly with gallium- as radiolabel, with higher affinity and more advantageous pharmacokinetics. when used with high-resolution and rapidthroughput multimodal whole body imaging of modern pet/ct cameras, these tracers provide the current state-of-the art ssr imaging. we will also discuss the clinical value of metabolic tracers, such as [ ] -hydroxytryptophan) for amino acid metabolism imaging. the clinical merits and indications of these tracers will be explained. the continuously evolving quest to develop tracer for other receptor systems expressed on nets will be illustrated, e.g. bombesin, vip, cck and glucagon-like peptide receptor ligands. finally, the role of imaging as selection for metabolic and peptide receptor radionuclide therapy will be discussed. gastroenteropancreatic neuroendocrine tumours (gep-nets) are a heterogeneous group of neoplasms that arise from cells of the diffuse neuroendocrine system and may present with a wide spectrum of clinical presentations. their prognosis is mainly related to their biology, proliferation and differentiation. the main goals of imaging are the diagnosis and the staging of these tumours. the diagnostic challenge is very different in functional tumours where clinical presentation and laboratory parameters are of utmost importance and in nonfunctional tumours where imaging may show characteristic features such as hypervascularisation and calcifications. staging is also essential as locoregional involvement and distant metastases (such as liver metastases) may change the therapeutic approach and are major prognostic factors. multimodal workup included morphological imaging modalities with ct and mr imaging, and endoscopic ultrasound was the most useful and functional imaging. the latter includes somatostatin receptor scintigraphy, fdg pet and more recent functional tools such as pet using ga and f-dopa. imaging may also play a role in assessing prognosis in combination with tumour differentiation and tumour proliferation, obtained from pathologic examination. last, imaging is useful in evaluating tumour response after treatment. although surgery remains the only potentially curative therapy for patients with primary gep-nets, other available treatments include chemotherapy, interferon, somatostatin analogues, and targeted therapies. imaging criteria rely not only on changes in tumour size, but also on internal tumour changes. the purpose of this session is to bring the audience up to date with a novel mri method capable of measuring brain perfusion. this method is called arterial spin labelling (asl) and provides a complete non-invasive means to quantitatively assess cerebral blood flow (cbf). as with any mri technique, asl relies on proper setting of many technical parameters to provide an adequate image quality, with minimal influence from potential artefacts. a recent position paper, co-signed by members of the perfusion study group from the international society for magnetic resonance in medicine and the european cost-funded action on 'asl in dementia', has established a series of simple guidelines to help promote this technology in clinical practice. these guidelines will be provided within this session. the created cbf maps will then be analysed, and a simple description of the main features and clinical characteristics of these physiological images will be given to the audience. in particular, a proper difference in the meaning of cbf in several diseases will be highlighted, as it can be either causative, such as e.g. in stroke and cerebrovascular diseases, where a reduced cbf leads to a breakdown in tissue homeostasis, or consequential, as in e.g. dementia, in which a reduction of cbf is the mere reflection of impaired metabolism, combined or not with an underlying brain atrophy. following this session, the radiologist or clinician should be able to better appreciate asl-based sbf maps in several neurological conditions. perfusion is an important parameter to assess the status and liability of organs and tissue. typically, exogenous contrast agents are administered to measure this quantity. arterial spin labelling (asl) is capable of estimating perfusion without the use of exogenous contrast media by labelling inflowing blood magnetically. this labelling process can happen either in a short time over an extended region or over a longer time period, but in a localised area. these techniques are called pulsed asl (pasl) or continuous asl (casl), respectively. for quantification, a critical parameter is the time the labelled blood takes to flow from the region of labelling to the imaging region. this time is typically called bolus arrival time (bat) or arterial transit time (att). it is important to either reduce the influence of this transit time on measured signal intensity or directly estimate it. applying proper bio-physical models, the perfusion-weighted data can then be converted in absolute measures of perfusion. more sophisticated methods allow measuring of the main feeding artery of each imaging voxel (vascular territory mapping) or assessing other parameters beyond perfusion, allowing more detailed assessment of tissue status and function." the use of asl in cerebrovascular disease j. hendrikse; utrecht/nl (j.hendrikse@umcutrecht.nl) obvious applications of arterial spin labelling in clinical mr protocols are cerebral blood flow measurements in patients with acute or chronic cerebrovascular disease. in patients with acute stroke, the cerebral blood flow measurements may indicate the infarct core, with severely decreased perfusion and the infarct penumbra, with decreased perfusion but still viable brain tissue. in chronic cerebrovascular disease, arterial spin labelling cbf measurements show the regionally impaired haemodynamics distal to a carotid obstruction. with adequate collateral blood flow, these areas may be relatively small and with a failure of compensatory mechanism the cerebral blood flow may fall below a critical level. in acute stroke patients, asl mri may show compensatory hyperperfusion in stroke regions after the recanalisation of an occluded artery. other applications of arterial spin labelling are clinical mr protocols in children. in addition to cbf-weighted images, asl mri has also the ability to measure timing parameters: typically, the time it takes for the magnetically labelled arterial blood to flow from the arteries in the neck to the brain tissue, which may be delayed in patients with cerebrovascular disease. furthermore, asl mri has the ability to visualise the (collateral) perfusion territories of the brain feeding arteries in many clinical applications. in patients with cerebrovascular disease, a pitfall may be the absence of label in brain regions due to delayed (collateral) flow, which also may result in high asl signals in (collateral) arteries. learning objectives: . to appreciate the different roles of asl in cerebrovascular diseases. . to become familiar with the different types of calculated images obtained from multi-time points and selective pulses asl. . to become familiar with the limitations and pitfalls of asl. the use of asl in non-vascular brain disease m. smits; rotterdam/nl (marion.smits@erasmusmc.nl) arterial spin labelling (asl) is a non-invasive mri technique with which cerebral blood flow (cbf) can be measured quantitatively. although the first publications of asl mri date over years back, it is only now that asl is commercially available on mri systems from all major vendors and is making its way into clinical practice. the main areas of interest for current and future nonvascular clinical application of asl mri of the brain are dementia and neuro-oncology. asl is proposed as a diagnostic alternative to fluorodeoxy-dglucose (fdg)-pet in the workup of dementia patients. asl has several advantages over fdg-pet. crucially, it can easily be added to the routinely performed structural mri examination. feasibility studies show that asl provides reliable cbf maps in dementia. in patients with established alzheimer's disease and frontotemporal dementia, hypoperfusion patterns are seen that are similar to hypometabolism patterns with fdg-pet. current and future studies in this field will need to demonstrate the validity of asl in the diagnostic workup of the individual patient, early in the disease process. studies on asl in brain tumour imaging indicate a high correlation between areas of increased cbf as measured with asl, and increased cerebral blood volume as measured with dynamic susceptibility contrast-enhanced perfusion imaging. the major advantages of asl for brain tumour imaging are the fact that cbf measurements are not influenced by breakdown of the blood brain barrier, as well as its quantitative nature, facilitating multicentre and longitudinal studies. the analysis of a sinonasal tumour requires mri for differentiation between tumour and secretions or polyps, to assess the local extension and spread into adjacent compartments (skull base, intracranial, orbit, deep facial compartments) and to depict perineural spread. t , t and high-resolution postcontrast series preferably in three, but at least in two different planes, are needed. ct provides additional information of bony structures and serves as a road map for surgical navigation. a fascinating benign sinonasal tumour is inverted papilloma, which has a high recurrence rate if not removed in toto and may harbour carcinoma in < % of patients. ct may show the attachment of the tumour as an area of increased sclerosis, which directs the surgeon to plan the operation. in patients with recurrent inverted papilloma, providing information to the radiologist about its presumed localisation is important, the more so in patients suffering from polyposis. malignant tumours are uncommon and of a large histologic variety. about % arise in the maxillary sinus; % are squamous cell carcinomas. they are often large at diagnosis, because clinical symptoms of nasal obstruction or headache are non-specific. diagnosis is by biopsy. in view of the rapid development of advanced endoscopic techniques, preoperative imaging is extremely important to assess accurately tumour extension. the skull base and dura as well as the lamina papyracea should be scrutinised for tumour locations to foresee an additional cranioendoscopic approach to a standard endonasal endoscopic approach. finally, imaging may play a role in differentiating non-nasal disease, e.g. primary bone tumours, from mucosal disease. to discuss risk/benefit considerations in medical imaging from the perspective of patients and highlight some of the pre-requisites to ensure trust and confidence. this presentation will draw on the work of the european patients' forum on patient safety, quality of care, health literacy, and patient empowerment, to explore what are the key factors to consider in radiation risk analysis, from a patient's perspective, recognising that 'one size does not fit all'. it will examine what constitutes quality information in this environment, effective dialogue between the patient and the radiologist, and the interdisciplinary team, informed consent and transparency of data. it will also highlight some of the particular challenges regarding vulnerable patients, and those requiring on-going treatment due to chronic disease (s). the justification for medical procedures is based on the assumption that the benefit outweighs the risk. this presentation looks at the radiation risk factors from recent high-profile papers (pearce et al., ) and publications from international bodies such as beir and icrp. at lower radiation doses (< msv), quantifying, understanding and communicating radiation risk to staff, patients and referring clinicians presents a range of challenges. advances in health information systems will facilitate more precise dose risk relationships. there are a number of approaches to stochastic radiation risk assessment such as organ-/age-based assessment and effective dose equivalent assessment. at-risk groups, such as children and pregnant women, need special focus. eu directives demand special attention for high-dose interventional procedures and ct scans. these techniques, particularly if repeated, require further risk assessment regarding potential deterministic effects such as erythema, hair loss, and radiation-induced cataractogenesis. this paper looks at strategies in consenting, monitoring, and follow-up of such high-dose effects. the practical implications of the new occupational icrp eye dose limits for interventional practice are also examined. with the advent of higher field scanners in clinical practice and the construction of 'mri compatible' implanted devices, the list of the do's and don'ts while performing an mri examination changes constantly. in this presentation basic safety guidelines and rules will be explained regarding static magnetic field effects, time varying magnetic field effects, radiofrequency field effects and acoustic noise effects both with regard to the patient as well as the personnel using the equipment. due to the advances in medical technology the list of possible 'safe' and 'unsafe' items changes almost daily. therefore it is very important to have all the information about the patient's condition and implants prior to the mri procedure in order to asses possible contraindications in advance. while until a couple of years ago cardiac pacemakers and neurostimulators were contraindicated in the mri environment, the advent of 'mri compatible' pacemakers and other implanted devices introduces challenges in patient safety. in fact these devices are only safe in certain configurations and also in a lot of cases specific mri scan sequences and rf antennas are only allowed. following the european emf directive, where the mri part is derogated, the protection of staff working with em fields also became a topic of debate. what are the possible risks for staff working with mri magnets and how can one implement practical rules for the safe use of the mri equipment. the field of image-guided ablation has expended recently with new ablation techniques like micro-wave, irreversible electroporation, cryoablation. nevertheless, after each treatment, whatever the technique used, we will leave in place in the treated organ, a scar instead of the tumour. follow-up of these treated areas are done by radiologists using ct of mr or contrast enhanced ultrasound. standardisation of terms has been done in by an international committee presided by dr. goldberg. this allows reporting criteria identical from one center and from one country to another. furthermore, it allows a more homogeneous literature and evaluation of success. nevertheless, techniques are not all the same and the cellular and tissue damage in the tumour and around it in the healthy parenchyma are not identical and does not have the same evolution over time. thermal ablation is frequently used to treat liver, lung and bone tumoural lesions because of its safety, efficacy and tolerability. one issue is the lack of reliable imaging modality to assess treatment efficacy and to detect early relapse. several papers have demonstrated that fdg pet/ct is a useful tool to follow-up ablated liver tumour, detecting residual disease easily and earlier than conventional imaging. few reports evaluating the usefulness of fdg-pet/ct in the follow-up of ablated lung lesions are available, but prospective studies show promising results, with a high negative predictive value of this technique. very few data are available on bone lesions and further studies are needed to assess the role of fdg-pet/ct in the follow-up of these lesions. in general, the in vivo typical aspect of completely ablated lesion on fdg-pet/ct is a ring shape, diffuse, peripheral, homogeneous fdg uptake at the treated lesion; on the other hand, the presence of heterogeneous and focal uptake is more frequently related to disease relapse. suvmax (standardised uptake value) is not a reliable indicator. it is higher in case of disease relapse than in completely ablated lesions, but also in case of important inflammatory reaction. finally, the best time point to evaluate ablation efficacy still remains to be defined, but fdg-pet/ct should be performed at least months after treatment to avoid dubious or inconclusive findings due to inflammatory reaction. the aim of thermal ablation treatment is to generate an area of thermocoagulation whose diameter is larger or at least equivalent to that of the tumour. this necrotic scar usually shrinks with time, but most often very slowly. therefore, the criteria of response based on size measurement cannot be applied. the pattern of thermal ablation is similar whatever the thermo-ablation technique used. on ct imaging, thermal ablation areas are well circumscribed and oval shaped. the criterion commonly used to assess the efficacy is the absence of enhancement in the thermal ablation necrosis, which corresponds to tissue devoid of viable tumour. on mr imaging, the thermo-ablation areas are typically hyperintense on unenhanced t due to the presence of proteinaceous material, and hypointense on t , explained by the dehydrating effect of thermal damage. the residual tumour is typically round in shape and located at the periphery of the necrotic area or in contact with large vessels. mr imaging allows earlier detection of residual liver tumour than ct imaging. neurodegenerative diseases, cancer and heart disease perhaps summarise the major challenges to medicine in this century, a challenge in accurate diagnosis and also in treatment. recent technological advances in medical imaging technology mean that these challenges can be dealt with more efficiently. anatomy can be studied faster, safer and with higher spatial resolution and precision. in addition to anatomy, the study of function has become feasible; more sophisticated hardware and software are being used to get the required data to map out the functionality and understand how this relates to anatomy to offer a more comprehensive view of normal development and explain pathological conditions. at the same time, researchers and clinicians are urged to create a platform for reciprocal communication to facilitate translation of the research findings to a graspable clinical benefit for the patients. this translational platform will allow flagging the major clinical questions, inform current and future research experiments and at the same time help utilise recent research findings in the clinical setting. this session will review state-of-the-art mri techniques and document different approaches to translational imaging. it will also allow understanding the obstacles and remedies in implementing translational medicine. many neurological diseases are associated with progressively reduced cognitive function. recent evidence suggests that proficient cognitive function depends on an appropriate interaction between large-scale cognitive control networks in the brain. it is hypothesised that damage to white matter microstructure, as found in many neurological diseases, disrupts the integrated operation of these networks and results in impaired cognitive function. diffusion tensor imaging (dti) has been used to investigate alteration to white matter micro-structure and resting state fmri (rsfmri) has proven to be a useful technique to assess brain function in multiple cortical systems. however, the combined use of these techniques in the clinical setting has not yet taken hold. in this presentation, i will demonstrate the flexibility of these mri techniques in assessing brain structure and function and address their feasibility of use in a clinical setting. imaging in oncology is a growing field within radiology. different tomographic techniques are used either isolated or as multimodality-multivariable imaging, as there is an increasing need to combine morphological and functional information. different cutting-edge modalities, such as pet-ct, dw-mr, ce-mr and mr-spectroscopy, are standard in most clinical scenarios. new images have advantages providing excellent soft-tissue contrast and multidimensional functional, structural and morphological information. the development of new diagnostic imaging research areas, mainly in the field of oncology, cardiology and neuropsychiatry, will impact the way medicine is performed today. both clinical and experimental multimodality studies, in humans and animals, will have to demonstrate an efficient use of the imaging information provided by the modalities to affect the future of medical imaging. we will comment on the impact of continued technological developments in medical imaging on patients with cancer (diagnosis, staging and follow-up) and the challenges of imaging technologies from research into clinical reality. the recent advances and developments in measurements and biomarkers which have led to a greater understanding of cancer will be commented, together with the morphologic, metabolic and functional oncological imaging research and clinical practice. finally, the role of the radiographer in advanced oncological imaging techniques will be evaluated. when imaging the head and neck region with ct or mri, teeth are always present. we frequently encounter radiolucent or radiopaque lesions of the jaw on scans performed for other indications. familiarity with typical dental conditions is necessary to subject the patients to the right therapy. to correctly evaluate and describe those lesions, this special focus session on dental imaging gives the radiologist a tool to report dental lesions. the anatomy of panoramic and ct images will be reviewed in the first talk. technical details as well as pitfalls will be presented. in the second talk, the focus is on pre-and post-surgical imaging with ct where typical lesions of the jaw will be presented. an insight into imaging the teeth with mri will be given by the third speaker. the discussion on the impact of radiologists on dental images will complement this special focus session's theme. panoramic radiography produces a single image of the maxilla, mandible, teeth, temporomandibular joints and maxillary sinuses. during the exposure, the x-ray source and the detector rotate synchronously around the patient producing a curved surface tomography. this horseshoe-shaped zone of sharp image is called the focal trough/image layer. advantages of this technique are a good overview of lower facial hard tissues, the convenience of an extraoral examination, a low patient radiation dose, and low costs. disadvantages are limited width of the sharply imaged layer, variable magnification, and oblique projection, especially in the upper premolar region. patient positioning is most important for obtaining a detailed panoramic radiograph. in addition to patient position errors, patient movement during exposure and metal objects can cause artefacts. also, ghost images caused by structures located between the x-ray source and the rotation centre can hamper diagnostics. cbct is a radiographic imaging method that allows accurate d imaging of hard tissues. during a cbct scan, the scanner rotates around the patient's head ( °- °) obtaining multiple sequential planar projection images by a d detector. the scanning software collects the data which is processed to create a volumetric data set with isotopic voxels with varying fov. disadvantages of cbct imaging are poor soft tissue contrast and artefacts. in addition to normal panoramic and cbct anatomy, after this presentation the audience will be familiar with the panoramic and cbct techniques as well as common errors and pitfalls of these techniques. pre-surgical and post-surgical imaging with mdct and cone-beam ct a. gahleitner; vienna/at (andre.gahleitner@meduniwien.ac.at) multislice-ct and more recently cone-beam-ct have become an established method for anatomic imaging of the jaws prior to dental implant placement. commonly referred to as "dental-ct", these high-resolution imaging techniques have gained importance in diagnosing dental-associated diseases of the mandible and maxilla. especially in patients with inflammations, cysts, oro-antral fistulas, odontogenous sinusitis, impacted teeth and dental fractures, new indications have emerged. unfortunately, most radiologists have had little experience in this area and many of the dental-ct findings remain undescribed. hence, we will determine the ct appearance of frequent dentalrelated diseases of the jaws and demonstrate typical pre-and postoperative findings. this presentation reflects our experiences from years of performing dental-ct with . investigations, closely working together with dentists and oral surgeons. learning objectives: . to learn about the typical findings from pathological conditions of the jaw, which confront dentists and oral surgeons. . to understand these findings in cross-sectional imaging like multislice and cone-beam ct. . to learn about the usual treatments for these conditions in order to know how they are used in dental radiology. dental mri s. rohde; dortmund/ de (stefan.rohde@klinikumdo.de) this lecture will discuss the potential of dental mri in the field of inflammatory and neoplastic disease of the periodontal space. experimental and preliminary clinical data from high-field dental mri will be presented with a special focus on the visibility of critical anatomical structures on high-resolution sequences. the results will be compared to high-resolution cone-beam ct. based on representative cases, we will review the main causes of thoracic non-traumatic vascular emergencies (acute aortic syndrome, pulmonary thromboembolism and haemoptysis); mdct angiography has become the firstline imaging test for the diagnosis of these entities. as acute aortic disease is the most common fatal condition in patients with chest pain and prompt recognition and treatment are of paramount importance, we will review the spectrum of acute aortic pathology focusing on the distinctive findings of each entity (classic dissection, intramural haematoma and penetrating aortic ulcer) and upgrading the clues for their diagnosis. acute pulmonary embolism (pe) remains a common clinical challenge. mdct pulmonary angiography has become the first-line imaging study in the diagnosis of pe because of its speed, accuracy, low interobserver variability and ability to provide alternative diagnoses. we will review the role of mdct in the evaluation of acute thrombotic pe: findings of acute pe (including how to evaluate the severity of an episode of pe at ct pulmonary angiography) and some potential pitfalls. massive haemoptysis is a life-threatening condition that is associated with a high mortality rate. haemoptysis usually involves bleeding from the bronchial arteries or, less frequently, from nonbronchial systemic arteries. haemoptysis of pulmonary arterial origin is rare, estimated at less than % of haemoptysis cases. mdct angiography permits noninvasive, rapid, and accurate assessment of the cause and consequences of haemorrhage into the airways and helps guide subsequent management. acute respiratory failure can have multiple underlying causes including infection, fluid overload, immunological diseases or exacerbation of preexisting lung disease. since the clinical symptoms are nonspecific, imaging plays an important role. the first imaging method is mostly the chest radiograph, easy to access and to obtain, but non-diagnostic in many cases. (hr)ct offers more possibilities to define the differential diagnosis. the option of this interactive workshop will be to get familiar with the spectrum of diseases that can cause acute respiratory failure and learn about key findings in radiography as well as ct to reduce the differential diagnosis. the interaction between preexisting lung disease, clinical information (e.g. chemotherapy, rheumatoid arthritis, copd) and imaging findings will be discussed using clinical case studies. options and also limitations of imaging findings will be illustrated. the following scenarios will be taken into account: acute cardiac failure and various appearances of oedema; acute immunological-toxic disorders including druginduced lung disease and inhalational injuries; exacerbations of preexisting lung disease including fibrotic and obstructive lung disorders; severe infections causing respiratory failure and their complications. contrast-enhanced ultrasoud (ceus) and elastography are evolving us techniques that have already found their ways into clinical practice (ceus) or will do so in the next few years (elastography). ceus is performed with nd generation us contrast media, but these contrast media are not available worldwide. in many european countries, sonovue® (bracco, it) is the only available contrast, consisting of micro-bubbles less than the size of red blood cells. in contrast to ct and mr contrast media, these bubbles are strictly intravascular. for us imaging, low mechanical index us techniques are used to see the resonance of sound by these bubbles. ceus is excellent for the differential diagnosis of focal liver lesions based on vascularisation and specific contrast uptake (imaging the wash-in and wash-out of us contrast over time with excellent temporal resolution) and increases us sensitivity to liver metastases in colorectal cancer patients. however, ceus is only useful when there are excellent us conditions. in pancreatic us, ceus allows to differentiate adenocarcinoma from neuroendocrine tumours based on vascularisation (us contrast uptake). whereas elastography (strain elastography or shear wave elastography) is established for the diagnosis of liver fibrosis, there are no general recommendations for using these various techniques to measure the "hardness of a focal lesion" in focal liver and pancreas lesions. diffusion-weighted mr imaging (dw-mri) provides information on tissue cellularity, extracellular space tortuosity and integrity of cell membranes by measuring the motion of water molecules in tissues modified by flows (blood vessels, glandular ducts,.). and interaction with cellular components. integrated in conventional-mr acquisition protocol, dw-mri increases sensitivity for lesion detection. the apparent diffusion coefficient (adc) can be calculated using a mono-exponential relationship between signal attenuation and b-value. this fitting is influenced by microcapillary perfusion. a more sophisticated approach (intravoxel incoherent motion, ivim) would enable estimation of parameters that separately reflect tissue diffusivity and microcapillary perfusion. using ivim-analysis, the derived quantitative parameters describe tissue diffusivity, perfusion and perfusion fraction. by providing qualitative and quantitative information, dw-mri is applied in oncology to characterise malignancy, including lesion aggressiveness, and to monitor treatment response. perfusion imaging is a useful tool to achieve information concerning tissue vasculature, microvascular permeability and interstitial space characteristics. dynamic contrast-enhanced techniques (dce-ct, dce-mr) are based on the analysis of the contrast agent (tracer) biodistribution in tissues. quantitative parameters are obtained using pharmacokinetic models, as transfer constant ktrans, extracellular-extravascular space fraction ve, vascular tissue fraction vp and the rate constant kep. quantitative dw-mr and dce-mr parameters are increasingly used as imaging biomarkers to predict tumour response and/or to monitor the effects of therapy, especially when anticancer agents with novel models of action are used (anti-angiogenic, tyrosine kinase inhibitors and several others). therefore, these new imaging biomarkers may have a pivotal role in correctly evaluating tumour response and stratify and managing cancer patients. elderly and the young can all present special problems that require a greater depth of understanding to obtain a diagnostic study. this session will help you in your quest for reliable high-quality investigations for all. a. coronary cta in patients with severe arrhythmias and high heart rate c. loewe; vienna/ at (christian.loewe@meduniwien.ac.at) besides all advances in scanner technology, heart rate remains a critical issue for coronary cta, and motion artefacts due to cardiac function represents still the most frequent reason for limited diagnostic image quality in cardiac cta. thus, the target heart rate is defined depending on the scanner system used and ranged between below bpm and below bpm. however, in a number of patients heart rate control is not possible or not successful (i.e. children, emergencies, heart transplant recipients), whereas a relevant proportion of ctas are performed in patients with heart rates higher than the target frequency. during this presentation, the possibilities for heart rate control by beta blocker injection and beyond will be discussed and possible workflows will be presented. furthermore, examination strategies for patients with high heart rates will be presented including reverse dose modulation, high-pitch scanning, systolic scanning and more. finally, tools for reconstruction and assessment of patients scanned at higher heart rate will be introduced. different to high heart rates and even more critical with regard to image quality is arrhythmia. due to the complexity of cardiac synchronisation, a ct suite is not the place for cardiac resynchronisation, whereas the indication to cta has to be reevaluated depending on referring diagnosis and severity of arrhythmia. since there are patients undergoing cta because of arrhythmia including patients prior to ablation treatment, strategies for examination and image reconstruction in case of arrhythmic patients have to be established and will be presented. cardiac computed tomography (ct) has become a widely available diagnostic tool used in a range of heart conditions. the commonest application of this technique is in the evaluation for coronary artery patency (coronary ct angiography) in patients with chest pain. when coronary arteries are heavily calcified, or post-coronary angioplasty with stent implantation, diagnostic problems can occur. in these circumstances, the evaluation of the coronary arteries on ct is hampered by the occurrence of high-density artefacts caused by calcifications and stent struts. these artefacts may preclude the appropriate assessment of the coronary lumen. the presence of motion artefacts in the dataset or image noise in very large patients may exacerbate the problem. in this scenario, accurate patient selection and preparation remain key to ensure that the diagnostic yield of the cardiac ct study is good. optimisation of scan parameters (kv), contrast injection protocol and use of appropriate postprocessing techniques (e.g. dedicated convolution filters) play an important role in daily clinical practice. recent technical developments include dualenergy scan techniques and gemstone spectral detector systems that acquire simultaneously high and low kilovoltage datasets. this is done to achieve tissue differentiation. in principle, by using monochromatic image reconstruction, the effect of high-density artefacts may be decreased. using a similar principle, high-density structures can be subtracted from the image. the introduction of iterative reconstruction algorithms may play a role in that these algorithms are theoretically more accurate in the modeling of physical noise and tissue geometries. lack of movement artefacts is one of the major prerequisites for good image quality in cardiac mr. it is crucial to have the minimum possible image acquisition time. it is important to make both te and tr as short as possible. data acquisition should be synchronised with patient's ecg or pulse. special attention should be paid to good quality of ecg recordings (good contact of electrodes with the skin, electrodes positioning, using of dielectric pad). in patients with arrhythmia, prospective ecg synchronisation should be used instead or a retrospective one, or special protocols may be applied for arrhythmia correction. breathing artefacts are usually not a problem for cardiac mri, because most sequences are acquired during a single breath-hold. using ssfp and parallel imaging allows obtaining a complete set of cine mr images through the whole heart in - short breath-hold periods. the technique of realtime cardiac mr is a good way to perform successful examinations even in difficult patients, especially the ones with heart failure. other systemic mrartefacts (aliasing, chemical shift, magnetic susceptibility, off-resonance) should be recognised and diminished or eliminated by the operator. these artefacts are more prominent in case of t systems. late-enhancement studies with gd are very dependent on the correct selection of the ti time. phase-contrast mri requires venc calibration in cases of valve stenoses. to meet the challenges and the benefits of cardiac mri, one must balance the constraints of signal-and contrast-to-noise ratios, spatial and temporal resolution, scan time and image quality. scoliosis, a common spinal deformity in teenagers, especially girls, progresses during their growth until the end of puberty. therapeutic choices, which are either medical -with corset -or surgical, depend on this progression. follow-up is mainly clinical. however, radiographs are often indicated to give precise information: ap and lateral views of the spine in standing position remain the basis of this follow-up. obtaining a good image quality at all levels of the spine has been a technical challenge for a long time, and the question of radiation dose is also a main concern, as these radiographs require a high radiation dose. fortunately, radiographic techniques have greatly improved over the past few years: from the conventional or cm film to digital screens and now flat screens, we now obtain better quality images with less radiation. today, eos system using xenon chamber is the best to give good ap and lateral views. it allows d reconstructions with minimal radiation dose. on these radiographs, measures can be performed, essentially cobb's angle. it helps in evaluating progression of the curves and making surgical decisions. other imaging techniques are requested only in particular situations as in the presence of neurological symptoms or general disease such as neurofibromatosis, or in young children with malformative scoliosis and sometimes preoperatively. ct is best to study the vertebrae; it objectivates vertebral malformations (agenesis, hypoplasia, blocks). mr will be requested to study the cord, nervous roots, craniospinal junction and the soft tissues. collaboration between the radiologist and spinal surgeon is essential. the key elements in the diagnostic process, as for most bone lesions, are: the age of the patient, the type of bone and bone segment involved, the location within the bone, the presence of signs of aggressiveness (type of osteolysis, limits, presence and type of periosteal reaction). conventional x-rays should never be absent from the initial workup of a focal bone lesion in a child, as they provide essential features for differential diagnosis and, in most cases, eliminate the need for other more costly techniques. the need and priority for other imaging techniques, such as ct, mri and bone scintigraphy, are decided on a case-by-case basis and sometimes a combination of these techniques, which complement each other, is needed. it is the radiologist's responsibility to recommend histological analysis of a lesion via biopsy whenever the imaging aspect is not characteristic or indicates signs of possible malignancy. learning objectives: . to recognise the most common benign bone tumours and pseudo-tumours. . to understand the differences between benign bone tumours, pseudotumours and malignancies in children. . to understand imaging modalities that could help in the differential diagnosis of benign bone tumours and pseudo-tumours in children. a- : a.c. offiah; sheffield/uk (amaka.offiah@nhs.net) the nosology and classification of genetic skeletal disorders subdivides conditions into groups defined by molecular, biochemical and/or radiographic criteria. conditions are associated with mutations in at least one of different genes. some of these conditions are rare or even extremely rare (occurring in fewer than in , of the population). clearly, the general paediatric radiologist cannot be expected to correctly recognise and diagnose all of these conditions. the approach is to perform a set of radiographic images dependent on the patient's age/size and to develop a standard system for reviewing these images. the standard set of images for a patient over to years of age consists of ap and lateral skull, ap chest, ap pelvis, lateral thoracolumbar spine, ap one upper limb, ap one lower limb and dp left hand. variations on this routine dysplasia skeletal survey will be discussed and a standard system for interpreting the images will be presented using radiographs of the commoner skeletal dysplasias as examples. in the last few years, interest in body composition (bc) analysis, which is the quantification and characterisation of relative amounts of muscle, fat, bone, and other vital parts composing the human body, has grown rapidly for clinical, research and epidemiological purposes. studying body composition has gained great importance for the comprehension and decoding of a multitude of patho-physiological processes (e.g. obesity, diabetes and endocrine diseases and also gastrointestinal, renal, nervous, infectious diseases, etc). and physiological and para-physiological conditions as in athletes or growth and ageing processes. although the main imaging techniques, which include dualenergy x-ray absorptiometry (dexa), computed tomography (ct), and magnetic resonance imaging (mri), offer a differentiated and attractive analysis of bc, they still need to find a definite position in clinical practice. through 'programming' low birth weight is associated with increased rates of coronary heart disease, stroke, hypertension & non-insulin dependent diabetes. prediction of morbidity and mortality from body composition, particularly fat, stimulates the importance and application of accurate and precise methods for quantitative assessment of body composition; the gold standard for which is cadaver analysis, but other methods have to be implemented in vivo. body composition alters in many chronic diseases, starvation cases, metabolic syndromes, and senescence stages, and is a key component of health. several techniques are available which vary in simplicity and complexity of use. each make assumptions and may not measure body composition directly, but make predictions from other measurements made. skinfold thickness measurements assess regional fat and are quick and simple to perform at all ages. reproducibility is good, but less so in obesity. there is some limitation of reference data to express results as standard deviation scores (sds). body mass index (bmi; weight/height ) is widely used as an index of relative weight expressed as sds for gender, age and ethnicity. waist circumference (wc) is a measure of central fat, and can also be expressed as a ratio to hip circumference. bioelectric impedance analysis (bia) measures body impedance to a small electric current, which estimates total body water (tbw) from which can be derived fat free mass (ffm). tbw can also be measured by neutron activation. air displacement plethysmography (bod pod) is a new method for body composition assessment. learning objectives: . to understand normal body composition in children and adults. . to learn how body composition can be altered by disease. . to appreciate the clinical relevance of assessing body composition. . to understand the non-imaging methods available for measuring body composition, with their advantages and limitations. computed tomography: what does it measure and how? j. damilakis; iraklion/ gr (damilaki@med.uoc.gr) ct allows measurement of total body fat and enables differentiation of subcutaneous from visceral adipose tissue. body fat can be estimated using the conventional technique of manual planimetry. using planimetry, the user delineates manually the boundaries of subcutaneous and visceral fat in each ct image. however, this method is labour intensive, time-consuming, and, therefore, not easily applicable in routine clinical practice. another ct method used for the estimation of abdominal fat is the semiautomatic measurement of adipose tissue area. abdominal fat is assessed in a ct slice by using a fixed range of hounsfield units to define the area of adipose tissue. a limitation of this method is that the attenuation range of fat may vary among individuals. the stereological volume estimation method is based on the cavalieri's principle. according to this principle, the volume of an object can be measured by cutting it into equally spaced slices and measuring the area of the object on each slice. to measure body fat using stereology, a square grid of test points is randomly superimposed on each ct section. all points lying inside the fat tissue region are selected by the user and the software automatically calculates the total number of points hitting the fat. stereology provides the possibility for efficient fat volume assessment. however, research studies are needed to optimise the stereological estimation of fat and compare stereological measurements with those from other adipose tissue measurement ct techniques. learning objectives: . to appreciate the role of ct imaging in body composition analysis. . to learn how to measure visceral and subcutaneous adipose tissue using ct. . to understand the advantages and limitations of ct techniques (planimetry, thresholding, stereology) in evaluating adipose tissue. mri: current and future applications d.c. karampinos; munich/ de (dimitrios.karampinos@tum.de) mri has been emerging as an ionising radiation-free imaging modality to measure fat distribution in the human body and fat content in different organs. the present talk will first introduce mri methods to measure fat distribution, including the well-established t -weighted imaging and the emerging dixon imaging approaches. the challenges related to the data acquisition and image post-processing of the two imaging approaches in the study of fat distribution will be reviewed. examples will be shown from the current use of mri in investigating adipose tissue distribution alterations or differences in patients with metabolic disorders, including obesity and diabetes. quantitative mri methods enabling the measurement of fat content with high spatial resolution will be then presented, with a focus on water-fat separation methods. the technical challenges associated with the establishment of mri-based fat content, as an accurate and reproducible imaging biomarker will be discussed. the selection of pulse sequence parameters and image reconstruction algorithm in a state-of-the-art water-fat separation experiment will be explained. applications will be shown from the growing literature of mri-based fat quantification in abdominal organs (e.g. liver) and in skeletal muscles, aiming to show the great premise of quantitative water-fat mri in quantifying fat content with high spatial resolution in different body parts. at the end of the talk, the potential of quantitative water-fat mri will be discussed in the emerging application of measuring bone marrow fat content and its association with bone health and metabolic disorders. the key role of dual-energy x-ray absorptiometry (dxa) in the management of metabolic bone diseases is well known. the role of dxa in the study of body composition and in the clinical evaluation of disorders which directly or indirectly involve the whole metabolism as they may induce changes in body mass and fat percentage is less known or less understood. dxa has a range of clinical applications in this field, from assessing associations between adipose or lean mass and the risk of disease to understanding and measuring the effects of pathophysiological processes or therapeutic interventions, in both adult and paediatric human populations as well as in pre-clinical settings. dxa analyses body composition at the molecular level that is basically translated into a clinical model made up of fat mass, non-bone lean mass, and bone mineral content. dxa allows total and regional assessment of the three abovementioned compartments, usually by a whole body scan. since body composition is a hot topic today, manufacturers have steered the development of dxa technology and methodology towards this. new dxa machines have been designed to accommodate heavier and larger patients and to scan wider areas. new strategies, such as half-body assessment, permit accurate body scan and analysis of individuals exceeding scan field limits. although dxa is a projective imaging technique, new solutions have recently allowed the differential estimate of subcutaneous and intra-abdominal visceral fat. the transition to narrow fan-beam densitometers has led to faster scan times and better resolution; however, inter-or intra-device variation exists depending on several factors. in cancer patients, disease-free survival is a good indicator for tumour response, but for many common cancers, treatment of disseminated disease is often noncurative. the increased duration of survival is related to changes in tumour size after treatment. however, the anatomical determination of tumour response has some limitations, especially when non-cytotoxic targeted therapies are used. with these new treatments options, the lack of progression may be associated with a good improvement in outcome, even in the absence of major shrinkage of tumours. new imaging biomarkers are therefore needed to assess therapeutic response. molecular imaging is now playing a prominent role in the monitoring of cytostatic targeted therapies. pet-ct, dynamic contrast enhancement studies or diffusion-weighted imaging are the most promising ones. the aim of this session is to present the state of the art in tumour response assessment with regard to new therapeutic regimens. in early oncologic times, treatments of different malignancies were reported in different ways. c. gordon zubrod and others first articulated the model of multicentre clinical trials and argued for standards to be agreed for included and excluded subjects, the method of assigning treatment and in measuring response. nowadays, we have better knowledge of tumour biology; however, we continue using the classical response criteria (who and recist) and overall survival as a primary goal for effective treatment in most of the malignancies. in the last few years, different criteria for responses have been reported to evaluate the disease and a lot of discussions between authors have been reported. now, we can use the recist criteria v . , implement the percist criteria and also forget the immunotherapy criteria. furthermore in the last few years, different treatments such as vaccines, antiangiogenics and targeted therapies have increased our arsenal in the treatment of different malignancies such as kidney cancer, prostate cancer, breast cancer and so on. another problem for our patients is that those diseases have a long evolution after the first-line treatment. we have to be able to define in each disease the treatment line and type of treatment to determine which evaluation criteria are necessary and which is the objective of treatment, pfs or os. we have to be aware that if we are not able to define such topics, we may be losing active treatments. recist . are the criteria most often used in studies to evaluate response to treatment in solid tumors. on each follow-up examination, response is defined by a combination of unidimensional measurement of 'target lesions', qualitative evaluation of 'non-target' lesions, and presence/absence of 'new lesions'. ct and mri are the preferred modalities for recist evaluation, and evaluations must be performed using the same modality, in the same plane. we will review anatomic locations which should be avoided, and how to deal with intercurrent events resulting in the impossibility of measurement. cases when recist seems inadequate will be discussed, including evaluation of bone metastases, and focal therapy. finally, though these criteria were developed for drug trials, they may provide a frame for reading examinations and writing reports in routine practice. reproducible, objective and quantitative criteria help to define or in the obese patient. ct is especially helpful in those with suspected retroperitoneal pathology, in the investigation of immunocompromised patients and those with complex post-operative problems. ct guidance may be required to facilitate percutaneous abscess drainage, particularly for deep pelvic collections. in addition, ct plays a vital role as a 'problem-solver' when ultrasound has failed to fully answer the clinical question posed. this short presentation will utilise clinical studies to illustrate the benefits of ct in the investigation of the acute abdomen, both as the initial imaging modality and as a second line tool following ultrasound. learning objectives: . to understand the advantages of ct as a primary imaging modality for children with acute abdomen. . to learn about clinical scenarios in which ct is relevant. . to appreciate the use of ct as an adjunct to ultrasound. abdominal trauma: us is better v. miele; rome/it (vmiele@sirm.org) ultrasonography (us) is a reliable technique whose advantages are rapidity, portability and accuracy in depicting intraperitoneal fluid without interrupting resuscitation and without radiation exposure. in the haemodynamically unstable paediatric patients, it represents the first line together with the x-ray evaluation. us for trauma has become more standardised and is worldwide known with the acronym of fast (focused abdominal sonography for trauma) or e-fast (extended-fast) used in depicting also pleural and pericardial effusions and pneumothorax. nevertheless, many parenchymal injuries are not correctly visualised at baseline us and some traumatic solid organ lesions can occur without hemoperitoneum. in case of haemodynamically stable patients, who have suffered a low-energy trauma, the greater time available allows the use of specific us contrast agents, enabling a better identification of traumatic organ injuries. contrast-enhanced ultrasonography (ceus) has a greater sensibility and specificity in the identification of parenchymal traumatic lesions, both in the first evaluation and follow-up, and could avoid unnecessary radiation and iodinated contrast medium exposure. in case of haemodynamically stable paediatric patients, who have suffered a high-energy trauma, us is not recommended as first-line investigation, because ce-mdct should be performed first. only in the follow-up, ceus can be considered an alternative to ct. in conclusion, e-fast and ceus should be considered as a useful tool in the assessment and monitoring of paediatric trauma. this examination can be performed at the patient's bedside, representing a useful alternative to ct in the paediatric traumatised patients and in the follow-up of a known abdominal injury. learning objectives: . to understand the differences in the diagnostic paths of paediatric patients. . to learn about the diagnostic efficacy and limitations of ultrasonography. . to become familiar with the use of the contrast-enhanced ultrasonography (ceus). abdominal trauma: ct is better m. raissaki; iraklion/gr (mraissaki@yahoo.gr) ct has been considered a sensitive, specific, and accurate test in the identification and grading of injuries, especially in the severely injured child. it should be overemphasised that traumatised children differ from adults: haemodynamically stable children may actually be actively bleeding and rapidly deteriorate. conversely, children have smaller calibre vessels, stronger vasoconstriction, and stronger solid organ capsules; bleeding may stop spontaneously, organ rupture is more difficult and delayed rupture is rare. this is why few children will undergo laparotomy or trans-arterial embolisation. the goal of imaging is to clear the abdomen in multi-traumatised children, identify those that may rapidly deteriorate because of clinically silent active bleeding, increase the surgeon's confidence level and define short-and long-term medical management. due to children's increased radiosensitivity, all ct scans should be indicated based on appropriate early clinical evaluation, assessment of risk factors for abdominal injuries and evaluation of the closely monitored child. alara includes the availability of outside scans upon admission, avoiding non-contrast scans and multiple phases and applying age-and weight-dependent exposure parameters. ct should not be performed as follow-up unless there is clinical deterioration. ct has the advantage of rapidly identifying and delineating with high resolution solid organ, vascular, mesenteric, bowel injuries and pending oligaemic collapse (hypoperfusion complex or shock bowel). important fractures and thoracic injuries may be simultaneously revealed with ct. the objective documentation of intraabdominal injuries is extremely important in suspected child abuse. evaluation of mr imaging in a patient with an intramedullary lesion should focus on key features: a) the location of the lesion on the cross-sectional area of the cord, best evaluated on axial images, b) the length of the lesion evaluated on sagittal images, c) the presence of cavitation and cysts, d) signal intensity on t -wi, e) the presence of enhancement and enhancement type, and f) associated leptomeningeal enhancement. the knowledge of the presence or absence of the brain lesions is important information for narrowing the differential diagnosis. the clinical picture and the onset of symptoms will help the differentiation between the neoplastic and inflammatory and vascular lesions. in this lecture, the imaging characteristics and typical patterns of intramedullary lesions will be presented. a diagnostic algorithm, which includes imaging, clinics and csf analysis, will be discussed. a common clinico-radiological situation is the differentiation between spinal infection, degenerative changes and osteopaenia, and spinal tumour. in most cases, conventional ct and/or mri will allow a certain differentiation, and additional biopsies are mainly used to obtain tissue for bacteriological or histopathological classification. advanced mr techniques such as dwi may sometimes help in increasing diagnostic certainty, but are seldomly used in day-to-day practice. spinal infections typically present around the intervertebral disc, but may also present as isoloated spondylitis without disc involvement, and sometimes the infection quickly spreads to the epidural space. degenerative changes my sometimes mimic infection due to strong gadolinium enhancement. the main differential diagnosis of spinal tumorous processes is osteopaenic changes (oedema), and ct may be helpful in such cases, for example to demonstrate degenerative gas in the vertebral body. rare causes of spinal abnormalities such as atypical pathogens and rare tumours may cause diagnostic problems; often the fastest road to a diagnosis is biopsy in those cases. the correct clinical diagnostic approach to spine disease is mandatory in the selection of patients to be treated. the anatomy and the relationship between different structures must be known in detail to understand the source of the pain and so to treat it percutaneously. mr, ct, dynamic x-ray and nm bone scan in selected cases can be used to understand the reason for the pain. mr with t stir sequence or t fat supp technique is mandatory to show bone marrow oedema and to decide on which metamer to perform the treatment. ct is often necessary after mr in primary and secondary spine tumours. disk disease can be seen either on ct or mr. two major treatments must be considered: discogenic pain and vertebrogenic pain, and for this reason disk treatment and vertebral treatment. disk treatment includes many mini-invasive systems to use; however, no treatment has been shown to be superior to the others, with good clinical results in - % of the cases, even at long-term follow-up. vp and assisted techniques are available for the treatment of not within a few minutes following ablation, ct of the ablation zone will demonstrate an extensive area of ground glass opacity (ggo). a ggo of at least . mm peripheral to the tumour is predictive of complete ablation. as early as hours post-treatment, the entire ablated region usually appeared as a well-demarcated homogeneous dense opacity on ct that corresponded to necrotic tissue and its surrounding rim of granulation tissue. this zone of ablation is the "baseline post-ablation imaging" for follow-up. then, a relatively slow involution of the ablation zone will occur with various patterns, including nodular, fibrosis, disappearance, cavitation and atelectasis. morphologic features of local tumor progression are an increase in the overall size or a change in the shape of the ablation zone (even without enhancement). it is generally considered that an ablation volume that does not increase in size on subsequent imaging after the baseline post-ablation imaging is a complete ablation. the relatively slow decrease in size of the ablation zone renders ct morphologic evaluation responsible for late discovery of local tumour progression. pet/ct is able to discover incomplete ablation earlier than ct at a stage the disease remains small. patients who have very early evaluation with pet/ct are at risk of either false-positive result due to early inflammation process, or false-negative result due to early inflammation masking active tumor foci. or months after ablation can be a reasonable time for pet/ct. learning objectives: . to consolidate knowledge on the imaging aspects of successful ablation. . to become familiar with the main pitfalls of post-ablation imaging. . to learn about the imaging aspects of most common complications. a- : c. follow-up imaging of thermal ablative therapies for kidney tumours d.j. breen; southampton/uk (david.breen@uhs.nhs.uk) image-guided ablation of kidney tumours has been increasingly set to become the standard of care treatment for smaller (< cm) renal tumours, yet ablation remains a non-extirpative technique and is therefore paramount, in this curative setting, that imaging follow-up should reliably confirm complete tumour eradication. early on in radiofrequency ablation (rfa), control remained problematic and inadequate treatment was evidenced by residual marginal crescents of viable, enhancing disease. in the current era of more definitive ablation with multipolar techniques, cryo-and microwave ablation (mwa), this pattern of treatment failure should rarely, if ever, be encountered. typically following adequate ablation imaging should confirm a completely nonenhancing tumour and a related cortical ablation zone. whilst nonenhancement is an accepted surrogate of tumour non-viability, most practitioners look for additional collateral features such as the 'halo' sign, appearing as a soft tissue ring in the adjacent peri-renal fat, usually a sound marker of complete tumour eradication. follow-up data have shown that cryoablation (cra) can yield robust outcomes, but often incurs notable haemorrhagic change and a 'rind-like' feature around the ablation zone. cra also appears to induce faster involution of the treated tumour, whereas rfa and mwa can induce a persistent granulomatous mass which only very slowly involutes over a number of years. late local recurrence is increasingly rare at around - %, but can occur as nodules of enhancing disease as late as - years after the initial treatment. subtraction mr can be useful, but to date pet and perfusion techniques still lack resolution and specificity. multiple percutaneous image-guided therapies are currently available for thermal ablation of bone tumours. thermal sources for these treatment modalities include high-intensity ultrasound, laser, microwave, radiofrequency, and cryotherapy. the predictability of thermal ablation is adequate to limit collateral damage and complications, however, is limited by biologic and anatomic variability of tissue. clinical evaluation is essential in symptomatic tumours. close imaging follow-up with ct, mri, bone scan, and pet plays a vital role in the management of the post-thermal ablation patient and detection of complications. recurrences or regrowth can be considered for repeated thermal ablation if the lesion is discovered early, before the tumour geometry, location, or distribution become unfavourable. the imaging features could vary with the different ablation method. the radiologist reporting the follow-up imaging should be familiar with different ablation methods. mri with dynamic contrast enhanced imaging and subtraction allows to detect recurrences in hypervascular tumour. contrast enhancement on t -weighted mri imaging seems to be predictive of clinically unsuccessful ablation. on ct scan, bone reconstruction can be visualised but is a slow process, particularly in adults. pet scan is an excellent morphologic and metabolic image to follow-up the tumours. however, the inflammation produce after ablation can be misleading, particularly during the first three months. imaging follow-up plays an essential role in the management of the post-thermal ablation patient and detection of complications. interventional radiology (ir) is an evolution of radiology that treats many diseases, originally treated by traditional surgery. the clinical importance of ir has been demonstrated over the years by performing procedures that offer the benefit of therapeutic treatments, competitive from the point of view of a shorter hospitalisation time. indeed, thanks to ir, patients are treated in a less invasive manner, hospitalisation is limited and thus promotes the containment of social costs. to maintain standards clinical oversight needs to focus upon correct organisational, functional and technological appropriateness of practice. to achieve correct ir performance, it is necessary to guarantee that all the instrumental guides (fluoroscopy, ultrasound, computed tomography equipment) function optimally, the procedural equipment including guides/catheters/stents, etc. are chosen according to their technical features and their characteristics are appropriate for use, incorporating cost considerations. as the complexity and variety of interventional procedures grow, there is an increasing need to ensure the appropriate training of specialised ir staff. consideration of dedicated education and continued professional development options for radiographers working in ir is of priority. the italian association of interventional radiographers (aitri) is very important in sharing guidelines in a multidisciplinary environment to standardise and harmonise the knowledge and skills of the team in hybrid theatre and interventional suites. interventional radiology technology is rapidly expanding and, to maintain safe and efficient practice, careful consideration of current and potential future organisational needs and the training of staff within an ir suite require detailed consideration. the lecture will discuss where new and current knowledge about radiation risk and effects are coming from. a brief overview of the different sources of knowledge, including hiroshima and nagasaki life span study, will be given. the most important stakeholders involved in research and formulating guidelines for radiation protection, with special emphasis on the new main publication from the international committee on radiological protection (icrp), and the impact on radiographic practice will be discussed. new tissue weighting factors for breast glandule gives a significant higher risk for females today, especially in the pubertal age. on the other hand, they have the risk for hereditary effects significantly decreased. estimation of risk, both in general and in the individual, will be discussed. regarding staff protection, there is a new recommendation for annual dose limit for the eye lens. the previous annual dose limit was msv, and this is now recommended to be decreased to msv averaged over five years with no single year exceeding msv. the new recommendation can be a limiting factor in interventional radiology and cardiology, if sufficient protection is not used. learning objectives: . to understand the importance of radiation protection in interventional radiology for patients and staff. . to appreciate the latest recommendations for staff radiation protection in interventional radiology. . to appreciate the important role of the interventional radiographer in radiation protection. a- : the improvement of patient care and aims for the highest possible levels of service are to the forefront of the modern "interventional suite". role development experiences will be explored, considering need, effect and management of development. service demands, the need to extend those provided, the limited number of radiologists, financial constraints and the requirement for continuous professional development at state level provide much stimulus for professional radiographic role development. increased job satisfaction, reduced waiting lists and improved patient experience all provide largely positive results it appears. resistance from radiologists, multidisciplinary team members and radiographers themselves however, can provide unnecessary barriers to change. once abridged the blurring of professional lines and insecurity within multidisciplinary team members may also occur. various experiences would suggest that only by allowing ownership of change by all the clinical stakeholders involved, thus empowering them, leads to successful development. on examination clear guidelines and distinct protocols must be established, taking account of current workload, to create the ideal scenario for good multidisciplinary team ethos, where members have overlapping yet clearly defined roles. professional accountability, responsibility and an understanding of those role's are all key, but without correct training, including support and resources, no regime for role development can be achieved successfully. clinical audit not only ensures the delivery of high quality and effective care in line with best practice, whilst ensuring cost effectiveness, but also provides the confidence for all involved, especially the interventional patient. learning objectives: . to understand the factors influencing change in professional roles within interventional radiology. . to learn about the potential impact of enhanced professional roles and a multidisciplinary team approach on service delivery and patient outcomes. . to understand the need for clear protocols and guidelines along with appropriate training and audit of practice when implementing such changes. there have been many attempts to develop gene reporters for mri, however these give relatively modest image contrast that can be difficult to detect. i will describe in this talk a reporter that gives intense and positive contrast in mr images (up to ~ x increase in signal), which can also be used with radionuclide imaging, thus combining the sensitivity of radionuclide imaging with the spatial resolution of mri. the contrast obtained is directly related to the degree of gene expression and is readily reversible, thus allowing longitudinal studies of changes in expression. the transplantation of pancreatic islet into the liver is an excellent example of successful cell therapy. transplanted islet visualisation in vivo using a noninvasive imaging method, for example mri, is necessary to prove technical success. monitoring of transplanted islets in vivo and long-term tracking their fate using mri requires their labelling by a suitable contrast agent in vitro prior to transplantation. under encite project we successfully performed animal experiments, which proved therapy potential of labelled pancreatic islets. labelled islets implanted into the rat liver were viable and induced long-term normoglycemia in diabetic rats. mri proved their viability and even distribution in the host tissue. these results allowed performing a clinical study on a group of patients, which were transplanted by iron labelled pancreatic islets from cadaver donors. the successful pilot experiment enabled introducing of transplantation of native islets into routine clinical practice. search for better alternative sites, improved immunosuppression and alternative insulinproducing cells require multimodal and multifunctional molecular probes. currently we test polymer meshes as artificial transplant sites, effect of mesenchymal stem cells as a supportive net for beta cells, and novel cellular probes combing h, f labels and fluorescent ones. optical imaging in the clinic j. dijkstra; leiden/nl (j.dijkstra@lumc.nl) recently a lot of developments have been done in the field of optical imaging. new sensitive devices made it possible to use both visible and near infrared light as diagnostic tool and guidance tool. the advantage of near infrared is that the penetration depth in tissue is much better than for visible light, allowing to see deeper. by using techniques like augmented reality, the data from infrared, both anatomical and functional, can be presented to the operator in real-time. optical imaging is used e.g. as an evaluation tools for mammography where the optical spectral properties of the tumor cells is used to monitor the effectiveness of chemo treatment. the spectral properties of the breast are measured using a laser at different wavelengths and a camera to create tomographically a d volume. dedicated near infrared probes which bind to certain tissue types make it possible to look for e.g. remaining tumor tissue in resection margins during surgery. this techniques also allow to visualise structures which should be avoided during procedures like nerves. by adding multispectral imaging, the endogenous contrast can be used. different tissues have their own absorption spectrum which can be shown as additional information, for instance the ration hb/hbo . optical imaging also allows for acquiring images at near microscopic resolution real time in-vivo by using optical coherence tomography. this modality is being tested to provide information about the presence of certain cell types very fast where otherwise histology is needed. mesenchymal stem cells constructs for image-guided cell therapy in myocardial ischemia and digestive fistulas o. clément, e. blondiaux, g. rahmi, l. pidial, a. silva, f. gazeau, c. wilhelm, g. autret; paris/fr (olivier.clement@inserm.fr) regenerative medicine has recently emerged as a potential therapeutic tool. a number of preclinical and clinical trials have been conducted in many diseases ranging from diabetes to myocardial infarction and neuro-regenerative diseases that assess the feasibility and benefits of injecting stem cells. however, outcomes relating to graft survival remain generally unsatisfactory, whether the process of injection is direct, intravenous or catheter-guided. to overcome such issues, tissue engineering has potential to improve cell engraftment and therapeutic response including functional parameters. this work aimed to evaluate mesenchymal stem cell constructs for image-guided cell therapy in myocardial ischemia and digestive fistulas. we tried to options: . fibrin patches based on fibrinogen monomers polymerised with thrombin and seeded with cells. . constructs based on d multilayers of confluent cells sheets. tissue constructs were labelled with iron oxide particles and evaluated in a model a myocardial infarction for the fibrin patches or digestive fistula for the d constructs. mri at . was performed at various time points after treatments using a high resolution coil. fibrin patches could induce a therapeutic effect by increasing the left ventricular ejection fraction compared to sham. d constructs induced an increased number of fistula healing and enhanced micro-vasculature density compared to controls. mri of labelled stem cell constructs allowed a good evaluation of the models and showed increased therapeutic efficacy. the proposed paracrine mechanisms will be discussed. is cell imaging relevant for the clinic? lessons to be learned from preclinical research u. himmelreich; leuven/be (uwe.himmelreich@med.kuleuven.be) non-invasive imaging of therapeutic cells has become a popular field of research over the last decade. this interest was mainly based on the hope that the location, migration but also function of immune, stem and other cells can be visualised over time in individuals. the development of novel contrast agents and mechanisms for mri but also other imaging methods has resulted in exciting basic research findings. in particular, the application of relatively biotolerant iron oxide based nanoparticles has fostered the hope for direct translation into clinical research and general practice. however, pre-clinical research has also highlighted several limitations of nanoparticle based cell imaging by using mri including the generation of unspecific contrast, difficulties to quantitatively image engrafted cells, unambiguous contrast, adverse effects on cell biology, limitations for longitudinal follow-up or the lack of functional information. such shortcomings are traditionally overcome in preclinical research by combining mri with other imaging modalities like bioluminescence imaging or positron emission tomography. our research focuses on the optimisation of cell labeling strategies for robust, sensitive and potentially quantitative visualisation of stem and progenitor cells in therapy models in vivo to assess cell behavior after engraftment. the sensitivity, stability, toxicity and adverse effects on the cell biology by the labeling procedure were studied for iron oxide based particles. the potential of gdchelates and f labeled compounds for cell labeling has been assessed in vitro and in vivo. based on our preclinical research finding the potential of future applications in patients will be explored. cardiac ct is becoming the imaging modality of choice for an increasing number of clinical indications, not only to rule out coronary artery disease but also to evaluate cardiac morphology and function, and to determine patient outcome after coronary artery revascularisation. however, as with any other imaging tools, appropriate interpretation of cardiac ct examinations is required to assess the clinical value of this newly established diagnostic imaging modality. this process requires performance of thorough cardiac ct acquisition protocols, detailed knowledge of standard cardiac anatomic and physiologic terminology, as well as appropriate postprocessing, reading and reporting. in particular, radiologists need to recognise and be aware of the imaging findings that may confound and lead to interpretation errors. this lecture will summarise the practical aspects of postprocessing, reading and reporting of non-invasive cardiac ct examinations. the value and limitations of every available ct postprocessing technique including two-dimensional multiplanar reformations, curved multiplanar reformats, maximum intensity projection (mip) and volume rendered images will be explained. moreover, hints for improving reading results by recognising technical causes for various artefacts in cardiac ct will be elucidated and reading approaches to diminish false positives, false negatives and inaccuracies when assessing coronary artery stenosis will be suggested. cardiac magnetic resonance (cmr) is a complex imaging technique due to the intrinsic anatomical and technical peculiarities of the exam. these include the non-orthogonal cardiac orientation within the chest cavity requiring dedicated acquisition planes and the complex respiratory and cardiac motion to which the heart is subject and requiring a combination of ecg-gated and breath-hold sequences. potential additional anatomical pitfalls also include normal structures and variants like the moderator band, papillary muscles, and the presence of prominent crista terminalis or myocardial trabeculations, whose recognition is mandatory and may mimic in some cases a pathological condition. technical issues of cmr concern the continuous intracavitary inflow of protons and the associated "slow-flow" artifacts (limiting visualisation of endomyocardial border in some cases) pitfalls related to ecg gating, like inadequate synchronisation or the t-wave swell phenomenon; and finally a series of specific artifacts intrinsically related to the use of different pulse sequences that may interfere with image quality. an additional, more complex issue to consider is also the widespread diffusion of high-field magnets which have further enhanced those aspects. knowledge of the spectrum of those cmr peculiarities is mandatory to approaching and providing a correct diagnosis according to the main clinical request. the present lecture will review the most important anatomical and technical pitfalls of cmr examination and offer, when possible, practical solutions to overcome those limitations. shoulder imaging and intervention are becoming increasingly important in clinical practice. this session considers the indications, techniques, imaging findings and relative merits of diagnostic ultrasound and mri of the shoulder. the indications, techniques and results of us-guided interventional procedures are also considered. a panel discussion will deal with controversies in shoulder imaging such as the accuracy of us and mri in assessing rotator cuff tears, tendinosis, impingement and muscle atrophy, and the efficacy of us-guided interventions. audience participation in the discussion will be welcome. the shoulder is an anatomic area that is very commonly evaluated with musculoskeletal ultrasound. ultrasonography is widely recognised as a reliable means of assessing rotator cuff disease with accuracies reaching % for fullthickness tears and - % for partial-thickness tears. diagnostic accuracy depends mostly on the skills and experience of the examiner. a comprehensive ultrasound (us) examination requires, first of all, sound knowledge of the anatomy. a specific scanning protocol must be adopted in every us examination in patients with shoulder disease, because focal symptoms do not correlate with the location of the disease. the greatest importance of ultrasonography in rotator cuff assessment lies in its dynamic character. several dynamic manoeuvres can reveal pathologies such as subacromial impingement. last, but not least there are several scanning pitfalls such as anisotropy. anatomy key structures as well as the us technique and scanning protocol will be presented in this lecture. dynamic manoeuvres will be described and demonstrated with videos. scanning pitfalls will be emphasised. advantages and disadvantages of the us examination compared to other imaging modalities will be discussed. guidelines concerning the mr examination technique of the rotator cuff will be presented in this session. on intermediate-weighted mr images, tendinopathy and partial and complete tears of the rotator can be differentiated with high diagnostic accuracy. indications for mr arthrography, especially for the detection of small articular sided partial tears in athletes, will be presented as well. established classification systems for the description of rotator cuff tears will be discussed. besides characterisation of rotator cuff lesions, especially in view of therapeutic decision-making, recognition of the underlying pathomechanism is necessary. therefore, the role of imaging is to detect different structural findings that are suggestive of a possibly underlying impingement syndrome. in primary impingement syndromes imaging abnormalities of the rotator cuff, the overlying bursa and the coracohumeral arch represent the centre of imaging findings. primary extrinsic impingement is caused by structural abnormalities of the coracoacromial arch, whereas secondary extrinsic impingement is related to glenohumeral instability. types of internal impingement (posterosuperior and anterosuperior impingement) are secondary to rotator cuff and/or capsular dysfunction. posterosuperior impingement can be diagnosed on mr arthrograms by identification of the socalled "kissing lesion" pattern, with corresponding lesions of the undersurface of the rotator cuff, posterosuperior labrum, greater tuberosity and superior glenoid. imaging abnormalities of this condition will be discussed. the shoulder is one of the joints in the human body that is most subject to a number of pathologic conditions, both in young and in elderly subjects, such as subacromial-subdeltoid bursitis, calcific tendinopathy, and degenerative conditions. being inexpensive, readily available, and radiation-free, ultrasound is the imaging modality of choice to guide interventional procedures around the shoulder. thanks to its high resolution and multiplanar capabilities, ultrasound can be used to guide needles precisely in the tendons of the rotator cuff or within the joint space, both gleno-humeral and acromio-clavicular. this approach can be used to perform a number of different procedures. when dealing with bursitis, a needle can be guided within the subacromial bursa to aspirate fluid and to inject anti-inflammatory drugs. in case of calcific tendinopathy, one or two needles can be used to dissolve calcium deposit and drain it, providing patients prompt relief. in selected patients with overuse tendinopathy, ultrasound can be used to guide intratendinous injection of platelet-rich plasma that has been reported to be helpful in stimulating tendon healing. although minimally invasive, these procedures should be performed in an ultrasound ward with a high degree of sterility, as risk of infection can be concrete. multi-detector computed tomography (ct) offers new opportunities in the imaging of the gastrointestinal tract. its ability to cover a large volume in a very short scan time, and in a single breath-hold with thin collimation and isotropic voxels, allows the imaging of the entire oesophagus, stomach, and the whole chest and abdomen with high-quality multiplanar reformation and threedimensional reconstruction. preparation of the patients by fasting from solid food approximately hours prior the examination is important. proper distention of the oesophagus and stomach by oral administration of effervescent granules and water, and optimally timed administration of intravenous contrast material are required to detect and characterise the disease. preoperative staging of oesophageal and gastric carcinoma appears to be the main indication for mdct and may replace endoluminal ultrasound (eus) in the staging of advanced cancers. the use of various reconstruction techniques, including virtual gastroscopy (vg) using a volume-rendering (vr) technique, is promising for the detection of early gastric cancer. the application of the texture analysis technique to distinguish between the different types of gastric and esophageal tumors is still evolving. finally, the introduction of fdg pet, in combination with mdct, has resulted in further optimisation of the diagnostic workup of oesophageal cancer, as well as specific types of cases of gastric cancer. by providing morphologic and functional information in the same setting, this technique has come to be the modality of choice, when available. the diagnosis of oesophageal and gastric cancer is usually based on endoscopic findings accompanied by biopsy. however, staging is a matter for diagnostic imaging and is the major determinant of disease management. this should be discussed within a multidisciplinary forum (multidisciplinary team [mdt] meeting) in which the radiologist plays a crucial part. accurate stagingusually based on the tnm staging criteria -is essential and the radiologist's report should reflect this pivotal role. the tnm staging of oesophageal cancer and gastric cancer will be discussed in detail. the phases in staging is essentially a filtering process which seeks to initially exclude distant metastasis and/or advanced local disease, initially by optimally protocolled ct scanning, if ct shows advanced disease, treatment is palliative, but even under these circumstances imaging will help determine the method of palliation. conversely, if ct demonstrates localised disease, f fdg-pet scanning for oesophageal cancer and for selected cases of gastric cancer is indicated. if this, too, shows no nodal or distant metastasis, accurate t staging with eus will help determine whether the patient proceeds directly to surgery or undergoes neo-adjuvant chemo/radiotherapy prior to surgery, or in the case of oesophageal cancer may be suitable for emr. the role of staging laparoscopy in gastric cancer will also be discussed. in summary, the radiologist and nuclear medicine physician are crucial in determining treatment. their reports are lynch pins in the mdt discussion of patient management. it is therefore essential that the imaging report should optimally inform this discussion. learning objectives: . to learn about the latest tmn staging in oesophageal and gastric cancer. . to appreciate the imaging criteria for local, nodal and metastatic disease, and understand the accuracy of imaging staging. . to become familiar with the structure of a perfect imaging report. a- : c. assessment after treatment a.m. riddell; london/uk (angela. riddell@rmh.nhs.uk) it is now established that for the majority of patients with oesophageal and advanced gastric cancer, there is survival benefit from the use of neoadjuvant therapy. therefore, there is a requirement for imaging to accurately restage the tumour and to assess the response to neoadjuvant therapy, to provide prognostic information and to direct future management. restaging following therapy is challenging, as differentiating treatment-related fibrosis/oedema from viable tumour is problematic with both ct and endoscopic ultrasound. the t and n staging accuracy for both modalities falls following neoadjuvant therapy. inconsistencies in measurements due to alterations in the degree of gastric/oesophageal distension can also limit the accuracy of recist criteria to determine the response. functional imaging techniques such as pet-ct offer an improved method for assessing response. alterations in the standardised uptake value (suv) occur much earlier than changes in size; therefore a metabolic response can be detected sooner, allowing for more rapid alterations in treatment strategies. acute complications following oesophagogastrectomy generally occur within the thorax and are either related to a leak at the anastomosis/mediastinitis or respiratory complications such as pneumonia or a pleural effusion. intra-abdominal collections may develop following oesophagogastrectomy and gastrectomy. late complications following both procedures are often due to tumour recurrence: locoregional such as lymph node recurrence or at the anastomosis; metastatic spread such as haematogenous spread or via the peritoneum or pleura. currently, there is no consensus on the most appropriate timing or frequency of postoperative imaging. a. an overview of pulmonary artery hypertension n.j. screaton; cambridge/uk (nicholas.screaton@papworth.nhs.uk) pulmonary hypertension is defined by increased mean pulmonary arterial pressure > mmhg at rest or > mmhg during exercise. ph causes significant mortality and morbidity, but commonly presents with non-specific clinical signs and symptoms resulting in significant delay in accurate diagnosis and specific treatment. untreated ph is progressive with increased pulmonary vascular resistance leading to right ventricular failure and ultimately death. the current dana point classification of pulmonary hypertension is clinically based. it groups diseases with similar pathophysiological mechanisms and therapeutic approaches. groupings include conditions characterised by diffuse small vessel narrowing (group and group '), ph secondary to left sided cardiac disease (group ), chronic hypoxic pleuro-parenchymal disease (group ), chronic thrombo-embolic pulmonary hypertension cteph (group ), and a miscellaneous group of diseases with either unclear or multi-factorial aetiologies (group ). in the dana point classification, small vessel diseases are subdivided into group which primarily affect the pulmonary arterioles and group ' affecting the capillary/venous pulmonary circulation (pulmonary capillary haemangiomatosis and pulmonary veno-occlusive disease). the differentiation of group from group ' diseases is important since in group ' arteriolar dilatation treatments can cause life-threatening pulmonary oedema. group is synonymous with cteph with other causes of large vessel obstruction (vasculitis and pulmonary artery tumour) being considered as group disorders. recent advances include an increased understanding of molecular mechanisms underpinning pah, facilitating targeted therapy development, a rapidly expanding role of surgical pulmonary endarterectomy in proximal cteph, and recognition of imaging as a potential therapeutic end point. ct allows depicting pulmonary hypertension (ph) and helps identifying its cause, therefore playing a crucial role in the diagnostic workup. ct features of pulmonary arterial hypertension include dilatation of the pulmonary artery trunk, with a diameter greater than or equal to mm, a ratio to the aortic diameter greater than : and a segmental artery-to-bronchus ratio greater than : in at least three pulmonary lobes. on ecg-gated ct, right pulmonary artery distensibility shows the best diagnostic value with % sensitivity and % specificity for a cutoff value of . %. among the various causes of secondary ph, ct is especially useful for detecting signs of chronic thromboembolic pulmonary hypertension, including wall-adherent thrombi, bands, webs or chronic arterial occlusion, mosaic lung attenuation and systemic collateral supply. ct shows signs of pulmonary edema, such as thickening of the interlobular septa, centrilobular ground glass opacities, mediastinal lymph node enlargement and pleural effusion in ph caused by pulmonary veno-occlusive disease, left heart diseases or mediastinal fibrosis. signs of lung parenchyma diseases may be indentified on ct; ph is a late complication in patients with pulmonary fibrosis, sarcoidosis or chronic obstructive lung disease, but may affect systemic sclerosis patients with limited lung parenchyma involvement. congenital cardiac abnormalities with untreated right-to-left shunting resulting in eisenmenger syndrome, such as ventricular or atrial septal defect and patent ductus arteriosus are easily recognised on ct. conversely, signs of peripheral pulmonary arteriovenous shunting in portopulmonary hypertension and ph caused by hepatopulmonary syndrome are more difficult to assess. learning objectives: . to learn about the ct diagnosis of pulmonary artery hypertension. . to become familiar with the causes of pulmonary artery hypertension on ct. a- : c. mri in pulmonary artery hypertension j. biederer; heidelberg/ de (juergen.biederer@med.uni-heidelberg.de) for the assessment of pulmonary arterial hypertension (pah), the dedicated min mri protocol would comprise a free breathing and noncontrastenhanced examination, short t -w sequences, dynamic contrast-enhanced perfusion imaging, a high-resolution angiogram, a d breath-hold acquisition, dynamic steady-state free precession or gradient echo sequences of the heart and a study of myocardial late enhancement. the morphologic sequences show typical features of pah: right atrial/ventricular dilatation, enlargement of the pulmonary trunk/main pulmonary arteries and peripherally attenuated pulmonary vessels. incidental infiltrates, nodules or masses of the lung, mediastinum and chest wall would be covered. the first pass contrastenhanced perfusion imaging demonstrates an increased mean transit time/decreased pulmonary blood flow, but a relatively homogeneous lung perfusion (important to differentiate from cteph, where multiple segmental perfusion defects would be expected). the cardiac part shows right ventricular mass, wall thickness and functional changes correlating with elevation of pulmonary arterial pressure: distortion of the interventricular septum, area change of the pulmonary trunk, right ventricular volume/stroke volume as well as pathologic right/left ventricular end-diastolic volume indexes. late enhancement of the right ventricular wall would correlate with myocardial fibrosis. furthermore, optional experimental velocity-encoded sequences (ideally for multidirectional flow visualisation, " d flow") show a decreased pulmonary artery blood flow velocity, increased retrograde flow and inhomogeneous velocity profiles. in conclusion for the near future, given the availability of scanner time and appropriate experience of the team, thoracic mri is probably the most comprehensive and effective single examination for the diagnosis and follow-up of pah. in the era of organ-based radiology, the group of diseases known as multisystemic malignancies represents an obvious challenge to both radiologists and oncologists. the necessity to match the growing possibilities of different imaging modalities with widespread multisystemic pathology and clinical sufficiency resulted in upgrading well-known diagnostic algorithms. the precise knowledge of clinical staging systems and classification, and the pathologic and physiologic mechanisms of the disease pathways are important for planning imaging modalities and specific protocols. perfect imaging of multisystemic malignancies now includes not only traditional anatomic-based modalities, but also much more often different types of whole body scanning such as ct, mri, pet and their combination in spect/ct, pet/ct and now even pet/mri. new imaging modalities and growing possibilities of traditional imaging techniques obviously influence the current clinical guidelines for this disclosure: c. heussel: consultant boehringer ingelheim, grifols, novartis. a common language between radiologists and clinicians, so that the latter can make an informed treatment decision based on sound conclusions to become familiar with the conditions necessary to implement them to understand the limits of their application to learn useful lessons from these criteria for routine clinical practice chairman's introduction a. palko this requires a thorough knowledge of the relevant brain anatomy, choice of appropriate structural and molecular imaging modalities and interpretation of mri and pet/spect in the most prevalent disorders in a structured fashion the presentation has the objective of giving the audience a synthesised panorama of our country situated in north america, with more than million inhabitants and a large prehispanic history based on the aztec, toltec and maya cultures, and also of interesting facts related to the three centuries of colonial existence under the spain influence and dominium until the war of independence. after , mexico has never had an international war; the last was against the northern border country, the united states. also, we will give important data on how in the mexicans rebelled against a -year dictator, general porfirio diaz. the mexican revolution was a long and cruel war, but later opened the doors to democracy and a complex developmental era began that is still in process. some demographic and contributions to health, science and culture issues will be mentioned and also the works of mexican scientists, writers and philosophers who have been presented awards including the nobel prize. in relation to well-known mexican artists such as diego rivera and frida khalo, some facts of their works will be presented. interventional radiology in oncologic patients g. elizondo-riojas; monterrey/ mx (elizondoguillermo@hotmail.com) interventional radiology (ir) is becoming an increasingly prominent subspecialty in the care of oncologic patients. its role extends from initial diagnosis to minimally invasive treatment of the malignancy and its complications. image-guided biopsies are increasingly performed using minimally invasive techniques. also, an integral part of care of these patients is vascular access as a means of medication, chemotherapy or parenteral nutrition, and interventional radiologists can place required the devices with well-established safety and efficacy. ir also plays a substantial role in the therapy of oncologic patients, through local tumor treatments such as transarterial chemo-embolisation and locoregional control with radiofrequency/cryo ablation, as well as management of complications of malignancy such as pain, obstruction (biliary, ureteral, etc) ., venous thrombosis and drainage of thoracic and abdominal collections. in mexico, ir is a growing subspecialty, and more medical students nowadays want to be radiologists and eventually become interventional radiologists. this is a paradigm shift. more and more radiologist wants to be involved with patient management and to be more than just "observers" in the process of patient care. we have to be prepared to offer this opportunity to our residents; otherwise other specialties will "have to fill the empty space" that we have left. interventional oncology has all the advantages to fulfil this opportunity. it is our chance to contribute to the advancement of medical care. learning objectives: . to appreciate the role of interventional radiology in the management of oncologic patients. . to learn how interventional radiology changes the quality of life for patients with cancer. . to understand the impact of interventional radiology procedures in the outcome of some neoplastic diseases. interlude: origins and development of radiology in mexico m.e. stoopen-rometti; mexico/mx (mstoopen@clinicalomas.com.mx) mexican radiology started in , just a few months after the discovery of xrays. during the last few decades, as well as in other countries, there has been a great development both in public and private sectors, which will be described in this interlude. learning objectives: . to learn about the history of radiology in mexico. . to learn about the development of radiology in mexico. . to learn about the present and future of radiology in mexico. modern issues in oncologic ultrasound j. mexico/mx (jtanus @hotmail.com) advances in ultrasound (us) technology allow confident characterisation of masses. these include harmonic imaging, compound imaging, power doppler, faster frame rates, higher resolution transducers, three-dimensional ( d) us, us contrast agents and, more recently, elastography and fusion imaging. highfrequency transducers provide superb spatial and soft-tissue resolution, permitting substantially improved differentiation of subtle lesion, margin resolution, and lesion conspicuity in the background of normal tissue. elastography features such as size ratios, shape, homogeneity, and maximum lesion stiffness complement conventional us in the analysis of lesions. ultrasound contrast agents have overcome some of the limitations of doppler ultrasound techniques with demonstration of irregular branching central or penetrating vascularity within a solid mass raising suspicion of malignant neovascularity (neoangiogenesis). ultrasound contrast agents can provide important information in the assessment of lesions to be treated by locoregional therapies, which include ablation (feeding vessels), trans-arterial chemo/radio-embolisation, detecting viable tumour persistence following this treatment; facilitation of needle positioning in cases of poor lesion delineation, and assessment of local tumour progression. it facilitates needle positioning in cases of incomplete or poor lesion delineation on unenhanced ultrasound. us contrast agents are an important key during the evaluation of the immediate treatment effect of ablation and guidance for immediate re-treatment of residual tumour. learning objectives: . to understand the role of ultrasound in the management of oncology patients. . to learn how ultrasound is used in large oncology centers. . to learn about the modern concepts of ultrasound in oncology patients. the aim of this lecture is to give an overview of the techniques for imaging inflammatory bowel disease (ibd) of the small bowel and the colon with either ultrasound (us), multidetector row computed tomography (mdct) or with magnetic resonance imaging (mri) and compare the different modalities for its strength and weakness. optimal imaging of the bowel begins with the preparation phase. the small bowel has to be distended for a concise examination. this is mainly done orally, which is named enterography. a solution of . % mannitol seems to be the one preparation technique mostly used for small bowel distension. another technique is the enteroclysis, application of contrast after intubation of the small bowel. the comparative advatages and disatvantages of the two preparation methods will be discussed. this intraluminal contrast gives a neutral contrast in ct and a biphasic signal in mr. the colon can be prepared in a fashion similar to colonoscopy meaning total cleansing. another possibility is the so called fecal tagging whereas the stool will be contrasted with an additive to standardised food. therefore no cleansing is needed for preparation. imaging parameters will be discussed for mr and ct. the aim of imaging for the bowel should be to establish the following: ) presence, severity, and extent of disease; ) activity of the disease and ) extra-intestinal complications. us, mr and mdct have proven to be a good tool to evaluate the extent, the activity of the disease and the presence of extraluminal complications. pros and cons will be discussed when to use which technique. learning objectives: . to understand state-of-the-art mri, ct and us protocols for imaging ibd. . to appreciate the comparative advantages and disadvantages of enterography and enteroclysis protocols. . to learn about protocol modifications when evaluating the colon.a- : b. small bowel disease j. stoker; amsterdam/nl (j.stoker@amc.uva.nl) classification of small bowel crohn's disease is helpful for assessing disease activity and treatment monitoring. similar to clinically based classification, also imaging-based classification systems have been developed, of which some have been externally validated. important imaging features for determining disease activity include bowel wall thickness and vascularity/enhancement; at mri, also wall oedema plays a role. stenoses, fistulas and abscesses are important sequels. for assessment and monitoring of small bowel disease the breast imaging reporting and data system (bi-rads®) for mammography of the american college of radiology (acr®) consists of several components, a standardised lexicon of terms to be used during reporting, a -step coding system for the mammographic density as a surrogate parameter for the mammographic sensitivity, and a group of assessment categories ranging from to for structured communication regarding the recommended further management. the goal of bi-rads® is to improve the quality of breast imaging reporting and communication. in addition, by providing structured reports it facilitates regular quality assurance measures. the bi-rads® atlas for mammography is currently in its fourth edition and was released in . the upcoming th edition is expected soon and will be incorporated into the course as it becomes available. the breast imaging reporting and data system (bi-rads) was first developed by the american college of radiology for standardising the reporting of mammography. since its first publication in , new editions have also addressed breast ultrasound (us) and mr imaging examination. the new (fifth) edition will be published soon. on the form, this new version was designed to include a web-based format. substance includes updates in lexicon descriptors, e.g. masses, calcifications, associated features (now comprising surrounding tissue with stiffness assessment), and special cases. us descriptors will be reviewed in the lecture. of note, guidance on how to link bi-rads descriptors with management recommendations has been added in the report section. a new approach to outcome assessment (audit section) is being proposed for screening us. the appropriate use of descriptors is expected to increase the accuracy of imaging interpretation. for relevant patient management, us analysis is to be integrated with other available imaging, as well as with clinical data. learning objectives: . to learn about the bi-rads lexicon. . to understand the usefulness of bi-rads system. a- : c. mri k. pinker-domenig, p.a.t. baltzer; vienna/at (katja.pinker@meduniwien.ac.at) dynamic contrast-enhanced magnetic resonance imaging (dce-mri) of the breast is a well-established non-invasive imaging technique. it has clinical application in the screening of high-risk patients, diagnosis and staging of breast cancer, monitoring neoadjuvant chemotherapy and post-treatment follow-up. to standardise the reporting of dce-mri of the breast and minimise false-positive results without compromising sensitivity, the american college of radiology (acr) introduced the breast imaging-reporting and data system (bi-rads®) mri lexicon in . bi-rads relies on the combined analysis of morphological appearance and lesion enhancement kinetics. it is widely used for reporting dce-mri of the breast and is applicable at any given field strength. it aims to provide an up-date report on the bi-rads® lexicon and instill confidence in using bi-rads® descriptors. the bi-rads® system may be placed in a broader clinical context to highlight its value for standardised reporting of dce-mri of the breast. mr-guided focused ultrasound is a new therapeutic modality which can allow selective destruction and or heating of tissues in deep body areas under close image guidance control. these talks will introduce the field and allow you to understand the areas of current work and areas of future development in this rapidly expanding field. session objectives: . to become familiar with mr-guided focused ultrasound. . to understand the advantages of focused ultrasound. . to learn in which areas focused ultrasound is evolving successfully. description of technique c. moonen; utrecht/nl (c.moonen@umcutrecht.nl)high-intensity focused ultrasound (hifu) is the only clinically viable technology that can be used to achieve a local temperature increase deep inside the human body in a non-invasive way. mri guidance of the procedure allows in situ target definition and identification of nearby healthy tissue to be spared. in addition, mri can be used to provide continuous temperature mapping during hifu for spatial and temporal control of the heating procedure and prediction of the final lesion based on the received thermal dose. the primary purpose of the development of mr-guided hifu was to achieve safe non-invasive tissue ablation. the technique has been tested extensively and is now accepted in the clinic for ablation of uterine fibroids. mr-guided hifu for ablation shows conceptual similarities with radiation therapy. however, thermal damage generally shows threshold-like behaviour with necrosis above the critical thermal dose and full recovery below. mr-guided hifu is being clinically evaluated in the cancer field. this presentation will cover the basic technologies for treatment of stationary tissues, and some advances towards treatment of mobile abdominal organs. for state-of-the-art mr liver and pancreatic imaging, a field strength of at least . t is required. all non-blood pool gadolinium chelate-based contrast agents are suitable for dynamic liver and pancreatic mri. all gadolinium chelates should be routinely administered at a rate of - ml/s followed by a ml saline flush at - ml/s using a power injector. to obtain hepatobiliary phase imaging in addition to dynamic phase imaging, the use of liver-specific contrast agents is required. gd-eob provides the highest hepatocyte enhancement, but an overlap between delayed phase and hepatocyte phase has to be considered during dynamic evaluation. the hepatocyte phase can be considered adequate when contrast is detected in the intrahepatic bile ducts. hepatobiliary phase imaging benefits from a gradient echo high flip angle, depending on magnet field strength. in the absence of liver function impairment and biliary obstruction, contrast-enhanced mr cholangiography can be obtained with gd-eob at - minutes, and with gd-bopta at - minutes. when the differential diagnosis is primarily between solid benign lesion vs. metastasis, the use of a liver-specific ca is recommended, due to the ability to diagnose fnh confidently. the combined interpretation of dynamic and hepatobiliary phase improves diagnostic accuracy of mr imaging for the detection of hcc.hemangiomas and intrahepatic ccc result in relative hypointensity in the late vascular phase after gd-eob administration. when combined with t weighted mrcp, contrast-enhanced mrc allows morphologic and functional assessment of the biliary system. gastrointestinal stromal tumours (gist) are rare malignant tumours arising within the wall of the gut from the interstitial cells of cajal, which act as pacemaker cells controlling peristalsis. the crude annual incidence of clinically detected gists is approximately cases per million in europe. the median age at diagnosis is approximately years, and % of patients are older than years. however, a small number of cases do occur in younger people and children, and these are usually syndromic gists. gists can occur at any site of the gut from the oesophagus to rectum, although they can also arise in the extra-gastrointestinal abdominal and pelvic locations, so-called e-gists. the commonest location is the stomach ( %), followed by the small intestine ( %) and rectum ( %). diagnosis is by biopsy, with spindle cell or epithelioid morphology, and immunohistochemical staining for cd (the protein product of the kit gene) and/or dog- receptors. approximately, % of gists have mutations in the kit gene, % have mutations of the pdgfra gene, and the remaining % have no mutation (wild-type) or rare gene mutations. early stage disease is managed by surgery, followed by years of adjuvant imatinib (a tyrosine kinase inhibitor with activity against kit and pdgfra receptors) for cases at high risk of relapse. advanced metastatic disease is treated with imatinib, with a median duration of response of approximately years. at disease progression, second-line treatment is with sunitinib, with a median duration of response of approximately months. imaging plays a major role in the detection, characterisation and staging of gastrointestinal tumours (gists). imaging of gists depends on the mode of presentation and the local availability. endoscopic ultrasound and computed tomography (ct) are the most widely used imaging methods. gists have a spectrum of radiological appearances depending on tumour size and site of origin, but often show certain key radiological features. multiphase multidetector row ct is necessary to detect and stage gists and multiplanar imaging is important so as to define the likely organ of origin of the mass, which may be a challenging task. the clinical diagnosis of a gist is based mainly on imaging, as biopsy runs the risk of tumour rupture or seeding of the biopsy tract. in patients with an unresectable primary tumour or metastatic disease at presentation, biopsy confirmation is important before starting medical treatment. ct or/and mri are used to assess primary tumour extension, local invasion and the presence of metastases, with main sites of metastasis being the liver and the omentum. gastrointestinal stromal tumours (gists) are treated with targeted therapy regimes. these treatment strategies are based on the suppression of tumour vasculature using the tyrosine kinase inhibition pathway. drugs like imatinib and sunitinib exhibit specific changes in tumours and metastases that can be detected on cross-sectional imaging. these changes differ significantly from standard treatment effects of cytotoxic chemotherapy. while the latter leads to destruction of tumour cells and thereby to a reduction of tumour size, the former will cause a decrease in vascularity and thereby in attenuation and enhancement of lesions while the size may remain unchanged. radiologists have to be aware of these specific patterns of response to treatment. this presentation will include a review of the choi criteria which have been developed for the assessment of gist lesions under treatment. also, current imaging strategies focusing on time-resolved ct imaging (ct perfusion imaging) will be presented. the aim of this lecture is to provide a practical approach to response imaging in gist patients integrating both existing and novel strategies. radiological response evaluation using anatomical imaging was developed in the s in early phase ii clinical trials. potential new anti-cancer agents were assessed by measuring tumour size before and during therapy and a predetermined reduction in size constituted response. radiological response represents an objective surrogate for patient benefit and was the primary end point in early phase ii studies. if a pre-defined percentage of patients achieved the required response, then the agent proceeded to large phase ii and iii studies where clinical time to progression or progression-free survival comprised the primary end point. between and , there were extraordinary advances in cross-sectional imaging techniques which became widely available. different imaging techniques were introduced piecemeal with different co-operative groups adding different requirements, so meaningful comparisons from one trial to another could not be made. in , the recist criteria were introduced requiring specific imaging stipulations and a minimum baseline tumour size to unify the different criteria and enable meaningful comparisons from one study to another. further advances continue to be made regarding anatomical and functional imaging. not all anti-cancer agents are cytocidal and many studies now use time to progression or progression-free survival defined by radiological imaging as the primary end point. certain tumour types respond in a unique manner requiring the employment of specific response criteria in these tumour types to adequately demonstrate response. some of these tumour types and their specific response criteria will be discussed. learning objectives: . to learn about the role of radiological response evaluation in clinical trials. . to understand how radiological response evaluation has evolved and continues to evolve. . to understand which response criteria are utilised in specific tumour types. emerging biomarkers for response assessment: pros and cons m.c. roethke; heidelberg/ de (m.roethke@dkfz.de) non-invasive response assessment of new specific anti-cancer therapies is an emerging field of oncologic imaging. in the past few years, large efforts were undertaken to develop new functional biomarkers to enable an earlier diagnosis, improved risk stratification and treatment monitoring of oncological diseases. imaging biomarkers reflect changes in tumour biology, which can be differentiated into certain categories (e.g. cell density, tumour heterogeneity, iron concentration, elastic properties, or cellular receptors). in a first step, alternative qualitative and quantitative imaging biomarkers for the different modalities (mri, ct, pet) are elucidated. for magnetic resonance imaging, new techniques with potential for treatment response monitoring such as diffusion-weighted imaging, susceptibility weighted imaging, and elastography will be assessed. then, quantification of iodine uptake of contrast-enhanced ct as an imaging biomarker will be addressed. novel pet imaging strategies for therapy monitoring will be mentioned with focus on receptor targeting tracers (e.g. psma, dotatoc). furthermore, the role of pet-ct/mri is mentioned to facilitate functional techniques in oncological imaging with imaging biomarkers. the potential clinical use of prior introduced biomarkers is demonstrated for several cancer entities (e.g. hcc, prostate cancer, malign melanoma, multiple myeloma, glioblastoma). advantages and disadvantages of the illustrated imaging biomarkers are discussed during this part of the lecture. the goal is to image the right patient, at the right time, using the right test and with the right interpretation so that patients will be advised on the right therapy. ultimately, the aim is to maximise patient outcomes at an affordable cost. this session will cover the state of the art in ebr, cer, and hta and demonstrate how the results are used in imaging decisions. the application of evidence-based medicine to medical imaging was relatively delayed in comparison to other specialties. radiologists should be aware of the necessity to justify radiological examinations and imaging-based interventional procedures on the basis of the best available evidence. diagnostic tests can be evaluated with studies exploring their value in terms of: . technical performance; . diagnostic performance; . diagnostic impact; . therapeutic impact; . patient outcome; . societal impact. notably, this is a one-way logic chain. while improvement at the upper levels implies improvement at some of the lower levels, the vice versa is not always true (e.g., a better diagnostic performance does not always imply a therapeutic impact or a better patient outcome). moreover, different degrees of recommendations are based on different levels of evidence, with experts' opinion as the lowest level of evidence and meta-analyses of high-quality homogeneous studies and multicentre studies being the best level of evidence. the quality of studies needs to be evaluated in terms of internal and external validity, the former regarding study implementation (protocol violations, reference standard, measurements, and readers' independence), the latter regarding study planning (study design, subjects selection, methods, and statistical analysis). biases influencing the internal validity limit the intrinsic value of study results, while those influencing external validity limit the generalisability of study results to clinical practice. finally, high-quality research must be planned to build the evidence in favour of radiological procedures, especially for new technologies which tend to enter the market without any preliminary demonstration of efficacy. state-of-the-art methods in ebr, cer, and hta relevant to imaging u. siebert; hall i. tirol/at more and more, clinical guideline developers and reimbursement decision makers need to base their work and decisions on solid evidence. this presentation will introduce the key concepts and principles of evidence-based medicine (ebm), comparative effectiveness research (cer), and health technology assessment (hta) and their application to the field of radiology. in particular, it will be discussed which role observational studies, clinical trials, and decision-analytic modeling play in ebr, cer, and hta and when each of these study types is needed to assess imaging technologies. specific topics include the assessment of bias, optimising cutoffs and multiple test strategies, making causal inferences, explicitly weighing benefits, risks and costs, and considering ethical, legal and social implications (elsi). the excitement generated by the discovery of x-rays led to the early adoption of this technology in many hospitals around europe. over the last three decades, the explosion of new medical imaging technologies together with the recognition by clinicians of the value of these for their patients has meant that imaging budgets have increased exponentially. evidence based radiology (ebr) is a relatively new approach designed to inform clinicians of the most appropriate technique to use in a given clinical scenario. the comparative effectiveness research (cer) is used by healthcare systems to develop a strategic approach to rationalise the availability of imaging investigations. the health technology assessment (hta) funding approach is a mechanism to assess new and emerging imaging technologies in a systematic timely manner. the uk has a nationally funded healthcare system which is designed to deliver equitable care for the population free at the point of delivery. the challenge for the uk has been to ensure the highest quality service by delivering the most appropriate technology and care for patients in a timely manner. the national institute for clinical excellence (nice) is an independent body which gives guidance on new drugs and medical devices. this body requires robust scientific evidence on which to base their decisions. to create the evidence base, the imaging studies need to be of a certain standard to be included (consort, stard, etc). an example from oncologic imaging will be used to show how ebr influences daily practice. acute abdomen and abdominal trauma are both emergencies with potential fatal sequela when under/ misdiagnosed. imaging plays a crucial role in the diagnosis and management. us is the imaging technique of choice in most cases, as it is rapid, portable, lacks radiation and there is no need for sedation in young infants. us accuracy can be enhanced by iv contrast agents administration. however, limitations may occur in obese children, in deep structures, or because of gas filled bowel loops. on the other hand, ct provides excellent visualisation without limitations by obesity, gas, or deep lesions. however, there is always the radiation exposure risk of the radiosensitive paediatric patient, while sedation is required in non co-operative young children. the aim of this session is to understand "which" is the modality of choice and "why", for the most common paediatric abdominal emergencies. the acute abdomen: ct is the answer a. paterson; belfast/uk (anniezunz@gmail.com)abdominal pain is a common symptom in children, and whilst the majority will have a self-limiting condition, in some the pain may indicate an acute medical or surgical condition that requires prompt investigation and treatment. in the paediatric setting, the primary imaging modality for those with an acute abdomen is ultrasound. however, there are certain patient groups -often older children or adolescents -in whom ct plays an important role. it is well acknowledged that ct is non-operator dependent: an important factor for children presenting to emergency departments outside children's hospitals. the speed of a ct exam is valuable in the acute setting, as is the capability to obtain images without having to touch the tender abdomen of a distressed child. ct offers a global perspective of the abdomen, and image quality is not hindered by the presence of excessive bowel gas, an abnormal body habitus only porotic fragility fractures, but also of primary and secondary spine tumours such as traumatic fractures. a wide variety of lesions in and around the orbita can impair eye movement. ct and mr imaging is frequently used to confirm or exclude lesions in and around the orbit in patients with impaired eye movement. first of all, it is very important to know the exact clinical history of the patients. ct is excellent for confirming a mass; however mri is more sensitive and arrives often at a single most likely diagnosis. characteristic imaging features may help distinguish among lesions that have overlapping clinical presentations. this review focuses on some of the common orbital masses. in this lecture common benign and malignant lesions will be discussed. vascular lesions include capillary (infantile) haemangioma, cavernous haemangioma, and lymphangioma. benign tumours include optic nerve sheath meningioma, schwannoma, and neurofibroma. malignancies that are reviewed include: lymphoma, metastasis, rhabdomyosarcoma, and optic glioma. in addition, benign and malignant lesions affecting the eye movement from outside of the orbit (e.g. from the paranasal sinuses, nasopharynx and skull base) will be discussed. trigeminal neuralgia is a unique form of facial pain, defined by the international association for the study of pain as a sudden, unilateral, brief, stabbing, recurrent pain in the distribution of one or more trigeminal nerve branches, triggered by a specific event. additional diagnostic criteria of the international headache society include paroxysmal, stereotypic pain attacks and absence of associated neurological deficits. it is due to trigeminal nerve compression at the root entry zone, a transitional zone between central and peripheral myelination, where the myelin sheath is thinner and more prone to compression and electrical stimulation. the most common cause is vascular compression, by aberrant loops of the sca, aica, pica or vertebro-basilar dolichoectasia, leading to a neurovascular conflict. small size of the prepontine and cpa cisterns has been identified as a predisposing factor. occasionally, expansile lesions of the skull base or cns compress the root entry zone and lead to atypical trigeminal neuralgia. mr imaging is the modality of choice to evaluate these patients and requires specific tailored protocols to depict its causes, including d heavily t w images reconstructed in the three planes and an angiographic sequence for the depiction of neurovascular conflicts. as neurovascular contacts are quite common in the general population, strict imaging criteria for the diagnosis of nvc have emerged. the offending vessel needs to cross the nerve perpendicularly and deviate or indent its course at the rez. a pictorial review of the most common causes of trigeminal neuralgia and trigeminal neuropathy will be presented. learning objectives: . to understand the causes of trigeminal neuralgia and its clinical symptoms. . to learn about the relevant anatomy and appropriate imaging methods. . to become familiar with the relevant radiological images, differential diagnosis and limitations of the method. c. facing problems of the face: facial pain, tics and palsy b. verbist; leiden/nl (b.m.verbist@lumc.nl) facial impairments may be due to neuropathies of the trigeminal and facial nerve. this lecture will focus on the seventh cranial nerve. this nerve is a complex, mixed nerve with motor fibres, parasympathetic fibres, sensory fibres to the external auditory canal and special sensory (taste) fibres to the tongue. the most common presentation of facial nerve dysfunction is facial palsy. the paresis or paralysis of facial muscles may be caused by several conditions such as developmental abnormalities, infectious or inflammatory disease or tumoural lesions. other presentations of facial nerve dysfunction are facial tics or hemifacial spasm and pain. the indications for imaging, choice of imaging modality and possible imaging findings in case of facial palsy, facial tics and facial nerve-related pain will be discussed. in this presentation, the procedures used to create the standard will be reviewed and some of the most recent changes discussed. as a collaboration of professional societies and companies, integrating the healthcare enterprise (ihe, www.ihe.net) seeks to establish methods wherein health computer systems can communicate to achieve specific functional objectives. these involve development of methodologies to implement standards such as dicom, hl , etc. in this presentation, the procedures used to create the ihe profiles will be reviewed. patient dose tracking is rapidly growing in the usa as a large number of commercial products are available in response to user demand. these products leverage the work that has gone into the development of the dicom dose structured report and the proliferation of digital imaging systems. medical physicists are crucial team members as they are most capable of performing patient dose measurements and calculations. noteworthy is that the 'toolbox' of the medical physicist is expanding. informatics with patient dose tracking tools invariably includes familiarity at some detail with dicom structure and tags, dose structured reports (sr), sr readers, ris content and orders, modality work list broker content, modality (imaging) specific content as well as pacs, archive rules for storage/retrieval and emr. patient-specific information and the highly detailed specifics of the individual dose prescription are necessary for advanced estimates of information. a dose tracking system (a qa tool outside of the normal imaging device that uses content provided by dose sr and other information systems) usually strives to elevate the quality of the dose estimate, using informatics tools such as patient and organ models, dosimetry engines, and predictive statistics. general use cases for dose tracking will be presented including advanced dose estimations for individual patient exams and qa review for both ct and fluoroscopy. examples of tracking of prescriptions by patient, protocol/procedure and operator for determination of patient dose history, 'outliers', continuous improvement (using dmaic tools) or for meeting regulatory or accrediting bodies will be included. with the introduction of digital radiology, it is possible to have automatic systems to collect and archive patient dose data individually, in addition to demographic, geometric, and other procedural parameters, as part of the dicom header or through other dicom services. these automatic systems mean significant benefits for patient dosimetry and quality control. different approaches were used, depending on the availability and level of implementation of the dicom standard, including extracting the technical information from the headers, using the radiation dose structured reports (which contain accumulated dose over several irradiation events), analysing the mpps messages sent by the modalities to the radiology information system, and implementing optical character recognition techniques on saved screen images. before issuing a formal patient dose report, the medical physicists should verify and correct all patient dose data. all these approaches allow managing more information and provide better capacity to audit the full imaging procedure and to help with the optimisation. the current level of technology allows doing so at a reasonable cost and with a great benefit for the clinical practice. automatic detection of abnormal patient doses or mistakes in the technical parameters used and their prompt correction is possible. diagnostic reference levels will be effortlessly reviewed with such systems. some examples of pitfalls and possible optimisation actions will be presented. pathology. finally, the perfect assessment of clinical course of the disease and possible outcomes, the understanding of tumor response criteria and therapyinduced changes are significant for image interpretation in patients with multisystemic malignancies. multiple myeloma is a haematologic disorder characterised by the infiltration and proliferation of monoclonal plasma cells mainly in the bone marrow. the main symptoms are hypercalcaemia, renal impairment, cytopaenia/anaemia and bone disease -summarised as crab-criteria. symptomatic multiple myeloma is consistently preceded by asymptomatic premalignant stages called monoclonal gammopathy of undetermined significance and smoldering multiple myeloma. these stages also present with detectable monoclonal protein and/or monoclonal plasma cells in bone marrow, but do not show any end organ impairment. staging of multiple myeloma is based on the measurement of the monoclonal protein in serum and urine as well as the assessment of impairment of haematopoiesis, renal function and mineralised bone. in the last decade, the development of novel therapeutic agents has led to an increase in response rates and survival time of patients with multiple myeloma, which further stresses the value of response assessment by imaging. cross-sectional imaging like mri, ct and pet-ct is currently replacing conventional radiological surveys in the initial workup and follow-up of patients with monoclonal plasma cell diseases. recent studies demonstrate the added value of mri in improving initial staging by unravelling a diffuse infiltration of bone marrow by plasma cells, a focal pattern or a combination of both. also, pet-ct can detect response to therapy earlier than conventional response criteria. furthermore, recent studies revealed that a complete remission of myeloma confirmed by mri or pet-ct goes along with a better prognosis compared to a complete response based only on serological parameters. learning objectives: . to learn about the role of clinical staging systems and imaging in work-up and classification of plasma cell disorders with a focus on multiple myeloma and to learn about parameters of disease activity. . to understand the pathophysiologic mechanisms of multiple myeloma and to learn about the effects of abnormal plasma cells on bone remodeling cells. . to become familiar with response criteria and therapy-induced changes and to compare the significance of imaging and serological markers for response evaluation in multiple myeloma. . to learn about recent studies on imaging based response assessment and prognostic significance.a- : over the last years, there have been numerous efforts to classify lymphoid malignancies, culminating in the who classification introduced in . so it is clear that malignant lymphomas require a sophisticated diagnostic approach based on clinical features, morphology, immunophenotyping and genetic analysis. it is essential that such an approach underpins the clinical management of these diseases, many of which are amenable to cure. the same situation is present in radiology staging procedures -it has a multidisciplinary approach. a wide range of staging procedures can include all radiology methods from cxr and chest and abdominopelvic computed tomograph to whole body mri, scintigraphy and pet-ct scans. lesions of the regional lymph node system. distant spread of mm is thought to transpire intra-and extravascularly (along the external vessel lattices) supported by specific mechanisms of extravasation and protection of mm cells against the local immune system. to date, many imaging methods, such as ultrasound, computed tomography (ct), magnetic resonance imaging, and positron emission tomography/ct, are used in the diagnosis of mm and its metastases, whereas lymphoscintigraphy has become an important tool for workup of the lymph node drainage patterns. radiological challenges include improved staging and re-staging of multi-systemic mm. accordingly, current imaging guidelines and the role and applications of the different imaging modalities in advanced mm (stage iii / iv) are reviewed with regard to their usefulness in patient management. however, standardised imaging protocols and integration of functional information to morphological imaging are needed in the future to allow for improved detection and guidance of treatment in patients with mm. the original atlanta classification of acute pancreatitis established in is outdated. in , an international working group has modified the atlanta classification for acute pancreatitis to update the terminology and provide new morphologic classifications.the revised classification of acute pancreatitis identified two phases of the disease: early (first week) and late (after the first week). acute pancreatitis is subdivided into two types: interstitial oedematous pancreatitis and necrotising pancreatitis. if the diagnosis of acute pancreatitis is established by abdominal pain and by increases in the serum pancreatic enzyme activities, a contrast-enhanced ct is not usually required for diagnosis in the early phase. this revised classification introduces new terminology for pancreatic fluid collections. depending on the presence or absence of necrosis, acute collections in the first weeks are called acute necrotic collections or acute peripancreatic fluid collections. once an enhancing capsule develops, persistent acute peripancreatic fluid collections are referred to as pseudocysts and acute necrotic collections, as walled-off necroses. all can be sterile or infected. this classification of acute pancreatitis allows a consistent, worldwide classification and should avoid confusion in the terminology of pancreatic fluid collections. the role of diagnostic imaging in chronic and inflammatory pancreatitis and ipmns is to detect structural changes of the ducts and of pancreatic parenchyma, assess the functional integrity of the gland, detect associated complications, and assist in management. these goals are generally fully achieved using mdct and mr imaging. in this lecture, the advantages and limitations of each technique will be discussed and illustrated. key features allowing differential diagnosis of both entities will be underlined. autoimmune pancreatitis (aip) was first described in and represents a rare form of immune mediated chronic pancreatitis which is characterised by a marked infiltration of lymphocytes and plasma cells into pancreatic tissue. whilst the majority of cases present with diffuse gland involvement, approximately % of patients demonstrate either segmental or focal involvement of the pancreas. clinical presentation is very variable with patients describing a range of symptoms; severe pain however is uncommon. imaging plays a central role in the diagnosis and management of aip and knowledge of the radiological appearances, which can vary significantly due to the various degrees of fibrosis and inflammatory infiltrate, is critical. cardinal features include focal or diffuse pancreatic enlargement with the loss of normal lobular architecture. in addition, pancreatic duct involvement as demonstrated by single or multiple focal strictures with limited more proximal dilatation is common. whilst these appearances may suggest a diagnosis of aip correlation with clinical history, serology and histopathology is mandatory to accurately diagnose atypical cases. in contrast, paraduodenal pancreatitis is a specific and rare form of segmental chronic inflammation characterised by fibrous tissue formation in the "groove" area between the duodenum, head of pancreas and lower common bile duct. the pathology was first described in and has since been subdivided into pure, segmental and non-segmental forms. whilst the radiological diagnosis of both aip and paraduodenal pancreatitis remains challenging, the presentation will emphasise relevant imaging strategies as well as provide tips and tricks for accurate diagnosis. bone scintigraphy is often used in combination with ct and clinical information for staging and therapy response evaluation of bone metastases. magnetic resonance (mr) imaging is a technique that is known to be valuable both for detection of bone metastases and for evaluation of their response to treatment. integrated positron emission tomography with computed tomography (pet/ct) combines the functional data from pet with anatomic information of ct in a single examination. f-fluoride is a non-specific pet tracer that has recently shown very high sensitivity for bone metastases. for accurate response evaluation, it is important to recognise not only changes in bone metastases, but also the effects of treatment on unaffected, normal parts of the bone marrow. these latter therapy-related benign changes could otherwise be misinterpreted as disseminated disease or vice versa. despite the fact that the methods mentioned above, alone or in combination, have improved the assessment of bone metastases, they often remain non-specific. a specific diagnosis can in many cases only be achieved with a bone biopsy. this refresher course will give you an overview of the present imaging methods for detection and characterisation of bone lesions. indications for, and how to perform, ablation and cementoplasty of bone metastases will be presented. optimal algorithms for treatment follow-up of bone metastases and advantages and disadvantages with different imaging methods will be reviewed. also, future development of diagnostic and therapeutic methods will be included in the presentations, e.g. potential role of emerging pet imaging probes for assessing bone malignancies. author disclosure: h. ahlström: research/grant support; astrazeneca. a. diagnostic approach: the role of hybrid imaging k. herrmann; würzburg/ this presentation will include a short review of the epidemiology of primary and metastatic bone disease, its classification and staging definitions. then we discuss the role of radionuclide imaging in initial and subsequent treatment strategy considerations. another aim is to provide guidelines for the appropriate use and applications of diagnostic radionuclide approaches for the assessment of bone diseases. the focus of this presentation will be on the role of radionuclide imaging in metastatic prostate, breast and lung cancer in the context of other available diagnostic imaging approaches. the ability of hybrid imaging technologies to achieve improved diagnostic accuracy will be emphasised and when appropriate the potential role of emerging pet imaging probes for assessing bone malignancies discussed. if feasible, the availability and effectiveness of radionuclide-based therapeutic approaches for malignant bone diseases will be reviewed. skeletal metastases are associated with significant morbidity and have high impact on health costs. effective palliative treatments are available, but less than % of patients respond in terms of tumour markers, pain or quality of life and all treatments are associated with some toxicity. it is generally accepted that the current imaging techniques are inadequate for measuring treatment response in a clinically relevant time frame such that skeletal metastases are usually considered as non-measurable disease in clinical routine and trials. there is therefore a need for non-invasive, objective methods to evaluate treatment response of skeletal metastases at earlier time points to guide clinical care. in addition to imaging structural changes in bone (e.g. x-ray, ct), it is now possible to image a number of biological characteristics of bone metastases including tumour metabolism, tumour cellularity, and osteoblast and osteoclast activity. methods currently available, or being actively investigated, include m tc-mdp spect/ct, f-fluoride pet/ct, f-fdg pet/ct, c/ f-choline pet/ct, rgd-spect/pet and dw-mri. some of the tracer methodologies will potentially be applicable for and complementary to pet/mri. the timing of follow-up imaging after commencing treatment is still under investigation. it is likely that tumour-specific methodologies may reflect response/non-response earlier than bone-specific methods due to a prolonged flare in reactive bone in responding metastatic sites. learning objectives: . to understand the techniques and quantitative parameters used for treatment response evaluation. . to learn the optimal timing for follow-up imaging after treatment. . to become familiar with common pitfalls in the interpretation of posttreatment mri and pet. panel discussion: how to differentiate between bone metastases and chronic inflammatory or treatment-induced changes : structured reporting is an opportunity and a challenge at the same time. expert speakers of this session will present and discuss their views on the subject, contributing to the clear vision of the members of audience on this complicated subject, convincing us to take this technique as an opportunity to improve and standardise the quality of our reporting activities. even though structured reporting (sr) has been appreciated as one of the effective methods of electronic reporting, there are not that many real applications to facilitate utilisation of this technique in practice. why it is so?the radiologists want a flexible tool to create clear, concise reports in a reasonably short time. like all template-based reporting systems, the application must be tailored to preferences of multiple users in a radiology department. the report templates should also be flexible in terms of length to avoid inappropriately detailed reports. the final appearance of the report is critical for acceptance by clinicians. accordingly, the technical challenges of development of sr systems may be grouped as: "user interface", "database structure", and "output file/document" problems. flexibility to comply with different user preferences will lead to both user interface design and database structure challenges. one of the major technical challenges is the transformation of structured data to a final text to be acceptable and applicable for the referring physician. also, exporting of reports to dicom format using dicom-sr concepts is still a challenge. multi-media formats are becoming more and more popular specially in sr reports. effective communication of radiology results is critical to high-quality health care. to that end, the radiological society of north america (rsna) has developed a library with more than best-practice radiology reporting templates. the templates are freely available online (www.radreport.org) and have been accessed more than , times. the new ihe "management of radiology reporting templates" (mrrt) profile builds upon the web's hypertext markup language version (html ) to provide an international standard for describing and exchanging reporting templates. this presentation will describe the rsna report template library, introduce the international mrrt standard for report templates, and describe opportunities for esr members and affiliated societies to participate. the broad goals of this initiative are to improve the quality of radiology reports, increase the productivity of radiologists, promote the use of practice guidelines, and advance data-driven health care.learning objectives: . to explore a library of 'best practices' radiology reporting templates. . to become familiar with international standards for radiology reporting templates. . to understand how structured reporting will promote the use of practice guidelines and will advance data-driven health-care. structured reporting in europe: the esr initiative o. ratib; geneva/chthe rsna has initiated a new initiative called radreport with the aim of helping promote standard reporting and improve reporting practices by creating a library of clear and consistent report templates (http://reportingwiki.rsna.org/). this project also supported in part by the national institute of biomedical imaging and bioengineering (nibib) has already gained the participation of numerous countries around the world, contributing with reports in different languages. these report templates are "structured" in the sense that they incorporate reusable knowledge, or meaning, to the clinical reporting process. structured interpretation and reporting for detecting significant prostate cancer is crucial, as it allows comparing inter-observer interpretation variability, reduces this variability by stimulating discussion of the individual scores, enhances communication with the clinicians in a uniform way, facilitates quality assurance plus research, and in this way improves patient outcome. in this presentation, the pi-rads scoring system will be shown and discussed, and fast effective structured reporting using computer software will be presented. in a rapidly ageing society, dementia prevalences are sharply increasing. furthermore, knowledge about disease mechanisms is constantly evolving. a more nosologic approach in the workup of dementia is required to improve prognostication, patient management as well as select appropriate treatment. a. anatomy of the limbic system t.a. yousry; london/uk (t.yousry@ucl.ac.uk)the term "limbic" has been used in many different connotations. to understand the anatomy, we have to define the term first. the clearest definition stems from broca: "le grand lobe limbic". defined as such, the limbic lobe is composed of main structures: the limbic and the intralimbic gyri. . the limbic gyrus is composed of the subcallosoal gyrus anteriorly, followed in an arc by the cingulate gyrus, isthmus and parahippocampal gyrus. the latter is composed of parts: i) a posterior narrow segment, the superior surface of which is called subiculum, and ii) a more voluminous anterior segment, also known as the piriform lobe. the latter consists of the anterior part of the uncus and the entorhinal area. . the intralimbic gyrus arches within the limbic gyrus. it is divided into parts: i) anterior (prehippocampal rudiment); ii) superior (indusium griseum); and iii) inferior (hippocampus). the latter consists of lamina rolled inside each other: the cornu ammonis and the dentate gyrus, with the cornu ammonis consisting of neuronal fields (ca - ). to understand the function, we have to understand the connections that define this function. the interplay with the insula is of particular importance. in summary, we will review the mri characteristics of the components of the limbic lobe, their connections, and their function, using . t, t and . t high field imaging. at the end of this lecture, you will know definition (s) of the limbic lobe, its major subdivisions, connections and functions. structural neuroimaging is increasingly used in the diagnosis of neurodegenerative diseases. in a memory clinic setting, computed tomography (ct) and magnetic resonance imaging (mri) have become the most important paraclinical diagnostic tools. recent guidelines on the diagnosis and management of disorders associated with dementia state that neuroimaging should be performed at least once during the diagnostic workup. in the past, neuroimaging had been performed to exclude potentially treatable causes of dementia. however, this exclusionary approach of neuroimaging in the diagnosis of dementia has been replaced by an inclusionary approach driven by conclusive evidence that ct and mri can substantially support the clinical diagnosis by the demonstration of a distinct atrophy pattern and the assessment of vascular (co)morbidity. for these purposes, the use of visual rating scales allows a fast and reproducible assessment of global/regional cortical atrophy and vascular white matter pathology. more recently, new imaging markers such as cerebral microbleeds and superficial siderosis haven been identified and linked to alzheimer pathology. in addition, next to structural mri, advanced and quantitative mri methods have been introduced in the clinical setting. mri perfusion techniques such as arterial spin labelling (asl) demonstrated a diagnostic value in memory clinic patients, particularly in those showing no or inconclusive pathology on structural mri. de (horst.urbach@uniklinik-freiburg.de) cognitive decline is a clinical symptom in many neurodegenerative disorders. mri is typically performed in the initial diagnostic workup of these patients. as a structural imaging modality, mri is often unrevealing while several molecular imaging modalities show disease-associated alterations: f-fdg-pet shows reduced glucose metabolism in neocortical association cortices, mainly in the posterior cingulate, precuneus and temporoparietal regions. since glucose metabolism is normally higher in the posterior cingulate and precuneus, reduced glucose metabolism must be specifically sought for or highlighted by voxel-based analysis. dynamic imaging of swallowing from the mouth to the duodenum is the only clinical test providing a fast and accurate diagnostic overview of the upper gi function. swallowing disorders are divided into those with dysphagia that means problems during eating or drinking, with suspicion of aspiration as an important subgroup and those with sensations like the globus syndrome or noncardiac retrosternal pain. oropharyngeal dysphagia in patients with neuromuscular diseases or postoperative conditions can be evaluated precisely and within the same session as oesophageal motility disorders, stenoses or postoperative problems are detected and in a complementary fashion to endoscopy, ph monitoring and manometry. the exclusive ability of dynamic imaging of swallowing is to diagnose combined disorders of the pharynx and the oesophagus, to detect and localise subtle and multiple benign stenoses of the upper gi tract and to combine visualisation of pathophysiology and bolus transport as well. radiologic evaluation concentrates on functional units, which represent the visible "moving areas" of deglutition, where evident pathologic conditions can be found: ) oral cavity and tongue, ) soft palate, ) epiglottis, ) hyoid and larynx, ) pharyngeal constrictors, ) pharyngoesophageal segment, ) oesophagus and stomach. this lecture intends to explain the common causes of dysphagia, the approach to a tailored examination and the use of an advanced imaging technique. further, the basics of taking the swallowing history and the structure of reporting swallowing disorders along the seven functional units will be provided. hoarseness is defined as a rough or noisy quality of voice. the possible causes of hoarseness are manifold, ranging from benign diseases such as the common cold to malignant tumours. therefore, hoarseness as a symptom should always be taken seriously. evaluation of a patient with hoarseness by a head and neck specialist will always starts with a history and a physical examination including laryngoscopy. after these examinations, a diagnosis can be established in most patients without additional imaging. the aetiologies of hoarseness are: ) inflammation/infection, ) trauma, ) neoplasms, ) others (including medical conditions, e.g. hypothyroidism) and ) vocal cord paralysis. this presentation will focus on imaging of vocal cord paralysis. vocal cord paralysis should not be considered a diagnosis, but as a symptom of (possible) underlying disease. when vocal cord palsy is discovered, this is frequently a reason to perform cross-sectional imaging. the radiological workup needs to include the full course of the vagus nerve. moreover, reporting radiologists need to be able to recognise the radiological signs of vocal cord palsy in the absence of hoarseness, since many patients with cord palsy are asymptomatic (up to %). the anatomy of the vagus nerve and its laryngeal branches that innervate the intrinsic muscles of the larynx will be discussed. then the (expected) course of the vagus nerve and recurrent laryngeal nerve will be shown. finally, radiological signs of vocal cord palsy will be discussed using examples from daily practice. in contemporary medicine, endovascular techniques often replace conventional methods of treatment, including surgery, in the management of an increasing range of diseases. this requires development of specific methods assessing the effectiveness of treatment and able to detect complications that may be much different from those occurring in conventional therapy. this is first of all seen in the endovascular treatment of patients suffering from vascular lesions and neoplastic diseases. commonly, there is a high initial technical success of endovascular treatment, but the durability is not always satisfactory and needs to be further examined. endovascular treatment is bound to the risk of specific complications, often without clinical symptoms. therefore, there is a need to use imaging follow-up of these patients.interventional radiologists are aware of the importance of those issues and should play an important role in developing and deciding about the follow-up scheme and choice of the best imaging methods for these patients. besides indepth knowledge of the treatment, radiologists are also familiar with the possible complications and can offer an optimal, minimally invasive and cost effective diagnosis and treatment. the potential imaging methods for follow-up after angioplasty (pta) and/or stenting include doppler ultrasound, cta and mra. although excellent images can be obtained by cta and mra, the drawbacks of these studies for routine follow-up are obvious. (access to ct and mr equipment, cost, contrast medium, ionising radiation, etc). in the uk, the most common imaging method is duplex ultrasound. there is no established role for the use of ultrasound contrast agents. ultrasound is freely available in the majority of centres, is inexpensive and complication free. follow-up by ultrasound may be undertaken by radiologists, although more usually by technologists in vascular laboratories. in practice, few patients are followed up by any imaging study at all unless they develop symptoms. the main category of patients who undergo some form of imaging follow-up after pta/stenting are patients with surgical bypass grafts, who are usually routinely followed up whether they have undergone angioplasty/stenting or not. in view of the fact that this is a lecture on follow-up after intervention, procedural complications are excluded from the discussion. the complications of angioplasty and stenting are similar. the main early complications are restenosis or reocclusion. other early complications are related to the arterial puncture site and include haematoma, dissection, occlusion and pseudoaneurysms. the main late complications are restenosis and reocclusion. treatment of restenosis and reocclusion usually involves additional angioplasty or stents using conventional or drug-eluting balloons or stents. cutting balloons, atherectomy and cryotherapy have a controversial role based on limited evidence. imaging follow-up of oncologic patients after embolisation procedures j.i. bilbao; pamplona/es (jibilbao@unav.es)the term "embolisation" groups different procedures in which imaging guidance is fundamental for both performing the procedure and for the early detection of complications. it has been demonstrated by several studies that "tumoural response" is tightly associated with survival, thus it is of major interest to measure accurately how the tumour is modified by the treatment. the "world health organisation" -who -first published the objective criteria for the evaluation of response. these criteria were slightly modified in and named as "response evaluation criteria for solid tumors"-recist. accordingly, "complete response" (cr) corresponds to the disappearance of the tumour, "partial response" (pr) is defined when there is a decrease in the tumour size equal to or higher than % and "progressive disease" (pd) is applied when there is an increase equal to or higher than %. "stabilisation of the disease" (sd) corresponds to measurements in between > % (pr) and < % (pd). tumours (until five nodules in the same viscera), no matter if necrotic or viable, were evaluated with one measure (its largest diameter). a new version (" . recist) has introduced modifications, among which is the number of nodules ( instead of ) that needs to be evaluated. in some circumstances, when morphologic criteria (recist and who) fail to predict the outcome, then new functional criteria have been recently established. "mrecist" (which only measures the tumour that uptakes contrast) is an accurate classification for hcc. new methods "beyond recist" have been established for some specific treatments such as targeted therapies. in this session, the audience will have a comprehensive overview of the elastography, its principles and current state-of-the-art clinical applications. the session will deliver an overview of currently available methods of both ultrasound elastography (use) and magnetic resonance elastography (mre), their physical principles, technical and equipment requirements as well as the influence of various factors for obtained results. following this session, the audience will have insight into clinical applications of elastography in liver, prostate and brain diagnostics. the strengths and weaknesses of elastographic modalities will be compared and discussed under consideration of more established imaging modalities. the session will present potential developments of use and mre and possible future clinical applications. panel discussion will aim at defining the place of elastography in current and future diagnostic algorithms. the well-known sensitivity of the palpating hand for detecting lesions is related to the incredibly wide range of values of the shear modulus in the body, spanning over eight orders of magnitude. elastography was invented to exploit this high variability of constants by introducing the shear modulus into the image contrast of medical ultrasound or magnetic resonance imaging. today, both ultrasound elastography (use) and magnetic resonance elastography (mre) are in clinical use, predominantly for staging liver fibrosis or discriminating tumours in the breast, liver, and prostate. the talk reviews the physical principles, technical requirements and current methods of both use and mre. furthermore, the strengths and weaknesses of elastographic modalities are compared and discussed under consideration of more established imaging modalities. an outlook of elastography is given, highlighting the real-time feasibility of use and the sensitivity of mre to tissue pressure. suspicion of prostate cancer (pca) exists in the case of elevated psa serum values and/or suspicious findings on digital rectal examination (dre). systematic biopsy (sb) will be done to confirm or rule out pca. unfortunately, an elevation of psa serum values often is caused by benign changes, e.g. prostatitis or adenomyomatosis and not every cancer is palpable on dre. furthermore, sb may miss clinically significant disease in up to %. therefore, a more targeted approach would be desirable and imaging of pca is currently under strong effort. one possibility for visualisation of pca is the representation of tissue elasticity. usually, cancers have higher cell and vessel density than the normal surrounding tissue and therefore are associated with increased stiffness. in contrast to dre, where only the posterior parts of the prostate can be reached, rte does not have this problem, since all anatomical regions can be evaluated. furthermore, this noninvasive technique is time-and cost-effective and targeted biopsy or focal therapy of the prostate can be done under real-time conditions. another important issue is that rte can be performed by both urologists and radiologists. nevertheless, rte is of limited value in the detection of small cancer lesions and there may be problems in visualising pca with predominantly gleason pattern . the lack of data about inter-and intraobserver variability and of multicentre studies (now in process) are probably the reasons why rte has not been included in the guidelines of urological societies so far. cerebral tissue structure is altered in many neurodegenerative diseases, but also during physiological processes like maturation or ageing. structural changes directly affect the mechanical tissue properties. magnetic resonance elastography (mre) is an imaging technique capable of assessing biomechanical brain parenchymal properties non-invasively. viscoelasticity can be quantified by analysing the propagation of mechanically elicited shear waves in the investigated tissue. thus, mre could be a helpful tool to detect physiological or pathological processes influencing the cerebral tissue integrity. learning objectives: . to appreciate the advantages and limitations of a virtual palpation of the brain. . to become familiar with mechanical wave induction and detection in cerebral measurements. . to learn about current and potential clinical applications. . to learn about the relationship between microscopic and macroscopic, as well as focal and globally diffuse biomechanical brain parenchymal alterations. are based on the generation of shear waves inside the liver tissue with specific mr liver drivers and ultrasound probes. subsequently, phase images revealing the displacement of the waves will be investigated for healthy and pathological (alcoholic fibrosis) livers. moreover, the development of phantoms mimicking the stiffness of biological soft tissues (liver, muscle, …) will be also presented. the viscoelastic (elasticity and viscosity) properties of healthy and fibrotic livers were measured, and the relevance of these parameters, used as a liver diagnostic marker, was analysed. in parallel, an increase of the wavelength as a function of the stiffness was obtained for the developed set of phantoms. the characterisation of the elastic properties is an excellent marker to differentiate minor and major fibrosis. a summary of the advantages and disadvantages of us and mr elastography techniques will be presented. a- : the breast lesion excision system (bles) has evolved as a breast radiological technology over the last decade and is now in widespread use in europe and across the world. it was designed as a large biopsy device, but more recently due to its unique capability to obtain a single large breast tissue specimen in only a few seconds by utilising a radiofrequency cutting and cauterising wave, it has increasingly been explored in the therapeutic setting. it is easy to use under ultrasound or mammographic guidance with procedures taking a similar length of time to that of a vacuum biopsy, and with patient preparation and anaesthetic essentially identical. the technical aspects of performing these procedures will be detailed as well as its use specifically to perform excisional biopsies. this is limited by patient and lesional factors, all of which will be discussed in more detail. following a bles, the post-procedure appearances need to be considered so that follow-up imaging can be interpreted accurately. the risks and complications of this procedure are outlined as well as a discussion made of the latest papers in this field that may suggest future applications and developments. learning objectives: . to understand the mechanism of the bles technique. . to become familiar with the post-bles aspects of various types of imaging. during the past few years, it has been shown that there is not a single disease entity called "breast cancer". there are different subtypes that entail diverse recurrence risks. this is the first issue to take into account, and patients will be stratified accordingly before any follow-up is planned. imaging findings in a patient treated for breast cancer will depend on the type of treatment: breast conserving therapy (bct), mastectomy (and all the reconstructive techniques), radiotherapy and minimally invasive techniques. to differentiate between fat necrosis and other common post-treatment changes from relapse, it is important to know the timeline when all these changes take place and also schedule the follow-up imaging procedures accordingly. other important issues to take into account are the limitations and indications of the different modalities (mammography, ultrasound and mri). although ultrasound and mammography have traditionally been used in the follow-up of these patients, mri is being used more and more often due to its superior multiplanar capabilities and the functional information not supplied by the other techniques. blood clot formation in the veins is referred to as venous thrombosis (vt). vt is most common in the deep veins of the legs and pelvis. if the thrombus dislocates to the pulmonary arteries this is referred to as pulmonary embolism (pe). the combination of deep vein thrombosis (dvt) and pe is referred to as venous thromboembolism (vte). the diagnosis of dvt and vte is essential, as vte is potentially fatal. furthermore, undiagnosed, non-fatal dvt may result in negative long-term pathologies (e.g. post-thrombotic syndrome). major risk factors for the development of vte are thrombophilia, history of dvt, age > , surgery, obesity, long travel, cancer, immobility and pregnancy. in addition, underlying congenital abnormalities or compression syndromes (may-thurner syndrome) are also important to diagnose. diagnosis of vte is not always easy. primary diagnosis includes clinical (wells score) and lab testing (d-dimer). first line imaging testing is done by leg ultrasound (us). if pe is suspected ct is indicated immediately. ct of the pulmonary arteries (ctpa) may be combined with indirect ct venography (ctv). the question of when to combine ctpa and ctv still remains. a favourable approach is to add ctv in a negative ctpa. recently mr venography (mrv) is emerging as comprehensive imaging tool especially in patients with chronic dvt to assess the extent and underlying causes. in this respect imaging without radiation dose with high spatial delineation of vascular abnormalities facilitates intervention for thrombolysis or interventional therapy including recanalisation and stenting. alternatively, intravenous unfractionated heparin is used in situations when a quick reversal is necessary. b) non-medication: early mobilisation has been widely adopted to activate the muscle pump. if early mobilisation is not possible, compression stockings and/or pneumatic compression boots are applied to better empty the veins. . treatment: a) anticoagulation: the main goal of anticoagulation is to prevent progression and recurrence of dvt. typically, anticoagulation is started with lmh, followed by vitamin k antagonists (coumadin). the first dvt is generally treated for - months; recurrent dvt for months. b) compression therapy: compression stockings ameliorate pain and leg swelling of acute dvt. in addition, the development of a post-thrombotic syndrome (pts) should be reduced. c) thrombolysis: the main goal of thrombus removal is prevention of pts. in addition, thrombolysis is used for severe ilio-femoral dvt, especially with phlegmasia coerulea dolens. there are different ways of thrombolysis: systemic thrombolysis was abandoned because of high bleeding complications. catheter-based thrombolysis has shown to reduce the bleeding risk, but it is quite time and resource consuming. the latest development is pharmaco-mechanical catheter thrombus removal which combines thrombolytic drugs with a mechanical maceration. recent studies (cavent,torpedo) have shown a reduction of pts compared to anticoagulation. the use of inferior vena cava filters (ivcfs) is a controversial method for the prevention of pulmonary emboli. there are large variations in their usage; often with ten to one hundredfold differences in placement between different countries. ideally, ivcf usage is based on sensible protocols derived from clinical experience and trials. in practice, this is less frequently the case. indications, contraindications and questionable indications (mainly during venous thrombolysis) will be discussed. familiarity with ultrasound guidance will be stressed. the key technical steps in the insertion and removal of ivcfs will be discussed. the question that this refresher course is aimed to answer is whether novel it tools may actually help in improving quality and workflow efficiency in daily radiological practice. in fact, since the early installations of pacs, it tools have often been considered as mere productivity tools rather than enabling technologies for fostering quality in medical care. the three distinguished lecturers in this course will address the following topics: improving quality and efficiency of computerised order entry through decision support, improving quality and efficiency of reporting through structure and templates, and improving quality and efficiency of dose management through exchange between modalities and registries. they will cover all aspects of the radiological workflow: from the selection of the most appropriate diagnostic procedure, to the efficient reporting of diagnostic and non-diagnostic data (such as radiation dose information and contrast media information). they will demonstrate how newly adapted it tools may provide assistance throughout the radiological workflow, with potentially enormous gains in terms of patients' safety and total quality management. session objectives: . to appreciate the need for it tools to ensure quality control. . to understand how to collect data concerning radiation dose. a. improving the quality and efficiency of computerised order entry through decision support p. mildenberger; mainz/ de (peter.mildenberger@unimedizin-mainz.de) electronic communication of requests for radiological procedures is a logical and relevant extension to integrated information systems in health care. this communication of orders and the entry (oe) into ris improve the workflows. but, quality improvements require additional efforts to link order-entry-solutions with health knowledge for ordering the appropriate imaging procedure. these clinical decision support (cds) tools should be based on generally accepted and implemented criteria, e.g. evidence-based medicine. it is known that the acceptance of such systems, if successful and efficient implementations are given, is very good. it concepts for cds are well known and standards for classifications are available, but semantic interoperability is still a developing area. actually, clinical information systems provide different levels of integration of oe and cds. further developments could be an implementation of interoperability profiles and the representation of cds knowledge in webbased services of non-commercial organisations. b. improving quality and efficiency of reporting through structure and templates e. neri; pisa/it (emanueleneri @gmail.com)the report represents the final stage of the diagnostic imaging process, for which the radiologist is responsible. this process is driven by an accurate selection of the diagnostic tests, based on the clinical question. therefore, the report should be an expression of this path, depending on the question asked by the clinician. the structure of a radiology report for the same clinical problem and the same method of investigation varies from country to country and between different radiologists. this diversity can be a problem in the global world where a patient can have imaging tests from different places with different reporting languages, or when patient data is transmitted and shared between different centres. there is, therefore, the need to standardise the report as much as possible; finalising its structure and contents to a specific clinical problem, and possibly using a standard terminology. an attempt to solve this problem is in progress with the joint initiative on structured report esr-rsna (http://www.rsna.org/reporting_initiative.aspx), aimed at improving reporting practises by creating a library of clear and consistent report templates. the main expected benefits of structured reporting templates are report uniformity and improvement of communication between referring physicians. literature evidences that structured reporting can also quicken report turnaround, and reduce costs and lexical errors; thereby improving the quality and safety of radiological practise. the european directive on medical exposures requires the assessment and evaluation of patient doses, especially in procedures involving high doses to the patient. in the current draft of the new directive on basic safety standards, some requirements on patient dosimetry in diagnostic and interventional radiology have been reinforced: x-ray systems should provide dosimetric information with the capability of being transferred to the examination report (for all ct and interventional systems). diagnostic reference levels (drls) shall be reviewed regularly. these requirements will push the industry and the users to develop better strategies to evaluate patient doses, transfer these values to patient reports (contributing to the patient exposure tracking system) and also use available software to process these dosimetric data and perform some automatic analysis. this analysis should: a) include periodic calibration factors for patient dose quantities, b) include automatic detection of high dose values (especially relevant for interventional procedures), c) include statistical analysis to update drls and compare with the existing ones and d) suggest corrective actions to fulfil the quality assurance programmes and the clinical audit requirements. dicom radiation dose structured reports represent a significant advantage, but more efforts will be necessary for the automatic process of the relevant data contained in the report to verify that the radiological risk is acceptable and to suggest, if appropriate, corrective actions to improve clinical practice. without these last steps, patient dosimetry efforts and european regulations for radiation safety could only have a moderate impact. gastrointestinal tumors include a variety of lesions, with the most frequent being adenocarcinoma of the small and large intestine, small bowel lymphoma, neuroendocrine tumours (nets) and gastrointestinal stromal tumours (gists). according to histology, location and initial imaging staging, those lesions may require completely different therapies: surgery alone, adjuvant chemotherapy (cht), neoadjuvant cht followed by surgery or a combination of neoadjuvant chemoradiotherapy (crt) followed by surgery. after treatment, imaging follow-up is mandatory. the most common post-operative findings after small bowel or colonic resections will be presented, together with clues for early detection of recurrence. in those cases which underwent adjuvant cht, imaging findings and diagnostic criteria related to the use of either cytotoxic or cytostatic drugs will be shown, to facilitate the interpretation of radiologists in assessing response to therapy. finally, tissue changes following neoadjuvant crt will be discussed, particularly in rectal cancer. the possibilities of different imaging modalities in assessing either complete or partial response to therapy will be presented, with a critical analysis of different imaging findings. unlike infectious enteritis, in patients with crohn's disease (cd) the changes in clinical activity have poor correlation with changes in inflammatory lesions after medical treatment. mucosal healing (mh) and improvement in patient's symptomatology are usually considered the main objectives of medical treatment. however, improvement in clinical symptoms is not always associated with mucosal healing and the persistence of severe inflammatory lesions augurs a more aggressive disease course in terms of flare-ups, hospitalisation requirements and the need for surgery. endoscopy is the gold standard for the assessment of luminal lesions in cd, but given the good correlation shown between endoscopy and cross-sectional imaging in assessing the activity and severity, imaging is increasingly introduced as a tool to monitor medical treatment. there is some evidence indicating that crosssectional imaging can be considered a responsive and reliable tool as it detects meaningful changes in patients' status over time after therapeutic interventions. besides this, cross-sectional imaging is capable of monitoring the colon and the small bowel and penetrating lesions that cannot always be assessed by endoscopy. becoming familiar with the radiological changes during and after treatment will be key in the decision-making process. degeneration is routinely defined as a morbid change of cells, tissues and even organs. in the msk system degeneration occurs in bones, joints (synarthrosis and diarthrosis), muscles and tendons. primary degeneration implies ageing as well as overuse, based on mircrotrauma. the latter may be related to constitutional, professional or sports-related factors. a distinction between ageing and overuse is not possible with the use of histology and imaging. only the intensity of the changes, which may be inadequate in relation to the age of the patient, may give a hint. on the contrary, the term "secondary degeneration" should be used for clearly defined events or diseases, which finally will result in the degeneration of tissues. the causes are inflammation (ra, infection), metabolic disease (e.g. cppd) or trauma. other ways to classify degeneration of the msk system are related to the anatomy. accordingly, degeneration of the particular parts of the peripheral or axial skeleton, muscles and tendons has to be discussed. one has to have in mind that bones, cartilage, muscles and tendons form a complex unit. the single elements of these complex units jointly undergo degenerative deterioration, e.g. bone, cartilage, fibrocartilage and capsule are jointly involved in case of oa of the knee joint. the same holds true for the synchondroses of the spine. at the rotator cuff not only the tendons, but also the underlying bone and the muscle tissue are changed. (o.m.vandelden@amc.uva.nl) there are many different types of embolisation, but all share many basic principles. the specific approach, technique, materials and equipment used may differ depending upon many variables such as the type of vascular territory involved (venous, arterial, small vessels, large vessels), clinical setting (elective or emergent procedure), and type of underlying disease to be treated (e.g. tumor embolisation, acute haemorrhage, vascular malformation, arteriovenous fistula, etc). pre-procedural imaging and planning are essential for most embolisation procedures and intra-procedural navigation can be done with fluoroscopy, dsa, and new d-techniques. with proper imaging workup prior to embolisation procedure time, radiation dose and contrast load can be significantly reduced. embolisation materials can be roughly divided into liquid agents (alcohol, glue, polymers), particulate agents (pva, calibrate microspheres, drug-eluting beads) and coils (detachable, non-detachable) and plugs (amplatzer®-plugs). plugs and coils are deployed at the exact site of destination and usually easy and safe to use. particles and liquid agents reach their site of destination by flow guidance and can be more difficult and unpredictable in their use. complications include puncture-site complications (thrombosis, dissection, haematoma), systemic complications (contrastinduced nephropathy, allergic reactions) and specific embolisation-related complications (non-target organ embolisation, end-organ ischaemia, postembolisation syndrome). when used with expertise and proper experience, most types of embolisation have good results with acceptable complication rates. although there has been advancement in imaging techniques, several pitfalls in the assessment of diseases of the pancreas, small bowel and rectum remain. the differentiation and the correct characterisation of tumours and inflammatory disease in some cases may be difficult. also, atypical presentation of common tumours or uncommon tumours persists and is a challenge. problematic situations are constantly represented as intestinal bleeding and unexpected findings. in this session, we point out the procedure of the best imaging modality and protocols of challenging cases. a. pancreas c. triantopoulou; athens/gr (ctriantopoulou@gmail.com)pancreatic imaging and interpretation of focal lesions remains a challenging issue. despite the advances in imaging techniques and the dedicated protocols that are now in use, a possibility for pitfalls remains. these pitfalls are related either to the inability of the early recognition of a lesion, or to the wrong characterisation of a "mass". both pitfalls are of great importance, taking into account the lethal pancreatic cancer and the possible complications that may follow an unnecessary pancreatic surgery. there are many variants of the pancreatic parenchyma, the ducts or even vessels, and deep knowledge of the pancreatic anatomy and embryology is needed, to be able to recognise these variations in imaging. a pancreatic variant may not only be misinterpreted as a severe pancreatic disease (e.g. ectopic spleen vs neuroendocrine tumour), but may also be the underlying cause of pancreatic inflammatory lesions (e.g. ectopic pancreas on the duodenal wall causing groove pancreatitis). other pitfalls are related to the atypical presentation of a common lesion (e.g. isoattenuating pancreatic adenocarcinoma), the presence of uncommon tumours, diffuse or multifocal diseases, the co-existence of two different entities and the heterogeneous or overlapping appearance of cystic lesions. in any challenging case, a focused methodology should be applied and the diagnostic procedure based on predefined imaging parameters, taking also into consideration the clinical and laboratory findings. excluding pancreatic cancer should be the first goal and every attempt should be made to differentiate between surgical and non-surgical cases. endoscopy is the gold standard in small bowel diseases, but it is a challenging technique: the pillcam is a commonly used tool and multidetector ct is a diffuse investigation technique. ct enteroclysis/enterography is a worldwide tool in intestinal exploration. throughout the investigation of the small bowel, morphologic anomalies or embryonal development defects may be present (meckel's diverticulum, duplications of intestinal tracts). artefacts may be determined and there may be misunderstanding. mdct is the first tool in emergency diagnosis; imaging pitfalls are possible. unsuspected diseases may be revealed: we will see a case of gist in a patient affected by intestinal bleeding of an obscure origin. ct enteroclysis may find unknown or unsuspected diseases: correct technical parameters have to be respected, as artefacts may be derived from an improper acquisition technique, such as a limited distension or poor intravenous enhancement. in follow-up, mr is preferred to investigate the intestine, mainly in the young population. unexpected findings may be due to rare pathology: we met a case of endometriosis, with ambiguous presentations; patients affected by autoimmune pathologies (pyoderma gangrenosum or lupus erythematosus), with unknown inflammatory intestinal chronic diseases. in oncology, pet-ct imaging is the main diagnostic tool, but there are abdominal pitfalls that limit its effectiveness (intestinal hot spots, artefacts from peristalsis): the small bowel may frequently obstruct an easy diagnosis. in case of emergency, if the radiological diagnosis is not clear, the surgeon determines the management of the patient. in case of a defective study technique, a correct examination may be repeated. endoscopy or pillcam may help to detect mucosal lesions invisible at ct. mri plays a key role in the assessment of disease of the rectum and in particular for the staging (and restaging) of rectal cancer. in this session, the relevant mr anatomy of the rectum will be discussed. furthermore, the strengths and weaknesses of mri in the assessment of rectal cancer during primary staging and restaging after chemoradiotherapy will be highlighted using various rectal cancer cases. finally, we will discuss some rare tumours of the rectum (including gist and carcinoid). optimisation of ct and mr techniques has provided new challenges for imaging in the assessment of airway and lung diseases. phenotypic abnormalities which can be recognised on visual and quantitative evaluation of ct images in copd patients may improve the diagnostic accuracy, help optimise treatment and provide a framework for clinical trials. although expiratory ct helps assess air trapping due to small airway obstruction and excessive dynamic collapse of large airways, potential tricks in technique and interpretation of ct images must be known. owing to multiple and successive improvements over years, functional and morphologic evaluation of the lung using mri has become reality for clinical practice. already regarded as a favoured cross-sectional imaging in paediatric chest radiology, mri of the lung is gaining new indications also in lung diseases occurring in adults. the potential for high-resolution computed tomography (hrct) to uncover several morphological subtypes that come under the umbrella term chronic obstructive pulmonary disease (copd) is now more familiar to both radiologists and specialist respiratory physicians. a complete approach to the classification of copd would ideally assimilate several parameters through a combined visual-quantitative hrct analysis. the insight that some subjects given the label of copd have "pure" airways diseases or emphysema can be readily provided by a simple visual evaluation. the subtypes -centrilobular, panlobular, and paraseptal emphysema -can be reliably distinguished on hrct images. the visual assessment of bronchial abnormalities and accompanying smoking-related interstitial lung disease may also complete the phenotypic classification of copd. furthermore, it is now possible to objectively quantify the global extent of emphysema, gas-trapping, and bronchial metrics by two types of softwares which are now increasingly available on latest ct workstations. learning objectives: . to learn about the classification of copd. . to appreciate the role of hrct in the classification of copd. a- : this presentation will examine the role of expiratory ct in the diagnosis of small and large airways diseases. small airways disease or air trapping may be a feature of asthma, copd and hypersensitivity pneumonitis, and is typically characterised by a mosaic attenuation pattern on ct. tracheobronchomalacia and excessive dynamic airway collapse (edac) are large airways conditions that may be recognised by tracheal narrowing on inspiratory and/or expiratory ct. the typical clinical and ct manifestations of these diseases will be reviewed. additionally, a number of important caveats with regard to ct in small and large airways disease will be discussed: ) the comparative strengths and limitations of ct, bronchoscopy and lung function testing in making the diagnosis; ) the overlap in appearances that exists between diseased and healthy individuals; ) the variability in definitions of conditions such as tracheobronchomalacia. the optimal ct technique for imaging of airways disease, including the role of dynamic expiratory ct, will also be reviewed. for routine clinical applications, a standardised mri protocol has been widely implemented. it addresses the major challenges of mri of the lung, i.e. low proton density, susceptibility artefacts as well as respiratory and cardiac motion. beyond visualisation of lung morphology, which is done in an inspiratory breath-hold, functional imaging has become an integral part of the routine protocols: perfusion, blood flow, ventilation, respiratory motion, diffusion. for perfusion imaging, inspiratory or expiratory scans can be used, blood flow measurement should be performed during shallow breathing, and ventilation and diffusion are mainly assessed in inspiration. respiratory motion is best assessed by dynamic image acquisitions. continuous breathing can also be combined with triggering or navigators to improve image quality or derive functional maps. mri is recommended as the first-line cross-sectional imaging modality in paediatric chest radiology, including cystic fibrosis, complicated pneumonia, any pulmonary or cardiovascular anomaly, as well as the assessment of mediastinal masses in children and adults. mri is also challenging pet/ct in the staging of lung cancer (tnm) as well as ct and echocardiography in the assessment of pulmonary arterial hypertension. mrieven without contrast -is an alternative in the diagnosis of acute pulmonary embolism if iodinated contrast medium is contraindicated or in pregnant women. mri should also be used in complicated pneumonia, e.g. in immunocompromised patients as well as for the functional assessment of chronic obstructive pulmonary disease (copd) or severe asthma. the role of mri in interstitial lung disease stills needs to be determined. unlike the simple forms of cranisynostosis whose modalities and timing of surgical correction are well established, the management of faciocraniosynostoses requires a multispecialistic and repeated evaluation of the single patients. their evolution, in fact, depends on the specific genetic anomalies but also on the phenotype, which can vary in time, of their clinical expression which may lead to different degrees in severity of the associated functional disturbances (csf dynamics impairment, visual deficits, respiratory anomalies, etc). consequently these conditions may require several surgical steps the timing of which are dictated by the exact clinical diagnosis and the evaluation of the functional status at different ages. current management of faciocraniostenoses is thus based not only on the radiological studies but also on the functional assessment of the brain function (e.g. mri imaging studies, metabolic and cerebral blood circulation investigations, respiratory pathways volume evaluation, sleep recording). today's surgical multidisciplinary management may benefit a large variety of techniques which extend from free bone cranioplasty with intraoperative active fixed expansion to more dynamic and less invasive methods which exploit the physiological brain expansion in infants or the gradual cranial vault expansion mechanically stimulated by springs or distractors. in the present report, we will review the decision making process and the rationale on which, at the necker-enfants malades, we base the use of the currently available techniques for skull expansion and correction of cranial malformations according to the accurate dynamic radiological and functional evaluation at different stages of the disease. learning objectives: . to learn about the imaging patterns of faciocraniosynostoses. . to learn how and when to image. . to become familiar with associated brain anomalies. b. all about the paediatric pituitary gland m.i. argyropoulou; ioannina/ gr (margyrop@cc.uoi.gr) mr is the imaging modality of choice for the assessment of the pituitary gland (pg) and the hypothalamus. the normal adenohypophysis is bright during the first two months of life and appears isosignal to the brain parenchyma afterwards. the neurohypophysis is bright, provided that the child is well hydrated. the pg height decreases during the first year of life and then increases until puberty. adenohypophyseal deficiency has been associated with a small pituitary size, an ectopic neurohypophysis, a hypoplastic or absent pituitary stalk, hypothalamic gliomas, craniopharyngiomas and iron overload states. precocious puberty may be idiopathic, but it has also has been associated with hypothalamic hamartomas, craniopharyngiomas and hydrocephalus. diabetes insipidus may be secondary to histiocytosis x, germinomas and trauma causing pituitary stalk interruption. the hypothalamopituitary axis is evaluated by using sagittal and coronal t -weighted images without and with contrast administration. dynamic contrast enhancement may be useful in the evaluation of disorders responsible of diabetes insipidus. magnetisation transfer imaging may be useful in the evaluation of pituitary deficiencies or precocious puberty. familiarity with orbital and ocular anatomy is crucial to the understanding of disease processes of the orbit. lesions of the orbit may be divided into those which are intraocular and those which are extraocular. the underlying aetiology and geneses of many different lesions that can occur in the orbits vary depending on the location of the lesions and sometimes on additional nonorbital conditions. ct and mri play crucial roles in the evaluation of orbital pathology where often mri still is a complement to ct examination in the evaluation of orbital lesions familiarity with the radiologic appearance of common orbital lesions is important, as many of these lesions will not be seen on physical examination. after a very brief anatomic overview, most of this lecture will focus on the more common benign and malignant lesions as well as differential diagnosis involving the orbit and lachrymal gland in both the paediatric and adult population. the lecture will also present suggested imaging protocol and standard of care with respect to imaging. learning objectives: . to understand the embryology and imaging findings of the most common malformations. . to learn about space-occupying lesions and the differential diagnosis of tumours and inflammatory conditions. . to be aware of the role of conventional and advanced mr sequences in the diagnostic approach to lesions in the orbit. room e the hand and wrist injury to the wrist and hand is an important clinical problem. first, such injury is relatively common. second, the spectrum of abnormalities is complicated. the purpose of this presentation will be to learn more about the imaging appearances of soft tissue and osteoarticular injury of the wrist and hand using multiple imaging techniques. emphasis will be placed upon pathomechanics and characterisation of greater and lesser arc injury patterns of the wrist. the various posttraumatic instability patterns of the wrist will be also discussed. rheumatoid arthritis (ra), psoriatic arthritis (psa) and other inflammatory disease can be diagnosed and sometimes differentiated in the early stages of the disease on the basis of mri and/or pdus features of the hand and wrist. rheumatoid arthritis (ra) activity is closely correlated with inflammation. the synovial membrane is the principal site of inflammation in which the inflammatory process enhances capillary perfusion and permeability. doppler ultrasonography (dus), using the amount of colour pixels in the region of interest, and dynamic magnetic resonance imaging (de-mri) are both able to detect this inflammation in the wrist and hand. although these techniques are both capable of monitoring synovium inflammation modifications after ra treatment, pdus has become an essential tool for ra joint monitoring in routine practice in view of its sensitivity in the detection of synovitis, feasibility in outpatient clinics, and low cost. (ellopis@hospital-ribera.com) radiological study of the wrist and hand is challenging due to its complex anatomy with many small structures and the number of normal bone and soft tissue variants that might mimic injuries. moreover, many findings can be asymptomatic. their knowledge is important to avoid misdiagnosis. during this lecture we will also review the role of the different imaging modalities, such as plain films for wrist alignment and bone structures as well as the important role of us and mr in differentiating tumour from tumour-like conditions. we will become familiar with some specific radiological findings that allow us to make accurate diagnoses of soft tissue and bone lesions. learning objectives: . to learn more about the spectrum of intra and para-articular soft tissue tumours, and soft tissue tumour-like lesions. . to become familiar with us and mri findings of specific soft tissue lesions. room e oncologic imaging chairman's introduction c. pfannenberg; tübingen/ de (christina.pfannenberg@med.uni-tuebingen.de) with recent advances in cross-sectional imaging, the frequency of detecting "incidental findings" has markedly increased. incidental findings (=incidentaloma) are defined as unexpected, asymptomatic abnormalities detected by imaging performed for an unrelated reason. these incidentalomas have created a management dilemma for both radiologists and clinicians, particularly in the cancer patient in whom any mass warrants further evaluation. discovery of incidental findings often leads to a cascade of additional tests that is costly, provokes anxiety and exposes patients to the risk of unnecessary radiation and intervention. the workup of incidentalomas varies widely by physicians, and strategies for optimising patient management are only beginning to emerge. in this course, guidelines concerning a rational approach to some of the more common incidental abnormalities in cancer patients are presented. basic principles and tools for interpretation of incidental findings, common pitfalls and protocol issues regarding the differentiation of benign and malignant lesions as well as treatment-induced abnormalities will be addressed with a focus on the liver, lung and skeletal system. at the end of the session, the speakers will discuss the role of functional imaging techniques for characterisation of incidental lesions. session objectives: . to become familiar with basic principles and common pitfalls in interpretation of incidental findings in the lung, abdomen and skeletal system. . to learn how functional imaging (dwi, pet/ct, bs) can help in lesion characterisation. . to provide attendees with clear and practical messages for the management of the most common incidental findings in cancer patients. a. abdomen: common pitfalls and protocol issues a. ba-ssalamah; vienna/ at (ahmed.ba-ssalamah@meduniwien.ac.at) an incidentaloma is quite a frequent diagnosis in medicine, in general, and in radiology, in particular. by definition, an incidentaloma is an abnormality or a tumour found by coincidence without clinical symptoms or suspicion. these incidental findings have long posed challenges to physicians, and particularly to radiologists as well as health-care providers due to the resulting high costs. the scope and scale of these challenges have increased with the introduction of new technologies, in particular cross-sectional imaging, such as mdct and mri using sub-millimetre thin slices. incidentalomas still cause a management dilemma for clinicians, radiologists and even for the patients themselves. this dilemma is particularly pertinent to oncology patients. therefore, a precise knowledge of the broad spectrum of incidental findings is crucial. based on this knowledge, we can choose the appropriate radiologic examination that will provide a confident diagnosis. this way, needless uncertainty, for both the patient and the physician in charge, is avoided. furthermore, invasive procedures, such as biopsies, with the potential for complications, as well as cost-intensive follow-up examinations, can be reduced. furthermore, mri is usually used to detect the side effects of chemotherapy of the liver. simple steatosis is usually reversible, unless the liver receives a 'second hit' of damage from other causes leading to other manifestations, including chemotherapy-associated steatosis, sinusoidal obstructive syndrome, nodular regenerative hyperplasia, veno-occlusive disease, peliosis, pseudo-cirrhosis, and sclerosing cholangitis largely due to microvascular injury. finally, the effects of chemotherapy on peritoneal and mesenteric structures after performing intraperitoneal chemotherapy will be discussed. this presentation will discuss the complex topic of incidental findings in thoracic imaging of oncological patients. incidental findings will be presented with respect to their respective organs and anatomical regions. the presentation will emphasise the clinical relevancy of the findings as well as the specific of risk estimation in oncological patients. finally, the presentation will discuss the usefulness (or lack thereof) of current management guidelines for incidental findings, as designed for the general population. illustrative clinical scenarios will be discussed. radiographs and ct often detect incidental observations in bones, corresponding either to normal variants or benign conditions, which require additional imaging to rule out malignant conditions. mri most often provides nonambiguous explanation for these observations. beside this, mri, targeting either the whole body or only the axial skeleton, often including diffusionweighted (dwi) sequences, is increasingly used for bone tumour detection in many "osteophilic" cancers and haematologic malignancies, without need for contrast material injection and without irradiation, with unparallelled sensitivity to bone lesions. an important advantage and also a challenge for musculoskeletal radiologists are the all organ screening capabilities of dwi, demanding careful study of the huge information provided and knowledge extending far beyond bones in terms of organs and metastatic spread of different cancers. a second major advantage is its ability to detect lesion changes under therapy. however, benign lesions and non-neoplastic conditions, as well as artefacts may also lead to confusing observations on mri, and particularly on dwi. anatomical mr sequences as well as other imaging modalities are of utmost help to recognise these pitfalls. whole body mri including dwi sequences has to position itself among other diagnostic tools, bone scan, spect, and especially pet with its variety of tracers, in the currently evolving strategy of bone screening techniques, which will most likely vary according to the primary cancer. learning objectives: . to present most frequent x-ray and ct pseudo lesions and benign conditions and show how mri often enables straightforward interpretation of these abormalities. . to highlight the strengths of mri with diffusion weighted imaging (dwi) for malignant lesion detection and characterisation within bones, but also beyond bones. . to highlight the typical appearance of malignant lesions on projectional and cross-sectional imaging. . to learn to differentiate these lesions from normal marrow variations, stress lesions, bony pseudo-lesions and other benign conditions, and to become familiar with treatment induced changes within lesions and their environment. . to situate mri amongst other functional imaging techniques (pet, spect, ). colorectal cancer is the rd most commonly diagnosed cancer in the world with % of cases diagnosed in the developed world. with an estimated . million new cases clinically diagnosed and over , people killed worldwide by this disease on an annual basis, colorectal cancer is a true public health concern. survival of colorectal cancer is directly related to the extent of disease and specifically presence of liver metastases. imaging plays a key role in the initial staging of colorectal cancer and is the gold standard in evaluating extra colonic disease, primarily liver metastases. imaging is also widely used for therapy monitoring and staging. ct, mri, pet/ct and pet/mri are the key imaging modalities. in addition, tumour response therapy assessed with morphological and functional biomarkers is increasingly used by advanced gastrointestinal oncologic programs. furthermore, image-guided therapy is widely used for patients with unresectable lesions. liver colorectal metastases were the first liver metastases to be treated with image-guided ablation techniques. a gamut of ablative techniques exists ranging from transarterial embolisation, radiofrequency ablation, highly intensified focused ultrasound, brachitherapy etc. in short, diagnosing, characterising and finally assessing tumour response after neoadjuvant and image-guided therapy are all functions where imaging plays a pivotal role. this course is divided into three logical segments dealing with current treatment options, morphological and finally functional biomarkers and it is followed by a panel discussion which includes audience participation. a. current treatment options t.k. helmberger; munich/de (thomas.helmberger@klinikum-muenchen.de) hepatic metastases in colorectal cancer may occur in % to % of the cases. considering the general oncological (isolated hepatic tumour load; prognostic benefit), and technical (size, number, location of hepatic metastases; expected hepatic functional reserve) framework, surgical resection is still considered to be the method of choice -even if this statement never had been verified by rtcs. nevertheless, in clinical reality only - % of patients with liver metastases may qualify for resection. in consequence, the majority of patients need other or at least modified therapeutical pathways including adjuvant or neoadjuvant chemotherapy and more and more image-guided local ablative therapies. the latter encompass chemo-(transarterial chemoperfusion/-embolisation), thermo-(radiofrequency-, laser-, microwave ablation, high intensified focused ultrasound), and radio-ablative (radio embolisation, interstitial brachytherapy, etc). techniques. particularly, thermalablative techniques gained wide acceptance over the last years, since ample evidence could be presented showing that this method can be applied as primary and also complementary therapies in resectable and non-resectable metastatic disease. furthermore, recent data confirm that in multimodality therapy concepts, progression free survival and overall survival in patients with primarily unresectable and with unfavourable prognosis is comparable to surgery with -year survival rates more than %. minimally invasive, imageguided therapies will not replace surgical resection; however, these therapy modalities are eligible in a large number of cases and should be implemented consequently in multimodality treatment regimens according to the interdisciplinary consensus of oncologists, interventional radiologists, and surgeons. colorectal liver metastases are typically identified as low attenuation lesions on portal venous phase ct. planning of optimal therapy depends on accurate localisation and characterisation of all focal liver lesions and this is best achieved with mri, which should include diffusion-weighted imaging and use of hepatocyte-specific contrast medium. the typical mri signature of colorectal metastases includes low signal on t w, moderately increased signal on t w, irregular peripheral arterial phase enhancement with low signal on portal venous phase, restricted diffusion and absent hepatocytes. morphological tumour response can be assessed with either modality. size-based systems for assessing tumour response are widely applied, with recist . the most widely used at present. recist . includes several modifications that make it more user friendly than recist . . more advanced morphological criteria have been described for new targeted and molecular therapies, including overall attenuation, the tumour-liver interface and the appearance of the peripheral rim of enhancement. as hepatic surgeons become more aggressive in their approach to resection in patients with liver metastases, it is important to understand that disappearance or calcification of liver metastases after treatment does not necessarily equate to a complete pathological response. the timing of imaging is therefore critical in assisting the surgeon to remove all previously affected hepatic segments after chemotherapy. learning objectives: . to learn about the algorithm for detecting and characterising liver metastases. . to understand conventional imaging criteria for assessing tumour response. . to learn about the rationale for monitoring patients after radical and palliative treatments. conventional size measurement criteria remain the most widely used method to determine the response of colorectal liver metastases to treatment. however, tumour size reduction is assessed relatively late (e.g. weeks after treatment) and new targeted treatment may be effective without reducing tumour size. new functional imaging techniques can be applied to quantify different aspects of tumour biology and to develop response, and predictive and prognostic biomarkers. we discuss the use of diffusion-weighted mr imaging, dynamic contrast-enhanced mr imaging and fdg-pet imaging in the evaluation of treatment response in patients with colorectal liver metastases. digital breast tomosynthesis (dbt) is a promising technique for breast imaging based on a full-field digital mammography (ffdm) platform. the x-ray tube moves through a proscribed arc, and several low-dose projection images are acquired. images are reconstructed into a stack of mm slices. the radiation dose is about the same or slightly higher than for ffdm. the most important advantage of dbt is the elimination of superimposed tissue, which improves detection of lesions otherwise hidden by dense breast parenchyma and reduces the interpretation problems caused by overlapping tissue. dbt is suggested to improve sensitivity as well as specificity in women with dense breast. dbt is superior to ffdm in tumour size assessment and for mass visibility and cancer conspicuity. dbt is comparable to ffdm for evaluation of microcalcifications. dbt is superior for the detection of spiculated masses and architectural distortion. the potential to improve sensitivity and specificity is of interest for screening. an open question is whether dbt should be applied in one or two views. two-view dbt would mean a doubling of radiation dose, but a solution is the implementation of synthetic d images reconstructed from the d dataset of dbt. the synthesised images are created by summing and filtering the stack of reconstructed dbt slices. using synthetic d allows "combo mode" to be implemented in screening with the same radiation dose as for conventional d. results using dbt in breast cancer screening are very promising. different study design of published studies may explain the variations of results. magnetic resonance imaging of the breast provides a multitude of techniques for detection of breast cancer and differentiation of benign from malignant lesions. in the last few years, significant improvements of scanner hardware and equipment in terms of coils as well as sequence software have been achieved. to date, multiparametric quantitative measurement of perfusion, diffusion and tissue chemistry is possible at field strengths up to tesla. although higher field strengths and improved sequence protocols provide higher signal and allow faster imaging, specific technical issues have to be considered to avoid artefacts and pitfalls. this talk aims to demonstrate the possibilities and limitations of multiparametric and (ultra-)/high-field mri. furthermore, these new techniques are put into the broader clinical context to determine the potential value for breast lesion detection and differentiation. positron emission mammography is a recently developed imaging device using positron emission technology. after injection of a positron-emitting radiotracer, the radiation is detected by a dedicated high-resolution camera, providing high spatial resolution. commercially available systems include cameras where the breast is compressed and which display a d image similar in its layout to a mammogram and hanging breast devices where a d image similar in its layout to mri is produced. these devices have been compared to pet/ct and have shown higher spatial resolution, accuracy and sensitivity in detecting malignant breast lesions. the most widely studied tracer in pet/ct and in pem is fluorodeoxyglucose (fdg). fdg is a glucose analogue and has had a major impact on oncology imaging with pet/ct. breast carcinomas demonstrate increased glucose consumption and the intensity of the uptake has been correlated with prognosis, hormonal receptor and her status, ki and nodal involvement. current potential indications for pem are detection and characterisation of breast lesions, local staging in patients with known breast cancer and baseline assessment prior to neoadjuvant chemotherapy. future developments of pem will involve new tracers and fusion with other breast imaging modalities. fluorothymidine is a proliferation tracer that seems promising in early assessment of response to chemotherapy. fluoroestradiol has the potential to image oestrogen receptor status in vivo. the possibility of pem-guided biopsy is a very exciting development and has the potential to select lesions with the highest intensity of uptake. fusion with mri, us and mammography will increase the diagnostic accuracy. the presentation will focus on two frequent findings on mri with increasing incidence during ageing, notably "unspecific" hyperintense lesions on t /flair and cerebral mircobleeds (cmbs) on t * or swi images. clinicoradiologic studies suggest that these t /flair lesions are associated with increased risk of cognitive decline, stroke and death. the clinical relevance of these lesions probably depends on periventricular versus deep white matter location. radiologic-histopathologic studies correspondingly indicate that the degree of histopathologically confirmed demyelination also depends on the location of these t /flair lesions. cerebral mircobleeds (cmbs) are punctate hypointense lesions on t * or swi images. while multiple cmbs are associated with diseases such as cerebral amyloid angiopathy (caa), clinicoradiologic studies demonstrate increased incidence of sporadic cmbs in mild cognitive impairment (mci) and alzheimer dementia, indicating a degrading effect of cmbs on cognition. on the other hand, radiologic-histopathologic studies show that not all black spots on t * or swi correspond to cmbs. cmb mimics include micro-calcifications. the presentation will demonstrate typical manifestations of these "unspecific" t /flair lesions and cmbs during ageing and provide tips for the interpretation of these lesions during daily clinical routine. with the increasing use of mri, it has become more common to discover incidental brain findings. these findings may be non-specific or have some morphological characteristics and geographical distribution highly suggestive of a demyelinating disease. the term "radiologically isolated syndrome" (ris) was introduced by okuda in to describe those asymptomatic patients with radiographic abnormalities highly suggestive of multiple sclerosis (ms). sometimes, these lesions may even meet the mri criteria for dissemination in space, which are currently used to predict future development or conversion to ms in patients presenting a clinically isolated syndrome (cis). however, we must not forget that these radiological criteria should be applied only when the patient had experienced any clinical manifestations suggestive of a demyelinating disease. therefore, what happens in those patients in whom we found highly suggestive lesions but have never had symptoms? due to lack of knowledge about the natural history or evolution of this new syndrome, clinical and therapeutic management of these patients is not well established, nor is the risk of conversion to ms. recent data, however, indicate that the presence of gadolinium-enhancing cervical cord and cortical lesions significantly increases the risk of conversion to cis or ms. all these data have improved the characterisation of ris subjects and in our understanding of risk factors for initial symptom development. incidental findings on brain imaging are defined as previously undetected abnormalities of potential clinical relevance that are unexpectedly discovered and unrelated to the purpose of the imaging. incidental findings are increasingly detected in clinical practice, with screening, and in the research setting. data on the prevalence of these abnormalities are scarce, the clinical course of the findings is often unknown, and the management of such lesions is not clear. the prevalence of incidental findings can be expected to vary depending on the purpose of the imaging exams. with improvements in imaging technology (higher field scanners, new pulse sequences), the number of detected incidental findings will increase dramatically. another important point is the advent of imaging biomarkers. advanced techniques for postprocessing and analysis, such as automated segmentation of brain structures or voxel based morphometry, will lead to the discovery of imaging biomarkers. once the predictive value of these markers has been established, most mri studies of the brain, both in the clinical and research setting, will reveal information that might be relevant for the well-being of patients or participants. although still incidental, these findings can unfortunately no longer be considered unexpected. we will soon face large medical, ethical, and practical problems as a result of technical improvements. in this lecture, the most important incidental findings and their prevalence will be reported. the management of incidental findings in the clinical and research setting will be discussed. finally, recommendations for follow-up will be provided. the primary assessment of laryngeal cancer is provided by endoscopy, which is accurate in delineating both the superficial extent and functional impairment. imaging techniques are recommended to grade submucosal invasion, local and distant spread. in the last few decades, open and endolaryngeal surgical procedures as well highly sophisticated rt techniques have been developed to treat the tumour while preserving critical laryngeal functions. for example, early glottic tumours (tis-t ) can be treated by endoscopic laser excision or rt. the integration of information acquired by endoscopy and imaging is essential for proper treatment planning. the radiologist has to know that the site of origin (supraglottis vs glottis or subglottis) and local extent (superficial and submucosal) are the key tumour-related factors in planning the type of treatment. submucosal invasion has a variable impact depending on the location of the primary tumour. the key issues in glottic cancers include the invasion of: the posterior paraglottic space, crico-arytenoid joint, and anterior commissure and contralateral true vocal cord, and vertical spread towards supra and/or subglottis, thyroid or cricoid cartilage. the key issues in supraglottic cancers encompass the invasion of: base of the tongue, preepiglottic space, piriform sinus, vertical spread into glottis and subsequent cartilage invasion (observed exclusively in transglottic cancers). functionsparing surgical techniques -as supracricoid laryngectomies -can be applied in advanced cancers when sparing of the posterior commissure, the ipsilateral crico-arytenoid joint, the mucosa investing the upper cricoid and the outer perichondrium of the thyroid cartilage is demonstrated by endoscopic and imaging mapping. the detection of recurrent laryngeal carcinoma is often challenging. to better appraise the expected findings after initial surgery of laryngeal cancer, the classical partial laryngectomy and laser laryngeal surgery are briefly presented.(chemo)radiotherapy has become an important treatment modality for laryngeal carcinoma. the recurrence rate in t -t laryngeal carcinoma is reported to be between % and %. the difficulty in differentiation between radiation reaction (such as oedema, fibrosis and soft tissue and cartilage necrosis) and recurrent disease is discussed. a histologic study on wholeorgan sections of recurrent glottic carcinomas is also presented: recurrent disease presents with different tumour spread than does a primary carcinoma, resulting in difficulties in estimating the extent of tumour recurrence. currently, selected recurrences of early glottic carcinoma are, whenever possible, treated by larynx-preserving salvage surgery. in those cases, a precise evaluation of the recurrent tumour extent is mandatory for planning larynx-preserving salvage surgery. this is especially true in critical regions such as the preepiglottic space, infiltration into cartilage, contralateral tumour spread and subglottic tumour extension. the accuracy of crt-classification is relatively low: many patients with signs and symptoms suggesting tumour recurrence present with post-radiotherapy changes; small tumor foci, often localised in the subglottic region, are undetectable by imaging studies. tissue changes in the neck treated by surgery and/or radiotherapy (rt) make the detection of residual or recurrent tumour more difficult. clinical evaluation of the neck is also hampered by these changes. rt-induced fibrosis and postsurgical scar tissue make palpation of the neck very difficult. endoscopy is hampered by endolaryngeal oedema which can persist for many months post-rt. therefore, any (non-invasive) method helping in the detection of recurrence is welcome. patients at risk for local failure after rt can be successfully identified by a post-rt ct (or mr) study between to months after rt. the optimal time point to perform such a 'baseline study' seems to be about to months post-rt. patients with indeterminate findings are candidates for 'imaging surveillance'; i.e. follow-up imaging every to months up to a period of years after rt. however, ongoing studies suggest that metabolic imaging (fdg-pet) may detect local recurrences with a higher accuracy than 'conventional' anatomically based imaging techniques, such as ct and mr. ct and/or mr-findings in the treated neck are frequently inconclusive. at present, new techniques are available to detect recurrent cancer. pet-ct is widely applied in the post-treatment setting. also, advanced mr-techniques such as diffusion weighted imaging (dwi) and dynamic contrast-enhanced mr (dce-mr) can be applied. focusing on the larynx, the advantages of these techniques as well as the limitations will be shown using imaging examples from daily practice. during the last years, several hundred new radiotracers for pet imaging of cancer have been developed and tested preclinically. these tracers are based on proteins (e.g. antibodies and antibody fragments), peptides (e.g. g proteincoupled receptor ligands) or small molecules (e.g. radiolabelled protein kinase inhibitors or metabolic substrates). using these radiotracers, the expression and function of a variety of proteins can be studied in patients, including for example glutamine/glutamate metabolism, neutral amino acid transport, psma, grpr, sstr, cxcr , folate and alpha-v beta- integrin expression. some of these radiopharmaceuticals are potential companion diagnostics and may allow selection of patients for treatment with radiopharmaceuticals or targeted drug conjugates. preclinical and initial clinical data are promising for several new radiotracers, but the key challenge is the clinical validation and dissemination of these compounds. validation and dissemination have been hampered so far by regulatory hurdles as well as by the lack of standardised trial designs for the validation of imaging agents and the lack of accepted and appropriate end points to prove clinical utility. addressing these issues will be critical for the future clinical use of pet and other molecular imaging technologies. imaging targets in cancer range from simple size measurements to more specific biomarkers on functional, cellular, metabolic and molecular levels. as our understanding of basic tumour biology has advanced, techniques have been developed to exploit this information to produce increasingly specific molecular imaging tools. the biodistribution of these molecular imaging probes should be more specific in diagnosing and assessing cancer than the morphological information acquired using anatomical imaging alone. this lecture will discuss current and emerging functional and molecular imaging techniques using mri and their applications in oncology. functional measures of tumour blood flow and vascular permeability can be made using dynamic contrast-enhanced mri. diffusion-weighted imaging is a surrogate for the cellular content of the tumour and emerging methods can be used to probe features of the extracellular space such as tumour ph and stromal content. on the molecular level, cell surface expression of specific proteins and enzyme activity within the cell can be imaged; labelled probes have been developed which bind to these proteins and a new mr technique is being developed for assessing tumour glucose in a similar way to pet. hyperpolarisation methods are emerging to overcome the major limitation of mr: low sensitivity. one such approach is dynamic nuclear polarisation, which can probe carbon metabolism non-invasively in patients with cancer. functional and molecular imaging techniques with mri will increasingly be used in radiology in conjunction with anatomical imaging methods to improve diagnosis and prognosis, target biopsies, as well as predict and detect response to treatment. with mr-spectroscopy, diffusion-weighted imaging, pet-and spect-ct, as well as pet-mri, molecular imaging has already become part of clinical routine diagnosis. also, some contrast agents such as spio for mps-labelling and scavenger-receptor binding gadolinium chelates are clinically applied for liver and lymph node imaging. however, there are still a number of highly promising novel tools that are expected to emerge clinically in the near future. in this context, the first part of the talk will give an overview on how optical imaging basically works and which future clinical applications can be expected. in this context, raman spectroscopy, fluorescence optical reflectance imaging, fluorescence mediated tomography, photoacoustic imaging and cerenkov luminescence imaging will be addressed. the second part of the talk introduces molecular ultrasound imaging as a safe and preclinically wellevaluated method. besides the diagnostic use of molecular ultrasound contrast agents, which have been already evaluated in clinical trials, the potential of this method also for theranostic purposes will be highlighted. the assessment of the amount of arterial calcification with computed tomography is a standard method in the risk stratification of coronary heart disease. coronary calcium detection by ct has been shown to identify atherosclerotic plaque and to quantitatively assess coronary calcium. many studies have demonstrated the association between the degree of coronary calcium, the burden of atherosclerosis, and the risk for cardiovascular events associated with coronary calcium. the cac scoring can provide individual risk assessment and reclassify the low and particularly intermediate framingham risk cohort into lower-and higher-risk strata. the absence of cac is associated with a very low risk of future cardiovascular events, with modest incremental value of other diagnostic tests in this very low-risk group. the role of cac in the workup of symptomatic patients is under discussion and there is evidence that the absence of cac has a high negative predictive value for ruling out acute coronary syndrome. although cac measurement is highly standardised, some technical aspects have to be considered and all possibilities of dose reductions utilised. this presentation will show the technical and epidemiological fundamentals of cac scoring and discuss the clinical applications of the method. coronary cta has gained an important role in the evaluation of patients with chest pain, suspected of coronary artery disease (cad). in comparison to invasive coronary angiography, coronary cta with -multidetector ct has an excellent sensitivity for coronary stenosis. specificity for stenosis is less excellent, but has improved with newer ct generations. the strongest impact of coronary cta has been on the exclusion of stenosis in symptomatic patients at low-intermediate pre-test probability. in contrast to invasive angiography, coronary cta yields more information than just the severity of stenosis. coronary cta also allows determination of plaque types (calcified, non-calcified, partly calcified) and quantification of the atherosclerotic plaque burden. also, new developments point to the possibility to assess the haemodynamic significance of stenosis based on ct density measurements in the coronary arteries, or evaluation of myocardial perfusion. thus, coronary cta yields quantitative imaging biomarkers that could be used for risk stratification in the future. recent studies have shown that whether or not a patient has stenosis and plaque, as well as the type and burden of plaque, affects the risk of myocardial infarction and mortality. therefore, the impact of coronary cta imaging biomarkers reaches beyond the mere assessment of stenosis for symptomatic treatment and could potentially alter medical management of patients for improvement of long-term prognosis. a- : c. added value of carotid and peripheral artery imaging for risk assessment l. natale; rome/ it (lnatale@rm.unicatt.it) technical protocols optimisation of mra and cta of carotid and peripheral arteries is crucial for image quality and to complete the analysis of both lumen and vessel wall. furthermore, mra protocol needs to be integrated with dedicated pulse sequences in order to obtain vessel wall imaging that is already included in cta datasets. new ct and mr scanners allow this optimisation in terms of radiation exposure, contrast media dose reduction and spatial resolution. all these parameters will be discussed for both techniques. according to international guidelines, non-invasive imaging indications will be discussed, with particular attention to high risk population (e.g.: diabetic patients). some possible imaging biomarkers of plaque composition will be discussed. learning objectives: . to become familiar with optimal technical considerations for performing cta and mra of the carotid and peripheral arteries. . to understand clinical indications for carotid and peripheral cta and mra. . to discuss the role of cta and mra parameters including plaque imaging as biomarkers of cardiovascular disease. key: cord- - z vbldd authors: mühldorfer, kristin; speck, stephanie; kurth, andreas; lesnik, rené; freuling, conrad; müller, thomas; kramer-schadt, stephanie; wibbelt, gudrun title: diseases and causes of death in european bats: dynamics in disease susceptibility and infection rates date: - - journal: plos one doi: . /journal.pone. sha: doc_id: cord_uid: z vbldd background: bats receive increasing attention in infectious disease studies, because of their well recognized status as reservoir species for various infectious agents. this is even more important, as bats with their capability of long distance dispersal and complex social structures are unique in the way microbes could be spread by these mammalian species. nevertheless, infection studies in bats are predominantly limited to the identification of specific pathogens presenting a potential health threat to humans. but the impact of infectious agents on the individual host and their importance on bat mortality is largely unknown and has been neglected in most studies published to date. methodology/principal findings: between and , deceased bats of european species (family vespertilionidae) were collected in different geographic regions in germany. most animals represented individual cases that have been incidentally found close to roosting sites or near human habitation in urban and urban-like environments. the bat carcasses were subjected to a post-mortem examination and investigated histo-pathologically, bacteriologically and virologically. trauma and disease represented the most important causes of death in these bats. comparative analysis of pathological findings and microbiological results show that microbial agents indeed have an impact on bats succumbing to infectious diseases, with fatal bacterial, viral and parasitic infections found in at least % of the bats investigated. conclusions/significance: our data demonstrate the importance of diseases and infectious agents as cause of death in european bat species. the clear seasonal and individual variations in disease prevalence and infection rates indicate that maternity colonies are more susceptible to infectious agents, underlining the possible important role of host physiology, immunity and roosting behavior as risk factors for infection of bats. bats are among the most successful and diverse mammals on earth. approximately chiropteran species are found on every continent except antarctica and inhabit a multitude of diverse ecological niches [ ] . bats play essential roles in maintaining healthy ecosystems, as they act as plant pollinators, seed dispersers, and predators of populations of insects including harmful forest and agricultural pests [ ] . most bat species are listed in the iucn red list of endangered species and almost half of these are considered threatened or near-threatened [ ] . to estimate and prevent further population declines, research has been primarily focused on bat biology, ecology and behavior, while disease aspects were largely neglected [ ] . in the last two decades, the importance of chiropteran species as potential vectors of significant viral diseases especially in regard to zoonoses has received growing attention. besides bat rabies that has been studied for more than half a century, extensive research efforts identified a large number of microbial agents [ ] including important emerging zoonotic viruses detected in bats across the world [ ] [ ] [ ] [ ] [ ] [ ] [ ] . however, most studies are limited to the identification of microorganisms detected and investigations regarding infectious diseases and causes of death in bats are sparse [ ] [ ] [ ] [ ] . in europe, research is predominantly focused on european bat lyssaviruses [ , ] and coronaviruses [ , ] , but first indications of bat-pathogenic bacteria [ , , [ ] [ ] [ ] and novel viruses [ , ] isolated from deceased bats in germany and great britain were found. in this study, we provide new data on infectious diseases in european bat species, considering factors likely to affect the susceptibility of bats to infectious agents including effects of seasonality, individual and species-specific heterogeneities, and possible intra-and inter-species transmission dynamics. all bat species in europe are strictly protected under the flora-fauna-habitat guidelines of the european union (http://ec.europa. eu/environment/nature/legislation/habitatsdirective/index_en.htm) ( / /eec) and the agreement on the conservation of populations of european bats (www.eurobats.org) that prohibit invasive sampling of bats for research purposes. for the animals investigated in this study, carcasses of deceased bats found in germany were kindly provided by bat researchers and bat rehabilitation centers of different federal states. between and , a total of deceased bats of european vespertilionid species (i.e., family vespertilionidae) were investigated (fig. a , [ ] ). the bat carcasses originated from different geographic regions in germany, i.e. berlin greater metropolitan area (n = ), bavaria (n = ), brandenburg (n = ), lower saxony (n = ), thuringia (n = ), and baden-wuerttemberg (n = ), and were collected by bat researchers and bat rehabilitation centers. most animals represented individual cases that were found dead, injured or moribund near human habitation. thus, the species composition in this study predominately reflected the urban and suburban bat fauna, which is characterized by a disproportionate abundance of a few bat species (fig. a , [ , ] ). two groups of and adult noctules (nyctalus noctula), respectively, were collected from tree hibernacula destroyed during wood logging. a further group of deceased adult n. noctula originated from a colony that was trapped in a rain pipe in december. nine dead juvenile pipistrellus pipistrellus were collected from a nursery roost. if bats died in care or had to be euthanized for animal welfare reasons, the carcasses were immediately stored at uc and were shipped to the leibniz institute for zoo and wildlife research, berlin, germany, for diagnostic investigations. of all carcasses examined histo-pathologically, about % were suitable for bacteriological investigation. a lesser extend ( %) was also examined for selected viral agents at the robert koch institute, berlin, germany. in addition, a brain sample of each animal was submitted to the friedrich-loeffler-institute, wusterhausen, germany, for rabies diagnosis. a full necropsy was performed on each bat and all macroscopic findings including ectoparasite infestation were recorded. for histo-pathological examination, small slices of multiple organ tissues (i.e., lung, liver, heart, kidney, adrenal gland, spleen, intestine, pancreas, brain, tongue, larynx, salivary gland and pectoral muscle) and tissues conspicuous for pathological changes were fixed in buffered % formalin, processed using standard methods and embedded in liquid paraffin. sections were cut at - mm and routinely stained with hematoxylin-eosin (he). in addition, special histological staining methods were used depending on microscopic findings, i.e. for the detection of bacteria (gram or giemsa staining), fungi (periodic acid schiff or grocott's gomori methenamine silver nitrate staining), iron (prussian blue stain), mineralization (von kossa staining), connective and collagen tissue (trichrome staining). details on pathological results are published elsewhere [ ] . the causes of mortality were rigorously standardized with the primary cause of death identified for each bat as the most serious injury, disease or event subsequently fatal to the animal. to ensure independence of primary and contributing causes of death, the categorization was based on the severity of pathological findings. samples of lung, liver, heart and kidney, and tissues conspicuous for pathological changes (e.g. enlarged spleen) of bats were plated onto columbia ( % sheep blood), chocolate, gassner, and macconkey agar (oxoid, germany) and were incubated at uc (chocolate agar % co ) for - h. specific culture media and conditions for the isolation of yersinia, salmonella and anaerobic bacteria were used if appropriate. primary identification of bacterial strains was based on colony morphology, hemolysis, gram-staining, indol production, catalase and oxidase reaction. bacterial species identification was carried out using the relevant commercial api test system (biomérieux, germany). additional conventional biochemical tests [ , ] were applied to confirm api test results where necessary. in case of ambiguous biochemical test results, s rdna gene analysis was performed for final identification [ ] . salmonella isolates were characterized at the national reference laboratory for the analysis and testing of zoonoses (salmonella) at the federal institute for risk assessment, berlin, germany. identification and characterization of yersinia and pasteurella species have been reported earlier [ , ] . homogenized organ tissue of lung, liver, heart, kidney, spleen, brain and salivary gland of bats were pooled for each individual and used for rna/dna extraction and further molecular analysis by generic pcr assays detecting flavi- [ ] , hanta- [ ] , corona- [ ] , and influenza a-viruses [ ] . also, pcr assays specific for previously described herpesviruses [ ] from european vespertilionid bats were used. for this purpose, rna/ dna was isolated using the nucleospinh rna ii kit (macherey-nagel, germany) and the nucleospinh tissue kit (macherey-nagel), respectively, according to the manufacturer's instructions. because of limitations in sample volume, for out of the bats pcr assays could only be applied for different bat herpesviruses. internal controls were used for all pcr assays to test for inhibition. for confirmation, all retrieved fragments of bat herpesvirus-specific pcr assays were checked for sequence identity to previously published isolates [ ] . for detection of lyssavirus antigen in brain tissue the fluorescent antibody test (fat) using a polyclonal antirabies conjugate (sifin, germany) was used [ ] . fat-positive brain tissues were subject of virus isolation in murine neuroblastoma cell culture (na / ) using the rabies tissue culture infection test (rtcit) as described elsewhere [ ] . lyssaviruses isolated in cell culture were characterized using both a panel of anti-nucleocapsid monoclonal antibodies (mab) [ ] and partial sequencing of a fragment of the nucleoprotein gene after rna extraction using trizol (invitrogen, germany) essentially as described [ ] . genomic dna was extracted from organ homogenates using the nucleospinh tissue kit (macherey-nagel) according to manufacturer's recommendations. genetic identification of the bat species was performed by amplification and sequencing of a bp fragment of the cytochrome b (cytb) gene [ ] using primers fm up ( -ccc chc chc aya tya arc cmg art gat a - ) and fm down ( -tcr acd ggn tgy cct ccd att cat gtt a - ). in addition, for differentiation of the distinct pipistrellus species, p. pipistrellus and p. pygmaeus, a rapid multiplex pcr assay was performed as described by kaňuch et al. the bat data were categorized in regard to different explanatory numeric and factor variables, e.g. bat species, sex and age class. the variable 'age class' ranked between and with increasing age (i.e. neonates, juveniles, subadults, and adults) and was used as numeric variable. for endoparasitic analysis, we defined a level variable 'bat size' according to the body size of a certain bat species to reduce the degrees of freedom of the full model, i.e. large species (n. noctula, eptesicus serotinus, and vespertilio murinus), medium-sized species (e. nilssonii, plecotus auritus, myotis daubentonii, m. nattereri, and p. nathusii) and small species (p. pipistrellus, and m. mystacinus). to detect effects of seasonality, different activity periods were specified according to the date of sampling, i.e. hibernation period (november to march), post-hibernation period (april/may), maternity period (june to august), and swarming period (september/october). as dependent binary variable for the respective models we either classified the mortality cause being disease or not (i.e. trauma), or the presence-absence of bacterial, ecto-and endoparasitic infections. we formulated different hypotheses to test for individual and species-specific differences in disease susceptibility and infection rates: (a) disease-related mortality in bats is influenced by sex, age and species-specific differences, and degree of endoparasitic infection. (b) bacterial infection in bats is influenced by sex, age and species-specific differences, occurrence of traumatic injuries and cat predation. (c) ecto-or (d) endoparasitic infection in bats is affected by age, sex and species-specific differences. seasonal effects were not analyzed because of too many missing data points. because the long-term dataset was highly biased towards sampling procedure, preservation of bat carcasses and following diagnostic investigations, we split and filtered the full data into several subsets reflecting the different analyses (table ) . all statistical analyses were performed using the r software v. . . (r development core team , vienna, austria). we used the chi-square test for given probabilities to evaluate significant differences in the sex ratio among bats of different species. for hypotheses a and b, we used a generalized linear mixed modeling approach (binomial glmm using function lmer in library lme ) with bat species included as random effect. this variable had not been significant as fixed effect (results not shown), but from other studies we can assume that there are speciesspecific differences in susceptibility of bats to certain infectious agents and therefore included it as random effect. we further used generalized linear models (glm with logit link and binomial error structure; for datasets with bat species . individuals) to test for individual and species-specific differences in parasite infection rates (hypotheses c and d). we created a full model for each hypothesis (a-d) to examine multiple and interaction effects of the specified variables. to select the final model variables, we used a stepwise backward algorithm (function stepaic in library mass) based on akaike's information criterion (aic) [ ] . the daic of the final model was calculated relative to a random intercept model to demonstrate the effect size of the selected variables. results of the diagnostic analyses follow the full data splitting into several subsets (see section 'statistical analysis' in material and methods; table ). all sampled bats belonged to different genera (i.e. pipistrellus, nyctalus, myotis, eptesicus, plecotus, vespertilio, and barbastella) and european vespertilionid species (fig. a) . three bat species, the common pipistrelle (p. pipistrellus, n = ), the noctule bat (n. noctula, n = ), and the serotine bat (e. serotinus, n = ) constituted about % of all bat carcasses investigated in this study, whereas p. pygmaeus, nyctalus leisleri, myotis brandtii, m. bechsteinii, m. dasycneme, plecotus austriacus and barbastella barbastellus were represented in small numbers of to animals. the overall sex ratio was . males to female with significant species-specific differences (fig. b) . animals in their first year of life (neonates, juveniles, and subadults) represented one third ( . %, n = ) of bat samples (fig. c) . overall, we were able to assign a cause of death to % (n = ) of bats investigated in this study. two thirds of mortality were due to trauma (n = ) or disease (n = ), while almost % of bats had died of other non-infectious causes like pulmonary edema, dehydration and hypoglycemia (table ). in % (n = ) no significant pathological findings could be found. among the traumatized bats, additional mild (n = ), moderate (n = ) and severe (n = ) inflammatory organ changes were noted in one half ( . %) of individuals, and % of the bats revealed bacterial (n = ) and/or parasitic infections (n = ) ( table ) . of the bats considered as dying of disease, fatal bacterial (n = ), viral (n = ) and parasitic infections (n = ) were observed in %. besides, amniotic fluid aspiration was noted in a neonate noctule bat (n. noctula), and a juvenile common pipistrelle (p. pipistrellus) was euthanized because of severe forearm bone deformation. the remaining bats ( . %) revealed moderate to severe pathological changes of unknown etiology or unconfirmed bacterial or viral cause ( table ). based on the glmm analysis, significant age-and sexdependent differences (daic = . ) were detected between the general causes of mortality, disease and trauma ( table ). the disease presence in bat samples decreased continuously with increasing age. neonates and juveniles of both sexes were significantly more affected by disease than older age classes (table ; fig. a) . we also found a significant trend in diseaseassociated mortality between the sexes, with adult females displaying higher disease prevalence ( . %) than males ( . %) ( table ). no significant association was observed between a certain cause of mortality (i.e. disease or trauma) and severity of endoparasitic infection (daic = . , result not shown). the seasonal distribution of disease-related mortality cases (fig. b ) described a trimodal pattern, with peaks in spring (april), summer (june to august) and winter (december). the proportion of traumatized individuals also increased obviously during the summer months up to and including the swarming period, but was low during the rest of the year. about % (n = ) of bat samples were examined bacteriologically. among these, different bacterial genera with more than bacterial species were identified (table s ). predominant bacteria isolated were enterococcus faecalis ( . %, n = ), hafnia alvei ( . %, n = ), serratia liquefaciens ( %, n = ), and pasteurella multocida ( . %, n = ). in % (n = ) of bats no bacterial growth was observed at all. comparative bacteriologic and histo-pathologic analysis identified different bacterial species that were clearly associated with pathological lesions and/or systemic infection, found in % (n = ) of bats investigated bacteriologically ( table ) . members of the families pasteurellaceae (above all p. multocida) ( . %, n = ), enterobacteriaceae (various bacterial species) ( . %, n = ), and streptococcaceae (above all enterococcus spp.) ( . %, n = ) were predominant bacteria associated with disease. more than half ( . %, n = ) of bacterial infections were observed in bats with traumatic injuries. the glmm analysis revealed low sex-and age-dependent differences in bacterial infection (daic = . , result not shown). female bats ( . %) and adults ( . %) showed marginally higher prevalence of bacterial disease compared to males ( . %) and to other age classes ( . %), respectively. however, we found a strong influence of cat predation (daic = ) associated with bacterial infection in bats (table ). testing for human-pathogenic zoonotic viruses, no examined bat sample ( / ) was positive for influenza a virus, corona-, hanta-and flaviviruses, respectively. no inhibition of the pcr assays was notified. out of bats tested for rabies virus infection, serotine bats (e. serotinus) were positive for lyssavirus by fat and rtcit. the viruses were identified as european bat lyssavirus type (eblv- ) sublineage a, both using mabs and sequencing. applying bat herpesvirus-specific pcr assays, out of bats proved to be infected with of the previously described bat herpesviruses ( table ). the highest prevalence of % ( / ) was observed for bat gammaherpesvirus (batghv ) in common pipistrelle bats (p. pipistrellus), followed by bat gammaherpesvirus (batghv , . %) in nathusius' pipistrelle bats (p. nathusii) and bat gammaherpesvirus (batghv , . %) in noctule bats (n. noctula). co-infection with different bat herpesviruses were recognized in n. noctula ( . %) infected with batghv and batghv , and in one n. noctula ( . %) infected with batghv and batghv . batghv was not only detected in its initially specific host p. nathusii, but also in other bat species, i.e. n. noctula, myotis myotis and m. mystacinus. although the prevalence of batghv ( . %) and batghv ( . %) differed significantly within its migrating host n. noctula, no difference was observed between the sexes. two juvenile n. noctula were found to be infected with batghv . interestingly, for the sedentary bat species p. pipistrellus being infected with batghv , a considerably higher prevalence was observed in juvenile bats ( . %) resulting in an overall prevalence of % also without difference between adult male and female bats. ectoparasites (mites, fleas, and ticks) were noted in % (n = ) of bats, but a potential bias in ectoparasite numbers collected from dead animals in comparison to ectoparasite abundance on live animals has to be taken in account. female bats ( . %) were slightly more infested by ectoparasites than males ( . %), whereas in different age classes ectoparasite prevalence was almost balanced. the glm analysis revealed significant species-specific differences in ectoparasite infestation (daic = . , table ). most bat species revealed low ectoparasite prevalence (range . - . %), while almost % (n = ) of n. noctula were infested with mites and/or fleas (fig. a) . microscopic examination of organ tissues revealed endoparasitic infection in % (n = ) of investigated bats, involving different protozoan (families eimeriidae and sarcocystidae) and helminth parasites (trematodes, cestodes, and nematodes). helminthes were predominantly found in the gastro-intestinal tract of the bats, while in some animals, granulomatous organ lesions were associated with larval migration of nematode species. based on the glm analysis, clear age-and species-specific differences (daic = . ) were observed between infected and non-infected bats ( table ). the prevalence of endoparasitic infection in bat samples increased significantly with increasing age, whereas the increase in prevalence was more rapid between juveniles and subadults ( . %) compared to the older age classes ( . %). marginal differences were also observed between the sexes, with female bats showing slightly higher ( . %) endoparasite prevalence than males ( . %). regarding species-specific differences, large bats like n. noctula, e. serotinus and v. murinus revealed higher endoparasite prevalence compared to individuals of medium-sized or small vespertilionid species (table ; fig. b ). this study was based on a passive surveillance sampling strategy that inherently influences the composition of bats sampled for diagnostic investigations [ ] and might also effect the data presented on causes of death by ecological and anthropogenic impacts of urban landscapes [ ] . trauma and disease represented the most important causes of mortality in deceased bats from germany, which differ from results of previous studies [ ] [ ] [ ] where disease-related mortality often played a subordinate role. young bats and adult females were significantly more affected by disease, indicating that sex-and age-related disease prevalence in table . pathological findings and bacterial, viral and parasitic infections specified for the general causes of mortality, trauma and disease. bats are strongly correlated with the maternal season. this assumption is further supported by the distinct increase of diseaserelated mortality from june to august, which corresponds to the maternity period of central european bat species. similar seasonal prevalence patterns in bats have also been described for parasitic (e.g. [ ] [ ] [ ] [ ] ) and viral infections (e.g. [ , , ] ). in contrast, the increase of trauma-associated mortality cases from july to october resembles major behavioral activity patterns of european bat species (i.e. weaning, mating, pre-hibernal fat storage, and migration) [ ] and could therefore predispose bats to trauma. however, both seasonal peaks also coincide with time and locations where sick, injured or dead bats are more likely to be discovered as well as with the seasonal roosting behavior of bats adapted on urban habitats [ ] . the additional seasonal peaks of disease-associated mortality corresponded to the post-hibernal and the early hibernal period of temperate zone bats. currently, there is a lack of knowledge of bat immunology. it is known for other mammalian species that hibernation reduces the innate and adaptive immune response; likewise an increasing risk of infection could be assumed for hibernating bats [ ] . with the start of the hibernation season, large aggregations of bats originating from various colonies might enhance the risk of spreading infectious agents similar to maternity colonies. equally, the post-hibernal increase of disease-related mortality is suggestive for reduced immunity in association with prolonged fasting during hibernation. bacterial diseases have rarely been documented in bats. pasteurella spp., here identified in % of bats, were the predominant bacterial pathogens reported in european bats and infection appears to be strongly correlated with cat predation [ , , , ] . in our study, bacterial infections were confirmed in % of bats investigated bacteriologically. most of these bacterial isolates represented opportunistic pathogens that usually do not harm the host unless the immune system is weakened [ ] or preceding injury to natural host barriers (e.g. skin abrasion). primary bacterial pathogens like samonella enterica serovar typhimurium, s. enteritidis and yersinia pseudotuberculosis [ ] were identified in almost % of affected bats. some of the bacterial species (e.g. burkholderia sp., cedecea davisae and clostridium sordellii) are newly described in bats. nevertheless, bacteriologic analyses can markedly be influenced by post-mortem bacterial invaders, freezing and storage of bat carcasses and the inability to detect certain bacteria by routine culture methods, resulting in some bacterial species that might have escaped detection. we found a strong association between cat predation and bacterial infection in bats as almost one half of bats ( %) caught by cats were affected by bacterial disease. various bacteria can be transmitted via cat bites [ ] ; hence bats attacked by cats are likely to succumb to bacterial infection even if non-fatal injuries were present. this relation has been proven for p. multocida infections in european bat species [ , , , ] . on the other hand, bats already debilitated by disease might easier fall prey to predators like cats. consequently, bats may also act as vectors for zoonotic pathogens, as domestic cats could pass these infectious agents on to humans. such cross-species transmission events from bats to domestic animals are well documented [ , ] . for all tested human-pathogenic zoonotic viruses no infected bat could be detected in this study except lyssaviruses. bat rabies is the only bat transmitted zoonosis in europe that is known to have resulted in human cases [ ] . unlike in other mammals table . result of the final model variables corresponding to different analyses: (a) disease-vs. trauma-related mortality, and presence-absence of (b) bacterial, (c) ecto-and (d) endoparasitic infection. where lyssaviruses ultimately cause lethal rabies, in bats nonlethal lyssavirus infections may also lead to the development of immunity [ ] . with the detection of eblv- we confirm that this lyssavirus circulates among e. serotinus as previous studies showed [ ] . in germany, bat rabies is also caused by eblv- and bokeloh bat lyssavirus (bblv) [ , ] , but while the latter was recently isolated from m. nattereri, eblv- is associated with m. daubentonii and m. dasycneme [ ] . the apparent absence of eblv- and bblv in the sampled bats is likely due to the fact that lyssavirus infections have a very low incidence in bat populations [ ] and that the sample size was too limited, especially concerning the relevant species. there is a high prevalence for herpesviruses in different insectivorous bat species in germany (this study, [ ] ). most of the previously described bat herpesviruses have been detected in low numbers in more than one bat species [ ] . here, we observed a high species-specific prevalence among herpesvirusinfected bats, indicating that a certain type of european bat herpesvirus is primarily associated with a single bat species. this is supported by batghv and batghv that were again only identified in their initial hosts p. pipistrellus and p. auritus (both sedentary), respectively, underlining the typical strong speciesspecificity of mammalian herpesviruses. however, species' range overlap and close inter-species contacts in bat roosts may result in cross-species transmission and could explain the observed overcoming of the species barrier (this study batghv , [ ] ). interspecies transmission have also been discussed for other mammalian herpesviruses, i.e. bovine and equine herpesviruses (e.g. [ , ] ). furthermore, for rna viruses (i.e. rabies virus) phylogenetic distance between different host species and overlap in geographic range have recently been demonstrated as strong predictors of host shifts and cross-species transmission in bats [ ] . some of the bat species (i.e. n. noctula, p. pipistrellus, and p. nathusii) in this study appear to be more susceptible to herpesvirus infection. in n. noctula, different gammaherpesviruses (batghv , , ) with significant prevalence differences were recognized. such type-specific differences in prevalence between the phylogenetically distant viruses batghv ( . %) and batghv ( . %) within one bat species indicates co-evolutionary virus-regulated mechanisms. parasite infestation in wildlife often occurs without clinical effects, but severe infection can reduce host fitness either in terms of survival or reproductive success [ ] . most data on infection dynamics in bats came from parasite studies focusing on individual and seasonal variations in ectoparasite prevalence (e.g. [ ] [ ] [ ] ] ). host density, roost preference and movement pattern seem to be important factors explaining individual and speciesspecific parasite infestation rates in bats [ ] [ ] [ ] . in european vespertilionid species, female-biased parasite loads are most likely associated with host physiology and differences in roosting behavior [ , ] . we also found species-specific seasonal variations in ectoparasitic infestation, with n. noctula and m. daubentonii showing higher ectoparasite prevalence in spring and autumn compared to the breeding season (data not shown), which is in accordance with zahn and rupp [ ] . additional findings of our parasite analyses are distinct variations in ecto-and endoparasite prevalence in relation to bat species. bats primarily roosting in trees or nest boxes were more frequently infested with ectoparasites like n. noctula ( %) and m. daubentonii ( %) compared to other species (range - %) investigated in this study. high ectoparasite loads have generally been described in bats preferring enclosed roosts like burrows and cavities [ , ] , suggesting that structural characteristics and the microclimate of roosting habitats influence ectoparasite survival and re-infection of bat hosts. this assumption is in accordance with results of pearce and o'shea [ ] who found differences in ectoparasite prevalence and intensity in eptesicus fuscus in relation to environmental factors (i.e. temperature and humidity) of different roost sites. in contrast to these results, the endoparasite prevalence in european vespertilionid bats seems to be correlated with the body size of the bat species [ ] . species-specific variations in diet and prey selection could possibly effect endoparasite prevalence in insectivorous bats [ ] , as larger bats feed on insects of a wider size range including hard-bodied prey [ , ] . this assumption is supported by the clear prevalence increase in subadult and adult bats compared to low endoparasite infection rates in juveniles primarily feeding on milk. a multitude of publications is restricted to pathogen presence or absence in different chiropteran species; here we demonstrate the impact of diseases and infectious agents on bats themselves. alongside to trauma-associated mortality and undefined mortality cases, disease aspects represented one third of mortality causes in investigated bats of european vespertilionid species. by comparing pathology and bacteriology results, we were able to detect different bacterial species that were clearly associated with disease in bats. at least % of all bats had died due to bacterial, viral and parasitic infections. finally, we found clear seasonal and individual variations in disease prevalence and infection rates, indicating an increased susceptibility to infectious agents in female bats and juveniles during the maternity season. our data emphasize and provide the basis for disease related studies in bat species on population level to elucidate the complexity of the ecology of infectious agents and host species likewise. table s bacteria isolated from bats found in germany. (doc) the status of the world's land and marine mammals: diversity, threat and knowledge ecosystem services provided by bats a review of the global conservation status of bats emerging diseases in chiroptera: why bats? ecological and behavioral methods for the study of bats. nd edition the isolation of hendra virus from pteropid bats: a natural reservoir of hendra virus isolation of nipah virus from malaysian island flying-foxes fruit bats as reservoirs of ebola virus bats are natural reservoirs of sars-like coronaviruses bats as a continuing source of emerging infections in humans isolation of genetically diverse marburg viruses from egyptian fruit bats bats, emerging infectious diseases, and the rabies paradigm revisited veterinary advances in the investigation of wildlife diseases in britain causes of morbidity and mortality in british bat species and prevalence of selected zoonotic pathogens. thesis for msc in wild animal health infectious and emerging diseases of bats, and health 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dog and cat bites european bat lyssavirus transmission among cats human rabies due to lyssavirus infection of bat origin first isolation of eblv- in germany novel lyssavirus in natterer's bat bat rabies serological survey of herpesvirus infections in wild ruminants of france and belgium new hosts for equine herpesvirus host phylogeny constrains cross-species emergence of rabies virus in bats behavioral adaptations to parasites: an ethological approach relationships between roost preferences, ectoparasite density, and grooming behaviour of neotropical bats roosting habits of bats affect their parasitism by bat flies (diptera: streblidae) ectoparasites in an urban population of big brown bats (eptesicus fuscus) in colorado when parasites become prey : ecological and epidemiological significance of eating parasites the implications of food hardness for diet in bats prey consumed by eight species of insectivorous bats from southern illinois the authors would like to thank berliner artenschutz team-bat-e.v., f. key: cord- -d se v authors: aguiar de sousa, diana; van der worp, h bart; caso, valeria; cordonnier, charlotte; strbian, daniel; ntaios, george; schellinger, peter d; sandset, else charlotte title: maintaining stroke care in europe during the covid- pandemic: results from an international survey of stroke professionals and practice recommendations from the european stroke organisation date: - - journal: eur stroke j doi: . / sha: doc_id: cord_uid: d se v introduction: the coronavirus disease (covid- ) pandemic has been placing an overwhelming burden on health systems, thus threatening their ability to operate effectively for acute conditions in which treatments are highly time sensitive, such as cerebrovascular disorders and myocardial infarction. as part of an effort to reduce the consequences of this outbreak on health service delivery to stroke patients, the european stroke organisation has undertaken a survey aimed at collecting information on the provision of stroke care during the pandemic. methods: cross-sectional, web-based survey, conducted from march through april among stroke care providers, focused on reorganisation of health services, the delivery of acute and post-acute stroke care and the availability of personal protective equipment. results: a total of stroke care providers from countries completed the survey, most of whom worked in europe (n = , %) and were stroke physicians/neurologists (n = , %). among european respondents, ( %) reported that not all stroke patients were receiving the usual care in their centres and ( %) estimated that functional outcomes and recurrence rates of stroke patients would be negatively affected by the organisational changes caused by the pandemic. the areas considered as being most affected were acute care and rehabilitation. most professionals had to adapt their activities and schedules and more than half reported shortage of protective equipment. discussion: strategies to maintain availability of stroke care during the covid- outbreak are crucial to prevent indirect mortality and disability due to suboptimal care. conclusion: european stroke organisation proposes a set of targeted actions for decision makers facing this exceptional situation. stroke, health services, covid- , stroke care date received: april ; accepted: may the coronavirus disease (covid- ) outbreak has been spreading rapidly around the world and places an overwhelming burden on emergency systems, health-care facilities and health-care workers. also, governmental instructions together with the response pattern of the population, due to fear of infection in medical facilities, have led to a situation where several elective consultations and procedures have been postponed. resource restrictions for urgent health conditions, such as stroke and myocardial infarction, may have a significant impact on mortality and morbidity, potentially even larger than that of covid- disease itself. an increase in mortality associated with treatable conditions has been demonstrated during previous viral outbreaks such as the influenza a h n virus, in which a greater surge in hospital admissions was associated with significant increases in the mortality attributable to other diseases, including stroke and acute myocardial infarction. , currently, data are not available regarding the impact of the pandemic on access to and delivery of stroke care in europe. as part of the ongoing effort to reduce the consequences of covid- on health service delivery to stroke patients, the european stroke organisation (eso) has undertaken a survey aimed at collecting information on the current provision of stroke care. these results should provide relevant information to direct activities aimed at promoting the best possible care to all stroke patients during the pandemic and support stroke physicians and other professionals working in the field. a cross-sectional, web-based survey for stroke care providers was administered from march through april . as of march , , covid- cases and , covid- -related deaths had been reported in the european region, with , deaths reported from italy alone and a total number of infected people in spain already exceeding those reported in china. a -item draft survey instrument comprising closed-ended questions and two open-ended questions was developed by the first and senior authors and circulated and tested among the members of the executive committee of eso. refinements were made as required to facilitate better comprehension and to organise the questions. the provider used for survey application and server capacity was lamapoll (http://lamapoll. de). the final survey was made accessible through a link that was distributed by email to all eso members and advertised in the official social media channels of the society. eso is pan-european scientific non-profit organisation of stroke researchers and physicians, national and regional stroke societies and lay organisations that was founded in and had members at the time of this survey. before starting, participants were informed of the purpose of the study, target respondents and confidentiality. participants had to confirm they wished to submit their final responses at the end and a message acknowledging successful completion of the questionnaire was sent by the server. cookies and ip address analysis were used to identify potential duplicate entries from the same user, which were avoided by preventing users to access the survey twice. total completion time was recorded. the survey was anonymous and confidentiality of information was assured. participation in this survey was voluntary and was not compensated. the data that support the findings of this study are available from the corresponding author upon reasonable request. we focused on the reorganisation of health services and reallocation of professionals, delivery of acute and post-acute stroke care and availability of personal protective equipment. open-ended text fields for comments and suggestions were also included. a copy of the questionnaire is provided in full in the data supplement. demographic data were self-reported by the participants, including occupation, type of hospital and geographical location (country). the world health organisation definition of the european region was applied. responses from stroke care providers working outside of the european regions were excluded from the primary analysis but provided as supplementary material. because italy and spain were the most affected countries in europe at the time of the survey, sensitivity analyses excluding respondents from other countries were performed for items related with delivery of stroke care and availability of personal protective equipment. data are presented as numbers and percentages. data analysis was performed using microsoft excel for mac . the survey was completed by participants from countries on six continents. no responses were excluded. table shows the demographic and occupational characteristics of the european participants. the distribution per country is described in supplementary table . most of the respondents were stroke physicians/neurologists (n ¼ , %) from europe (n ¼ , %) and working at tertiary hospitals (n ¼ , %). the remaining participants were interventionalists ( %), rehabilitation physicians ( %), allied health-care professionals ( %) and other professionals working in the field (resident physicians, emergency physicians and intensivists; %). among european respondents, ( %) were stroke physicians/neurologists (table ) and ( %) reported having treated patients with stroke and covid- . a summary of the responses from the participants working outside europe is provided in the data supplement. among stroke care providers working in the europe, ( %) reported that not all stroke patients were receiving the usual care in their centres, with ( %) estimating that this was happening in more than one quarter of patients (table ) . of the respondents from italy and spain, ( %) reported being unable to provide the usual care to all stroke patients ( figure ). two hundred sixty-six european participants ( %) estimated that functional outcomes and recurrence rates of stroke patients would be affected by the changes in stroke care related to the covid- outbreak. the areas of stroke care considered as being the most affected by the current situation were rehabilitation (n ¼ , %) and acute stroke care (n ¼ , %). for topics related to medical management of stroke patients, only responses from stroke physicians or neurologists (n ¼ ) were included ( table ) ; ( %) european participants reported that stroke code pathways were affected at their centres. also, ( %) reported that their centre avoided admitting patients whenever possible and ( %) described lack of beds for stroke patients, while ( %) had been forced to direct stroke patients to other hospitals. about one in five physicians reported that several basic ancillary examinations were no longer available (n ¼ , %). closure of transient ischemic attack clinics was described by european respondents ( %). for questions related to endovascular treatment, we considered responses provided by stroke physicians/ neurologists and interventionalists. in europe, ( %) reported problems in endovascular treatment, particularly that this was not possible when there was a need for intensive care (n ¼ , %) or that they were no longer able to provide this treatment at their centre at all (n ¼ , %). the most common comment included in the openended text fields was the perception of a clear drop in hospital admissions for stroke and later arrival during this period. about two-thirds of european participants reported changes in their working situation, either related to new activities or modifications in the work schedules ( table ) . sixty-two ( %) described extended working hours due to a lack of personnel and ( %) reported the need to contribute to new tasks outside stroke care. almost one of every five professionals was doing most of the work from home. compared with those working in tertiary care centres, participants working in community hospitals were more likely to have similar schedules and activities as before the outbreak and less likely to work from home (supplementary figure ) . several specific strategies for team management were reported, including organisation of separate teams for different activities and rotation schemes that include periods of isolation. shortage of personal protective equipment was reported by more than half of the european respondents (n ¼ , %) and by % of the stroke care providers from italy and spain. this cross-sectional survey including stroke care providers from europe revealed profound changes in delivery of stroke care early during the early phase of the covid- pandemic. these answers reflect the participants' experiences and perspectives at a specific point in time during the outbreak in which the infection was rapidly spreading. only % of respondents considered that, in their centres, all stroke patients were still receiving the usual acute and post-acute care, and more than % estimated that functional outcomes and recurrence rates of stroke patients would be affected by the changes in stroke care related with the outbreak. european stroke care providers reported their most significant challenges centred on acute care and rehabilitation. although most professionals adapted their activities and schedules, substantial challenges were noted in maintaining the pathways for acute stroke patients and the quality of inpatient care, particularly in having sufficient resources to guarantee availability of beds for all patients and proper etiologic investigation. acute reperfusion strategies, secondary stroke prevention and rehabilitation, among several other interventions, have the potential to provide a significant and long-lasting benefit to patients with stroke, and the targets for the implementation of these treatments have been set before. [ ] [ ] [ ] [ ] even in a setting of increasingly limited resources, efforts to preserve essential services and to provide the maximum benefit for the population have to be made. importantly, rehabilitation should be also included in decisions regarding which health services are essential, as delayed access to these therapies can compromise health and functional outcomes. to overcome these demands, local and national health managers need to carefully plan the extent to which acute and post-acute stroke services should operate during the pandemic peaks and how their continuity can be maximised, mitigating the risk of a collapse of stroke care. also, multidisciplinary collaboration should be maintained to ensure a smooth workflow, as is the case with anaesthesia and intensive care for patients receiving endovascular treatment for ischaemic stroke. one in five respondents had treated patients with stroke and covid- . the current evidence suggests that covid- often triggers a strong inflammatory reaction that may predispose to ischaemia and thrombosis. as this combination of diseases is likely to become more frequent, it is important to ensure that effective reperfusion therapies and stroke unit care are also available for these patients, in parallel with a safe and efficient medical environment. also, proper evaluation should not be delayed, regardless of concerns about possible infection, and confirmed patients with covid- should be transferred to the designated medical institutions for further optimal medical treatment when indicated. shortage of personal protective equipment was a common concern, as it was reported by more than half of the respondents. stroke teams are in the frontline of the pandemic, meeting patients in the acute setting with unknown covid- status. establishing protected stroke code pathways , will contribute to adequate acute management of stroke patients with confirmed infection or unknown covid- status. furthermore, as highlighted by several respondents, it is crucial to communicate with the community so that people know they can continue to safely seek appropriate care when stroke symptoms ensue, and that this is critical. this report was not meant to be all inclusive of the undergoing changes and difficulties faced by health-care providers, hospitals and other health-care systems to maintain delivery of stroke care during this outbreak but to raise awareness among decision makers of the importance of preserving capacity to provide appropriate care to stroke patients during the covid- pandemic. after consideration of these key issues, eso reinforces the following recommendations: • stroke care is an essential health service and should be prioritised in the strategic planning to manage the demands related to the response to the covid- outbreak. • the general population should be informed that stroke is an emergency and treatment is still available, so they must continue calling emergency services immediately in case of suspected stroke. public education campaigns can be an effective way of raising awareness. • acute stroke teams are frontline workers. patients with unknown covid- status should be evaluated under 'protected stroke code' and therefore access to appropriate personal protective equipment is mandatory for all team members as well as clear protocols for individual protection. • stroke registers and researchers must deploy resources to evaluate the effects of the covid- pandemic on case volumes, time metrics and clinical outcomes. we suggest that the available data sources are used to assess changes in number hospital admissions, baseline characteristics, in-hospital workflow metrics, treatment rates and functional outcomes of stroke patients during this period. • as the outbreak progresses over time, countries should be able to increasingly resume post-acute care services for stroke patients, such as rehabilitation. as the current situation is likely to influence the organisation of health care for the next months or years, this may require the adaptation and expansion of rehabilitation facilities and staff to meet the communities' needs, both now and in the longer term. moreover, strategies to develop and implement telehealth services should be promoted and supported by health authorities. this study has several limitations. first, most participants were from tertiary centres and countries are not equally represented, limiting the scope of the conclusions and the generalisation of these findings. second, the survey was carried out during a short period and lacked longitudinal follow-up. because of the increasing dissemination of the infection, health-care strain may worsen and the long-term effects remain unknown. third, we were unable to distinguish whether the respondents worked in the same hospital or region or in different regions. finally, response bias may exist, and non-respondents may either be more likely to work in the most affected regions and not have the time to participate, or work in the least affected regions. strain of health services during the covid- pandemic has been causing major disruptions to stroke care in europe, with likely serious and long-term implications. shortage of personal protection equipment has been common among stroke care providers. efforts to maintain stroke teams and safe provision of stroke care, including reperfusion treatments, should be prioritised. the author(s) declared no potential conflicts of interest with respect to the research, authorship or publication of this article. the author(s) received no financial support for the research, authorship, and/or publication of this article. not required. the corresponding author (ecs) affirms that this is an honest, accurate and transparent account of the study being reported; that no important aspects of the study have been omitted; and that any discrepancies from the study as planned have been explained. all authors had access to the data in the study and can take responsibility for the integrity of the data and the accuracy of the data analysis. contributorship das and ecs initiated and coordinated the study. das, hbw, vc and ecs contributed to the development of the survey. das performed the principal analysis and drafted the manuscript. all authors interpreted the data and revised critically the manuscript for important intellectual content. a novel coronavirus from patients with pneumonia in china impact of the fall influenza a (h n ) pdm pandemic on us hospitals estimated global mortality associated with the first months of pandemic influenza a h n virus circulation: a modelling study coronavirus disease (covid- ) situation report - (data as reported by national authorities by stroke unit treatment. -year follow-up effect of treatment delay, age, and stroke severity on the effects of intravenous thrombolysis with alteplase for acute ischaemic stroke: a meta-analysis of individual patient data from randomised trials endovascular thrombectomy after large-vessel ischaemic stroke: a meta-analysis of individual patient data from five randomised trials action plan for stroke in europe covid- : operational guidance for maintaining essential health services during an outbreak: interim guidance world health organization what is the evidence for physical therapy poststroke? a systematic review and meta-analysis clinical course and risk factors for mortality of adult inpatients with covid- in wuhan, china: a retrospective cohort study protected code stroke: hyperacute stroke management during the coronavirus disease (covid- ) pandemic european society of minimally invasive neurological therapy (esmint) recommendations for optimal interventional neurovascular management in the covid- era the authors acknowledge the advice and useful comments of prof. martin dichgans in the development of this survey. the authors thank daniela niederfeld for her administrative support. supplemental material for this article is available online. key: cord- -bo pr s authors: nan title: medical conferences in the post-covid world: a challenge, and an opportunity date: - - journal: eur radiol doi: . /s - - - sha: doc_id: cord_uid: bo pr s nan the current covid- pandemic has struck humanity like a biblical or historical plague, with a rapidity rarely seen even during those previous events. from apparent beginnings in the far east at the end of , it has spread throughout the globe, causing over million confirmed cases (with a much-higher probable prevalence) and over , deaths at the time of writing (april , ). protective measures (personal and societal) to reduce the impact of the pandemic have impacted greatly on daily life in most parts of the world and have already produced enormous economic consequences for individuals, businesses and countries, with more to come. uncertainty regarding the duration and geographical spread of the pandemic, the time needed to control it and the future availability of a cure or vaccine makes precise future planning impossible in virtually all aspects of life. we truly are living in "interesting times". what is already evident is that the life changes we have experienced in recent months will be with us in some form for a long time, and that life after the pandemic will be different from before. we will have to adapt to a "new" normality in behaviours and relationships, informed by this experience. in some ways, current protective advice has roots in long-distant learning. during the - bubonic plague in england, the inhabitants of the derbyshire village of eyam were persuaded by their clergyman, rev. william mompesson, to quarantine themselves for months to prevent the disease (which had spread from london to eyam via fleas trapped in cloth) from spreading further. more than of their population of died, whole families were wiped out, but the disease was contained, failing to reach nearby large populations in sheffield and manchester [ ] . healthcare professionals have been particularly affected by covid- , managing the difficult task of caring for those affected by the disease, and leading the way in advising governments on necessary actions. there is much we still do not know about the causative coronavirus, and the spectrum of disease it causes. as time goes on, we are learning that, while most cases present with flu-like symptoms (or suffer none at all), and a small proportion progress to severe respiratory distress, many other body systems and organs may be affected [ ] [ ] [ ] . robust data about pathologic changes induced by the infection are still incomplete, and the pathophysiology of the disease process is not yet fully understood. and, of course, healthcare professionals have suffered a disproportionate death rate from covid- , resulting from their direct role in patient contact and care. societies and patients owe a great debt of gratitude to those colleagues whose lives have been lost in the service of others. radiology has been affected by the pandemic in many ways. our departments have rapidly implemented workflow changes to ensure timely service for acutely presenting patients, to protect other patients and staff from infection and to facilitate hospital-wide pathways designed to separate covid- patients from non-infected ones [ ] [ ] [ ] . like all involved clinicians, radiologists have had to learn the features of the disease as they apply to our practice, including typical and atypical radiologic findings, appropriate use of imaging techniques (depending on available resources) and a correlation of findings with other clinical and laboratory data [ , , ] . the effects on scientific societies have been dramatic. planned activities have had to be changed at short notice, new covid- activity has had to be arranged quickly and most have been obliged to alter, postpone or cancel planned meetings. nor is this confined to academia. the first large gathering cancelled in europe was the geneva motor show, scheduled for march to , . one of the earliest medical conferences affected was the european congress of radiology, which was planned for march to , (and which will now take place in an online-only format from july to , ). many other conferences have been similarly affected; no one yet knows when face-to-face conferences can safely resume, and what restrictions may still be necessary when they do. the main goal of any medical society is to promote and facilitate collaborative work and growth; this goal cannot be met if members cannot meet (face-to-face or virtually). a recent editorial by evens outlines the many aims of professional meetings: education and research, policy-making, career and professional skills development, marketing by participating companies and interaction among colleagues at the meeting itself and at associated social functions [ ]. how can we achieve all, or most of these aims if it is not possible to meet in person? it is feasible to switch from an on-site to an online meeting format and still meet most of the goals of a "conventional" medical conference, although it is challenging. some societies are already attempting this: the american college of cardiology held its virtual meeting from march to , , and the european society of radiology and european association of urology will hold online conferences in july . many of the necessary techniques are well established. recording and streaming of conference sessions for later on-demand viewing has been offered by some societies (including the esr) for some years. live webinars are common educational tools (and have been offered by the esr on covid-related topics since march ). however, a full congress is a more-complex proposition, involving a variety of session types, aimed at diverse types of attendees, with many different forms of interaction between speakers and delegates. future online (or hybrid) conferences must find or invent new methods of allowing discussions and interactions involving remote participants during all sessions, to create the same sense of community and collective learning that can be felt during successful on-site events. this will not be easy, but making this necessary shift successfully will establish a new standard for what defines medical conference success in the future. the covid- pandemic has changed how humans interact with each other, and these changes are likely to persist, at least for some years. if scientific societies wish to remain relevant to their members and successful, they must adapt to this new reality. scientific meetings must change their structure to survive, and successful societies in the future will be those which embraced this new circumstance, and adapted early. future conferences will be different from those of yore, and we must learn not to judge them by old standards. the mould is broken; the new, online ecr will be a different meeting, with a different structure and different outcomes. it will be the first, and the best (so far) of a new generation. eyam: the village that sacrificed itself to the plagueand the lessons it can teach us. the independent covid- and the digestive system neurologic manifestations of hospitalized patients with coronavirus disease cardiovascular complications in covid- adapting to a new normal? key operational principles for a radiology service facing the covid- pandemic covid- patients and the radiology department -advice from the european society of radiology (esr) and the european society of thoracic imaging (esti) the radiology department during the covid- pandemic: a challenging, radical change ct features of sars-cov- pneumonia according to clinical presentation: a retrospective analysis of consecutive patients from wuhan city diagnostic performance of ct and reverse transcriptase-polymerase chain reaction for coronavirus disease : a meta-analysis evens rg ( ) the impact of a pandemic on professional meetings. radiology: imaging cancer publisher's note springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations key: cord- -wu tuvy authors: katz, jonathan b.; shafer, amy l.; eernisse, kenneth a.; landgraf, john g.; nelson, eric a. title: antigenic differences between european and american isolates of porcine reproductive and respiratory syndrome virus (prrsv) are encoded by the carboxyterminal portion of viral open reading frame date: - - journal: vet microbiol doi: . / - ( ) -b sha: doc_id: cord_uid: wu tuvy antigenic differences between european and american isolates of porcine reproductive and respiratory syndrome virus (prrsv) were revealed by serologic analysis of a recombinant protein derived from prrsv open reading frame (orf ). the hydrophilic carboxyterminal amino acids encoded by the orf of a european (lelystad) isolate of prrsv were expressed as a recombinant fusion protein (bp -p) in a baculovirus gene expression system. sera from gnotobiotic swine exposed to prototypic reference european and american isolates of prrsv and sera from conventionally reared european and american swine convalescing from naturally acquired prrsv infections were used to characterize the bp -p protein. sera from gnotobiotic and conventionally reared swine exposed to european isolates of prrsv were significantly more reactive (p < . ) with bp -p than were the corresponding american prrsv antisera using the indirect immunoperoxidase monolayer assay (ipma). prototypic european, but not american, prrsv antisera also recognized bp -p using western immunoblotting and radioimmunoprecipitation assay (ripa) procedures. however, gnotobiotically derived antiserum to an atypical american-origin prrsv was reactive with bp -p by both ipma and western immunoblot. despite a predicted potential for n-linked glycosylation, studies with tunicamycin and peptide-n-glycosidase f (pngase f) indicated that bp -p was not n-glycosylated in either insect cell cultures or trichoplusia ni larvae infected with the recombinant baculovirus. sera from rabbits inoculated with bp -p failed to neutralize both the european (lelystad) and american (atcc vr- ) reference isolates of prrsv and did not react by ipma with prrsv-infected cell cultures. taken together, the data suggest that the carboxyterminal portion of prrsv orf encodes a non-neutralizing viral peptide that is partially responsible for the serologic differences noted between european and most american isolates of prrsv. f ( pngase f) indicated that bpo -p was not n-glycosylated in either insect cell cultures or trichoplusia ni larvae infected with the recombinant baculovirus. sera from rabbits inoculated with bpo -p failed to neutralize both the european (lelystad) and american ( atcc vr- ) reference isolates of prrsv and did not react by ipma with prrsv-infected cell cultures. taken together, the data suggest that the carbox- in the few years since it was initially described (keffaber, ; lindhaus and lindhaus, ; paton et al., ) , the disease complex now known as the porcine reproductive and respiratory syndrome (prrs) has become an economically significant swine health problem throughout europe and north america (goyal, ) . the viral etiology of prrs was confirmed both in the netherlands (meulenberg et al., ; terpstra et al., ) and in the united states with the isolation of two previously unknown prrs viruses (prrsv), a prototypic european prrsv (lelystad isolate) and a prototypic american prrsv (vr- isolate). these to nm enveloped rna viruses wensvoort et al., ) share common antigenic determinants but may be differentiated using both polyclonal antisera and monoclonal antibodies (nelson et al., ) . serologic analysis of european and american swine naturally exposed to prrsv has confirmed that geographically based antigenic differences do exist between most north american and most european isolates of prrsv; these differences are consistent with those first noted between the lelystad and the vr- isolates bautista et al., ) . there is some evidence, however, that antigenically intermediate prrsv or possibly both prototypic antigenic types may exist within the north american swine population (bautista et al., ) . prrsv has been tentatively classified as an arterivirus, along with equine arteritis virus (eav) and two other proposed members of that genus: lactate dehydrogenase elevating virus (ldv) and simian hemorrhagic fever virus (shfv) (plagemann and moennig, ) . prrsv contains a to kb single strand positive polarity polyadenylated genome (plagemann and moennig, ) and utilizes a nested ' coterminal mr.na gene expression strategy (meulenberg et al., ; conzelmann et al., ) . the prrsv genome contains open reading frames (orfs), including the large ' terminal orfs la and lb encoding the putative viral polymerase and the ' terminal orf encoding the nucleocapsid protein (meulenberg et al., ; conzelmann et al., ) . prrsv virions are believed to contain two other structural, presumably envelope, proteins (nelson et al., ) . it has not been determined which of the orfs through encode these proteins or the identities and functions of the other proteins encoded by these orfs. recombinant dna technology permits the expression of individual gene segments in order to better understand the antigenicity of their protein products and their roles in the host-virus relationship. we report here the expression and serologic analysis of a markedly hydrophilic portion of prrsv orf (lelystad isolate). this segment was of additional interest because comparison of ldv, eav, and prrsv orfs through revealed that the orf protein products of these viruses would be the most divergent in size but potentially the most universally highly n-glycosylated of all the proteins encoded by those orfs (den boon et al., ; conzelmann et al., ; godeny et al., ) . these features suggested to us that prrsv orf might encode a glycoprotein mediating virus-host cell interactions; therefore, it might be a target of host defense mechanisms subjected to selective evolutionary pressure. we hypothesized that it might account in part for the geographically oriented antigenic differences between the majority of european and north american prrs viruses, and that was the basis for this study. the lelystad and vr- isolates of prrsv were provided by c. terpstra, central diergeneeskundic institut, the netherlands, and by d. chladek, boehringer ingelheim inc., usa, respectively. a british reference prrsv isolate, humberside , was a gift from s. edwards, central veterinary laboratory, united kingdom. prrsv isolates and were recovered in this laboratory from midwestem american swine tissues. antisera to the isolate had been found uniformly cross-reactive with numerous north american prrs virus isolates; isolate was of interest because it was completely fastidious for replication in swine pulmonary alveolar macrophages. all viruses except were propagated in cultures of marc- cells, which are a prrsv permissive cell line subcloned from ma- cells (kim et al., ) . the identity of all viruses was confirmed using prrsv-specific monoclonal antibodies in an indirect immunoperoxidase monolayer assay (ipma) format. gnotobiotic swine were used to prepare antisera to each prrsv isolate. each lo-dayold piglet was intranasally inoculated with lo to lo tcid,, of one virus isolate, and the resultant convalescent antiserum was collected between to days post-inoculation. twenty field origin sera derived from clinical cases of prrs in swine from northern germany were generously provided by t. blaha, tierartzliche hochschule, hanover, germany. fifteen american field origin prrsv seropositive swine sera were also obtained from d. kinker, iowa state university, ames, ia, usa. each serum originated from a separate swine herd experiencing prrs, and these sera represented samples from herds located in states in the eastern, southern, and midwestem united states. the ipma ( holm-jensen, ) was used to evaluate serial -fold serum dilutions for reactivity against both the lelystad and the prrsv isolates. these viruses had been inoculated h previously onto separate marc- cell monolayers. immunodetection was accomplished using protein g-horseradish peroxidase (hrp) conjugate and aminoethylcarbazole substrate (zymed, so. san francisco, ca, usa). a prrsv serum neutralization (sn) procedure was used to evaluate selected sera as recently described (yoon et al., ) . recombinant orf antigen, cesium chloride purified prssv (nelson et al., ) , and ultrafiltered concentrates of cell cultures infected with the lelystad isolate were used as antigens in western immunoblotting following denaturing polyacrylamide gel electrophoresis in % to % or % to % gradient gels (laemmli, ; burnett, ) . immunodetection was again accomplished with protein g-hrp conjugate and an insoluble tetramethylbenzidine (tmb) substrate (kirkegaard and perry inc., gaithersburg, md, usa). a radioimmunoprecipitation assay (ripa) recently described for prrsv (nelson et al., ) was used with the same antigens to evaluate sera from the gnotobiotic swine inoculated with prrsv isolates. a hydrophobicity/hydrophilicity plot (kyte and doolittle, ) of the putative orp protein revealed a markedly hydrophobic amino terminus followed by the remaining predominantly hydrophilic % ( of amino acids) of the protein (fig. ) . we postulated that this latter portion might be externalized and, therefore, available to interact with host cell receptors or antibodies and that the amino terminus perhaps represented a biosynthetically cleaved leader peptide or otherwise cryptic segment not directly interactive with the host. to focus our examination on the hydrophilic body of the protein, we excluded the hydrophobic segment through a subunit cloning strategy. total rna was extracted from a cm* cell monolayer infected with the lelystad isolate of prrsv. a -mer oligonucleotide complementary to the viral rna sequence (meulenberg et al., ) nucleotides downstream from the ore termination codon (positions , to , inclusive) was used to prime viral cdna synthesis in a ~ reaction (maniatis et al., ) . a base pair (bp) cdna segment was then amplified through cycles of the polymerase chain reaction (pcr) (saiki et al., ) using a ' primer located between positions , to , and a ' primer ( , to , ) to which a pst i recognition sequence had been appended (fig. ) . following xho i and pst i digestion and gel purification, the pcr product was directionally ligated into a baculovirus polyhedrin gene transfer plasmid, pacsg-his-nt (pharmingen, inc., san diego, ca, usa). the resulting plasmid contained the carboxyterminal amino acid-encoding portion of orf fused in frame to a ' vector sequence encoding amino acids including a polyhistidine domain. the resulting fusion polypeptide had a calculated mass of , daltons ( kda) . following dna sequence verification, recombinant plasmid dna ( ug) was cotransfected with bsu -digested acnpv baculoviral dna ( . ug) (pharmingen, inc.) into a spodopteru frugiperda sf- cell culture using a liposomal transfection reagent (dotap, boehringer marmheim, indianapolis, in, usa). clonally purified baculoviruses expressing prrs virus-immunoreactive material were identified by ipma. separate pcrs of cell culture fluids from monolayers infected by these cloned viruses were used to confirm the presence of the orf gene sequence and the absence of the nonrecombinant baculovirus polyhedrin gene sequence. the latter pcr employed primers spanning the polyhedrin gene insertion site (invitrogen, inc., san diego, ca, usa). one rdna baculovirus, bp , was used for recombinant protein production. recombinant protein (bpo -p) was produced by infecting both sf- cell cultures and -day-old trichoplusia ni larvae according to standard procedures ( o'reilly et al., ) . ] asparagine amidase (boehringer mannheim, inc.) in a further effort to evaluate the n-glycosylation status of the protein. the polyhistidine sequence within bp -p enabled efficient affinity purification of both cell culture and larval origin bpo -p using immobilized nickel ion chromatography as previously described (janknecht et al., ) . purified larval and cell culture origin bpo -p was adjuvanted (fatunmbi et al., ) with avridine (pfizer, inc., groton, ct, usa) and used to immunize four rabbits each on four biweekly occasions. sera from these animals were then evaluated by ipma, sn, western blotting, and ripa for anti-prrsv reactivity. bf'o -p expression was detected in the cytoplasm of infected sf- cells using gnotobiotically-derived lelystad isolate antiserum in an ipma format (fig. ) . bpo -p expression was noted within h post-infection (hpi) and persisted at least to hpi. comparison of bp infected and uninfected cell culture proteins by sds-page revealed the presence of a new protein in the infected cell cultures (fig. a) . the size of this protein was consistent with the kda mass predicted from the amino acid sequence of the fusion protein encoded by bp . the prrsv-specific identity of this protein was confirmed by western immunoblotting (fig. c ). bpo -infected insect larvae contained a similarly sized immunoreactive protein within hpi (figs. b, c). protracted incubation of affinity purified larval bp -p with pngase f did not result in a detectable reduction in apparent bpo -p molecular mass (fig. c ). there was also no detectable decrease in the molecular mass of bp -p from tunicamycin treated cell cultures relative to untreated controls (fig. c ). the ipma, sn, ripa, and western immunoblot procedures were used to evaluate rabbit antisera developed against both larval and cell culture origin bpo -p. none of these sera exhibited respectively. lane : molecular mass markers (kda) panel c: immunoblot of affinity purified larval bpo -p protein with and without prior digestion with pngase f (lanes and , respectively). immunoblot of sf- cell culture origin bpo -p protein produced in the presence or absence of tunicamycin (lanes and , respectively). lane : cell culture control infected with recombinant baculovirus containing vector-only transfer plasmid sequences. prrsv sn activity at a : dilution. all sera were ipma-reactive at : dilution using bp -infected sf- cells but failed to react at a : dilution with isolate-infected marc- cells. these sera also did not react at the : dilution with lelystad isolateinfected cell monolayers fixed between and hpi. two of the rabbit antipeptide sera were reproducibly reactive by western immunoblot with a diffuse ( to kda) band of antigen found in homogenates of marc- cells infected h previously with the lelystad isolate (fig. ) . repeated ripa and western blot attempts using these sera were unsuccessful in identifying a virus-specific protein in purified virion preparations. antisera to gnotobiotic lelystad, humberside , and virus isolates reacted specifically at : dilutions with nitrocellulose blotted bpo -p, while antisera to vr- and virus isolates were totally nonreactive at the same or lower dilutions (fig. b) . ripa results were consistent with these findings. these sera were also evaluated by ipma against lelystad and infected marc- cells and bpo -infected sf- cells (table ) . antisera to the lelystad and humberside isolates were differentially reactive with cells fig. . western immunoblot analysis of sera from two rabbits inoculated with bpo -p. lane : molecular mass markers &da). lanes and : blots of rabbit anti-(bp -p) sera, showing reactivity against to kda proteins found in cells infected with prrsv (lelystad isolate) hours previously. lane : blot of rabbit serum (same as used in lane ) against uninfected cell antigens. lanes and : blots of hyperimmune anti-bpgf-p rabbit serum and gnotobiotic anti-prrsv (lelystad) swine serum, respectively, against the bpo -p immunogen. hyperimmune rabbit serum also detects small ( - kda) breakdown products of bp -p resulting from its catabolism in infected insect cells. 'german sera significantly more reactive (p < . ) with lelystad and bp viral antigens than the corresponding american sera and significantly (p<o.ol) less reactive with antigen than with the other viral antigens. damerican sera significantly more reactive (p < . ) with viral antigen than the corresponding german sera and significantly (p < . ) more reactive with antigen than with the lelystad and bp viral antigens. knot determined. infected with the lelystad isolate and also recognized bpo -p in infected sf- cells. antisera to the vr- and isolates were differentially reactive with isolate infected cells. antiserum to the isolate was nearly equally reactive with cell cultures infected with either of the prrsv and with the bp virus (table ) . convalescent sera from field origin prrsv-infected german and american swine were differentially and antithetically reactive by ipma. a two-way analysis of variance was used to assess the serologic differences. the geometric mean titer (gmt) of the german sera was significantly greater (p < . ) against lelystad and bp viral antigens than against the viral antigen. conversely, american sera were significantly (p < . ) more reactive with viral antigens than either of the other two antigens (table ) . taken together, the data demonstrate that immunoreactive epitopes within the carboxyterminal portion of the protein encoded by lelystad prrsv orf are expressed during the course of european prrsv infections, and that this polypeptide sequence is at least partially responsible for the serologic differences observed between european and american prrsv isolates bautista et al., ) . antiserum to the macrophage-fastidious and antigenically intermediate isolate was reactive with bpo -p, further suggesting the importance of the orf protein in the serologic differentiation of prrsv isolates. the similarity between the ripa and western immunoblot reactivity patterns of bp -p using european and american prssv antigens and gnotobiotically derived convalescent swine antisera suggests that perhaps both linear and conformational orf epitopes may be involved in the antigenic differentiation of european and american prrs viruses. our data support the hypothesis (bautista et al., ) that coexisting high lelystad and atcc vr- prrs virus antibody responses found in some american swine may result from infections with a virus of mixed antigenicity as opposed to dual infections with prototypic american and european-like prrsv isolates. like the homologous predicted ldv and eav orf proteins, the prrsv orf protein contains the greatest number of sites for potential n-glycosylation of all the proteins encoded by orfs through (den boon et al., ; conzelmann et al., ; godeny et al., ) . n-linked glycosidic moieties have frequently been found important in mediating viral infectivity, in directing soluble and cell-associated host immune responses, in protecting critical viral protein epitopes from immune attack, and in interacting with viral polypeptide components to ensure correct viral glycoprotein conformational structure (rademacher et al., ; li et al., ; grigera et al., ) . attempts to produce observably deglycosylated and nonglycosylated forms of bpo -p using both tunicamycin inhibition and enzymatic deglycosylation approaches indicated that the recombinant protein did not contain n-linked glycosidic modifications. these findings must be interpreted cautiously. despite the theoretical potential for a high degree of n-linked glycosylation, the carboxyterminal portion of the native orf protein may in fact not be glycosylated. perhaps more likely however, the aminotetminal peptide sequence omitted during bp construction may have represented a signal sequence required to direct nascent orf into the er-golgi complex for glycosylation during insect larval and sf- cell infections. although insect cells and tissues are generally assumed to be glycosylation-competent, differences between mammalian and insect glycosylation patterns have been observed (rademacher et al., ; shimokawa and smith, ; o'reilly et al., ) and could also account for the data. the addition or removal of very small n-linked glycosides to the bp -p protein might have been undetectable given the molecular size resolution of the western immunoblotting, but this is unlikely because viral n-linked glycosidic moieties are usually substantial in apparent molecular mass (rademacher et al., ) . regardless of the foregoing considerations, the orp of the lelystad isolate clearly encodes epitopes recognized immunologically by the swine host during the course of both experimental and natural infections. monospecific bpo -p rabbit antipeptide antisera reacted by immunoblot but not by ripa with a diffuse band of antigen ( - kda) found repeatedly in virus-infected cell cultures. this material was specific to virus infected cells, but was apparently not of virus structural protein origin, because it was not detected by either ripa or immunoblot using purified virus preparations. the difference between the ripa and the western immunoblot performance of the rabbit antipeptide antisera using unpurified infected cell culture material may be due to the presentation of the relevant reactive linearized nonstructural virus protein epitopes under the conditions of the denaturing immunoblot but not under the milder conditions of the ripa. monospecific bp -p antisera were not neutralizing, but the functional significance of this finding is unclear because other workers have suggested that antibody responses to prrsv may not necessarily be protective and might even participate in antibody-dependent enhancement of infection (choi et al., ) . the failure of monospecific rabbit bpo -p anti-peptide antisera to react with prrsv-infected cells is consistent with the ideas that the natural orf translation product may be conformationally (epitopically ) different from the recombinant bp -p fusion protein, or that the native protein may be a nonstructural, perhaps soluble, virus gene product. lactate dehydrogenase elevating virus, eav, shiv, and apparently prrsv all possess three structural proteins including a nucleocapsid protein encoded by the ' terminal orp (nelson et al., ) . the other two presumed prrsv structural proteins are kda and kda in apparent molecular mass (nelson et al., ) and are thus far removed from the - kda band identified here. although the precise identity and functional role of the prrsv orf gene product thus remain enigmatic, a differential virus serotype-specific serologic response is clearly directed against it by swine exposed to prrsv. serologic survey for lelystad and vr- strains of porcine reproductive and respiratory syndrome (prrs) virus in u.s. swine herds characterization of swine infertility and respiratory syndrome (sirs) virus (isolate atcc vr- ) electrophoretic transfer of protein from sds-polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein a antibody-dependent enhancement of sirs virus replication isolation of swine infertility and respiratory syndrome virus (isolate atcc vr- ) in north america and experimental reproduction of the disease in gnotobiotic pigs molecular characterization of porcine reproductive and respiratory syndrome (prrs) virus, a member of the arterivirus group enhancement of antibody response of turkeys to trivalent avian influenza vaccine by positively charged liposomal avridine adjuvant complete genomic sequence and phylogenetic analysis of the lactate dehydrogenase elevating virus ( ldv) virology porcine reproductive and respiratory syndrome lelystad virus, the causative agent of porcine epidemic abortion and respiratory syndrome (pears), is related to ldv and eav effect of glycosylation on the conformational epitopes of the glycoprotein of vesicular stomatitis virus i. detection of antibodies against hog cholera virus and bovine viral diarrhea virus in porcine serum. a comparative examination using cf, pla, and npla assays rapid and efficient purification of native histidine-tagged protein expressed by recombinant vaccinia virus reproductive failure of unknown etiology enhanced replication of porcine reproductive and respiratory syndrome ( prrs) virus in a homogenous subpopulation of ma- cell line a simple method for displaying the hydropathic character of a protein cleavage of structural proteins during assembly of the head of bacteriophage t glycosylation is necessary for the correct folding of human immunodeficiency virus gp in cd binding ratselhafte schweinkrankenheit molecular cloning: a laboratory manual differentiation of u.s. and european isolates of porcine reproductive and respiratory syndrome (pius) virus by monoclonal antibodies baculovirus expression vectors: a laboratory manual lactate dehydrogenase-elevating virus, equine arteritis virus, and simian hemorrhagic fever virus: a new group of positive-strand rna viruses primer-directed enzymatic amplification of dna with a thermostable dna polymerase expression of prostaglandin endoperoxidase synthase- in a baculovirus system experimental reproduction of porcine epidemic abortion and respiratory syndrome (mystery swine disease) by infection with lelystad virus: koch's postulates fulfilled antigenic comparison of lelystad virus and swine infertility and respiratory syndrome (sirs) virus a modified semm neutralization test for the detection of antibody to porcine reproductive and respiratory syndrome (prrs) virus in swine sera we thank tom glasson, gene hedberg, wayne romp, melanie harris, and nancy platter for illustrative services and careful manuscript preparation. key: cord- -rncleqqy authors: ramírez, j. martín title: long-lasting solutions to the problem of migration in europe date: - - journal: a shift in the security paradigm doi: . / - - - - _ sha: doc_id: cord_uid: rncleqqy this is the following of a previous publication on the refugee crisis in europe and its security challenges. here we suggest some long-lasting solutions to the problem of migration. these may be summarize in four points: dealing with the countries of origin and of transit, adequate control of borders, and positive measures to facilitate the integration of the newcomers in their countries of destination. in a previous publication on the refugee crisis in europe and its security challenges, i concluded that a global problem like this one could not be solved without an adequate orderly, and controlled immigration policy, creating systematic and controlled arrival and integration programs, because the mental structure of european societies is not prepared to face a disorderly increase in migration flows (ramirez (ramirez , . it is thus vital to regulate the arrival of migrants if we want to achieve their real integration in our culture (espaliú berdud ) . here i will try to go forward, pinpointing the terrible damage made by the massive escape of young people from countries that desperately need them in their struggle for development; their great problem is precisely the continuous loss of human capital. besides of that, who emigrate are not the "poor among the poor", but people with certain economic means to be able to afford the trip and contacts in the place they are going to. what is more important, according to me, is to argue that the best way would be to encourage migrants to stay home, preventing massive uncontrolled displacements, is addressing the factors that drive emigration of their own origin countries. to stop unwanted migration, the developed countries must promote a profound democratic and economic stabilization and development. we must always remember that the first right of every human being -after the right to live-is the right not to emigrate and to have the opportunity of living peacefully and prospering in our own home, as pope pius xii wrote in the apostolic constitution exsul familia: "all men have the right to a family living space in their place of origin". the right to emigrate is only subsidiary to the main right to have a family living space in the place of origin, when this cannot be assured. migration has been a sensitive and contentious topic for ages. escaping violence, war, poverty and environmental disasters, more people than ever are migrating worldwide. since it is not a cyclical, but a structural phenomenon, which means that whatever the method put forward migration cannot be stopped. currently, according to the uno estimates, million people live outside their country of birth ( . % of the world's population). in , about % of the world's . billion people lived abroad (unhcr ) . this figure grows due to inequality, climate change, conflicts, and the interconnectivity that facilitates the movement of people. migration itself affects values, identities, cultures, assimilation capacities of societies, and, far from being a problem in se, it may be a solution to many problems; e.g., benefits the demographic catastrophe present in the aging western societies. but it doesn't always engender positive changes; it may also show important disadvantages. this is why immigration must be orderly, capable of duly regulating the massive arrival of people; otherwise our social protection systems will not resist. the illegal immigration requires proper vetting to identify criminals and terrorists-the crime rates of the foreign immigrant population are significantly higher than those of the natives-, and sometimes, to put in metaphorical terms, surgical excision procedure without anesthesia will frequently induce pain. no country needs a trojan horse. but we will leave this so interesting aspect for another parliament. although mass emigration to developed countries is a global phenomenon -it happens even within each country, internally displacing for instance from rural areas to urban ones: china is a clear example-, the present chapter will be focused mainly on what would be the better long-lasting solutions to its present situation in europe, one of the leading destinations in the world in terms of migratory flows, with million migrants, russia included, according to the un's department for population. immigration is an irresolvable problem at short-term. migration policies should follow a long-term vision, addressing economic, security and sociological points of view (de la cámara ), creating systematic and controlled arrival and integration programs of security, trade, development and employment issues. but, being a global problem, migration will only be solved as a consequence of a frank and sincere joint co-responsibility between all countries of origin, transit and destination, sharing the burden of dealing with both regular and irregular migration whenever possible. this was the intention of the un global compact for safe, orderly and regular migration, signed at the end of by representatives of the countries of the united nations (uno) met in marraquesh, in a non-mandatory and rather controversial document that says, in an indirect manner, that for the entry to be safe, orderly and regular, the conditions of entry must be safe and not subject to the mercy of smugglers, seekers' and workers' entry profiles should be differentiated, and entries should be legal. its "objectives" were peppered with vague declarations, platitudes and split differences, such as the fight against the mafias that deal with human beings, the defence of the rights of immigrant workers or a change in narrative about migration towards a more positive approach. partly in the spirit of other global agreements like the paris climate deal, it encouraged states to co-operate on tricky cross-border matters without forcing them to do anything, and urged governments to treat migrants properly, but also to work together on sending them home when necessary. at least, it may help build the trust between "sending" and "receiving" countries that is the foundation of any meaningful international migration policy (economist ). let us offer some recommending ways of improving migration at four different levels: going to the roots in the countries of origin, dealing with the countries of transit, having an adequate control of borders, and suggesting some measures in the countries of destination. i have critized elsewhere (ramirez ) the saving buenist attitude of a migrationist maximalism that wants to open the doors, the ports, the windows and throw all the walls, in favor of a weberian realism (the ethics of convictions versus the ethics of reason) (leguina ): even if we would welcome forty or fifty million africans a year, africa will continue to have the same population. it would be good if they start helping africans in their own countries and avoiding as much as possible the reasons why they want to emigrate (ramirez ) . we want the potential migrants to have a better future in their countries. as ousman umar repeats like a mantra, "the solution is in the country of origin, not in the destination. you have to feed minds (well); if you feed the mind you are satisfying hunger for more than a hundred years" (umar ) . some african prelates within the catholic church are calling attention to the most forgotten aspect of this debate: the terrible damage made by a massive escape of young people from countries that desperately need them in their struggle for development. in a book released recently, cardinal robert sarah, currently prefect of the congregation for divine worship and the discipline of the sacraments, declares his personal position on immigration: "all migrants arriving in europe are crammed, without work, without dignity. is this what the church wants? the church cannot cooperate in this new form of slavery into which mass immigration has become" (sarah a, b; indelicato ). the european union (eu), thus, has to structurally turn over economic resources for the development of africa (and of middle east), tackling the problems that are the main cause of migratory movements, analysing them and offering real help in their own home. consequently, their priority has to be to invest in the countries of origin, because migration control has to be paid with money: financing, companies, information, facilitating their institutional, social, political and economic conditions in order that nobody will be obliged to leave their home, and creating sources of employment in those countries of origin. some political party has proposed in its program that the eu should prepare a sort of "plan marshall" for africa, similar to the one usa did for the post-war europe. according to the spanish royal academy (rae) dictionary, buenismo is the attitude of who reduces the seriousness of conflicts, acting with excessive benevolence and tolerance. this "goodism" is a demagogy destined to hold power through emotional blackmail, quite different to a real goodness, born of charity or philanthropy (robles, ) . cardinal robert sarah is a native of guinea guinean who grew up under a harsh marxist dictatorship and became archbishop at the age of with the task of guiding the diocese of conacry, when still in his country there was the socialist regime of sekou touré. so, i think he knows what he's talking about. in the forties of the last century, america passed the economic co-operation act, better known as the marshall plan, because its inspiration from a speech at harvard university by george marshall, america's secretary of state. the marshall aid, aimed to revive europe's war-ravaged economies, encouraged the europeans to quash inflation and to narrow their deficits while eventually dismantling price controls and import barriers. these reforms had enormous benefits. before fear of inflation and taxation prompted german farmers to feed their harvests to their cattle, rather than it is not enough with assigning cooperation funds. as de la concha stresses ( ), a proper management of migration flows requires working as much as possible in cooperation with the countries of origin taking into account, as appropriate, the various causes of migration (economic, security, political prosecution, climate, etc.). thus, a more ambitious measures are required, such as generating opportunities for the local population and offering fiscal incentives to the investments of european companies in those regions that seem to be condemned to diaspora. this move will also offer new markets for european products and services, as well as opening of eu markets to exports from these countries. and, what is more important, besides of contributing to the economic development of those countries, europe has to improve their social and political quality of life, which is precisely what they try to find in europe. our main objective, therefore, has to be to strength the production base and the creation of jobs in the countries of origin, the provision of basic services (health, education) to the local populations, and the literacy, ensuring that students in their countries have the tools to decide their future and thus avoid the temptation to migrate to europe, avoiding future victims (de la cámara ). either we offer them opportunities in their own countries, or they will come to ours looking for them zalba ( ). the best practical means for achieving it are probably inter-governmental agreements with the origin countries, giving them a generous economic aid. this is the stance taken by the visegrad group (v ) (check republic, hungary, poland, and slovakia): instead of uncontrolled massive immigration, we have to act in their origin countries; instead of importing problems, exporting help in situ. this attitude would also be "much more inexpensive", as is honestly admitted by juho eerola, a finn of the nordic freedom, or by the then slovak prime minister, peter pellegrini, during his first visit to brussels: "what we have to do is to invest in the countries where the problem is originated. each euro spent in northern africa is more efficient than spent in the migrants who arrive massively to eu" (eerola, april ). but, there is need to build up conditions to create secure environment within the transit african countries bordering the mediterranean sea. the north african governments are responsible for effective and transparent governance aimed at management of emigrants flows through their territories. eu's money poured into their accounts are wasted as they do not cooperate as they can. another effort to stem the flow of migrants to europe is "to save and protect the lives of migrants and refugees", motivating "operations of urgent evacuation" returning them to their origin countries through a repatriation programme that encourages those who have made it to northern africa to go home voluntarily, rather than risk sell it to the cities for money that might be diluted by inflation or seized by the government. its true significance laid not in the cash it provided but in the market-friendly policies it encouraged. to receive aid, european governments had to commit to restore financial stability and to remove trade barriers. we have a similar problem within our own countries, migration towards big cities is up siding the living in small rural areas; we have to stop it offering them "the needed modern technology to have the best of both worlds. it is not an easy problem, but i feel a deep analysis needs to be done so we understand more of the dynamics behind the movements"! (lindhard, ). a rickety boat across the mediterranean. this is what uno, eu and african union (au) jointly agreed in abiyan ( nov. ): people who turn back get a free flight-cutting out the need for a perilous return journey across the sahara. the programme has repatriated some , migrants to various west african countries, which barely scratches the surface because it reckons there are about m of them in the african shores, waiting for their risky jump to europe (economist ). the spanish government did this unilaterally in , after . immigrants arrived to the canary islands in open boats, known as cayucos: some direct forms of operative cooperation with those countries through which migrants come or transit, sending there some specific police units for working jointly with the local security forces with the aim of restraining subsequent invasions from other western african countries. since the devolution of immigrants to their origin countries is one of the best deterrent action to avoid the "calling effect", governments have to launch adequate information campaigns in their own origin countries to discourage potential migrants. the eu member states are africa's largest donors, supplying more than half the aid the continent receives. africa exported twice the value of goods to the eu as it did to china in . but it is not just a matter of addressing the factors that drive emigration. europe and africa share something much more fundamental: a future dependent on one another. as moroccan king mohammed vi pointed out in , solidarity between europe and africa has to be "built on shared responsibility and mutual dependence." the european continent is a global player worthy of genuine partnership of equals with africa that priorities concerning mutual interests through rapid funding in education, health and infrastructure for africa's youth would contribute to global growth. europe's investment must be bolder in terms of financing, policy and governance reforms than what is currently on the table (cole ) . economic development, government reforms, institutional strengthening, will result in common prosperity. europe, poor in natural resources and in demographic decline, desperately needs to contribute to the unblocking of the future of africa. up to now, the -economically important-european efforts have to qualify as at least disappointing. the eu needs to encourage, promote opportunities in africa, move from a policy built around aid to cement our relationships on trade and investment (palacio a) . cooperation between states that produce migrants and those that receive them can help to streamline migration flows. the european union has to structurally turn over resources for the development of africa and middle east, which are the main source of migrants nowadays, through agreements with extra communitarian countries for stopping irregular immigration towards europe. and this requires patience and diplomacy, treating the partner governments as equals. the optimism of the "refugees welcome" campaign in -wir schaffen esled towards an uncontrolled flood of refugees which destroyed the perception of order and stability. in june , there was a great turn in the attitudes of the rulers regarding immigration, with a broad consensus at the european council around much more restrictive. this practical agreement intended to hand over the decision-making capacity over who enters and who does not in the "fortress europe", strengthening the policies of controlling the external borders; fighting against traffickers; and more aid in the countries of origin instead of endless aid in europe. there is a need to strengthen collaboration with transit countries. it is necessary to prevent ships from leaving their points of origin. in cases where this is not possible, the practice should be implemented that those rescued at sea should be assisted and returned to the point of origin or departure of navigation. this is the specific task of frontex, the european border and coast guard agency: provide technical assistance and support to the countries of origin and transit to help strengthen border controls, even if it cannot replace national competence; it is converted into an authentic border police, but not in charge of rescuing. i will to come back to this point later. according to the ngo african center for strategic studies, there is a migratory flows that move . million people from one place to another from and within africa in the hands of traffickers; % of them are minors, and only % of those . million, will try to reach europe; the remaining % travel between african countries. in this context, i point out at a report by the department for human rights at the un, marking the horror of criminal smuggling networks which lead to humanitarian scandals such as: enslavement; imprisonment; rape; prostitution; the sale of organs; and camps, whose living conditions are beyond imagination. what alternative do citizens of many african countries have to come to europe? if the european states renounce their ethical convictions, delegating their responsibility, mafias and some connected "heroic" ngos will occupy the scene doing the dirty work. those still determined to reach europe may have to pay large sums to people-trafficking gangs and risk their lives in the sea (we may say the same about america, substituting the word "sea" by "desert"). the more borders, bureaucratic obstacles, closed routes and prohibitions the immigrant finds, the more profitable for the trafficker. no one moves as many people on the planet as immigration traffickers, nobody determines human flows as much as their implementation in a given territory, nobody has their flexibility to change plans and adapt to changing circumstances and nobody gets so many benefits as they do. their criminal business is more lucrative nowadays than drug or arms trafficking. some of those traffickers have changed tactics. before, they put hundreds of immigrants in old overloaded wooden boats with the aim of trying to reach europe. nowadays, after cashing from them, the immigrants are taken in safe boats from african territorial waters, until they know that the ship of some ngo is a few miles away. then, once in international waters, they put on life jackets, crammed them in zodiacs, which can barely sail a few hours and warn by radio that there are shipwrecked people in the area. the more rescue boats they have, the more immigrants will try to cross the mediterranean…, and the more deaths, drowning in the sea, those claimed heroes of the goodism will provoke. a ngo director denounced on television that although he had managed to rescue people from a zodiac, another had perished. what nobody asked was if he did not realize that by telling those who leave libya that there are boats a few miles away they are contributing to the death of hundreds of people who, for one reason or another, cannot be picked up. trafficking in human beings to the european destination is more profitable than drug trafficking: it moves around the mediterranean, according to the latest un report published last year, about billion dollars. just take the account: boats for people are used for , crammed, standing, and almost unable to move for hours. at , e per head, it gives , e per trip. it is paid in two parts: the first , e to take them from their country of origin to the ports of libya and the other , to move them to european port, in an offshore mothership, and then in small boats that launch into the sea in a point located by gps where they can be picked up by the rescue boats of the ngos, or by merchants, fishing or recreational. if they do not find this type of boats in the area, they make a distress call with a satellite telephone with the coordinates. if the trip is frustrated and they are returned to their countries, they try again when they have collected the money. and several thousand have not even had that opportunity because they have lost their lives. with that turnover, there is no problem in paying the corresponding bribes in each place (fernández arribas ). "the good trafficker tries not to lose the immigrants who have been his clients. his goal is to get them to europe and earn money. if they lose, e.g., a truck with immigrants between agadez and algeria or a boat that sinks with people, is not a problem, because they have already paid; but if they lose kg of cocaine or two boxes of kalashnikov, then they do have a serious problem. that is the difference". and how are these people's traffickers? "these gangsters are criminal entrepreneurs, but entrepreneurs. they must have many contacts, be credible, creative, charismatic, with the power of conviction, knowledge of the routes, the laws, the latest news… which make them very fast, elastic and flexible. they also participate in other businesses; f. inst., when the vehicles that bring immigrants back through the sahara return, they never make them empty: they carry weapons or drugs back to take advantage of the trip" (musumeci and di nicola ) . fernández arribas ( ) offers a very detailed description of how tens of thousands of human beings are stranded in territories dominated by mafias, in their expected way towards europe. although every corner of the world retains its specificities, the human traffic mafias analyzed by the un reflect some common patterns. all of them have recruitment agents from the main groups that are victims of extortion. a second level is that of the local mafias, who know the land and generally pay the collectors. one more step is that of small-scale service providers: trucks, boats and other means of transport. the most dangerous level is that of the big international mafias, who control the entire process and also add links to large international crime groups and use immigrants as a way to earn money quickly and easily. infrastructures built for the exploitation of the mineral resources of the desert, oil in libya and uranium in niger, facilitate trips through the desert. according to reports from the spanish police, nigeria and libya are the most active and most established bases of immigration mafias in central africa, with ramifications in other countries, where sudanese mercenaries act unscrupulously. in the waiting time, they live in abysmal conditions, suffering very serious violations of the human rights, such as abuse, harassment, violence, robbery, kidnapping, extortion, common torture and rape. in north africa they have their point of convergence in agadez, a town north of niger, or in sehba, south of libya, where control is exercised by heavily armed tribal groups. from that point of confluence, they are transferred to the coastal cities of the tripolitana region, where trips are negotiated and where part of the population collaborates with their garages as a temporary shelter at an abusive but obligatory price for those who see the end of your nightmare construction, agriculture or any job in order to get the money. the boats used are manufactured in libya and the fishermen always have on board some drum of gasoline, pure gold for the weak boats of the new slaves of the st century. in libya, each link in the chain receives its commission, especially those who seek protection and security, from militias to corrupt authorities. this action-paying poorer ones to set up vast holding-pens for humans in unhuman conditions-is a big business for a few behind the misery of many others, and involves something which would not be tolerated at home, but it seems somehow acceptable in these situations because it is out of sight. stopping these traffickers, which is the solution that many politicians in europe offer to regulate immigration, is almost impossible; they are always ahead of a european immigration policy, without the necessary coordination. a clear example has been the failure of the military operation eunavfor med sophia (from to ), which ambitious goal of dismantling the mafias that traffic with people from the libyan coasts to europe has not been fulfilled, partially because the closing of the italian ports to illegal immigrants has left it out of play. due to the fact that operation sophia no longer uses ships but only unmanned aerial vehicles, it cannot continue rescuing people at sea. federica mogherini, at that time high representative of the union for foreign affairs, explored a new mission, with more modest goals: to train the libyan coast guard to control their own waters and avoid the departure of irregular immigrants; but there were also no practical results. this shows the external and humanitarian action of the eu, as it really is: "lack of criteria, commitment and agreement on immigration issues…. public opinion will continue to show ships coming from the libyan coasts and criminal organizations will increase their profits" (de ramón-laca ; see also espaliu , espaliú berdud . as accurately pinpointed by bitzewski (personal communication), though, it is not the eu administration to blame but the heads of states not being willing to take the action. they point their fingers out towards the eu but, when the eu puts proposals on the table, they get reluctant to contribute to the common effort. it starts with the frontier countries, crying but doing very little to protect their own borders, and goes to the rest of europe raising any reason not to commit themselves to the program. the eu should be tougher on the north african governments, corrupted and ineffective. a carrot and stick should be one of the ways to start discussion with them. it's their obligation to control migrants movement within their territories but they make money on it! turkey and egypt are perfect examples of the game. when they want they can stop trafficking! but, obviously, the money is the main reason for action. the north african countries are interested in this business and it is up to us to give them a signal "do not be too smart". we deal with state controlled trafficking and we are very naïve not forcing these governments to take the responsibility. we can help them but we cannot work for them. a key move to avoid the "calling effect" is to guarantee security and economic agreements -migration control has to be paid with money-, with those countries migrants transit through in their way to europe, preventing them from leaving its coasts and returning to their country of origin those people whose asylum in the eu has been denied. in november the eu established the emergency trust fund for africa (eutf for africa) with an allocation of e . million covering algeria, tunisia, morocco and egypt, aimed "to contribute to safe, secure, legal and orderly migration from, to and within the region and support an effective management of migration flows that protects human rights" turkey is the main starting point for the arrival of potential refugees from middle east to europe. there are more than millions syrian refugees, and almost another one in lebanon. the repatriation agreement signed by brussels and ankara in march -entering , million euros annually in the turkish coffers + visa free travel for some turkish citizens-has been a quite good example of efficacy, convincing them to keep refugees from europe's shores: more than , migrants-three fifths of the detainees came from afghanistan and pakistan-have been detained in turkish territory since the document entered into force (turkish directorate general of migration management ). according to gerald knaus, leader of european stability initiative (esi), the entry of refugees into greece was reduced "by % and even higher in the number of deaths". and, in a more global view, the more than one million irregular migrants (mainly refugees) who arrived in europe in have fallen to . in . turkey thus has played a leading role as a dam to contain migration outside europe. in the future, eu will have to invest more in integration and public awareness programmes in turkey. erdogan himself has proposed resettling at least some of the refugees in a safe zone he wants set up in northern syria. all of this is legally possible. syrians in turkey do not enjoy formal refugee status, which would protect them from deportation, but "temporary protection", which does not. according to the high commissaire of united nations for refugees (acnur, in french), a turkish meltdown would send economic shockwaves or new surges of migrants onto the european mainland. erdogan has encouraged such fears (july nd, ), threatening the suspension of the migration deal over the eu's sanctions. another point for the arrival of potential refugees from africa is the maghreb. those eu southern members with close ties to it have an special interest in encouraging the reform of state structures, as well as working to reduce the socioeconomic disparities and lack of opportunities that remain the public's most pressing problems in those countries. two successful cases have been the attempts of cooperation of the eu with morocco and tunisia, as well as a. very important job in the control of migratory flows and the fight against mafias, terrorism and drug trafficking is due to the creation of a coordination authority for the gibraltar strait. by way of example, there is a positive counter-terrorism cooperation carried out by several eu countries working closely with northern africa on security, providing training and equipment for counter-terrorism and conducting some joint operations. morocco might be a capable security partner as the authorities closely monitor its population and controls its religious sphere. it has been successful in preventing attacks and obtaining information that can benefit its european partners, but its counter-terrorism efforts fit within a framework of conserving rather than transforming the state's unaccountable relationship with its subjects, which relies on a repressive political system and resists outside calls for reform. nato has also awarded us$ billion to turkey. four years later, just when this book is going to press, the crisis has returned. turkey does not seem proactive anymore. tayyip erdogan has called for more european support for ending the war in syria and for receiving millions of refugees. and as a pression measure, on february , , turkey decided to open its borders for migrants to move to the european union. this has brought together more than , migrants along the border with greece. three weeks later, though, following the strategies of the other countries to avoid a further spread of covid- , ankara has announced just the opposite: the closure of the eu's borders. the spanish authorities have observed with relief a downward trend in arrivals in mid-january , with a significant reduction of a % of the figures of irregular immigration: the entries, an average of about . a month, confirm that the flows have been considerably reduced since then and that they have fallen to levels below those registered the previous year, when spain became the main european gateway for irregular immigration. this proactivity of morocco, deploying agents to reinforce the fight against the mafias, stop the exits by sea and employed its coastguard in the rescues on the high seas, has been highlighted both by spain and by an internal report of the european commission as the most effective tool to contain migrants. this change of attitude coincided with the announcement of the visit of king philip vi to rabat, which was finally held on february , . on that trip, the spanish monarch asked mohamed vi to go "beyond" in the control of irregular immigration. later, the fisheries and agriculture agreements, signed by eu and rabat with an injection of million euros to contain migratory flows, have also been key to boost moroccan collaboration. besides of that, spain has approved to add the almost million that rabat asked last year to contain the exit of immigrants; that is to purchase surveillance equipment "to combat irregular immigration, immigrant trafficking and trafficking in human beings", as well as for fuel, maintenance of patrols, diets and salaries (palacio b) . the moroccan and sub-saharan return agreements are also praised as a "deterrent factor" to avoid the "call effect". morocco moves many sub-saharan people to the south, leaving them lying in the middle of the desert, or locked in small cells inside an illegal detention center in arekmane ( km from melilla) and then deported to their countries. tunisia has also made significant advances in its security policies, but it has yet to find balanced ways to deal with its porous borders and the disproportionately large number of radicalized people. unfortunately, the attempts to solve the problem of migration with libya have been rather frustrating. libya is a too fragile state, a territory without government since a few years ago, which is key in the irregular migratory flows towards italy and, in the late times, increasingly diverting the flow of west african migrants to spain, via new algerian routes (fernández-sebastián ). nowadays it is the largest and more serious migration corridor, due to the use of mafia "facilitators", as we had previously commented. in this context, some libyan militias behind much of the people-smuggling migrants across the mediterranean claim that eu and, more specifically, italy have given them money and equipment to improve the coastguard stopping migrant boats from setting sail. this may explain the falling of the number of migrants crossing the sea. finally, the european leaders seem delighted with the cooperation with egypt, where immigrants no longer arrive due to the decision of this country to prevent them from leaving its coasts. if all the countries of the south did like egypt, there would be no immigrants at sea. a punctual example of joint collaboration of countries of both sides of the mediterranean sea is the cross-border military exercise neptune, with the strategic objectives of unifying efforts and coordinating action plans among the mediterranean countries, especially in terms of detention and combating the movement of foreign terrorists through ports on both sides of the mediterranean. thus, passenger and freight transport vessels were included in the controls and the use of interpol databases could be activated participation. this activity has been developed between several ports such as tangier med, tangier city and bni-ansar (nador). in sum, the eu should remain committed to encouraging and supporting the most significant pressing remaining security challenges faced by those countries, such as: ( ) the reduction of the social frustrations, economic inequality, lack of opportunities, and governance problems that increase the likelihood that people will join extremist groups and recruit radicalized individuals; and ( ) the reform of state structures, especially in relation to: (a) improving security governance based in an adequate culture and professionalism of the security forces (f. ins., training of police officers, courts judges and staff, customs and coast guard officials); and (b) developing systematic approaches to prevent further radicalization and addressing conditions that facilitate it, such as: exploring better ways to handle radicalized individuals than large-scale incarceration; distinguishing between committed jihadists and those who are more open to reintegration into society; and developing programmes to promote religious education and awareness, gearing them towards pupils and their families from an early age, so relevant to the treatment of radicalized individuals and the prevention of further radicalization (dworkin and el malki ). although the priority has to be focused on the countries of origin and transit, the fact is that we have to face a massive amount of irregular migrants who are arriving to our borders, because a chaotic, uncontrolled immigration cannot be allowed. the main objective of eu policy in this topic should be to encourage legal and orderly immigration and deter illegal flows. what should be the european criteria for who is welcome to europe and who is not, to avoid an uncontrolled migration into our continent? how could be accomplished the aims to become a historic leap in the consolidation of a european area of freedom of movement without internal borders? a spanish writer, de prada ( ), has recently reminded the clairvoyant solutions offered by thomas aquinas already in the middle age on the obligations and limits of hospitality, making clear that the help demanded by those who suffer should not be confused with their unconditional reception. he refers to several possible types of peaceful immigrant: who passes through our land in transit to another place; who comes to settle in it as an outsider; and who wants to fully join the nation that receives it "embracing their religion" (their culture, we would say nowadays). he always puts the notion of common good, which requires a desire to integrate into the life of the host country; and he is inclined not to admit them until examining their degree of "affinity" with the nation that receives them. and, as a measure of legitimate defense, we should reject those immigrants considered hostile, understanding as such not only those who have the purpose of perpetrating crimes or violence, but in general those who harbor intentions contrary to the common good of the nation that receives them (see also : ramirez ) . the decision on who is welcome and who is not does not belong to the humanitarian people, even less to the people-smugglers, but to each state. security and protection of the borders is competence of each state member. without borders there are no states and no one else can supplant a state's right to manage and protect them, treating migrants humanely but also firmly, swiftly returning those who arrived illegally or whose claims to asylum have failed. even if the eu lacks competence over it, it does not preclude the convenience of an europeanization of the migration management, through a joint cooperation for specific purposes, even if it means losing part of the national authority over a very sensitive area of the territory. the political leaders of the eu must improve their coordination and develop common legislation on borders and immigration, creating systematic and controlled arrival and integration programs, focused on the new scenarios, such as the solution of the migratory crisis, with a common, effective, and decisive security policy, finding a balance between the implementation of measures that guarantee the security of the states, while respecting the rights of those migrants who no longer enjoy their protection in their respective countries of origin. the eu not only lacks competence over the management of the flow of refugees, but also lack of homogeneous response, as it is shown by of the dublin regulation (eu european parliament ), which has resulted an absolute failure, inept to establish a coordinated strategy with minimal effectiveness. the eu foreign policy remains hopelessly underpowered, limited to coaxing national capitals towards agreement and supporting their ad hoc initiatives (the economist economist a, b; niño ). it is good to remind again that it is not eu to blame but the states not taking actions. the dublin regulation ( ) determines the eu member state responsible for the examination of an application for asylum seekers of international protection under the geneva convention relating to the status of refugees of july , as supplemented by the new york protocol of january (eu ). according to this legislation, the first member state in which the asylum application is submitted will be responsible for the examination of the request for international protection and the asylum seekers have to stay there. this, the so-called "one stop one shop", burden the countries situated at the entry to europe, notably those in the south, and the individual preferences -that is, where people arriving into europe actually want to go to and where do they wish to live-are bound to not be properly taken into account. consequently, if they move later to another european country -known as secondary movements-, this second receptor may return them to the first state. this secondary migration penalizes those southern countries, because most refugees arrive to the coasts just as a transit towards other northern european countries, where many of the newcomers had linguistic and cultural or family ties. or with better job opportunities and welfare provision. as a matter of fact, most of them live already in germany. in front of this, the different eu member states have rather antagonistic approaches: (a) northern countries, preferential asylum for most of the refugees, are in favour of a quotes policy imposed by brussels' "eurocrats" for the reubication of refugees, and suggest an eu budget with more funds for those regions with higher number of asylum seekers. (b) mediterranean countries, plus portugal and france, suffering a heavy migratory pressure in their borders, have the feeling of being left alone to cope with immigrants, and reject the responsibility of attending by themselves everybody who arrives to their borders, because it would mean establishing a sort of sanitarian cordon with the rest of the communitarian block. their aim is a reform of the dublin regulation and the help of the other eu countries in the sharing of the irregular immigration arriving to the outer borders. for instance, the greek government and aid agencies argue that the eu must overhaul the dublin regulation, so that asylum-seekers are distributed more evenly. the current system, they say, is unfair. five countries-greece, italy, spain, france, and germanyreceived over three-quarters of europe's asylum applications in the first half of . greece has had to deal with times as many claims as hungary, a country of comparable size and wealth. the rules are also ineffective: eu money is not an adequate replacement for relocation. southern countries are also reluctant to expand frontex because this affects their rights to the sovereignty of control of the territory, but also concerns related to better registration of migrants…. this is the key problem! (c) the visegrad countries (v ) -poland, hungary, czech republic and slovakiaplus baltics (latvia and lithuania) and some recent addings (at least, austria, and partially italy), call for reform, and refuse to consider any binding sharing in , germany received . applications, almost double that of any other eu country, requesting the return of . refugees to other eu countries, but only managed to execute % of the returns of asylum seekers requesting from the eu. based on that, the german home minister, horst seehofer, wants to send tens of thousands of migrants to italy, and, on the contrary, italian home minister, feels legitimized to send those who arrive to the northern european countries; f.ins.,"france was to welcome , immigrants, but in reality accommodated only " (france has returned to italy , people in ). following the principle of solidarity, which is included in the lisbon treaty ( ), jean-claude juncker called on the member states of the eu to share refugee quotas according to the wealth of each country and population size, but it shattered and the european policy of immigration and asylum was marked by increased sovereignism in the states of europe. out of asylum-seekers. based on their right to security, they insist that their priorities are more about controlling europe's outer borders, in order to preserve the free movement of persons within the schengen space, than about managing the burden of refugees who cross them. what should be a reasonable attitude to avoid an uncontrolled moving of illegal migrants within the schengen area? the intended distribution of the burden of refugees imposing a quota system -i.e. every eu state had to admit a fixed amount of people-has not worked: only a % of the quota has been achieved by the different members. this system of quotes was firmly opposed by the central europe defence cooperation (cedc), arguing that the structures of their countries are not "ready" for supporting the social weight of the newcomers: it would be highly detrimental to the national security and sovereignty. they felt marginalized by angela merkel decision to keep germany's borders open to refugees at the peak of the crisis in , without any previous consultation; it seemed to them, the chancellor had turned them into transit corridors for undesirable migrants drawn by the promise of a cushy life there. they suggest as alternative to accept the free choice of country. following this rationale, the polish minister of foreign affairs in years - , witold waszczykowski, was backing the disagreement of his president, andrzej duda, to force the entrance of immigrants against the wishes of the polish people, asserts that "the security of poland is more important than the unjustified decisions of the european institutions on the issue of the refugees". viktor orban even dares to declare central europe a "migrant-free zone". the central european countries are pushing for the eu to get involved in the custody of the external borders and accuse the southern countries of letting irregular migrants pass unchecked through their territory to the rest of the eu, and ending up circulating in europe at will. for instance, sebastian kurz, at that time austrian kanzler and rotatory president of the council of the eu, told in a local newspaper "i am afraid that many are trying to avoid the high number of migrant records that they would have to do" (referred in abc, / , ) . his words suggested that national authorities sometimes turn a blind eye to not registering migrants and that, on the other hand, a community official who did not work directly for any state would not allow those distractions. the eu summit in brussels on june th , following a suggestion of angela merkel and horst seehofer, reached a transitory agreement based in two points: (a) bilateral agreements between different states, stopping secondary movements in their inner borders, i.e. sending the refugees back to the state in which their asylum application was first submitted, obviously with an economical compensation; and (b) more resources for setting up centres of transit (internment) or "landing regional platforms" with the task of reviewing asylum applications, "distinguishing between economic migrants and those needed of international protection". this move would speed up the deportations, disincentives dangerous adventures and reduce drownings at sea. this seems to partially accept sebastian kurz's proposal of a "copernican revolution" in asylum policy migratory model, as a new possible alternative similar to the one applied in australia: to implement the number of naval patrols for intercepting boats with refugees and confine them in islands like nauru o papua-new guinea": control first; then generosity. following this model means two things: first, taking tough action along borders, at sea and "upstream" (inside africa) to slash the number of asylum-seekers smuggled to europe; thwarting smuggling, the argument goes, would reduce drownings at sea, and reassure voters who might otherwise be tempted by the far right. second, striking deals with poorer countries to establish camps, run with un agencies, to receive, detain and, eventually, return migrants. it is a laudable but ill-defined goal, because it is unclear where to establish those camps. it was mentioned "in safe countries outside the eu". but where? some members want deals with north african countries to reduce departures, but it still unknown which non-european countries would be willing or able to host such centres. in front of this north african rejection, the danish prime minister at that time, lars lokke rasmussen, revealed that some communitarian countries are austria, which occupied the eu presidency that semester, hoped for a quick agreement to equip frontex with a supranational force of up to , members and to establish a landing platform for irregular migrants in north africa. algeria: just the day after the summit, algerian foreign minister abdelkader messahel said in an interview with the french broadcaster rfi: "i believe that europeans have enough capacity, means and imagination to manage these situations." "it is excluded that algeria opens any retention zone. "morocco: the next day, the moroccan foreign minister, naser burita, emphatically stressed that this country has always "rejected and rejected" in its territory the detention centers for foreign migrants. tunisia: for logistical and political reasons, it could be the ideal candidate to host this type of centers: its coasts are the closest to the maritime zone where the majority of migrants sailing from the western strip of libya are rescued; and, its democratic transition seems to be quite consolidated, being the country in the region that receives the highest volume of eu aid funds per capita. however, the tunisian ambassador to the eu, tahar cherif. abounded in categorical rejection by his political class and public opinion. "for tunisia, the creation of reception centers is a red line," says valentin bonnefoy, a researcher at the tunisian ftdes, a ngo specialized in migration. more than seven years after the revolution, this maghreb country does not have an asylum law, something that many observers attribute to a veiled strategy to avoid becoming a host country. egypt also rejected the possibility, although in a not-so-sharp way: "the eu migrant reception facilities in egypt would violate the laws and the constitution of our country," said parliament president ali abdel seeking to establish camps for failed asylum-seekers in a continental country, but out of ue, more specifically in the balkans. there is even a third possibility to consider: within the own eu border. jean-claude juncker, in his speech on the state of the eu, proposed to establish a more efficient control of the eu's external border, converting frontex, created in and reinforced in as a european guard of coasts and borders, into an authentic border police, but not in charge of rescuing. although still subject to the control of the national authorities, it is provided with broad competences in areas such as the control of irregular migratory flows from outside the eu, the return of irregular migrants to their countries of origin, the detection of illegal steps between countries of the club and the fight against the channels of transit in third countries. on november th , the eu decided to grow it from , s to a standing corps of , , with a % jump in funding next year, to m. e, for the first time, the eu will be able to dispatch gun-toting men and women clad in eu uniforms to patrol its fringes, without asking member-states to cough up guards. in this way, the surveillance capacity in european territory will be reinforced. this necessary hard-headed policy of tough border controls, swift return of illegal immigrants and encouraging would-be migrants to stay home obliges governments to work with others in the control of its migratory flows. europe cannot maintain the schengen system of internal free migration if it does not control its external borders. and, we have to admit it, it does not. the eu has so far failed in all its attempts to establish a common policy to address the problem; it has become impossible to have a level-headed conversation about managing migration in europe. the malta agreement (september rd, ) is a tentative deal, signed by a group of five eu interior ministers (germany and france, as well as finland, italy and malta), on a temporary scheme for a rapid relocation in other countries of asylum seekers, who are rescued from the mediterranean sea. it could open the way to a comprehensive overhaul of the dublin regulation by which the migrant's country of arrival is responsible for verifying the right to asylum of each person. it states that "the relocation member state will assume responsibility for the relocated person" and contemplates a quick procedure to say in advance the number of potential refugees each state is willing to host, setting a period of only four weeks to relocate rescued refugees in the mediterranean in other member states. this would allow the frontline first-arrival countries (the mediterranean), to be discharged from the responsibility of participating in their reception, quite heavy in periods of strong migratory pressure. in return, berlin and paris hope to achieve a reduction in the socalled "secondary movements" or movements of irregular migrants from the country of entry to other eu states. aal in welt am sonntag. in addition, aal said that his country already has millions of refugees from nearby countries, such as syria, yemen, iraq or sudan, so its reception capacity is at the limit. the eu ministers discussed on - - , in luxembourg this proposal to relocate asylum seekers rescued at sea, but they got a lack of solidarity of the whole european bloc, and the sole support of portugal, luxembourg and ireland. the agreement also speaks of a return protocol "immediately after landing", redoubling diplomatic pressure on migrant countries of origin, using "appropriate incentives to ensure full cooperation, including consular cooperation, from countries of origin", although at least it includes everyone passing a "medical and safety exam". it also underlines the need to "commit to improving the capabilities of the coastguards of mediterranean third countries". it is hoped that the deal will put an end to the game of pass-the-migrant spectacle of some countries squabbling over which should accept responsibility for small numbers of asylum-seekers, breaching in what until now has been a guiding principle: that irregular migrants must be dealt with by their country of first arrival. since participation in the system will be voluntary, much will now depend on how many countries will take part, how many asylum-seekers each will accept and whether penalties are to be imposed on those countries that refuse to co-operate (this last seems unlikely, given that such an idea would have to be approved by all the eu's members, including the countries liable to be affected (economist ). summarizing, there is a widespread perception that a series of instruments to control its external borders are not working well -the dublin asylum agreements, refugee quotas, frontex, return and development policies…-and that a lack of trust has grown not only between the member states, but also between the states and the european institutions, especially after the refugee reception crisis in . in few words, there is a deficient management of migration and a weak european solidarity on this issue (de la cámara ; de wenden ). the call thus is now for a joint cooperation towards a true common policy on asylum and immigration with a complete and efficient control of the eu's external border, reinforcing the role of frontex. everybody accepts this priority as the only possibility left for preventing a reinstatement of inner borders controls which would lead to the end of one of the most valued achievements of the eu, the schengen space. a positive step in this direction has just being announced, after the starting of their new mandate, by margaritis schinas, vice president of the european commission in charge of completing a new architecture that guarantees solidarity to the countries of the external border: the eu is going to make sure that there is a sufficient dimension of responsibility in terms of border control and returns. and as tangible measures in that direction, they promise an improvement of the management of external borders with , permanent staff; moving towards a federalization model of border management, this coming spring there will be the first community corps with their own boats and carry weapons, with the deploying the first european coastguards. the responsibility concerning the immigrants does not finish at the border. once accepted as our guests, we cannot forget about them. and, among our humanitarian obligations, we have to help properly integrating newcomers into our own society, avoiding their eventual fondness towards a multiculturalism, which only produces ghettos of their own culture, or wrongly thinking that the mere acceptance as migrants give them the right to have a claim to social welfare. and, on the contrary, who arrive without being invited cannot impose the host other people's rules. once they find a home in a new country, they have to accept the laws, rules, traditions and culture of the adopted society that welcomes them, learn the local language, mix with the natives, and be aware of the specific needs and priorities of their new home. in few words, they come to european territory with equal rights and obligations, but no more. whoever is not able to assimilate to their new society, should better return to their origin (ramirez (ramirez , . according to the dublin regulation, asylum-seekers are the responsibility of the first country to record their presence and, if they move later to another european country, this second receptor may return them to the first state, most of them arrive to the southern coasts heading north, trying to slip there unnoticed, as illegal paperless "invisible migrants", to avoid being picked up and returned to their first country of arrival. officials in the country of arrival can also use bureaucracy to slow the process of registration. and so, after six months, the new host state becomes responsible. this gives a strong incentive for foot-dragging. the recent malta tentative deal ( ) tries to partially solve this problem, allowing the frontline states to be discharged from the responsibility of participating in the relocation of the newcomers. once accepted as our guests, what to do with them? let us focus on the policy towards them in sweden and germany, preferential asylum of most migrants. strange enough, they are far from coincident at all. in sweden, immigrants received ample food and shelter, a generous welfare benefits, . times higher than the ones received by host taxpayers, as well as many facilities for family reunification, without any special focus on their social integration. even more, most of them are not allowed to work. this almost open bar policy, paying migrants to do nothing, has produced (a) an increment of the social expenses on the foreign population, with the consequent reduction of the benefits for the natives -the newcomers have priority to the swedish people; (b) a call effect on other tentative immigrants; and (c) increase of criminality and insecurity, which triggers the anti-immigrant feelings of host taxpayers who feel swamped. in my opinion, migrants should be encouraged to work, getting them language classes and education and offering job training, and introducing them into the labor market, meeting the main demands of the host country: f. ex, in nordic countries, anticipating a shortfall of geriatric nurses. in germany, the arrival of nearly one and a half million asylum seekers since polarized the country, with a consequent rise of a climate of political anxiety. it seems they have finally realized that the solution of the problem is to integrate into the labor market those who are already in the country, and relax the requirements to allow the entry of more workers. with this purpose, in december , the german government approved the fachkräfteeinwanderungsgesetz ("skilled workers immigration law"), an ambitious legislative package to attract qualified labor from non-eu countries and alleviate the pressing lack of workers in some sectors and regions of the country. this will extend the rules covering foreign graduates to vocationally trained workers, cautiously opening the door for rejected applicants for asylum, but who are already integrated into the labor market, and recruit more workers from outside the eu with medium and high education to find qualified work under certain conditions. germany economy enjoys enviable health, registering an unemployment rate of . %, the lowest since the reunification of the country. figures from the institute for employment research (iab), under the ministry of labor, estimate that there are about . million jobs vacancies, while posts actually registered in employment offices in november amounted to , ( , more than the same month of the previous year). however, one of the biggest concerns of the german businessmen, the so-called mittelstand, the motor of the exporting power and the backbone of the german economy, is the lack of skilled workers. unlike in other european countries, small and medium-sized german companies are distributed throughout the country, not necessarily concentrated in industrial centers. this often hinders the recruitment of workers who prefer to live in urban areas. there is a shortage mostly of workers with a level of professional training, such as plumbers, or drivers, and diploma courses, for example, of children's educators. elderly care and tourism, are also crying out for workers. this legislative text allows non-eu citizens to go to germany to seek work for six months provided they have the necessary training, know the language and have the means to survive during that time. in the case of persons with temporary permits, such as rejected asylum seekers, they must show that they have been working in germany for at least months and that they do not have a criminal record. the head of the employers' association, ingo kramer, recently indicated that , of the asylum seekers who arrived in are working or receiving some vocational training. in addition to the labour shortages, there are also demographic forecasts, which warn of a marked aging of the population. these projections will also affect eastern europe, where a good part of foreign workers in germany come from and whose arrival is expected to decrease in the coming years. according to thomas liebig, expert on migration issues at the oecd, "these countries are becoming places of immigration. we have to look for workers outside the eu". although the labor integration of immigrants advances, experts warn that it will take time. "it's a slow process; sometimes they are people who do not speak a word of german, and many of them come from afghanistan or somalia, where they have not had schooling" explains liebig ( ) . tangible achievements, indeed! milanovic and branco ( ) proposes an intermediate position between open to all or closed borders, under the name of "circular migration": to admit workers who cover specific positions, having salaries and work conditions similar to those of the locals, but with not vote rights, nor social benefits for children, retirement or free education; and at the end of their contracts, they will have to return to their countries. in few words, they will be partial citizens for a limited time, like it is already happening elsewhere: in gulf, singapore, iraq, usa, canada. perhaps one country may need extra workers and be in position of offering them stable working conditions; but this may not occur in other eu members. this is also suggested by alejandro portes, princess of asturias of social sciences award : offering a temporary, flexible and comprehensive program of visas to allow access to young people who want to come to work or study for a while, allowing a fruitful flow between countries. come in that way would also be much more economical, and would do a lot of damage to the traffickers, because those who come in that way would not have to pay them ( october ). psychological experiences of refugees and the response of the community in the lake chad region summa theologica (fathers of the english dominican province translation, prima secundae, question letter to the editor, the economist de la cámara m ( ) the eu migration challenge inmigración y cuestión religiosa, abc, ago de ramón-laca j ( ) operación sophia: paradojas de la acción exterior europea. análisis en minutos | nº a new european pact on immigration and asylum in response to the "migration challenge the southern frontline: eu counter-terrorism cooperation with tunisia and morocco the problem with eu foreign policy. too much historical baggage, the economist a migrant move in malta. the eu reaches a tentative deal to share out migrants rescued at sea. the economist taking european defence seriously: the naval operations of the european union as a model for a security and defence union, cuadernos europeos de deusto los buques de la operación sophia en el dique seco: elementos de un revés para la política común de seguridad y defensa regulation (eu) no / of the european parliament and of the council of maritime dimension in the fight against illegal migration on the western mediterranean route sbarchi? un nuovo schiavismo leguina joaquín ( ) migraciones sin respuestas alemania busca extranjeros para , millones de empleos, el país, dic how to stop migration towards big cities? research gate la inevitable migración hacia europa, el pais confessioni di un trafficante di uomini. reverse niño elena ( ) terrorismo e inmigración: cambiando el foco lazos y nudos con marruecos, el mundo europe's partnership with morocco. proyect syndicate apostolic constitution exsul familia nazarethana, castel gandolfo portes a ( ) princess of asturias of social sciences award buenismo ante el problema de los refugiados, migraciones en el siglo xxi: riesgos y oportunidades el terrorismo como desafío a la seguridad global the refugees issue in the frame of the european security: a realistic approach bondad y buenismo, abc, ago sarah r ( ) le soir approche et déjà le jour baisse turkish directorate general of migration management (turkish: göçİdaresi genel müdürlügü) ( ) migration management global compact for safe, orderly and regular migration. marraquesh global trends: forced displacement in he is member of the advisory board of the society for terrorism research and of the professors world peace academy. he has also chaired the complutense research group on sociopsychobiology of aggression and the departments of psychobiology at the seville and complutense universities, as well as being director of the rector office at the autonomous university of madrid. dr. martin ramirez was a humboldt and fullbright fellow but, let us close here our considerations, stressing that a revision of national and eu legislation is required, focused towards a cooperation with the countries of origin and transit; and that, instead of dreaming naively in a chimeric -non-existing-european eldorado, we have to promote the great possibilities that african continent has, knowing that, as the journalist lucia mbimio says, "we must not stop dreaming, but change the compass of dreams! returning from europe to africa is not a failure." ( , ). key: cord- -j za g p authors: kaden, v.; lange, e.; hänel, a.; hlinak, a.; mewes, l.; hergarten, g.; irsch, b.; dedek, j.; bruer, w. title: retrospective serological survey on selected viral pathogens in wild boar populations in germany date: - - journal: eur doi: . /s - - - sha: doc_id: cord_uid: j za g p the objective of this study was to retrospectively evaluate the occurrence of porcine parvovirus (ppv), aujeszky’s disease virus (adv), transmissible gastroenteritis virus (tgev), porcine respiratory coronavirus (prcv), porcine reproductive and respiratory syndrome virus (prrsv) and swine influenza virus (siv) in selected wild boar populations in germany (n = , ). commercial enzyme linked immunosorbent assay and hemagglutination inhibition tests were used for serological monitoring. the serosurvey revealed seroprevalence rates of . %, . %, . %, . %, . % and . % for ppv, adv, prcv, siv, prrsv and tgev, respectively. the seroprevalence rates differed between populations and age classes with the highest number of antibody-positive wild boars in older animals (> year old). no antibodies to tgev were found in baden–wuerttemberg and in mecklenburg–western pomerania (investigation period / ). in addition, sera collected in mecklenburg–western pomerania in / were negative for siv. even though the seroprevalence rates established for these viruses, except for ppv, were relatively low, wild boars may act as a reservoir for pathogens and a source of infection for domestic pigs and humans. based on the epidemiological situation, no risk of a spread of these viruses should emanate from wild boars, neither for wildlife nor for livestock. however, effective and science-based disease monitoring programmes should continuously be carried out in wild boar populations. wildlife diseases may represent a potential threat not only to local wildlife populations but also to domestic animals and humans. various studies have been carried out to analyse the prevalence of pathogens in wild boar populations and the role of these populations as reservoir for pathogens or a source of infection for domestic pigs. not only viral and bacterial agents have been diagnosed in european wild boars (e.g. markowska-daniel and pejsak ; albina et al. ; gortázar et al. ; vicente et al. vicente et al. , lipowski ; jacobson et al. ; vengust et al. vengust et al. , bonilauri et al. ; dezorzova-tomanova et al. ; ruiz-fons et al. martínez et al. ; kaci et al. ; de deus et al. ) but also parasites (e.g. takacs ; de-la-muela et al. ; fernandez-de-mera et al. ) . normally, the prevalence rates of infections in european wild boar (sus scrofa l.) correlate with the population density of the animals. during the last decades, populations have increased not only in germany (lutz and wurm ; kaden ; gethöffer et al. ) but also in other european countries (sáez-royuela and terilería ) . this development is one of the reasons why wild boars may become a more important potential reservoir for different infectious pathogens. in germany, several studies have been carried out to analyse the presence of different pathogens in wild boar populations. so far, the role of infected wild boars as a potential reservoir for pathogens or a source of infection for domestic pigs has been analysed in detail for classical swine fever virus (csfv; dedek et al. ; dahle et al. ; kaden et al. ; oslage et al. ; lutz and wurm ; kern et al. ; fritzemeier et al. ; schlüter and kramer ) . furthermore, serological investigations in individual wild boar populations in germany have revealed the presence of different other viral agents, e.g. aujeszky's disease virus (adv, pseudorabies virus; dedek et al. ; dahle et al. ; oslage et al. ; lutz and wurm ; müller et al. müller et al. , kaden and müller ; lutz et al. ) , porcine reproductive and respiratory syndrome virus (prrsv; oslage et al. ) , porcine circovirus type (schulze et al. ; knell et al. ) , porcine parvovirus (ppv; dedek et al. ; liebermann et al. ; lutz and wurm ) , swine influenza virus (siv; dedek et al. dedek et al. , polley et al. ; kaden et al. ) , bovine viral diarrhoea virus (dahle et al. ; schmitt and wittkowski ) and hepatitis e virus (kaci et al. ) . within the framework of studies on the oral vaccination of wild boars against csf in germany (kaden et al. ) , sera were collected for serological investigations on the presence of csfv. the objective of this study was to retrospectively assess the occurrence of infections with adv, prrsv, ppv, siv, porcine respiratory coronavirus (prcv), and transmissible gastroenteritis virus (tgev) in wild boar populations of different vaccination areas. the samples were collected from wild boars bagged in five german bundeslaender (federal states; fig. blood samples for the serological survey were taken by hunters from the pericardium or the thoracic cavity immediately after the animals had been shot. the samples were sent to the local veterinary diagnostic laboratories of the bundeslaender or to the friedrich-loeffler-institut fig. sampling areas (grey) vs areas with oral vaccination against csf (samples from mecklenburg-western pomerania) where they were centrifuged at , to , rpm, and the sera were stored at − °c until use. all samples were collected year-round in areas where oral immunisation against csf was carried out. however, most of them were taken during the main hunting seasons (from october to february). the samples were derived from animals of the following age classes: . % from young wild boars (≤ year old), . % from sub-adults (> to years) and % from adults (> years old); . % of the collected sera could not be assigned to a specific age class. commercially available enzyme linked immunosorbent assays were used in accordance with the manufacturers' instructions for screening of antibodies to the following viruses: adv (chekit aujeszky, idexx laboratories/ bommeli diagnostics, liebefeld-bern, switzerland), prcv and tgev (ingelvac® tgev/prcv-diagnostikum, boehringer ingelheim vetmedica gmbh, ingelheim, germany), prrsv (prrsv antibody test kit, idexx laboratories, ludwigsburg, germany) and ppv (ingelvac® ppv-diagnostikum, boehringer ingelheim vetmedica gmbh, ingelheim, germany). in contrast, the sera were tested for antibodies to siv using the hemagglutination inhibition (hi) test based on a slightly modified standard protocol of the office international des epizooties (oie diagnostic manual, http://www.oie.int/eng/normes/ mmanual/a_ .htm). these sera were checked for antibodies to siv h n , h n and h n as described by kaden et al. ( ) . serum dilutions of : to : , were tested and sera with no activity at a dilution of : were considered to be negative for antibodies in the hi test. the statistical analysis was carried out with the statistic programme sigmastat . (spss science software gmb, erkrath, germany) using the mann-whitney rank sum test. the significance level was set at p= . . antibodies to all investigated viruses were detected in the wild boar populations of the various bundeslaender (table ) . exceptions were mecklenburg-western pomerania, where the samples tested negative for antibodies to tgev and siv during the hunting season / , and baden-wuerttemberg, where no antibodies to tgev were found. the seroprevalence rates varied more or less among pathogens and individual populations. the highest seroprevalence rate was detected for ppv ( . %) followed by adv ( . %), prcv ( . %), siv ( . %) prrsv ( . %) and tgev ( . %). the highest ppv seroprevalence rate was detected in mecklenburg-western pomerania in , the lowest in baden-wuerttemberg with . %. the percentage of serum samples with antibodies to adv was relatively low to moderately high, ranging from . % (saxonay-anhalt) to . % (mecklenburg-western pomerania). table indicates the number of serologically positive wild boars per age class and pathogen. in general, the proportion of seropositive animals was significantly higher in older wild boars (> year old) than in the juveniles (< year old) except for wild boars with antibodies to tgev. consequent pathogen surveillance in wildlife may provide an effective epidemiological overview which allows to assess the risk of infection and of a spread of agents within the wild boar populations as well as from this wildlife species to domestic pigs and, in case of zoonotic agents, also to humans. therefore, continuous serological monitoring of wild boar populations is of national and international interest. the present investigations support these intentions and were performed with the aim to retrospectively examine the epidemiological situation with regard to various pathogens (adv, prrsv, ppv, siv, prcv and tgev) in selected wild boar populations over different years in germany. except for adv, these viruses have recently caused economically important infectious viral diseases in our domestic pig herds. as expected, antibodies to all six viruses were found in the investigated populations, however, with differences between the individual populations. it is not surprising that ppv is widespread in our wild boar populations (seroprevalence rate: . %), and that its seroprevalence rate differs significantly from those of the other viruses tested. most animals tested positive for ppv were derived from mecklenburg-western pomerania. our findings correspond to those established in this bundesland in the s (dedek et al. ; liebermann et al. ) as well as to data previously established in north rhine-westphalia (lutz and wurm ) , in lower saxony and in rhineland-palatinate (gethöffer et al. ). these high ppv specific seroprevalence rates in germany also agree with findings in other european wild boar populations (roić et al. ; ruiz-fons et al. ; vengust et al. ) . although found large differences between the ppv seroprevalence rates in local spanish wild boar populations, our serosurvey for germany does not generally confirm this, only in baden-wuerttemberg the ppv seroprevalence was lower than in the other populations tested. despite the high ppv seroprevalence in german wild boars, this agent does not seem to have a negative influence on the reproduction rate of wild sows as determined in mecklenburg-western pomerania (kaden et al. ) as well as in lower saxony and rhineland-palatinate (gethöffer et al. ). the second highest seroprevalence rate (averaged . %) was found for adv. generally, the differences in the seroprevalence rates between the bundeslaender were lower than for ppv. the proportion of wild boars with antibodies to adv ( . %), which were shot in the eastern part of brandenburg during the hunting season / , corresponds well with previous findings from (müller et al. ) . as the investigation areas were largely identical, the findings suggest a stable epidemiological situation. in contrast to these results in eastern brandenburg, oslage et al. ( ) found a very low seroprevelance ( . %) in western brandenburg. this underlines the differences in the epidemiological situation in this bundesland. whereas the seroprevalence rates for adv generally were on the same level in mecklenburg-western pomerania and brandenburg, the proportion of antibody-positive wild boars in saxony-anhalt ( . %), baden-wuerttemberg ( . %) and rhineland-palatinate ( . %) was significantly lower (p> . ). in the s, a similar seroprevalence rate ( %) was found in north rhine-westphalia (lutz and wurm ) , whereas investigations in lower saxony showed a very low adv prevalence of between . % and % in wild boars (dahle et al. ; gethöffer et al. ). the latter authors found % of the wild boars in the eifel region, rhineland-palatinate, to be antibody-positive to adv, the investigations in the pfalz region (palatine forest) showed a negative result (n= ). these findings do not correspond with our experiences in the pfalz region. we suppose that the higher sampling rate in our study better reflects the epidemiological situation in this population. considerably higher percentages of seropositive wild boars were detected in other european wild boar populations, e.g. between % and . % in spain, - % in slovenia (vengust et al. (vengust et al. , , . % in croatia (Župančič et al. ) and % in macedonia (gagrčin et al. ) . in contrast, the proportion of wild boars seropositive to adv seems to be relatively low in the netherlands (cromwijk ; elbers et al. ) . although coronaviruses, especially tgev and prcv, are important pathogens in commercial pig farms worldwide, only little is known on the epidemiological situation in european wild boars. in slovenia, vengust et al. ( ) found no antibodies against tgev in wild boars; however, % of the investigated samples were seropositive to prcv. a sero-surveillance study in feral pigs in the usa (saliki et al. ) also was negative for tgev. our serosurvey underlines that the occurrence of tgev ( . % antibodypositive animals) was generally lower than that of prcv ( . %) in the investigated wild boar populations. none of % animals with antibodies) . the prcv seroprevalence rates in saxony-anhalt ( . %) and in baden-wuerttemberg ( %) were significantly higher than the tgev seroprevalences in both bundeslaender. at present, it is not possible to assess the reason for this relatively high seroprevalence rate for prcv in saxony-anhalt. as prcv infections in pig farms do not play an important role in this region (tyrpe, pers. communication), livestock does not seem to be the source of infection for the wild boar population of saxony-anhalt. hence, further investigations should be carried out with the aim to evaluate the development of the epidemiological situation regarding prcv in the population including a risk assessment for wild boars and domestic pig farms. the low proportion of wild boars with antibodies against prrsv in this study generally corresponds with previous serological investigations in brandenburg, saxony-anhalt, north rhine-westphalia, lower saxony and rhineland-palatinate (oslage et al. ; lutz and wurm ; gethöffer et al. ). interestingly, a relative high proportion ( . %) of wild boars with antibodies against prrsv was found in saxony-anhalt. as prrsv is widespread in pig herds, a spread of this virus from domestic pigs to wild boar cannot be excluded. virus transmission from livestock to wild boar can take place by different routes. the main sources presumably are infected manure and slurry as well as kitchen waste (feeding of animal waste is banned in the eu) as the un-enveloped prrsv shows a high tenacity and survives in the environment for a relative long time. direct contacts between infected domestic pigs and wildlife are supposed to be very rare. in contrast to our findings, wild boars were found to be free from antibodies to prrsv in some european countries, e.g. in spain ruiz-fons et al. ) , in croatia (Župančič et al. ) and in slovenia (vengust et al. ) . likewise low seroprevalence rates were reported by albina et al. ( ) in french wild boars. infections with this arterivirus also occur in italy (bonilauri et al. ) . antibodies against siv (on average . %) were found in wild boars of all investigated areas except for mecklenburg-western pomerania in dedek et al. ( ) . the seroprevalence rates in the other bundeslaender presented here showed large differences, ranging between . % in brandenburg and . % in baden-wuerttemberg. the high proportion of seropositive animals in baden-wuerttemberg cannot be explained at the moment. our investigations indicate that especially siv subtypes h n and h n are circulating in the population with a dominance of subtype h n which is also predominant in domestic pigs in europe (van reeth ). however, antibodies against all three siv subtypes may occur in european wild boars (summarised by ruiz-fons et al. ). antibodies against the tested viruses were present in animals of all age classes. as expected, the seroprevalences generally were higher in the older animals (> year old) except those for tgev. we can only speculate on the origin of antibodies in the juveniles (≤ year old) as we do not have any information on the precise age of these wild boar piglets. based on the experiences with other diseases, it must be assumed that the antibodies of serologically positive juveniles (wild boar piglets) are of maternal origin during the first to months of life; later on, they are induced by natural infection. however, maternal antibodies to ppv may obviously persist longer, i.e. for up to months (johnson et al. ). in conclusion, our study shows that ppv, adv, tgev, prcv, prrsv and siv are present in german wild boar populations. the detected seroprevalence rates of most pathogens were relatively low, except that of ppv. the high seroprevalence of ppv suggests an endemic status of this virus in our wild boar populations. as ppv is widespread in european wild boars and domestic pigs, no risk emanates from wild boars at present. independently of this assessment, wild boars may generally act as a reservoir for pathogens and as a source of infection for domestic pigs. however, it is important to recognise that virus transmission between wildlife and domestic animals is not a oneway street. for example, wild boars may also become infected indirectly or directly through infected pigs. to assess the real epidemiological situation in wildlife and the risk of disease for wild boars and livestock, effective and science-based disease monitoring programmes in wild boars must be carried out, especially for economically important and notifiable agents that may affect wild boars and domestic pigs as well as humans. these surveillance programmes should not only analyse prevalences of pathogens but also include wildlife-biological parameters and management strategies as well as the molecularbiological characterization of pathogens circulating in wild boars and domestic pigs. the latter might become important for the evaluation of the epidemiological process. based on the experiences with csf control in wild boar populations (kaden et al. ) and the risk of an introduction of new or re-emerging diseases to europe, as e.g. african swine fever which just now circulates in domestic pigs, free-range backyard pigs and/or in wild boars in armenia, georgia and russia (www.oie.int), surveillance should include the whole territory of a country and should be carried out continuously, that is year-round. aujeszky's disease (ad) and procine reproductive and respiratory syndrome (prrs) virus infections in french wild boars from presence of prrsv in wild boar in italy serologisch onderzoek bij wilde zwijnen op de veluwe (serological investigations on wild swine in the veluwe region) antibody prevalence of hog cholera, bovine viral diarrhoea and aujeszky's disease virus in wild boar in northern germany epidemiological study of hepatitis e virus infection in european wild boars (sus scrofa) in spain helminths of wild boar in spain ergebnisse flächendeckender serologischer untersuchungen beim schwarzwild (sus scrofa) in einem bezirk der ddr. verhandlungsbericht des . internationalen symposiums über die erkrankungen der zoound wildtiere zum vorkommen von antikörpern gegen influenza-a-virus beim schwarzwild detection of lawsonia intracellularis in wild boar and fallow deer bred in one game enclosure in the czech republic serosurveillance of wild boar in the netherlands, - wild boar helminths: risk in animal translocations epidemiology of classical swine fever in germany in the s wild pigs reservoirs of aujeszky's disease reproductive parameters of wild boar (sus scrofa) in three different parts of germany natural aujeszky's disease in a spanish wild boar population the prevalences of brachyspira spp. and lawsonia intracellularis in swedish piglet producing herds and wild boar population observations on the epidemiology of porcine parvovirus detection of hepatitis e virus in archived german wild boar samples bekämpfung der klassischen schweinepest beim schwarzwild gefährliche verwandtschaften: schwarzwildein natürliches reservoir für infektionserreger und ansteckungsquelle für hausschweine? forschungsreport nr. . bundesministerium für ernährung die schweinepest in deutschland in den jahren und . forschungsreport nr. oral immunisation of wild boar against classical swine fever: concluding analysis of the recent field trials in germany classical swine fever (csf) in wild boar: the role of the transplacental infection in the perpetuation of csf diagnostic procedure after completion of oral immunization against classical swine fevere in wild boar epidemiological survey of swine influenza a virus in selected wild boar populations in germany incidence of classical swine fever (csf) in wild boar in a densely populated area indicating csf virus persistence as a mechanism from virus perpetuation serologische untersuchungen auf porcine parvovirus beim schwarzwild european wild boar (sus scrofa l.) as reservoir of infectious diseases for domestic pigs serologische untersuchungen zum nachweis von antikörpern gegen viren des seuchenhaften spätaborts der europäischen schweinepest und porzine parvoviren beim wildschwein (sus scrofa a long-term survey of pseudorabies virus infections in european wild boar of western germany the sero-prevalence of influenza virus amongst pigs and wild boar in poland torque teno virus (ttv) is highly prevalent in the european wild boar (sus scrofa) pseudorabies virus infections in european wild boar-a potential danger for domestic pigs? pseudorabies in the european wild boar from eastern germany pseudorabies virus infection (aujeszky's disease) in wild swine prävalenz von antikörpern gegen die viren der europäischen schweinepest, aujeszkyschen krankheit, und des "porcinen reproductiven and respiratory syndrome zum vorkommen von antikörpern gegen verschiedene influenza-a-subtypen bei haus-und wildschweinen in prevalence of antibodies to procine parvovirus in wild boars (sus scrofa) in croatia seroprevalence of six reproductive pathogens in european wild boar (sus scrofa) from spain: the effect on wild boar female reproductive performance a review of viral diseases of the european wild boar: effects of population dynamics and reservoir rôle serosurvey of selected viral and bacterial diseases in wild swine from oklahoma the increased population of the wild boar (sus scrofa l untersuchungen zur verbreitung von bvd-virusinfektionen bei schalenwild in bayern identification of postweaning multisystemic wasting syndrome in european wild boar (sus scrofa) contribution to the helminth infestation in wild boar (sus scrofa l.) in hungary avian and swine influenza viruses: our current understanding of the zoonotic risk presence of antibodies against aujeszky's disease virus in wild boar (sus scrofa) in slovenia a serological survey of selected pathogens in wild boar in slovenia antibodies to selected viral and bacterial pathogens in european wild boars from southcentral spain serosurvey of aujeszky's disease virus infection in european wild boar in spain prevalence of antibodies to classical swine fever, aujeszky's disease, porcine reproductive and respiratory syndrome, and bovine diarrhoea viruses in wild boars in croatia acknowledgements we thank silvia schuparis and sybilla welsch for technical assistance as well as r. dürrwald (impfstoffwerk dessau-tornau) and j. süß (formerly federal institute for risk assessment, berlin) for providing siv strains used for hi tests. furthermore, we gratefully acknowledge anette beidler, th. vahlenkamp and w. boehle for critical review of the manuscript and hunters for the collection of blood samples. key: cord- - ly to authors: skoczkowski, tadeusz; verdolini, elena; bielecki, sławomir; kochański, max; korczak, katarzyna; węglarz, arkadiusz title: technology innovation system analysis of decarbonisation options in the eu steel industry date: - - journal: energy (oxf) doi: . /j.energy. . sha: doc_id: cord_uid: ly to decarbonisation of the iron and steel (i&s) industry is crucial in the efforts to meet the eu ghg emission reduction objectives in … . promoting decarbonisation in this sector will necessarily require the identification, development, and diffusion of breakthrough technologies for i&s production. this paper uses an approach inspired by the technology innovation system (tis) to analyse the development of technology in the eu i&s industry and identify potential avenues of its decarbonisation. we have described key elements of the tis, analyse the functioning of these elements and their interactions in a more general context of innovation dynamics and policy design; the focus has been put on the role of actors and the identification of the main specific blocking and inducement mechanism in the tis to better explain its functioning. risks and uncertainties have also been discussed. we argue that deep decarbonisation in the i&s industry is feasible but its tis requires firm support, mostly political, to finance intensive r&d and reduce the business risk. to this end, all actors shall support more effectively the invention and implementation of new radical production technologies. the recommendations are mostly addressed to politicians although stressing the importance of collaboration of all actors. the i&s industry is on the good track reducing co emission by mtco between and however, observation of the efforts carried out in the i&s industry to increase energy efficiency, confirms that currently there is not enough incentive to encourage steel producers to autonomously undertake more sustainable and less ghg intensive production [ ] , [ ] . although increasing efficiency may lead to further savings, the industry has to make much effort to accelerate the development and diffusion of novel technologies to get out of the current standstill. this requires a technological change of a breakthrough nature [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] . a few breakthrough technologies have been proposed years ago [ ] , [ ] , [ ] , [ ] . although up to now, despite multiannual r&d works followed by the extremely long demonstration phase, these technologies are still not at the market phase [ ] . therefore it is of research interest to analyse the reasons for such a low rate of technology development. this article aims to analyse decarbonisation options in the eu i&s industry using the technology innovation system (tis) approach to deliver policy recommendations. apart from standard elements of tis, this analysis tries to embed considerations of sustainability (which include environmental, equity and competitiveness perspectives). therefore, the article has two purposes in the context of i&s industry decarbonisation: . identify the key elements of the innovation system. study the functioning of such systems as a basis for policy recommendations. this paper contributes to this task by harnessing the technology innovation system (tis) approach [ ] , [ ] , [ ] , [ ] to highlight the potential levers and barriers to promoting technological innovation in this i&s industry. tis is a useful heuristic framework that relies on the analysis of success or failure of a technology based on the performance of the surrounding technological system and trace development and di usion of innovations. tis approach enables to include socioeconomic elements to the analysis [ ] . twomey and gaziulusoy [ ] deliver an overview of concepts and frameworks for understanding and enabling transitions to a low carbon built environment. along with tis, they examine similar frameworks, e.g. multi-level perspective (mlp), strategic niche management (snm), and transition management (tm). a comprehensive review of the literature on innovative theory can be found in [ ] . diercks et al. [ ] stress that "…socio-technical understanding of innovation provides several appropriate analytical concepts that can help to shape our thinking and understanding of transformative innovation policy". the tis is considered as useful for policy recommendations and practitioners as it may provide for both sides novel insight in the understanding of functioning innovative systems. we use the tis approach to evaluate how the i&s industry is performing in a low-carbon transition with regards to some crucial aspects, e.g. a shrinking market in europe, and long transport distances to customers outside of europe. a hydrogen economy, smart grid development, including res booming, possible "green steel" market building. deep decarbonisation calls for an in-depth sociotechnical transition that also covers changes in users' behaviour, culture, policy, industry strategies, infrastructure, and science. if a tis analysis is to aim at improving policies or activities needed to a cost-effective and equitable transition, it must specifically account for sustainability and society implications. in-depth knowledge on innovation systems is needed to plan and control the transition process more effectively by providing the politicians with research-based conclusions [ ] . oda et al. [ ] investigated, based on an intensive and in-depth survey of steel-producing j o u r n a l p r e -p r o o f facilities and energy-efficient technologies in the steel sector. [ ] underline the importance of supporting the industry in their research efforts to make the innovations, often taken for granted, a reality from on. they stress how important it is to keep the values of the decision-making criterion, e.g. payback period, close to normal business conditions to exploit the possibilities offered by the prospective innovative technologies in this industry to the full, and the need to close the efficiency gap by making use of the policy options available. in the literature, a few analyses of national i&s industry development are to be found. two national cases of germany and sweden are worth commenting as the two countries have a specific position in the eu i&s industry -the former is the larger eu steel producer with traditional high technological culture, the latter boasts the high technologically advanced steel industry much focused on energy efficiency improvements. arens et al. [ ] claim that the german steel sector has two options to achieve its european co emissions reduction target for -if it strongly decreases its production output or technological changes are introduced. in this option, first, alternative steelmaking processes need to be developed including low-emission technologies and co -free processes, e.g. h-dr -eaf or electrolysis-based steel production with both routes based on co -free electricity. secondly, there should be a path on incremental co reductions in the short to medium term, e.g. heat recovery from bf slag, and from waste heat in eaf, production of high-quality steel from scrap-based secondary steelmaking, and the utilisation of by-products. arens and worrell [ ] examined the diffusion of six key energy-efficient technologies in the german steel industry and their impact on energy intensity. some novel technologies diffused at a quicker rate at the initial stage but since the s the rate levelled off. the diffusion rate depends much on developments in the sector, e.g. shutdowns or application of energy-efficient technologies. they estimated the primary energy efficiency potential of the selected technologies at . % for . in , the steel industry in sweden adopted a vision leading up to with three pillars -to lead technical development, to nurture creative individuals, and to create environmental benefits. these undertakings demonstrate importance on technological innovation with clear objectives in societal value creation and sustainability [ ] . the efforts of the swedish i&s industry to improving energy efficiency and implementing new technologies are very well documented [ ] , [ ] , [ ] . the main conclusion might be that the industry is aware of energy efficiency importance, but more effort is required as energy-saving potential remains untapped. regarding breakthrough technologies, sweden is proud of the hybrid project aimed at producing hydrogen by electrolyzing water and then using the hydrogen to make direct reduced iron (h-dri) [ ], [ ] . the eu reach is rarely addressed. recently, mayer et al. [ ] made a top-down macroeconomic analysis of a currently considered technological change in the eu i&s industry towards carbon-neutrality, based on techno-economic bottom-up literature. they analysed economic conditions (electricity and co price) for switches in the type of production process towards process-emission-free iron and steel technologies required for reaching climate targets, even though the current degree of alternative technology maturation varies considerably. they argued that if minimal criteria of cost-competitiveness are reached, macroeconomic benefits are feasible. yazdanparast et al. [ ] compared the gaps between service, product, and process innovations in one of the european steel companies. they claim that innovation success depends on cross-functional collaboration among members of the innovation network to share and leverage resources that are hardly ever available to one company. the research identifies internal and external success factors and challenges of each of the three innovations. vögele et al. [ ] assessed, using modelling, impacts of the global challenges, e.g. floor prices, tariffs, carbon price, on the i&s j o u r n a l p r e -p r o o f industry in the eu. they showed that moving eiis from the eu may increase global energy demand and co -emissions and bring disadvantages to developing countries participating in the value chain. they stressed the need for innovations to avoid relocations of production. the literature on transitions has so far not much addressed eiis [ ] . searching for tis analyses, we found that majority of them are based on the country-to-country and/or productto-product analysis whereas our case is classified as "sectorial". we cite a few recent examples. andreasen and sovacool [ ] used tis for analysing the hydrogen technological innovative system in denmark and the us focusing on fuel cell development. they conclude that although these two countries applied remarkably different tis strategy neither approach was effective. they also stressed the need for further research on tis as "better conceptual models of tis and innovation are needed". kochański et al. [ ] applied tis to analyse electricity smart meter in the eu. this an example of the tis approach focused on a single product (smart meter) discussed in the wide eu context. despite strong eu political support and the potential advantages for customers, the massive roll-out is slower than expected. they identified the major barrier at the level of implementation, e.g. insufficient regulatory framework which does not fully ensure interoperability, data protection, and security standards, or organisational effectiveness. clear presentation of benefits for the end-users has also failed. kushnir et al. [ ] studied the tis approach to adopting hydrogen direct reduction (h-dr) in the swedish steel industry. they identified several potential barriers, particularly in timely co-ordination with the power system regarding new infrastructure for h-dr and eaf electricity demand and noted that the i&s industry will have to compete for res-based electricity. they pointed out the necessity of political intervention to remove the barriers to make the swedish i&s industry the first nearly fossil-free industry in the world after . the paper is organized as follows (fig. ) . in section , the economic importance of the i&s industry for the eu economy, the need for the industry decarbonisation is underlined, and the reasons why the industry is hard for decarbonisation are given. in section , the methodology applied is outlined. the boundaries of the system under analysis are defined in section . section provides identification and discussion on the role of all structural components of the tis, namely infrastructure including physical, knowledge, financial and technology; actors; institutions and policies; institutions, and finally networks and interactions. in section , the mapping of the functional patters of the tis is done and then, in section , assessment of the functionality has been provided. section provides analysis and discussion of blocking mechanisms and inducements of innovation, and in section an outline of risks and uncertainties in the i&s industry is sketched. in section , conclusions and policy implications are presented. because of the complexity, the innovation process cannot be approached by solely focusing on standard measurable variables such as economic criteria, e.g. revenues, productivity, and quantities. understanding the dynamics of technological innovation requires the exploration and characterisation of the economic and social spaces in which the innovative activities take place and understanding of the interactions linking a wide range of actors and institutions [ ] , [ ] , [ ] , [ ] , [ ] . tis enables to identify key elements and then to analyse and assess the effectiveness of their functioning in multidimensional space which in majorities of analysis is simplified to fewer dimensions, e.g. technical and economic. tasks of this research covered: (a) mapping and description of key actors and institutions, and a qualitative assessment of how they interact at the supra-national, national, and sub-national levels, including a description of how the existing landscape evolved, and reasons for it, (b) mapping of innovation strategies, roadmaps and policy instruments across the innovation chain and their development and implementation, (c) identification of key barriers and inducements of innovation through desk research, and then adding supplementary comments on risks and uncertainties. there are some crucial trends (external factors) which make such tis approach in the i&s industry indispensable to make innovation feasible, e.g. global climate policy with the eu as a leader; "green" and circular economy; emerging hydrogen economy; transformation in the power system to the smart grid model which offers new symbiosis with industry, e.g. due to res penetration; increasing storage capacities, grid flexibility. there are also changes in seeking analogy between tis and the control theory, it can be stated that tis is like control system of timedependent dynamic nonlinear plant with highly distributed elements with a feedback. the plant is subjected to many, mainly stochastic noises which make its control highly unpredicted and difficult. some of the signal and j o u r n a l p r e -p r o o f the ways business operates nowadays, e.g. globalization, deregulation, advancements of information technology, and increasing customer demands which force the companies to continually deliver new products. we built on the existing knowledge but unlike other analysis of i&s industry, development our tis-based approach enables to a systematic analysis of the innovation system in a wider dimension than the standard methods, e.g. encompassing societal and environmental issues, networking, external factors. our paper tries to analyse the whole sector, namely the i&s industry in the eu. standard tis approaches are narrower, e.g. focused on the comparison of product development in different countries or industry in one country. to this end, following bergek et al. [ ] , we use a -stage approach for the tis performance assessment ( fig. specify key policy implications. the advantage of this paper is that it strictly follows the methodology of tis. first, we identify and described four key elements of tis, namely infrastructure and technology; actors; institutions; network and interactions [ ] . this approach highlights four constituent pillars which play a crucial role in innovation systems. then we analyse processes that are needed in each tis to stimulate technology development and diffusion. by mapping these processes (functions) over time, the insight into the dynamics of innovation systems can be understood. this step is based on the characterization of the seven functions specified in table and then detailed in section . the "functions" may be understood as key processes driving the development and diffusion of new technologies done in a wide sustainable context (table ) . a comprehensive latest literature review of the functions can be found e.g. in [ ] . [ ] , [ ] , [ ] tis is very flexible in methods of gaining the necessary information, e.g. by case studies, event history analysis, multiple data sources, and interviews [ ] . our research method was based on a systematic review of the literature (srl) since it provides context-specific insights based on systematic, transparent and reproducible review procedures [ ] . this choice is reasonable since the subject of our research, i.e. i&s industry has been very well described in academic literature, political documents, strategies and business reports which can hardly be supplemented by any complementary data acquisition. additionally, the eu dimension of our study would enormously extend the scope of potential interviewers and business confidentiality would make it tough to get data beyond the publicly available. the papers analysed were identified via scopus following the procedure described in appendix a. we also grabbed from the eu and tis national case studies -for the uk and italy made within the innopaths project [ ] . in the tis analysis the main technological drivers of transformation in the eu industry have been considered [ ] : . digitalisation (e.g. industry . ), . advanced materials development, . complex and global supply chains, . market competition and over-capacity, . life cycle design, pollution prevention, and product recyclability, . decarbonisation and energy efficiency, . evolution of customer requirements. the object of study is the innovation system in the i&s industry in the eu and its capacity to adopt new core technologies. this research is limited to nace . "manufacture of basic iron and steel and of ferro-alloys" which is broad and covers numerous technologies and products. the scope of its considered is rather limited to iron ore reduction technologies and does not include the production of semi-finished products of steel and manufacturing of hotrolled and cold-rolled products of steel. it is justified since most of the energy including electricity end-use in steel manufacturing is consumed by furnaces [ ] . the boundary for this analysis was confined geographically to the eu but essential externalities outside europe were considered to eliminate the risk of missing essential external factors. we consider that analysis of the i&s industry for the whole eu is feasible due to the high homogenisation of the whole surroundings secured by close integration of the mss forced by legislative and regulatory harmonization, common internal market rules, and the common framework of r&d. such an approach makes the result of our analysis useful both for the eu and ms levels. it is to note that lutz et al. [ ] observed that there cannot be a general approach for the transformation in the eiis since the capital structure may differ between countries and sectors. therefore, for some countries, analyses must be conducted individually. infrastructure and technology encompass four categories: physical, knowledge, financial, and technology. physical infrastructure includes artefacts, instruments, machines, etc.; financial infrastructure comprises subsidies, financial programs, and grants; and knowledge infrastructure encompasses knowledge, expertise, and strategic information [ ] . technology comprises the subset of knowledge about the full range of devices, methods, practices, and processes that enable the fulfilment of human purposes is a "specifiable and reproducible way" [ ] . silva and de carvalh [ ] provide an extensive account of the history of the steel industry. description of technologies in the i&s industry can be found anywhere, e.g. [ ] , [ ], [ ] , [ ] , [ ] , who summarise the present status of different kinds of steels and relative technologies (incumbent and emerging), review steels over past decades and also forecast of advanced steels in the future. over the last years, the i&s industry in the eu has undergone significant changes, e.g. the elimination of production in the traditional blast furnace (bf-ohf), the doubling of the share of production through the electric arc furnaces (eaf) technology (up to % now) and the almost complete transition to continuous casting [ ] , [ ] , [ ] . two main routes for steel production, i.e. basic oxygen furnace (bof) and eaf, are already well optimised thermodynamically, e.g. [ ] . since the s, steel specific energy consumption in the european steel industry has dramatically decreased by %. however, technical energy efficiency potential of … % still exists and will be essential to reduce emissions from existing, long-lived industrial assets [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] . specific co emissions in europe are some of the lowest in the world - . tco /t of steel, as compared with the global average of . tco /t in (global ccs institute), aiming at . tco /t of steel in assuming hypothetic diffusion of breakthrough technologies in combination with ccs [ ] . based on our literature research, ten ghg mitigation options, which can align the i&s industry with the objectives of the paris agreement, were identified several means of ghg emission reduction [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ ] , [ the iea lists several innovative technologies within the i&s industry [ ] : new smelting reduction process based on coal (trl - ); top gas recovery blast furnace (trl - ); top gas recovery blast furnace with coke oven gas reforming (trl - ); new direct reduction based on natural gas (trl ); direct reduction based on natural gas complemented with up to % electrolytic hydrogen (trl ); direct reduction based on hydrogen (trl ); direct use of electricity to reduce iron oxides (aqueous alkaline electrolysis, low-temperature °c) (trl - ); direct use of electricity to reduce iron oxides (molten oxide electrolysis, hightemperature > °c) (trl - ); carbon capture and storage (ccs) applied to commercial iron and steel technologies (trl - ); conversion of steel works arising gases to chemicals and fuels production (ccu) (trl - ); smelting reduction based on hydrogen plasma (trl ); electrolytic production of hydrogen. their market entry predicted in … is mostly delayed [ ] , [ ] , [ ] , [ ] , [ ] , [ ] . the technologies recently identified by european trl-technological readiness level. j o u r n a l p r e -p r o o f experts from the steel industry as innovative are improving energy efficiency beyond the state-of-the-art; new smelting reduction technologies; direct reduction technologies, based on natural gas; direct reduction technologies, based on hydrogen; direct use of electricity for iron ore reduction; use of biomass in steel production; more recycling of steel and other breakthrough solution paths for low-carbon steel production [ ] . the list of technologies which can be classified as "off-shelf" embraces a few items only, e.g. h-dr [ ] . this selection of reduction options which covers traditional stages of i&s production (e.g. coke making, sintering, bof, eaf) as well as a few breakthrough technologies, was done according to the potential to decarbonization till [ ] , [ ] . the best available technologies (bats) have the potential of emission reduction of … % in the eii, even when applied on a large scale [ ] , [ ] . current bat may contribute to short-term energy and ghg emissions savings in the i&s industry, but the switch to innovative technologies by the mid- st century is far more uncertain [ ] . other options for deep decarbonisation of the i&s industry are shifting to low carbon energy supply via either biomass, sustainable charcoal, nuclear energy, res-based electricity or ccs [ ] , [ ] , [ ] , [ ] . additionally, energy management can serve as an effective tool in improving energy efficiency [ ] , [ ] , [ ] , [ ] , [ ] , [ ] . the main conclusion from quantitative assessment has been that material and energy efficiency measures will help in decarbonisation process, but do not suffice to get the reductions needed. actors within an innovation system "typically include individuals and organizations operating at multiple scales" [ ] . different categories of actors within different sectors have different characteristics, motivations, and strategies which must be appropriately described and understood [ ] . the role of actors that contribute to the success/failure of the tis in the i&s industry is manifold and presents a complex array (table ) . role in i&s industry tis ec develops long-term social and economic strategies like climate-energy policy, industrial policy tries to improve co-ordination among complementary policies, e.g. innovation policy, tax policy, energy market regulation [ ] sets sustainable long-term priorities and objective strongly affected the i&s industry, e.g. political support for climate policy and the following decarbonisation of industry plays a crucial role in setting and the enforcement of climate-energy legislative framework (directives, regulations), e.g. eu legislation supporting deep industry decarbonisation, e.g. energy and material efficiency, eu ets introduces sectorial regulations and set market conditions via market regulations, e.g. taxes, import quotas initiates and finances r&d, e.g. finance i&s industry programs, establish innovation centres e.g. numerous programs supporting innovation policy to keep up with the world's innovation leaders launches short-term intervention programs in times of economic stagnation or crisis or natural disasters institutions are defined as a set of formal and informal rules, norms, decision-making procedures, beliefs, incentives and expectations that guide the interactions and behaviour of actors in an innovation system [ ] . institutions set the environment in which all actors operate [ ] . the ec has warned that "today, the european steel sector finds itself in a very difficult situation" [ ] , and to solve the issue, the ec has taken steps to strengthen the eu's defence against unfair trading practices. the ec called for immediate action for a european industrial renaissance in [ ] . the ec took planes to protect the industry from the unfair international competition including the steel industry, "…notably through modernisation and investments in innovation" [ ] . in , the ec reconfirmed: "energy-intensive industries, such as steel, chemicals, and cement, are indispensable to europe's economy, as they supply several key value chains" [ ] (table ) . trade improving the speed and efficiency with which the ec can begin and resolve trade defence investigations enabling better use of policy instruments to enforce trade defence intensifying discussions through the oecd with major steel producers like china, russia, and india to safeguard eu interests improving the access for eu steel exporters to "third markets" secure access to raw materials as crucial due to scarcity in the eu using eu funds to modernise the i&s industry, including retraining and j o u r n a l p r e -p r o o f development reintegration of redundant steelworkers into the labour market. engage high-level stakeholders in the industry to agree on a plan of action to modernise the industry implement the ec's "circular economy" strategy to reduce waste and increase efficiency in manufacturing energy compensate for energy prices as eu prices are higher than in most of its international competitors supporting energy-intensive industries within existing state aid rules reviewing the eu ets to ensure that the i&s industry remains competitive whilst also reducing emissions implementing the eu energy union source: own work. network and interactions describe the dynamic relationships and links between actors, but a "network" can also be seen as a higher form of "actors" organisation. interactions may take place within networks and can be analysed at the level of both networks and the level of individual contacts [ ] . analysis of networking usually poses a great challenge in tis research. even when the other components of tis are identified, to trace their interrelation is difficult both from the methodological and practical perspectives. the links may be multidimensional, overlapping, and tacit. even when properly recognised, there is neither a standard methodology nor metrics to measure their strength therefore narrative description or tracing connections in literature or patents must suffice. it is common to analyse an industrial sector as an isolated system with no or only a few links to the ambient constituted by other elements of the whole economy and society. however, combating climate change is such a global trend that it has enforced an interrelated transformation in other industries, e.g. a strong and immediate interdependence relationship between coal, power, chemical, and i&s industries [ ] . european companies, operating in a high-wage region, to compete try to create additional value, e.g., more and specialised variants of steel offered, highly sophisticated logistical services, and excellent after-sales service [ ] . analysis of networks in the i&s industry should embrace a wide range of actors and types of collaboration, e.g. suppliers and manufactures in the whole chain, customers, energy endusers, research institutions, multinational research, pilot and demonstration projects, innovative platforms, industrial and trade associates [ ] . r&d works in the i&s industry are often presented as cooperation among global players, e.g. top steel producers, novel technology providers, and leading research institutions. it is, however, to note that at the initial stage, the network also includes small companies, often not related to the specific sector, but with in-depth knowledge which can be supplementary to the steelmaking processes. the industry itself points at the overall immaturity of "collaborative solutions" and their frameworks [ ] . the discussion between the i&s industry and other actors can be led in two ways [ ] : • the institutional channels as the steel community is a part of society at local and global levels e.g. acting in corporate social responsibility, or participation in international initiatives, e.g. within un (global compact), business associations or business circles. • the science-based route related to the role of steel and analysed multidisciplinary as a mixture of soft and hard sciences. in this section, we will systematically analyse the seven functions of the tis in the i&s industry providing indicators of each of these functions (table ) . [ ] , [ ] , [ ] influence on the direction of research the top priority of climate-energy policy sends a clear signal to policymakers on the necessity of deep decarbonisation in i&s industry through technology innovation the high potential of material efficiency in the circular economy, e.g. scrap reuse, by-product utilisation the high potential of ghg reduction through breakthrough technology implementation low expectations of a breakthrough in core incumbent technologies a few identified technologies with limited energy and material efficiency improvements few emerging breakthrough technologies e.g. "green hydrogen", large use of res-based energy, electrowinning, dir, ccsu [ ] , [ ] , [ ] resource mobilisation well established research institutions, e.g. at universities, industrial institutes, and centres, with highly competent staff joint research centres of the academia and industry with public cofunding the co-operation of the main market players (manufactures) in r&d projects continuous sufficient inflow of educated staff at all levels of competence, e.g. trained workers, managers, engineers, scientists [ ] insufficient human and financial resources involved in the demonstrative phase of breakthrough technologies market formation mature market, without a niche, for steel products prone to economic cycles and crises unfair trade rules exercised by some manufactures make innovation technologies difficult to enter into the market evermore demanding environmental standards followed by penalty regulations accelerate r&d and market adoption market demand for improved products set by the main customers, e.g. automobile, construction sector ("learning by using") legitimation global political agreement on the urgency of ghg emission reduction broad consensus of all actors on the need to decarbonise the i&s industry actors, institutions are well established in multiannual historical processes low interest of public opinion in technology innovation in the i&s industry as such, but the high expectation to reduce environmental harm visibility of effects of environment protection measures build-ups of local support, e.g. reduction of air pollution eu and national environmental targets, e.g. ghg emission reduction, set indirectly the targets on i&s industry decarbonisation development of positive externalities acknowledged societal importance, national and local, e.g. retaining jobs flow of knowledge among different tiss, e.g. res-energy, chemical industry, cement industry, automobile, construction, resulting in mutual benefits strong complementarians among technologies, e.g. steel-hydrogen-res, electrolysis-res; in by-products utilisation, e.g. cement industry large potential contribution to the circular economy participation in global trend to the decarbonisation of eii realising the potential for ghg emission reduction and energy and material efficiency . assessment of the functionality of the tis . . knowledge development and diffusion innovation systems in the i&s industry are even more complex than in other sectors since the connections among actors and institutions occur at many stages of the innovation process in multiple sectors and countries and different scales, and are not "visible" to the public. additionally, the market is not directly driven by millions of customers as it happens in the case of many other products. most of the i&s large companies are involved in developing its since they are aware of the importance of ghg emission reduction and their key role in it. they recognise the need for co-operation at different stages of technology development, particularly in r&d and pilot phases despite clear business competitions. government's support is materialised in funding via r&d programs and creation of research centres which does not violate public aid rules and increases the attractiveness of the country to global manufactures [ ] , [ ] . the rate of innovation in carbon mitigation related technologies in the i&s industry has been increasing over the last decades what supports the thesis that climate policy is a strong driver of innovation [ ] , [ ] . the inertia of some tis components, not strictly of r&d nature, may amount to years, e.g. in political or standardisation decisions (see work plan of [ ] , and shall be accounted for in the tis assessment. the diffusion of technologies may therefore substantially vary. grubler [ ] , compared the development of four technologies used for steel production which competed simultaneously with decreasing and increasing market shares. the diffusion trajectories of the processes were diverse, with time constant ranging from less than two decades (replacement of the crucible process) to nearly seven decades (diffusion of eafs). the development of breakthrough technologies cannot take momentum also due to the complex and fast-changing externalities, e.g. energy and feedstock prices, fluctuation in demand for steel. our tis operates in a well-established industry facing a strong need for decarbonisation, although, at the same time, the industry faces limited options for breakthrough technologies soon. unlike many other industries, technological transformation should be mainly carried out with the active involvement of the industry through bottom-up initiatives rather than by new entrants or be directly transferred from academia to industry. the lack of new entrants in the mainstream business weakens the innovation drive among incumbent actors. the demonstration phase in the i&s industry is so risky that it cannot be jumped over or shortened radically. there are pieces of evidence of dropping or mothballing projects caused by economic hardships [ ] . research challenges are well recognised and are mostly initiated by environmental requirements and only partially by a limited demand for higher quality products [ ] . the eu and ms governments support r&d in the i&s industry but they have very limited possibilities to intervene directly into strictly business matters. the i&s industry in the eu can potentially remain a leader in its although competition from other countries is tough [ ] , [ ] . the i&s industry people are aware of the importance of the eu internal market, growing competition from globalised markets, the influence of financial markets on corporate strategies, and commodification. the development of its has suffered from the lack of sufficient r&d financing and resulting human resources involved. political support both at the eu and national levels despite different actions has not brought about the results expected by the i&s industry. therefore, pilot and demonstration stages remain a "bottleneck" in the whole innovation cycle. the industry raises several key business-oriented aspects essential for low-carbon innovation: . cost savings and competitiveness; . carbon price; . developing robust inter-industrial collaboration models; . reduced environmental externalities (delivering improved corporate sustainability reputation); . international competition for low-carbon products [ ] . at present, the energy consumption, ghg emission, and by-product utilisation in the i&s industry are serious political and public concerns that need to be addressed to avoid costs of pollution and make the technological processes sustainable. the i&s industry in the eu will be able to reduce its co emissions by % in an economically viable way until , compared to levels, while the steel market will grow by . % annually (to raise the eu crude steel production to mt in ) [ ] . the divergence between predicted ghg reduction possibilities and the production growth illustrates the market demand for breakthrough its. in response to these plans, the eu i&s industry declares that it will be able to reduce production's co emissions by % by and by … % by if breakthrough technologies are adopted [ ] . to end this, the i&s industry must undergo deep decarbonisation. the eu regulatory framework does not favour any of the emerging technologies postponing in this way possible competition for a more mature phase of technology development. the economic impacts of technological breakthroughs in the i&s industry will likely be nonlinear as system thresholds are breached and have knock-on effects. the market for its will emerge from the first success pilot installations which will be multiplied on other sites depending on the expected revenues, the scale of investment needed, and the threat of stray assets [ ] . . . legitimation steel as a product is very well recognised by people and enjoys high esteem [ ] , [ ] . i&s industry is highly valued by politicians as an economically and socially important and prestigious sector [ ] . steel is a key element of an industrial society and thus for meeting sustainable development goal [ ] . communities in which the steelworks are located also appreciate its importance to the local labour market and prosperity [ ] . the i&s industry tries to demonstrate its commitment to all aspects of "green economy", e.g. energy and material efficiency [ ] , [ ] , [ ] , [ ] . these efforts impact politicians although do not much affect the public therefore the i&s industry receives much less public attention as the environment polluter as compared e.g. with the power sector. the its in the i&s industry and other industries demonstrate in some respects close synergy, e.g. with power, automobile, construction, chemical, defence industries. the positive influence of each other results from common interests and objectives, e.g. these driven by res-based energy, material, and energy efficiency, complementary of technologies, easiness to adopt. the benefits of "spilling over" from one tis to another have not been yet fully recognised and valued although the symptoms of such synergy are "visible" and a prevailing tendency for the interdisciplinary research helps to create joint r&d programs and research centres. competition between the incumbent systems and the emerging ones is a key element in the development of tis. it is considered in categories of "blocking mechanisms" and "inducement" [ ] . the inducement mechanisms are understood as "believed in growth potential and government r&d policy" [ ] . presenting barriers and inducements against the functionalities increased the readability of the text. the vulnerability of r&d programs to external factors, e.g. economic crises ageing of current capacity leads to "greenfield" investments or at least to "brownfield" retrofits the main players in the eu steel industry recognise the decisive role of technological innovation in seeking global market advantage source: own work. r&d investments are needed, both in human resources and physical capacity, otherwise, rates of innovation and technology diffusion are apparent, and the support of different actors within the low-carbon innovation system is lacking. establishing industry and academia joint research programs and centres, e.g. in private-public partnership, gets the key actors closer [ ] . a high level of research in a country and public support helps to save domestic capacity from closing down [ ] . globalisation and overcapacity do not facilitate its in any eiis unless innovation secures high profits with short pay-back time [ ] , [ ] . low cost remains the decisive criterion in purchasing decisions on the global market with overcapacity of around million tonnes in , equivalent to almost % of global steel production capacity what effectively discourage innovative investments [ ] . there are several general trends in global development that give new impulses to technology change and innovation boost. these tendencies help to reject the traditional dogma of cost-effectiveness as a major criterion in technology assessment. on the other hand, periods of overcapacity are suitable for deep technological changes as there is no risk of losing important markets and customers assuming the company remains financially stable. additionally, local capacities will also weight, e.g. availability of qualified workforce, accessibility to res-based energy or reshydrogen, community acceptance for ccs [ ] , [ ] . the reduction of environmental burden due to innovative technology always builds better relationships between the investors and the community. all major actors of the i&s industry are active in the field of digitalisation this industry [ ] , [ ] integration of i&s technologies with other technologies, e.g. chemical, power industry, ict the r&d trajectories are set and it is hard to expect novel technologies to appear soon [ ] , [ ] . the progress in developing the currently researched breakthrough technologies is very slow and therefore requires public support at the eu and national levels [ ] , [ ] . more concentrated public financial support for r&d and then for breaking the demonstration and market barrier is a prerequisite if the i&s industry in the eu is to remain competitive especially in the light of the expected increase of costs due to the eu ets participation [ ] . r&d in the i&s industry is not considered highly innovative and, thus, is not a priority for governments in their national development strategies, neither for research institutions [ ] , [ ] even though these technologies are game-changers for deep decarbonisation. yet, the high value, traditionally attributed in the i&s industry to technological progress and innovation, allows the industry to remain highly advanced although progress in core technologies remains slow. it had been predicted that the currently used (standard) technologies would dominate until [ ] , [ ] . now it seems that the time horizon is likely to be pushed till … . a large number of technological projects in the i&s industry is co-financed from eu funds, e.g. the ulcos (ultra-low carbon dioxide (co ) steelmaking) is a consortium of european entities (all major eu steel companies, some engineering partners, research institutes and universities) [ ] , [ ] , [ ] . however, eu r&d programs seem to be too much dispersed, weakly focused on crucial decarbonisation challenges, and not tackled in a truly interdisciplinary way in a long-term perspective [ ] , [ ] , [ ] . the difficult economic position of the eu i&s industry in recent years -the dilemma of cost-cutting or the need for investments in new capacity possible unwillingness to finance industry operating at highly competitive markets at times of economic instability and hardship possible lack of access to investment capital especially in the difficult economic standing of the company the funding gap between trl - ("valley of death") [ ] , [ ] , inadequate availability of risk capital [ ] high capital involvement and low rate of return from financing high-emission investments lack of upfront clarity about the funding [ ] not fully recognised benefits of it in lca low r&d private funding insufficient to overcome barriers of pilot implementation at acceptable risk insufficient targeted economic incentives for ti, e.g. in the form of tax reduction, risksharing or loan guaranties from private and public financial institutions, governmental quarantines business risks and risks of perturbation in production deter large investments which may result in losing markets, crucial clients, or penalty payments lack of "green" criteria in public procurement [ ] low profits in incremental technological improvements the high share of energy in costs of final products (up to %) rising costs of energy make its more profitable the large share of public funds allocated for "green economy" envisaged in the eu financial perspective for - growing unwillingness or even ban on financing non-sustainable investments (like coal power stations) with a risk of extension on other eii, e.g. i&s industry the expected positive impact of eu ets on low-carbon technological up-take in i&s industry after (modernisation fund, innovation fund) technologies from environmental funds lack of new business models in i&s industries rewarding ti source: own work. very high investment costs in core technologies and long investment time in the unstable global market deter low-emission, high-cost transformation. financial incentives are indispensable to overcome the barriers for such investments to support private capital, e.g. via private-public-partnership [ ] , [ ] , [ ] , [ ] . for example, a demonstration hisarna technology plant of industrial size ( . … . mt/y) will require an investment of € … million [ ] . public intervention should complement and not overlap with existing eu and national funding programmes [ ] . there is a need for viable and robust business models for long-term and deep decarbonisation beyond incremental and short-payback measures. rules of support should be more flexible, e.g. simple two-stage application procedure, aligning the timing of support with funding needs ("funding against milestones"), ensure complementarity between different eu funds [ ] . the breakthrough technologies can hardly be classified as competing at this stage of their development when their actual costs and benefits are not yet properly assessed in the market scale [ ] , [ ] . their implementation will, to a large extent, depend on several criteria, sometimes local, e.g. availability of res-based energy and local feedstock, demand for waste heat, cost, and price structures in a specific country [ ] , [ ] . also, the time of implementation of process emission-free "high-cost" technology, e.g. or , plays its role as early implementation may increase the range of negative gdp implications in [ ] . the emission abatement in industry, e.g. in i&s, may turn out to be much more technologically challenging then emission reduction from energy use [ ] . the innovation diffusion in eiis exhibits a natural long time constant resulting from long decision cycle and investment phase even in pilot installations [ ] , [ ] . securing sufficient and stable financing would substantially reduce the duration of the premarket phases and lower the business risks. the ec intends to "support clean steel breakthrough technologies leading to a zero-carbon steel making process by " [ ] . the ec is aware of the costs of such large-scale innovative projects and plans to use for this purpose the funding from the european coal and steel community. financing options for modernisation opened by eu ets reform ( - ) seems to be the desired step but with some doubts about its potential for a truly deep technological transformation in the i&s industry. the i&s industry may in the future encounter a barrier of assessing to bank financing for high emission investments like it occurred in the power sector in coal-based technologies [ ] , [ ] . steel products are constantly evolving, driven in part by r&d conducted in collaboration with steel-using industries [ ] j o u r n a l p r e -p r o o f hydrogen" incremental improvements to core process technologies slow down pursuit for "breakthrough" technologies (lock-in) low use of res-based energy in technological processes, e.g. hydrogen res steel electrolysis high usage of coal in technologies where lower emission substitution is possible, e.g. as an energy carrier or feedstock not fully recognised impact of energy and material efficiency on operational costs high technological uncertainty between research and demonstration phases and then full commercialisation (scale uncertainty) "energy efficiency gap" -insufficient interest in energy savings in auxiliary technologies, e.g. waste heat recovery, electrical drives insufficient involvement in demand side management, e.g. in eafs a shift towards eaf would help i&s industry to move towards a circular value chain development of energy service market, e.g. energy management, energy audits, energy-saving obligation, enables quicker adoption of energy-efficient innovations [ ] , [ ] , [ ] high quality of domestic products reduces the risk of replacement of domestic products by import ("carbon leakage" consequence) [ ] the growing share of res in the eu power capacity may make res-based steel technologies economically viable [ ] natural obsolescence of some installations opens option for deep technology change the i&s industry, due to its characteristic is not predisposed to radical breakthroughs. the i&s technology although based on well-established knowledge when implemented is not straightforward replicable and on each site requires adaptation which creates additional risk. moreover, high technological integration entails that any change of one technological process requires changes to other parts of the process which increases the costs and the risk of technological incompatibility. the tis distinguishes between incremental innovations that track existing technological trajectories and radical innovations that lead to new technologies. the progress in the i&s industry mostly consists in the process innovation that tends to follow predefined technological trajectories through incremental innovation aimed at enhancing productivity [ ] . the radical case typically means investing in novel technologies for the basic conversion process or changes in feedstocks and low-carbon energy supply. for example, the eaf shift suitably combined with res-based energy could lead to the entire decarbonisation of steel production. since this deep decarbonization transition at present is proceeding very slowly, therefore, even incremental its can serve as an effective tool in ghg reduction [ ] , [ ] , [ ] , [ ] . through learning by doing, the practising engineers facilitated incremental innovations that triggered partial reinvestments. the leaders in the i&s industry employ many ancillary means of energy and material efficiency, e.g. waste heat utilisation, variable electric drives, advanced measuring and control systems, if only these are economically viable [ ] , [ ] , [ ] . however, these can only suitably complement mainstream measures. the variety of technologies under research, although not vast, will enable to compose the future technological structure in most suitable configuration depending on different j o u r n a l p r e -p r o o f geographical, social, economic, institutional environments criteria which characterize the different mss. for example, the mature technologies available for using biomass as fuel and feedstock in steel production are financially more attractive than the electrification of heat at electricity prices above approximately $ /mwh [ ] . electricity prices will be decisive in the choice between electricity-based decarbonization and ccs [ ] . innovations in the i&s industry are predominantly technological, with traditional organisational structures and business cases [ ] . i&s producers can use other options, e.g. consolidation and structural capacity reduction, customer orientation and commercial excellence, operational optimisation including material and energy efficiency [ ] , [ ] . long-lasting insufficient progress may demotivate actors from launching intensive r&d programs the crucial role of the world's largest steel companies may impede innovation process in some countries while supporting in others relatively slow deployment of bat due to lack of legal obligations or clear economic benefits [ ] the concentration of r&d programs on breakthrough technologies as the solemn options for deep decarbonisation the need for the i&s industry decarbonisation has been well recognised by all actors and is not questioned, see e.g. [ ] , [ ] , [ ] . since r&d directions in the i&s industry have been defined years ago, noting low pace of progress, and still pending pilot phase of key technologies, it can be assumed that further progress requires a strong involvement of key actors, i.e. the ec, ms governments and the i&s industry who shall play the leading role in acceleration the transformation. the other actors, i.e. the researches, are also important but innovative acceleration depends on the coherent action of the former group. that eu strong determination of decarbonisation helps to get synergy but has up to now turned out to be too weak to cause deep innovative changes. uncertainties around post-paris policy, unconvincing eu industrial policy which does not reflect the high climate ambitions and lack of economic incentives make main global i&s manufacturers reluctant to get fully involved in the it transformation. they remain unwilling to take the lead in r&d as long as eu financial incentives are apparent, and policies do not secure future stable regulatory framework. the push must, therefore, come from the actors who from different reasons have an interest in lowering ghg emission, namely the ec, ms governments, and society. for them, environmental protection is the common denominator enabling cooperation. relatively slow rate of i&s industry preparation to industry . as compared with highly innovative industries highly competitive global markets and weak faith in substantial benefits due to the fast implementation of breakthrough technologies conserve the "cost-efficient" criterion of technology development legislation insufficiently forces ecoinnovation developing or often-changing regulatory framework for less mature technologies, e.g. ccsu and hydrogen infrastructure [ ] ineffective use of eu ets revenues for lowemission technology transition in the steel industry [ ] i&s industry's objective to increase competitiveness may not coincide with cost optimal ghg emission reduction lack of interest to force the inclusion of external costs of energy into total costs of industry lack of agreed sectorial targeted energy and ghg reduction goals among big emitters, e.g. power, chemical, and steel incoherence of many eu politics and strategies aimed at supporting innovative technologies [ ] inadequate regulatory or legal frameworks efforts to remove distortion in the internal energy market to make it more competitive development of circular economy enforces better energy and material circulation, e.g. steel scrap, feedstock, by-material usage to support innovative technologies, e.g. tax realise, subsidies, public support to r&d steady risk of import substitution to the eu and difficulty with adoption common eu market defence and expansion strategies weak political, the organisational and economic power of few new entrants table . examples of blocking mechanisms and inducements in the i&s industry in the eu. the corporate style of management in large companies (multinational) impedes decisionmaking process and internal networking functioning [ ] corporate culture may not sufficiently support its difficulties to place deep decarbonisation projects as a priority in long-term strategies of i&s plants its are not a priority in the steel industry practice energy and material saving objectives not well integrated into operational, maintenance or purchase procedures low priority of improving industry corporate image as a "green" business (corporate social responsibility) entering of products into the market without obligation to trace "carbon foot" low interest of top management in implementing its especially in time of economic hardships or uncertainty unwillingness to undertake innovative projects by technical staff due to possible personal responsibility and the lack of financial gains or option for promotion a limited number of benchmarks mostly due to the unwillingness of industry to reveal energy, environmental and economic data [ ] natural trend of advancing technology towards more sustainable and competitive high pressure from employees and union trades on improving work conditions that can partly be accomplished by it implementation growing power of "green"-oriented parties on the eu political arena since the eu needs a long-term strategy to increase industrial competitiveness, a framework for building a new decarbonised industry and innovation policy strategy is required [ ] . the actual i&s industry decarbonisation options do not match the ambitious climate goals set by the ec [ ] , [ ] . the gap has been poorly assessed by politicians and the needed counteractions are not in place according to the i&s industry [ ] . the eu i&s industry is deeply embedded in the global business chain with the growing use of external subcontractors as part of their manufacturing operations, while changes of technological infrastructure on the national level require significant investments and, more importantly, a harmonised decarbonisation strategy of the whole national economy. this implies the need for tide coordination of actions among the key players in the long run. there are not "countervailing" policies which would intentionally impede the decarbonisation. the indirect impact is mainly by environmental legislation. the energy efficiency-related directives, e.g. energy efficiency directive [ ] , eco-design directive [ ] exert a weak impact on the i&s industry . the eu ets much helped to bring ghg emissions to corporate routines and culture and raised priority of the low-carbon technologies in the business hierarchy to strategically importance, e.g. by promotion energy management systems [ ] . similar to the analysis of the eu ets impact on the power sector [ ] , research on i&s industry interactions with res-based energy, chemical industry (res-hydrogen), and ccsu would bring interesting findings concerning r&d [ ] . unlike other sectors being under decarbonisation, eu legislation is less effective at enforcing the i&s industry decarbonisation by market rules, e.g. energy certificates for buildings, compulsory minimum energy efficiency regulations, appliance energy labels, car emission standards effectively help to support decarbonisation in other sectors. therefore, the eu should try to use indirect pull levers, e.g. public procurement and recent circular economy regulations affecting main steel-consuming industries, in particular the automotive, buildings and construction [ ] , [ ] , [ ] . similarly, to other eiis, energy costs are one of the main drivers of competitiveness for the eu i&s industry since energy costs represent up to % of total operational costs. intensification of markets for a quality product could be done by widening the scope of the eco-design directive, e.g. to better address co-design of products for material efficiency, durability, repairability, reuse, and recycling; public procurement, focusing on sustainable products and product design requirements [ ] , [ ] . assessing the necessary intervention in the post -covid- economy, it is recommended to increase funding for the acceleration of r&d and roll-out of pilot projects of low carbon technologies, also in the i&s industry, e.g. hydrogen-based steel making [ ] , [ ] . the number of industrial installations covered by the industrial emission directive (ied) and bat conclusions may serve as metrics of the effectiveness of eu regulation in establishing clean technologies and processes in the industry. the eea reports that there is continued progress in the number of installations covered by in the i&s industry, in total [ ] . a strong link between bats and reductions in air pollutants achieved was identified. as compared with what emissions would have been based on steel production data and emission factors from (per unit of steel produced), emissions of sox had been reduced by %, of nox by %, of pm by % and mercury by % [ ] . decarbonisation of the i&s industry does not encounter hostile coalitions in the eu, however, the roles of the two main actors, namely the ec and the manufacturers, have not been assigned. the i&s industry in the eu does not oppose decarbonisation being aware that the process is unavoidable [ ] . however, the i&s industry finds itself "squeezed" by the ghg reduction constantly forced by the eu ets, global competition, and perceived inability of the ec to protect the eu industry against unfair competition [ ] , [ ] , [ ] . the main global manufactures may threaten national governments with "carbon leakage" when seeking favourable economic privileges to continue or restore their business. the power of national governments seems to be inadequate and too weak to steer the i&s industry operating domestically towards more innovative transformation than it is due to the eu policies. therefore, the industry has tabled several postulates to the politician to empower the transformation and let this ue sector to survive [ ] , [ ] . many of them call for higher support to its through fostering the r&d works [ ] , [ ] . the eu steel companies were successful to achieve free allocation in phases i-iii of eu ets mostly due to effective lobbying. concerning participation in the eu ets the i&s industry constantly raises two issues -lack of abatement opportunities and inability to pass through costs. okereke and mcdaniels [ ] found that these arguments are only partially supported by evidence because of information asymmetry between the industry and politicians, inherent inappropriateness of adopting political decisions on environmental regulations, and privileged treatment in the eu ets, despite that all the key elements of the tis are in place and work towards the low carbon goal, the transition rate is low. seeking the obstacles, the ambiguous and week industrial policy towards eii of the eu can be blamed. this entails insufficient r&d public support to make a real breakthrough in its in the i&s industry. the global manufactures may benefit from the eu technology innovative system. the eu financed knowledge "leak" through global companies has also to be considered as a braking factor as the companies can control the rate of innovation according to their current business interest. on the other hand, using their r&d capacities, they also support eu research institutions [ ] . there are many policies and instruments the effect its in the i&s industry. they may embrace energy efficiency, e.g. energy audits, energy efficiency obligation schemes, voluntary agreements; energy policy, e.g. energy taxes; environmental policy, e.g. emission taxes and emission permits; financial incentives, e.g. subsidy-based "technology push" instruments, reward-based "market pull" instruments, and access to capital measures; regulatory measures, e.g. codes and standards; information and feedback provision [ ] , [ ] , [ ] . their interaction is complex and exacerbates the importance of policy coordination to secure the policies' effectiveness [ ] , [ ] , [ ] . the innovation system in the i&s industry unlike such systems in other non-eiis has not been driven by all customers pressing on radically improved products ("demand-side innovation") [ ] . there are customers demanding quality, but even they do not expect radical improvements due to innovative breakthroughs in technologies. the currently used high-emission technologies are capable to deliver the desired products at highly competitive prices. therefore, most customers show no direct interest in ghg reduction by the i&s industry and do not squeeze on the manufactures. the creation of the eu market for "green steel" is, therefore, if politically agreed, one of the fundamental steps to radically improve the economic viability of breakthrough technologies. public procurement (using shadow carbon prices as a bid criterion or setting limits on carbon intensity) and "green labelling" shall become the initial actions. then, more radical measures should follow, e.g. creation of investment incentives while ensuring "carbon leakage" protection by spreading carbon pricing globally, adjusting carbon prices at the border and stopping free allowance allocation, material "feed-in-tariffs", contracts for difference, applying consumption charges, promote the recycling of materials, also by extending producers' responsibility for the management of their end-of-life products. [ ] , [ ] . more attention must be paid to the industrial symbiosis and integration of i&s technologies with other technologies [ ] , [ ] , [ ] , [ ] . its from the i&s industry have very limited options to diffuse to other sectors because of their specific technology orientation. this limits the circle of their potential users ("market scale") and does not build up the inter industrial synergy. in the industrial symbiosis, the three most effective symbiotic measures for co abatement are bf gas recycling, coke oven gas recycling, and bf slag sold to the cement industry [ ] , [ ] , [ ] . participation in demand side management (dsm) ideally suitable for eafs should also be more exploited as a synergy with the power sector. conversely, it is predicted that radical technological change to the i&s industry will come from other industries, e.g. with res-based energy, "green hydrogen" technologies, electrolysis at affordable cost, and economically viable ccsu [ ] , [ ] , [ ] , [ ] . ccsu underscores coupling between the i&s industry and the chemical industry [ ] . at electricity prices below approximately $ /mwh, hydrogen use for greenfield steel production sites is more cost-competitive than applying ccs to conventional production processes [ ] . more extensive use of eafs would increase electricity demand. the eurofer [ ] predicts that the demand of the i&s industry for res-based energy in the eu amounts to approximately twh a year by (including for the production and use of hydrogen). this in turn would require public support for res and hydrogen industries to remain globally competitive [ ] , [ ] . power grids are getting pivotal for all industries facing the option of intensive electrification. eaf, h-dr technologies are only feasible when the availability and reliability of huge amounts of cheap energy are secured. h-dr gets cost-competitive with an integrated steel plant at a carbon price of € … eur per tonne co and electricity costs of €/mwh [ ] . this will take advantage for countries with well-developed res capacity and power systems meeting the standards of the smart grid. this requires closer than today long-term planning between the power sector and i&s industry in technical and co-financing aspects to retain energy security of the i&s industry [ ] . there is a threat that there may be competition for res-based electricity among different industries, e.g. transport, chemical. phasing-out nuclear capacity in some countries shall also be analysed in terms of being replaced by res-based energy especially in the light of options of use of hydrogen in steel production utilizing nuclear process heat [ ] . state ownership of the i&s industry and/or power enterprises poses an interesting challenge of large scale private-public partnership. novel technological options, e.g. hot briquetted iron from the dri combined with local res capacity may lead to a relocation of reduction/palletisation and steel production sites to avoid power grid congestions [ ] . the iea proposes identifying geographic clusters where key energy infrastructure can be built to support the decarbonisation of industry [ ] . lack of proper sectorial coordination between the i&s industry and the power sector may lead to a shift from process-related emission to combustion-related emission especially in countries with a high share of fossil fuels (coal, gas) in the energy mix [ ] . planning of the power system for a substantial increase in res is declared as one of the four key policy priorities to foster the eu energy transition [ ] . decarbonisation in the i&s industry may have a far-reaching impact on other industries, e.g. the global coal demand, and, consequently, poses societal problems in heavy steel and coaldependent regions ("fair transition") [ ] , [ ] . alongside efficiency improvements, this means that coal use in the i&s industry declines by around million mtce by [ ] . breakthrough transformation in core technology will entail deep changes in the entire supply chain, e.g. coal, gas, scrap, which will have to be restructured around these new methods [ ] . networking within the company or group in the same industry, between industries, and suppliers has been proved important for effective energy management [ ] . internal networking help to exchange ideas and discuss common strategies and future legislation, e.g. on the environment. industry calls for enabling and incentivization of cross-sector collaboration by supporting consortia with cross-sector technologies, e.g. to enlarge value chains, increase cross-industry cooperation, and to innovate horizontally applicable, integrated solutions and innovation that result in services replacing or complementing existing products. [ ] . public and political awareness needs to be increased by setting up communication channels, e.g. forums and platforms [ ] . okazaki and yamaguchi [ ] investigated possibilities of accelerating the effectiveness of the global voluntary sectoral approach for transfer and diffusion of energy-saving technologies in the i&s industry the authors claimed that the success depends on the four factors -sharing of information, the establishment of challenging numerical targets voluntarily, utilize communications, especially among experts, promoting more widespread use of technologies to achieve the set targets, and setting up and sharing of a common longterm vision. the findings show that non-technological factors, difficult to be included in the modelling, play important role in technology diffusion (see also [ ] ). the i&s industry in the eu is very well managed and supported by many organisations and platforms which work as lobby organisations, e.g. represent the industry in talks with the ec, co-ordinate eu wide activities, organise r&d programs [ ] , [ ] , [ ] . the i&s industry should more actively use the opportunity of building a positive atmosphere around the industry by participating in corporate social responsibility programs and "green" labelling the products to receive stronger public support for its decarbonisation needs [ ] . international collaboration and industry-government cooperation will be needed to further improve the energy and environmental performance of the i&s industry [ ] , [ ] , [ ] , [ ] . in some instances, coordinated action across multiple stakeholders may be required, e.g. sharing best practices driving innovation across industry groups [ ] . ti development in the i&s industry brings some long-term risks and uncertainties, some of them will become permanent barriers if timely not removed [ ] , [ ] . bachner et al. [ ] studied uncertainties in macroeconomic assessments of low-carbon transition pathways in the eu i&s industry. they show that effects strongly depend on technology choice, prevailing macroeconomic states as well as regional characteristics and to less extant on the underlying socio-economic development and the climate policy. based on our analyses, we provided a list of risk and uncertainties around technological innovation in the eu i&s industry (table ). the low share of costs of pollution in total manufacturing cost, e.g. these caused by the eu ets no progress in including "external costs" in eiis operation, e.g. in power sector "externalities are not priced") the continued gap between energy and environmental policies the inclination of some governments to "sacrifice" i&s industry at the price of meeting national climate objectives symptoms of "silos" among responsible central government bodies ("poor governance") technological risks socioeconomic economic risks the potential loss of market shares due to failures and delays in it expected implementation the economic failure of some key innovative technologies, e.g. ccsu the volatility of iron ore, energy carriers and scrap on global markets [ ] high and unpredicted costs of r&d work and transition to the demonstrative phase significant difficulties in obtaining commercial financing especially for basic and pre-commercial r&d [ ] large steel demand plunged following a change in the global economy volatility in financial markets and the vulnerability of emerging economies to a deterioration in financial conditions [ ] lack of "de-risking" instruments to reduce business risks the prevailing market tendency for "cheap products" over more costly "sustainable products" [ ] lack of reliable information, e.g. about future prices and costs, asymmetric information, principal-agent problems, learning by using [ ] the tendency to revert from grants to risk financing options for higher advanced developments [ ] lack of clear criteria and a highly transparent set of requirements, procedures, and decision making processes in the public financing schemes [ ] rising protectionism [ ] risk due to post covid- economic crisis [ ] the european business community has asked for the horizon' europe budget to be raised to at least € billion [ ] . through the analyses they identify some general trends which affect the i&s industry development through the diffusion of technologies. unused funding due to lack of deal flow [ ] the imposition of trade tari s by third countries (i.e. usa) that tend to increase the endeavour of countries like china, brazil or russia to export more steel to europe [ ] societal risk prolonged low public awareness on innovation benefits [ ] the public opinion focused on emissions of the power sector, not prioritising i&s industry public acceptance to import cheaper "dirty product" loss of legitimacy when public financial support is necessary [ ] source: own work. . conclusions and policy implication . . benefits of tis approach the main aim of the analysis was to apply the tis approach to highlight the actors, barriers, and inducements on the way of the i&s industry in the eu to deep decarbonisation. this paper identifies the key elements of its in this industry, and then analysis of the functions of innovation through the lens of the tis approach. it facilitates more in-depth insight into the highly complex and dynamic political, technological, economic, and societal environment at this climate-decisive moment. the eea warns "past trends for resource efficiency, the circular economy, and climate and energy are encouraging; the outlook is less positive" [ ] . our research, integrating a range of technological and non-technological aspects and thanks to identifying the involvement of many actors, their roles, and links among them in the functioning of the tis, should help to provide guidelines for political decisions. it contributes to a better understanding of sometimes divert interests among actors, to the prevention of duplication of efforts and to avoid excessive costs and stranded assets due to better coordination. this was indeed a challenging endeavour, yet worth taking because of its novelty. our tis analysis is close to the mission-oriented innovation system (mis) aiming to accomplish a widely shared societal challenge (mission) related to climate change. the notion of mis defined by hekkert et al. [ ] as "the network of agents and set of institutions that contribute to the development and diffusion of innovative solutions with the aim to define, pursue and complete a societal mission" should increase public awareness of the necessity of low-carbon transformation in the i&s industry and make the relevant r&d higher valued among scholars. power sector transformation to the smart grid concept, eii decarbonisation, stringent energy efficiency priorities, the digital revolution, closing material cycles are rather means than the individual initiative of completing the climate mission [ ] , [ ] , [ ] . we show that that deep decarbonisation in the i&s industry is feasible but requires firm support, mostly political, to finance intensive r&d and reduce the business risk. to that end, all actors need to come to a common strategy followed by an action plan to realise the decarbonisation vision. they shall support more effectively the further refinement and market implementation of breakthrough technologies. the major driving force for the low-carbon transformation in the i&s industry is the strategic vision of the all eu actors of radical reduction ghg emission through innovative technologies, and material and energy conservation [ ] , [ ] , [ ] , [ ] . j o u r n a l p r e -p r o o f technologies [ ] . policy, regulation, and energy price-induced technical change are welldocumented drivers for innovative technologies [ ] . public authorities at different levels, including local, are essential players in facilitating the dialogue among parties to set clear visions, mitigate divergent interests, balance costs, and benefits. the "climate" mission is not sufficient for all, especially for "steel dependent" communities, thus it shall be supported by presenting a fair cost-benefit assessment of transformation. the ec and the ms governments are firstly expected to support its by suitable legislation and regulations, e.g. setting realistic pollution limits, creation of "green steel" market, effective use of public procurement, setting material requirements for large steel consumers, reforming the eu ets, setting barriers against unfair steel import, world's regulation of scrap trade. the second desired support is to co-finance the decarbonisation under the state aid framework [ ] . in this respect, suitable energy prices and financing all stages of r&d, particularly important in the early stages of the innovation cycle, are highly required [ ] . there is a persistent state of the informational advantage of the i&s industry over politicians about costs and benefits of climate policy which in the long-term is harmful to both sidesdoes not enable to set effective and cost-efficient policies and adopt appropriate corporate strategy [ ] . more reliable data from the i&s industry and better access to public data banks are essential to eliminate the asymmetry of information among all actors [ ] , [ ] . modern means of data processing, e.g. big data, artificial intelligence, analytics, create opportunities for better economic effectiveness in the whole i&s business chain. the requirements of climate and environmental protection are getting stronger so that incumbent technologies are reaching their inherent limits and lose social acceptance. i&s industry will not be able to get closer to the eu reduction targets unless breakthrough technologies are introduced and prove their cost-effectiveness [ ] . decarbonisation consisting in material and energy efficiency will help, but it is not sufficient to get the reductions needed. other measures, e.g. organisational, managerial, logistical, business optimisation, can support but the technological breakthrough is a must. thus, national, and supra-nation support to technology research, development, and deployment play a key role. politicians should not allow breakthrough innovative investments to be placed outside of europe [ ] . the i&s industry is well suited to benefit from the increased priority of circular economy, e.g. recycle of scrap or utilisation of by-products or increased demand better products from other industries [ ] . reducing total steel demand via a shift to a more circular economy requires inter alia deep changes in the global scrap market [ ] , [ ] , [ ] . the rate of diffusion of breakthrough technologies in the i&s industry is not clear [ ] , [ ] , [ ] , [ ] , [ ] , [ ] . this analysis points to the need to accelerate r&d in the i&s industry to commercialise breakthrough core technologies and increase the role of material and energy efficiency. it also stresses the necessity of more res-based energy and importance of hydrogen technologies. in addition to a very much needed research focus on the technological side, more research on socio-technical drivers and barriers is needed to manage the transition process successfully [ ] . international collaboration and win-win industry-government cooperation will be essential to make a real breakthrough. domination of china as manufacture and the country's technological advances shall be carefully watched [ ] . the economics of the transition implies political involvement throughout the whole innovation cycle, from the preferred cost of capital, covering a large share of r&d costs, building energy infrastructure, e.g. electric energy, hydrogen, to the creation of competitive markets. the cost of deep decarbonisation in the i&s industry is high and therefore perceived unacceptable from a political point of view unless a trustful assessment of the long-term costs and benefits is presented and then accepted by the society which may take years. therefore, to make the transition viable, the future policies and regulations, e.g. environmental, market, should be far-reaching and encourage investors to employ these today breakthrough technologies and not to conserve the current state. the future economic framework should be conveyed to investors well in advance to avoid stray assets [ ] . many concerns shall be shown to the future impact of the eu ets on the i&s industry. the ineffectiveness of the eu ets as a driver for low-carbon technological transformation deters the industry to embark on large innovative projects. the new innovative eu ets built-in mechanisms are aimed to eliminate this insufficiency [ ] . it is postulated that benchmark used as a criterion for public support should be replaced by lca approach that can better value steel as a recycle product [ ] , [ ] . there should be an incentive for "degree of innovation" which should reward breaking innovations [ ] . our research also suggests that eu policy should be more focused on the development of knowledge-intensive and technologically complex industries, including. the i&s industry, as the main factor to retain industrial competitiveness. such a policy should enable rapid diffusion of technological innovations through the production and distribution networks of global corporations. this research is addressed to policymakers to elaborate an optimal policy strategy of technological change of the current functioning of the relevant technology-specific innovation system. it should lead to developing, modifying, and deploying the policy framework aiming at improving the functioning of tis by removing the blocking mechanisms. identification of structural elements of the tis, discovering the internal links among them and dynamics of functioning, control input signals, noises (barriers) and feedbacks in the tis have been done. the authors understand that more in-depth insight requires further research to build a coherent framework for it and contribute to meeting economic, sustainable, and societal objectives. further tis analysis and full risk assessment including technology and marketspecific risks would enable knowledge-based public funding granting for i&s projects, e.g. in modernisation or innovation funds of the eu ets. the research also provided a few recommendations for the managers, e.g. necessity for cost and risk management, the importance of building interfaces, and relations with supplier partners and customers [ ] . funding: the article has been prepared under project innopaths: innovation pathways, strategies, and policies for the low-carbon transition in europe. the project has received funding from the european union's horizon research and innovation programme under grant agreement no. . of "energy"; "environmental science"; "social science"; "multidisciplinary" and "business; management & accounting". this search was expanded via a snowballing procedure using the list of the literature of the collected contributions to gather additional sources. the literature on the tis was gathered by searching standard entries: "entrepreneurial activities"; "knowledge development"; "knowledge di usion"; "guidance of search"; "market formation"; "resources mobilisation"; "legitimacy"; "legitimation" and "positive externalities". additionally, considering a dynamic business characteristic of the i&s industry, the literature search was extended to cover the ec internet resources, political, and business websites. therefore, the feature of our analysis is its wide scope of the analysed sources and documents, e.g. advisory groups, eu and government-sponsored research projects, industry alliances, which are creating specifications. the slr used had some disadvantages, e.g. it relied only on one peer-reviewed database, the combination of the keywords used might miss some relevant papers and institutional insights. the role of the steel industry in a circular economy and its proposals ahead of the commission's launch of a revised package action plan for a competitive and sustainable steel industry in low and zero emissions in the steel and cement industries barriers, technologies and policies steel roadmap for a low carbon europe global warming of . °c an ipcc special report technical summary decarbonization of industrial sectors: the next frontier present 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decarbonisation. • tis analysis as a tool for granting public funds for i&s projects acknowledgements: we are in debt to ms ewa stefaniak for her friendly support and diligent proofreading of this paper. j o u r n a l p r e -p r o o f ☒ the authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.☐the authors declare the following financial interests/personal relationships which may be considered as potential competing interests: j o u r n a l p r e -p r o o f key: cord- - gimxteg authors: gourtsoyannis, john title: covid- : possible reasons for the increased prevalence of olfactory and gustatory dysfunction observed in european studies date: - - journal: clin infect dis doi: . /cid/ciaa sha: doc_id: cord_uid: gimxteg nan a c c e p t e d m a n u s c r i p t dear editor, leurs et al describe the interesting observation that the reported prevalence of olfactory and gustatory symptoms seems to be substantially higher in european covid- cohorts compared to east asia cohorts. most of the studies from east asia report a prevalence of around - %; - european studies report a prevalence of - %. - however the authors did not sufficiently explore the possible reasons for this observed discrepancy. i think that it is worth doing so briefly here. some have postulated that one possible explanation for this discrepancy is a cultural difference in the epistemology of how patients and medical staff experience or report these symptoms. whilst this is indeed possible, i feel that this is unlikely to account for such as striking difference in observed prevalence. there are two alternative and potentially more plausible and interesting explanations that i feel are deserving of more study. the first possibility is informed by the hypothesis that there are differences in ace receptor expression in the nasopharynx of east asians as compared to european populations. increased expression of ace in european populations may contribute to a higher risk of olfactory and gustatory symptoms. the other intriguing possibility is that this emerging symptom amongst european populations is actually related to mutations in the virus itself; mutations which may be resulting in a clinical difference. new genomic analysis shows that the spike mutation (d g; a g-to-a base change at position , in the wuhan reference strain) is found almost exclusively in europe . this analysis also shows that it is likely to have originated in europe and has now spread across the continent. if this correlates with the increasing prevalence of olfactory/gustatory dysfunction then the possibility is therefore raised that the european mutation profile represent a clinically distinct strain, associated with a different symptom profile to the reference wuhan virus. this possibility would also raise further lines of inquiry with regards to tracking the evolution of sars-cov- virulence and its neuroinvasive potential. i have no conflicts of interest or funding sources to declare a c c e p t e d m a n u s c r i p t clinical characteristics of coronavirus disease in china clinical features of patients infected with novel coronavirus in wuhan, china epidemiological and clinical characteristics of cases of novel coronavirus pneumonia in wuhan, china: a descriptive study neurological manifestations of hospitalised patients with covid- in wuhan ,china: a retrospective case series study alterations in smell or taste in mildly symptomatic outpatients with sars-cov- infection self-reported olfactory and taste disorders in patients with severe acute respiratory coronavirus infection: a cross-sectional study olfactory and gustatory dysfunctions as a clinical presentation of mild-to-moderate forms of the coronavirus disease (covid- ): a multicenter european study loss of smell and taste in combination with other symptoms is a strong predictor of covid- infection comparative genetic analysis of the novel coronavirus ( -ncov/sars-cov- ) receptor ace in different populations spike mutation pipeline reveals the emergence of a more transmissible form of sars-cov- . biorxiv key: cord- -kjs je u authors: martel, cyril; nielsen, gunnar d.; mari, adriano; licht, tine rask; poulsen, lars k. title: bibliographic review on the potential of microorganisms, microbial products and enzymes to induce respiratory sensitization date: - - journal: nan doi: . /sp.efsa. .en- sha: doc_id: cord_uid: kjs je u nan the regulation (ec) no / concerning feed additive applications in europe is based on a safety opinion made by efsa (european food safety authority). the regulation includes an annex ii, which provides the general requirements that should be satisfied by a dossier supporting an application for the use of a product as a feed additive. in particular section iii of the annex ii deals with safety of a feed containing an additive, and this is further subdivided into safety for target animals (section . ), for consumers of the products derived from the animals fed the additives ( . ), for users/workers exposed to the additive ( . ) , and finally for the environment ( . ) . in section . , it is stressed that the allergenic potential and sensitization via the respiratory route is of concern, and should be evaluated. annex iii of that regulation provides the specific requirements concerning specific intended additives depending on the technical nature of the feed additive. in this context, technological additives and zootechnical additives are of interest. thus, in spite of the guidance on toxicological safety component of a feed additive dossier [ ] , efsa is instructing that enzymes and microorganism should be assumed a priori to be respiratory sensitizers unless convincing evidence to the contrary is provided. in order to investigate the potential of microorganisms, microbial products and enzymes to induce respiratory sensitization when used as food and feed additives, we conducted a systematic literature search of all the information we could find linking microorganisms and enzymes to respiratory sensitization. we supplemented this search with references from important reviews in the field of enzymes and microorganisms and occupational health, as well as with interviews with some leading companies in these fields. finally, we conducted a survey of existing regulations on the subject in developed countries. the immune system has evolved to protect individuals from microbial pathogens as well as larger parasites. however, the immune system can sometimes react inappropriately to innocuous antigens. allergic reactions occur when an individual produces ige antibody against such innocuous antigens, or allergens. while other antibodies are free circulating in blood or extracellular fluid, ige is also found in tissues, where it is tightly bound to the mast-cell surface through the ige receptor. exposition to the allergen triggers a cascade of responses initiated by the activation of mast cells by igereceptor cross-linking [ ] . basophils and activated eosinophils can also act in a similar fashion. ige is thought to have evolved as a mechanism to fight off large multicellular parasites, like nematodes and trematodes, less vulnerable to other effector arms of the immune system (like phagocytosis or cd t cells) [ ] . indeed, the typical symptoms of type i hypersensitivity, like smooth muscle contraction, itching and generation of mucus, seem appropriate to dislodge such organisms. however, modern hygiene practices have considerably lessened the population's exposure to parasites in developed countries, and the focus on ige is overwhelmingly set on its role in the pathophysiology of allergy. it is estimated that almost half the populations of north america and europe have allergies to one or more common environmental antigens [ ] . why some antigens lead to ige production and type i hypersensitivity is not fully elucidated. certain antigens and routes of antigen presentation to the immune system are known to elicit preferably a th response charaterized by the production of ige. most allergens are relatively small, highly soluble proteins. the typical presentation of an inhalable allergen involves a transmucosal route and very low doses (as low as microgram per year for some pollen allergies) [ ] . myeloid dendritic cells are the dominant antigen-presenting cells in the respiratory mucosa, and they are known to be very efficient th inducers. exposure to low doses of antigen in general also tends to favor a th response in the absence of th -inducing co-stimulatory signals. unlike pathogens that are usually eliciting the same pathology in a large majority of the population (albeit with difference symptoms or severity), only some of the people who are exposed to common allergens make ige antibodies against them. in addition, there is a clear trend of increase in prevalence of atopic allergy, and of asthma in particular, in developed countries. environmental factors that may potentially explain these trends are changes in the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. exposure to microorganisms in early childhood, environmental pollution, allergen levels, and dietary changes [ ] . alterations in exposure to microorganisms are the most plausible explanation at present for the increase in atopic allergy. this includes exposure to pathogens, but also to a large number of commensal, non-pathogenic microorganisms early in life [ ] . epidemiologically, atopy is negatively associated with a history of infection with measles or hepatitis a virus, and exposure to mycobacterium tuberculosis, but severe respiratory syncytial virus (rsv) infection in children is positively correlated to the later development of asthma. the underlying idea behind this theory would be that th immune response early in life might reduce the likelihood of th responses later in life and vice versa. more recent versions of this theory focus on the role of tregs as opposed to inflammatory th [ ] . broad exposure to commensal bacteria early in life is associated with protection against ige-mediated hypersensitivity. the relationship between environmental pollution and type i hypersensitivity is unclear [ ] . some epidemiological studies have shown a lower prevalence of allergy and atopy in severely polluted cities than in generally cleaner ones. other studies on diesel-fuel particulates show that they trigger symptoms in patients. studies of the effects of exposure to secondhand tobacco smoke or occupational hazards on atopic sensitization have also led to discordant results. similarly, there is no clear evidence that the rising prevalence of allergy is due to any systematic change in allergen exposure or to changes in diet. the most common route of allergen exposure is inhalation. most often, it initially leads to mild allergies causing allergic rhinitis, with sneezing and a runny nose as main symptoms. this results from the activation of mucosal mast cells beneath the nasal epithelium by allergens such as the proteins from pollens [ ] . allergic rhinitis is characterized by intense itching and sneezing, local oedema leading to blocked nasal passages, a nasal discharge, and irritation of the nose as a result of histamine release. allergic conjunctivitis is a similar condition triggered by airborne allergens deposited on the conjunctiva. these reactions are annoying but cause little lasting damage when they are linked to seasonal allergens like pollen. more severe damage can come from activation of mast cells in the lower airways, which leads to allergic asthma [ ] . it is a chronic condition that requires treatment and can be lifethreatening. both allergic asthma and rhinitis can come from constant exposure to occupational allergens as it is the case for fungal enzymes and baker's asthma, but asthma can also be perpetuated even in the absence of further exposure to allergen. the airways become characteristically hyperreactive and factors other than reexposure to antigen can trigger asthma attacks. it is worth noting that, although asthma has potentially more severe consequences, quality of life studies indicate that rhinoconjunctivitis is perceived as worse the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. than bronchial asthma for the patient [ ] . this is probably linked to the fact that asthma is treated more efficiently and more strictly than rhinoconjunctivitis. type iii hypersensitivity reactions are caused by inhalation of soluble environmental antigens that trigger immune responses by precipitated antibodies (mainly igg) and lymphocytes; the characteristics of hypersensitivity pneumonitis (hp) are summarized from recent reviews [ , , , , ] . unlike allergic reactions, they are linked to igg rather than ige production. the pathology is caused by large aggregates of igg:antigen complexes, also known as immune complexes. these immune complexes fix complement and can bind to and activate leukocytes bearing fc and complement receptors; these in turn cause widespread tissue injury to small blood vessels in many tissues and organs, including the skin, kidneys, and nerves. this is thought to be the underlying pathophysiological mechanism in hypersensitivity pneumonitis (also called allergic alveolitis). immune complexes also form in autoimmune diseases such as systemic lupus erythematosus where, because the antigen persists, the deposition of immune complexes continues, and serious disease can result. hp has been described in various settings, often involving organic particles and environments where moulds abound: farming, bird-breeding for example. some chemical compounds have also been shown to be able to act as hapten and cross-link the host albumin to create an allergic particle. common industrial antigens causing hp include isocyanates (paint sprays), plastics (packing plants), mycobacterium avium complex (metal working fluids), aspergillus (agriculture), and thermophilic actinomyces (agriculture). genetic factors are important for development of hp and hp occurs seldom in smokers. the most likely reason why some inhaled allergens provoke igg rather than ige is probably linked to doses of exposure. occupational allergens associated with hp are generally linked with very high levels of exposure, such as in the case of metalworking fluids or farmer's lung. hp can present acute, subacute or chronic forms. the chronic form, with lung fibrosis, can be very severe, and even life-threatening. the -year mortality in chronic hp is as high as %, and in the subset of the fibrotic type of chronic hp, the -year mortality reaches %. in the acute form, respiratory symptoms are lung crackles, cough, dyspnoea, fever, myalgia, headache, and malaise. if the exposure continues, the acute form develops into the subacute (< month of exposure) and further into the chronic form (> month of exposure). during the continued exposure, cough and dyspnoea at exertion gradually become worse. in the acute form, high resolution computed tomography (ct) shows ground-glass lung opacities and micronodules that may progress with addition of air-trapping, emphysema and fibrosis in the chronic form. pulmonary function tests show low carbon monoxide diffusion capacity and typically a restrictive pattern. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. in spite of spores from naturally occurring fungi being described as a common cause of asthma and allergies, there is little description of occupationally induced fungal allergies. microorganisms have been involved in numerous outbreaks of hp in industry, agricultural work and in indoor climate; examples of involved organisms are listed in table . several outbreaks of hp have occurred due to aerosol exposure to water-based metalworking fluids contaminated with microorganisms. in several of these cases, mycobacterium species were considered the offending agents. mycobacteria are ubiquitous in the human environment, including in natural water, drinking water and soil. they are easily aerosolized due to their hydrophobic surface. growth of mycobacteria may have been promoted in metalworking fluids by the use of biocides and disinfectants that have diminished or eradicated competing microorganisms; mycobacteria are highly resistant to quaternary ammonium compounds, formaldehyde, morpholine, chlorine, chloramines, ozone and heavy metals [ , , ] . hp in farmers caused by exposure to mouldy livestock feed is known as farmer's lungs, which is associated with exposures to thermophilic actinomycetes and fungi [ , ] . workers engaged in packing of peat moss, an organic substance derived from sphagnum moss, have also developed hp from contamination by moulds. the peat moss contained high levels (~ x colony forming units/g) of monocillium species and penicillium citreonigrum [ ] . sphagnum vegetation may also contain high levels of mycobacteria as m. avium and m. intracellulare [ ] . hp may be due to microorganisms playing an intended role in production processes. hp in workers cultivating edible mushrooms, mushroom worker's lung, may be due to microorganisms [ , , ] . thus, mushrooms are cultivated on compost that can contain high amounts of thermophilic actinomycetes and moulds, having a role in the production of the compost. the actinomycetal spores can be found in high levels in the air during preparation of the compost, during the spawning (~ colony forming units/m ), i.e. inoculation with mushroom mycelium, or during other work in the mushroom house [ ] . additionally, mushroom worker's lung may be due to the spores from exotic mushrooms [ , , ] . however, no sensitization was observed to the white button mushroom agaricus bisporus in a the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. study in patients with mushroom worker's lung. use of airway protection during spawning prevented respiratory complaints [ ] . mushroom worker's lung and cheese worker's lung are among the types of hp where the microorganisms have a role in the production processes. although not commonly encountered, hp may be caused by unintentional growth of moulds and bacteria in buildings with excessive inside moisture. aspergillus niger is a common antigen accounting for development of serum precipitins and hp. stachybotrys species are commonly found in damp buildings with cellulose material, but it has not been implicated as a cause of hp overall, in cases where hp appeared due to unintended exposures to microorganisms, exposures have often been the related to wet or humid conditions that promoted the growth and thus, resulted in unusually high exposures. also in these cases, the work conditions may have promoted aerosol formation. thus, primary prevention should, where possible, limit wet or humid conditions or otherwise, growth should be controlled. if high exposures cannot be avoided, respiratory protection has to be worn. aerosol formation should always be minimized. where microorganisms have intended roles in a production, process specific risk management have to be introduced. both for unintended exposures and for exposures to intentionally used microorganisms, spore-producing microorganisms are often the cause of hp. the present document has been produced and adopted by the bodies identified above as author(s the present document has been produced and adopted by the bodies identified above as author(s in a case-reference analysis of a cohort of employees in a european detergent factory, chest disease was not increased significantly at ≤ . ng/m of proteases and eye and nose symptoms were not increased at ≤ . ng/m . the authors mention that only the protease level was measured although amylases and cellulose were also used, and that "irritant" dust and non-occupational reactions may have contributed to the findings above these levels [ ] . outbreaks of sensitization, upper airway symptoms and asthma still occurred in the detergent industry where strict exposure controls were not followed. in such a case, the protease level was in the range from to ng/m , several enzymes were used and amylase was found to be a more potent sensitizer than the protease [ ]. allergic reactions were observed in swedish consumers in the early period of use of enzymes in detergents, when products were dusty and enzymes were not encapsulated. a re-creation using the product and the consumer habits at the time suggested that exposure may have been as high as ng/m [ ] . recently, a study was performed in atopic filipinos who had a daily use of hand laundry for several hours; the study lasted up to years. the laundry contained a protease and an amylase. the laundering was with granular products or with bar products that were also used for personal cleansing. exposures were low (≤ . ng/m ) for the granular product, lower (≤ . ng/m ) for the bar product for scrubbing, and below the detection limit (< . ng/m ) for personal cleansing. none of the subjects developed the present document has been produced and adopted by the bodies identified above as author(s overall, taking into account the high number of cases of allergy in relation to exposure to enzymes and the low number of cases with hp, the type i allergy is considered the critical effect and it is inferred that protection against allergic airway diseases will also protect against hp. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. identification of key parameters: microorganisms (the relevant types, forms and species are organisms that are known to be used per se in feed and food) enzymes that are used in feeds and food sensitization (main emphasis on ige-related respiratory sensitization but also allergic alveolitis, and non-immunological hypersensitivity are covered). physiological/immunological mechanisms, distinction between the exposed organ and the target organ/symptoms (i.e. can respiratory sensitization occur via other exposures?) are discussed. it is important to emphasize the ubiquitous occurrence of microorganisms, their products and enzymes in all biological materials versus the deliberate addition in feed or food. in the allergen research of today many allergens from plants and animal sources are cloned and expressed in e. coli or pichia for research purposes. such applications are excluded (but documented) at an early stage of the literature search. to which extent are microorganisms, microbial products and enzymes respiratory sensitizers? to solve this problem three parallel approaches have been taken: • an epidemiological search. the literature on epidemiological data related to respiratory allergies and sensitization will be screened in order to identify descriptions of reactions to microorganisms, microbial products and enzymes. • a clinical/case based search. case stories and descriptions of isolated outbreaks of respiratory sensitizations and allergies will be documented • an allergen based search. literature will be searched for occurrences of microorganisms, their products or enzymes. currently available methods to test respiratory sensitization and their validity the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. • structure-allergenicity-relationships (prediction of allergenicity based on the primarysecondary-tertiary protein structure) • in vitro-models for antigen presentation and sensitization. • animal models of allergic sensitization the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. microorganism: microbiological entity, cellular or non-cellular, capable of replication and/or of transferring genetic material. includes fungi and bacteria (also under the form of spores). viruses, archea, protists, as well as microscopic plants and animals are generally also included in this definition, but have little relevance for this report, as they are not used as food or feed additives. microbial product: pool of organic compounds that are released from biomass growth and/or decay. includes (but is not limited to): humic and fulvic acids, polysaccharides, proteins, nucleic acids, organic acids, amino acids, antibiotics, steroids, exocellular enzymes, siderophores, structural components of cells and products of energy metabolism. enzyme: enzymes are proteins from animal, vegetal or microbial origin that catalyze chemical reactions. relevant enzymes for this report include (but are not limited to): amylases, proteases, trypsin, cellulases, pectinases, xylanases, papain, and lactases. it is agreed upon that this report will focus exclusively on microorganisms, microbial products and enzymes that are added deliberately to food and feed. additionally, those additives need to have been clearly and individually defined to be taken into consideration. the lists of microorganisms and enzymes (appendices and ) were compiled by using the sources listed in table : in order to narrow down our search, we established a list of relevant microorganisms and enzymes of interest. it was agreed that the focus of the report would be on ige-related sensitization, as well as allergic alveolitis and non-immunological hypersensitivity. for each item on these lists, the following pubmed search was conducted: item and (ige or hypersensitivity or rhinitis or conjunctivitis or asthma or urticaria or anaphylaxis or allergic alveolitis) when the outcome of the search was hit, the item was also searched in the allergome database. when the outcome was between and hits, the titles of hits were screened manually to identify articles susceptible to contain epidemiological, clinical or laboratory data. reviews were systematically included. after the title screening, abstracts were then viewed to determine which articles were relevant. when the outcome was larger that hits, the search was narrowed using the following strategies, in that order: use "item" instead of item include only hits containing the item in abstract/title include only hits containing the item in title we used some of the most authoritative reviews in the field of respiratory sensitization and/or industrial use of enzymes and microorganisms to backtrack through isi's web of knowledge the most important articles that they referenced. our subcontractor (gunnar nielsen) made a survey of existing legislation and regulations concerning enzymes and microorganisms in developed countries. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. in order to enhance our understanding of the relationship between food and feed additives and respiratory sensitization, we interviewed representatives of two of the leading companies in the field: chr. hansen a/s and novozymes. the details on these companies are listed in annex . these companies were first mailed a list of our questions (listed in annex ), after which they agreed to meet us, and in the case of chr. hansen a/s, to show us one of their production lines. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. . . state-of-the-art of the test methods the allergenicity of a protein can refer to two properties: either its capacity to induce ige thru an immune response or its capacity to cross-react with ige induced by a previous antigen [ ] . the first case involves interaction with and processing by antigen-presenting cells, but also contacts with epithelial cells that can modifiy the th /th /treg balance towards a given antigen. it is overall a very complex matter, and large parts of the theory behind it remain unexplained. it is therefore difficult to predict, and in-silico attempts to model this have been largely unsuccesful, although numerous hypothesis have been advanced. no common structural feature between allergens can be conclusively pointed at. the only common properties of allergens so far are that they can evade th -suppressing mechanisms and interact with dendritic and mast cells. current allergen prediction algorithms are better suited to predict ige cross-reactivity. past algorithms have focused on the association of unknown sequence with a single allergen motif or an allergen peptide [ ] . however, recent developments are now taking into account the fact that allergens generally contain multiple epitopes [ ] , that these epitopes are very often discontinuous [ ] , and that antibodies have interaction sites to a given antigen [ ] . taken together, these facts have led to algorithms focused on several motifs, with a marked improvement in false positive rates [ ]. as of today, in-vitro models are used to investigate the mechanisms of allergic diseases and the processes of each individual type of cell, rather than the allergenicity of a product. fundamental knowledge is still missing to create an in-vitro model of the various cell interactions that could mimic the human response to an allergen. there is, to our knowledge, no reliable, established in-vitro way to predict allergenicity. in recent years, the ue has supported a large project called sens-it-iv (http://www.sens-it-iv.eu/), whose goal is to develop such an in-vitro model. in , the members from the project published scientific articles on the topic. although promising, this research has not yet reached its goal. respiratory sensitization and the symptoms that it causes require a complex interaction between several cell types, including epithelial cells, dendritic cells, macrophages, cd + t cells, b-cells and mast cells. the variety of in-vivo models used for respiratory sensitization the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. has been thoroughly reviewed by verstraelen et al [ ] , and this section only briefly describes their findings. three main types of epithelial cells have been used as models: primary airway epithelial cells, alveolar cells lines, and bronchial cell lines. primary cells are of course the closest representation of the airway epithelium, but their scarcity and donor variation is a major problem for the reproducibility of the experiments. alveolar macrophages studies often use cells collected by bronchoalveolar lavages for ex-vivo studies. several monocytic cell lines also exist, but like with all cell lines, whether they accurately reflect the behavior of functional primary cells that they ressemble is subject to discussion. monocyte-derived dendritic cells (dcs) have replaced langerhans cells as primary dcs for in-vitro experiments. however, some very satisfying cell lines are also available, such as mutz- , which display all the antigen processing and presentation pathways from a functional dc. due to the importance of interaction and co-stimulation by different cell types, co-culture models have been subject of much interest. successful tri-cultures (macrophages/dc/epithelial cells) show great promise. d cultures, although in need of optimization, are also very interesting alternatives. currently, the most widely used experimental animal for modelling allergic responses in the airways is the mouse. numerous antigens can be used for sensitization, such as ovalbumin, dust mite and a. fumigatus. typical induction of asthma in murine models has requirements: a systematic immunisation to trigger th immunity and a repeated pulmonary allergen challenge. pulmonary challenge only without systemic immunity is not sufficient to elicit pulmonary symptoms [ , ] . the intraperitoneal route is the most common way to elicit th immunity, although repeated instillation in the upper airway can also be used for some antigens. although the mouse model is the most popular in the field of allergy research, it suffers from several flaws that limitate its predictive value. the differences in induction mechanisms, together with physiological and immunological differences between mice and humans might make it more suitable for clinically-oriented studies than for allergenicity prediction. relevant models to study allergenic properties would need to rely on sensitization from inhalation only, the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. and these are rare. interestingly, our discussion with researchers from novozymes a/s is corroborating this line of thought: animal models are out of favor in their labs, and remain only used for relative allergenicity experiments. it is worthwhile to note that in-house research at novozymes a/s has failed to show significant differences in relative allergenicity between different enzymes. this may be due to the fact that physical properties (size, formulation) matter more than sequences and functions, but it could also be another indication of the poor predictability of the mouse model. table summarizes the various models currently used in respiratory allergy research. models of non-respiratory endpoints such as allergic contact dermatitis e.g. the guinea pig maximization test are not included: the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. this section describes the results of the systematic literature search that was conducted on pubmed, together with the extra information that we found by back-tracking references from important reviews through web of knowledge. for each microorganism or microbial product from our list, we listed all the relevant information we could find in the following order: species were found to have relevant information, for a total of relevant articles. only species for which relevant information could be found are listed here. use in food or feed: a. globiformis is used to alter the taste of some food products, for example to metabolize bitter limonin of citrus. epidemiological studies: a study of agricultural workers from poland diagnosed with hypersensitivity pneumonitis showed that a. globiformis was the etiological agent in cases [ ]. case reports: n/a literature arguing against a possible sensitization by the agent: n/a use in food or feed: various strains of the fungus a. niger are used in the industrial preparation of citric acid (e ) and gluconic acid (e ). a. niger also is used to produce enzymes such as glucoamylase and pectinases. epidemiological studies: n/a case reports: a case of hypersensitivity pneumonitis to a. niger was reported in [ ], while several cases of allergic fungal sinusitis, rhinitis and bronchopneumonitis have been reported [ , , ]. it has been suggested that asthma and atopy may both represent a protective response against contained airway infection due to ubiquitous proteinase-producing fungi [ ]. literature arguing against a possible sensitization by the agent: n/a the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. a. oryzae is a fungus used in chinese and japanese cuisine to ferment soybeans or to saccharify rice, other grains, and potatoes in the making of alcoholic beverages. a. oryzae is also used for the production of rice vinegars. it can also be used to produce glucoamylase and alpha-amylase. epidemiological studies: n/a case reports: articles were found describing cases of respiratory sensitization linked to the use of a. oryzae in the food industry [ , ]. both cases involved a high seasonal concentration of spores in the homes of japanese brewers and respiratory symptoms. interestingly, one of these articles referred to the existence of at least other similar cases described in the japanese literature. literature arguing against a possible sensitization by the agent: n/a use in food or feed: b. licheniformis is used to degreade feathers to produce cheap and nutritious feed for livestock. also used as a probiotic in livestock. epidemiological studies: n/a case reports: n/a literature arguing against a possible sensitization by the agent: b. licheniformis spores were found to induce a th type response in-vitro in bal/c mice [ ] . use in food or feed: b. pumilis is used to degreade feathers to produce cheap and nutritious feed for livestock. epidemiological studies: in , bernstein et al [ ] found cases of machine operator's lung, a type of hypersensitivity pneumonitis. those cases all followed occupational exposure to metalworking fluid. at least of those patients presented serum precipitin tests positive to b. pumilis. case reports: n/a literature arguing against a possible sensitization by the agent: n/a use in food or feed: b. subtilis can be used in the commercial production of the japanese food natto as well as the similar korean food cheonggukjang. it can also produce amylase; and is used as a probiotic for livestock. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. epidemiological studies: a cross-sectional study of workers from a herb processing plant showed that they responded with a high frequency % to a skin prick test with extract from bacillus subtilis [ ] . use in food or feed: d. hansenii is generally the predominant yeast in the smear of bacterial surface-ripened cheeses such as munster. epidemiological studies: n/a case reports: a -year-old female was reported to have developed cough, fever and dyspnoea following repeated exposure to a home ultrasonic humidifier [ ] . precipitating test and lymphocyte proliferative response was positive for an extract of d. hansenii. literature arguing against a possible sensitization by the agent: n/a use in food or feed: e. faecium is a probiotic used in feeds for livestock. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. literature arguing against a possible sensitization by the agent: in the atopic dermatitis mouse model, e. faecium suppressed total ige production and induced il- and ifn-gamma production; intraperitoneal injection of strain t inhibited serum ige elevation and atopic dermatitis symptoms [ ] . use in food or feed: this organism is used to produce certain types of yoghurts, especially from nordic countries. use in food or feed: lactobacilli are used in the food for acidification, and/or enhancement of flavor, texture and nutrition. they can also serve as starters or complementary cultures for several varieties of cheese, fermented plant foods, fermented meats, in wine and beer production, sourdough bread and silage, and as probiotics in feeds for livestock. epidemiological studies: n/a case reports: n/a literature arguing against a possible sensitization by the agent: there is ample evidence indicating that lactobacilli (or at least some strains of lactobacilli) alleviate allergic symptoms. such properties were observed for l. brevis in the atopic dermatitis mouse model, where oral administration of the bacteria inhibited ige-production, and skewed the immune response towards th dominance [ ] . l. brevis-fermented kimchi also strongly alleviated symptoms of prurit, anaphylaxis and inflammation in a mouse model [ , ]. l. casei was shown to attenuate lung inflammation and th cytokine profiles in der p sensitized mice, hinting at immunomodulatory properties useful to prevent respiratory allergies [ ]. one study in-vitro in atopic dermatitis patients [ ] and one study in mice [ ] supported similar conclusions about l. fermentum. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. overtvelt et al showed that l. helveticus reduced airways hyperresponsiveness, bronchial inflammation and proliferation of specific t cells in cervical lymph nodes in the murine asthma model and characterized this strain as a probiotic acting as a th /possibly treg adjuvant that potentiates tolerance induction via the sublingual route. in the ova-induced allergy mouse model, nonaka et al [ ] showed that a l. pentosus strain induced il- and il- in-vitro and modulated the th /th balance toward a th -dominant state. in-vivo, serum ige levels were diminished and active cutaneous anaphylaxis reaction were suppressed. splenic il- production from splenocytes of ova-immunized mice was upregulated by oral administration of l. pentosus. in , a double-blind, placebo-controlled, crossover study in children showed that a combination of l. rhamnosus and l. reuteri was reduced symptoms of atopic dermatitis, especially in patients with increased ige levels [ ] . supplementation of l. reuteri during pregnancy was associated with low levels of tgf-beta and slightly increased levels of il- in colostrum [ ] . studies in balb/c mice argued for a role for non antigen-specific cd (+)cd (+)foxp (+) regulatory t cells in attenuating the allergic airway response following oral treatment with l. reuteri [ ] . oral treatment with the live bacteria greatly decreased allergen-induced airway hyperresponsiveness, but a similar effect was not observed with the killed organism [ ]. use in food or feed: l. lactis is used in the early stages for the production of many cheeses including brie, camembert cheese, cheddar, colby, gruyère, parmesan, and roquefort. other uses include the production of pickled vegetables, alcoholic beverages, and other fermented food-stuffs. it can also be used as a probiotic in feeds for livestock. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. epidemiological studies: n/a case reports: n/a literature arguing against a possible sensitization by the agent: two recent studies in mice indicate that treatment with l. lactis to togther with an allegen (blg) decreased il- production and enhanced ifn-gamma production by blg-reactivated splenocytes, suggesting a switch from th -to th -immune response [ , ] . symptoms after intranasal challenge were locally reduced, as evidenced by decreased release of il- in bronchoalveolar lavage fluids. use in food or feed: l. mesenteroides is a bacterium associated with the sauerkraut and pickle fermentations. epidemiological studies: n/a case reports: n/a literature arguing against a possible sensitization by the agent: in an in-vitro study in mouse splenocytes, a strain of l. mesenteroides proved to be a useful th stimulating agent, especially by upregulating ifn-gamma production [ ] . use in food or feed: p. pentosaceus is used as an acid producing starter culture in sausage fermentations, cucumber and green bean fermentations, soya milk fermentations, and silage. epidemiological studies: n/a case reports: n/a animal studies: duchaine et al [ ] showed that p. pentosaceus has a pro-inflammatory effect in mice similar to that of s. rectivirgula, the most common etiological agent of farmer's lung. this was in contrast of a most recent publication by masuda et al [ ] , where they showed that the sn strain of this bacteria increased the production of th cytokines in peyer's patches of allergic diarrheic mice. literature arguing against a possible sensitization by the agent: n/a use in food or feed: p. candidum is used in cheese-processing. epidemiological studies: in a study from , cheese-factory workers out of had airway symptoms, five had asthma requiring treatment [ ] . two-thirds of the symptomatic cheese-workers had precipitating antibodies; compared to only half in the non-symptomatic group. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. case reports: n/a literature arguing against a possible sensitization by the agent: n/a use in food or feed: p. nalgiovensis is the most frequently used starter for cured and fermented meat. epidemiological studies: a study of workers from a sausage factory showed that symptoms of hp such as sneezing, cough, dyspnoea, nasal obstruction, headache, and discomfort were significantly more frequent in workers exposed to p. nalgiovensis [ ] . case reports: a -year-old male pork butcher was reported to experience cough, tightness of the chest, and sibilant dyspnea for years [ ] . episodes were related to the handling of sausages and inhalation of the dust coming from the sausage casings at his work in the butcher's area of a supermarket. the patient had ige to p. nalgiovensis. literature arguing against a possible sensitization by the agent: n/a use in food or feed: p. roqueforti is typically used in the production of blue cheeses. epidemiological studies: n/a case reports: campbell et al [ ] described the case of a cheese factory worker with symptoms of cough, dyspnea, and malaise, and findings of bibasilar crackles, reduced lung volumes, hypoxemia, and bilateral infiltrates on chest roentgenogram. all symptoms resolved after she left the workplace. bronchoalveolar lavage revealed a high percentage of lymphocytes. antibodies to p. roqueforti were detected in serum and lavage fluid. literature arguing against a possible sensitization by the agent: n/a use in food or feed: s. cerevisiae is a top-fermenting yeast, and has been used for brewing and baking for centuries. it is also widely used as a probiotic for livestock. epidemiological studies: a study of subjects, including atopic dermatitis (ad) patients, patients with allergic rhinitis and/or asthma, and nonatopic controls found that a positive spt reaction (> or = + +) was seen in % of patients with severe ad, in % with moderate ad, and in % with mild ad or no history of ad. patients with rhinitis and/or asthma and nonatopic controls displayed a positive reaction in only and % of cases, respectively [ ] . case studies: in , a -year old baker was reported to have hydrorrhea, sneezing, nasal obstruction, wheezing, spasmodic cough, and dyspnea upon exposure to the dried form of the the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. yeast. the patient began to use conventional wet yeast and carried on normal work activity without symptoms [ ] . an interesting case was described in . the patient experienced generalized urticaria and asthma after eating pizza and bread, but only fresh from the oven, and had ige to s. cerevisiae [ ] . if bread, pizza and cakes were eaten more than one hour after preparation, no symptom would occur at all. why the allergen was active only in readybaked foods remains unexplained. literature arguing against a possible sensitization by the agent: n/a the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. this section describes the results of the systematic literature search that was conducted on pubmed, together with the extra information that we found by backtracking references from important reviews through web of knowledge. for each enzyme from our list, we listed all the relevant information we could find in the following order: epidemiological studies: doekes et al [ ] report a prevalence of % of respiratory symptoms among exposed workers in a factory producing premix for feed additives. % of those workers had measurables ige against phytase. zober et al [ ] performed a cross-sectional study on workers from an r&d facility with potential contact with phytase and found symptoms of conjunctivitis, rhinitis, or bronchitis in % of those employees. baur et al [ ] examined employes with occupational exposure to phytase either during largescale enzyme production (powdered form), enzyme laboratory analysis, or animal husbandry. symptoms were reported in % of the subjects, including dyspnoea ( % of the subjects), rhinitis ( %) and conjunctivitis ( %). % of the subjects exhibited ige-mediated the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. sensitization, and % had measurable igg levels. finally, caballero et al [ ] examined workers from two animal feed factories and found that seven of those were sensitized to phytase or -phytase. three of those workers had asthma symptoms, two had asthma and rhinoconjunctivitis, one had asthma and rhinitis, and one had rhinitis only. use in food or feed: accelerate the breakdown of starch, for example to enhance yeast performance in the baking industry. production organism: aspergillus niger, aspergillus oryzae, bacillus amyloliquefaciens, bacillus subtilis, bacillus licheniformis, bacillus stearothermophilus, microbacterium imperiale, trichoderma reesei, trichoderma longibrachiatum (alpha-amylase); barley, soybean (beta-amylase) epidemiological studies: there is a large number of studies of amylase-linked allergic reactions, especially in the bakery industry. in , a cross-sectional study of bakery workers in the netherlands found a positive association between positive skin prick tests to alpha-amylase and work-related respiratory symptoms [ ] , establishing conclusively the strong and positive relationship between alpha-amylase allergen exposure levels in bakeries and specific sensitization in bakery workers. this finding was confirmed years later in a study looking at flour samples from british bakeries [ ] . in , brisman et al [ ] took a closer look at a cohort of bakers and found new cases of chest symptoms, of eyes/nose symptoms, and of positive spt to a-amylase. they showed exposure-response relations for chest and eyes/nose symptoms and for sensitization, and an increased prevalence for chest symptoms in the most exposed workers. similar relations were confirmed one year later in bakery workers in belgium in [ ] . a large study of supermarket bakery workers in south africa revealed that only % of the wrokers had a sensitization to fungal amylase [ ] , underlining the effect of work environment regulations. by comparison, ige antibodies to fungal amylase are found in % of us blood donors [ ] . similarly, it has been observed that alpha-amylase from bacillus amyloliquefaciens did not cause sensitization in a study involving animal feed workers [ ] whereas other fungal enzymes were found to be sensitizers in the same settings. for the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. case reports: since the link between amylases and allergy is well-documented thru numerous epidemiological studies, case studies worth mentioning fall a bit outside of the classical baker's asthma. interesting case reports on amylase include a double sensitization to lysozyme and amylase in a baker with rhinoconjunctivitis and asthma [ ], a sensitization occuring from infrequent contact with the enzyme in a lab technician showing symptoms localized on her hands without nasal or respiratory effects [ ] , and one case of occupational allergic contact urticaria from fungal but not bacterial alpha-amylase [ ] . two cases of igemediated rhinitis have also been reported in hospital and pharmaceutical workers exposed to biodiastase, a prescription drug based on alpha-amylase [ ] . animal studies: it has been shown that amylase from bread crust and rolls crust kept between . and % of the antibody-binding capacity of amylase from dough [ ] . in-silico work: warren et al [ ] have developed a dynamic population-based model for the development of work-related respiratory health effects among bakery workers. the model predicts that non-atopic/non-sensitised workers had probabilities of developing moderate symptoms and progression to severe symptoms of respectively . % ( % ci . to . %) and . % ( % ci . to . %) per mg/m /year of flour dust. these probabilities were twice as high in atopic workers. they also predict that % ( % ci to %) of workers with severe symptoms are sensitised to wheat and % ( % ci to %) to alpha-amylase. allergens of a similar class outside of food and feed area: alpha-amylase from porcine pancreatic extract [ ] and mite amylase [ ] are both respiratory sensitizers. asparaginase ec: . . . -food use in food or feed: converts asparagine into aspartic acid. this prevents the formation of carcinogenic acrylamide during high temperature processes such as cookie manufacturing. production organism: aspergilly niger, aspergillus oryzae. epidemiological studies: n/a case reports: n/a animal studies: n/a in-silico work: n/a allergens of a similar class outside of food and feed area: asparaginase can also be used as a cancer chemotherapy agent. lee et al have reviewed allergy cases of this application and found that reactions have been reported to occur in up to % of treated patients, although serious anaphylactic reactions occur in less than % of patients [ ] . the present document has been produced and adopted by the bodies identified above as author(s). in accordance with the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. in-silico work: n/a allergens of a similar class outside of food and feed area: the major latex allergen hev b is an esterase [ ] . an important allergen from carica papaya pollen also has an esterase activity [ ] . epidemiological studies: sander et al [ ] examined bakers exhibiting symptoms of asthma, rhinitis and conjunctivitis. % of them were found to have specific ige to glucoamylase. case reports: quirce et al [ ] have described four cases of patients (three bakers, one enzyme processing plant worker) presenting symptoms such as cough, shortness of breath and wheezing. all four patients had ige against glucoamylase. in-silico work: n/a allergens of a similar class outside of food and feed area: n/a other routes of exposure: kanerva et al [ ] described the case of a chemical enzyme factory process operator presenting symptoms of itching and dermatitis. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. use in food or feed: glucose oxidase is used for the removal of glucose or oxygen from the foodstuff in order to enhance their stability during storage. production organism: aspergillus niger, aspergillus oryzae, penicillium chrysogenum. epidemiological studies: n/a case reports: simon et al [ ] reported the case of a -year-old man with granulomatous lung disease, employed in the extraction and purification of glucose oxidase from aspergillus niger two years before admission. the link between the symptoms and the enzyme has not been formally proved in this study. animal studies: n/a in-silico work: n/a allergens of a similar class outside of food and feed area: n/a hemicellulase use in food or feed: used in bakery for the enhancement of dough qualities (mechanical handling, stability) and for product optimisation (volume, consistency, storage life), as well as in the production of fruit juice and other beverages, in the production of spirits and in the alcohol industry, in wine production or as an additive to animal feed (to increase digestibility). production organism: aspergillus niger, bacillus amyloliquefaciens or subtilis, trichoderma reesei or longibrachiatum. epidemiological studies: in a cross-sectional study, elms et al [ ] analyzed the serum of bakery workers. % of those workers had ige against hemicellulase, and there was a significant correlation between specific ige and nasal symptoms. case reports: quirce et al [ ] have described four cases of patients (three bakers, one enzyme processing plant worker) presenting symptoms such as cough, shortness of breath and wheezing. two of those four patients had a positive skin-prick test to hemicellulase. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. epidemiological studies: horner et al [ ] examined patients showing symptoms of respiratory allergies and skin test reactivity to at least fungal allergens. % of those patients showed rast reactivity to invertase. lactase or galactosidase (beta) use in food or feed: lactase is used commercially to prepare lactose-free products, particularly milk. it is also used in preparation of ice cream, to make a creamier and sweetertasting product. production organism: aspergillus oryzae, kluyveromyces lactis, bacillus circulans. epidemiological studies: in , muir et al [ ] looked at volunteers from a lactase packaging plant and observed that % had positive results to skin prick test with lactase. the lactase skin test results correlated with nasal and eye symptoms, but not with symptoms suggestive of asthma. in , bernstein et al [ ] found similar results in a study involving pharmaceutical workers occupationally exposed to lactase. % of the workers were skin prick positive, and those were times more likely to have respiratory symptoms than workers with negative skin-pricks. use in food: lysozyme is used as a food preservative due to its antibacterial properties. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. epidemiological studies: in a double blind, placebo-controlled food challenge, concentrated lysozyme was allergenic in a skin prick test, but no patient reacted adversely in the provocation test to fined wine [ ] . the authors concluded that wines treated with fining agents at commercial concentrations did not present a risk to allergic individuals when filtered. similar conclusions were drawn about the contents of lysozyme in grana padano cheese [ ] . however; it has been suggested that lysozyme may present a harmful adjuvant in wine processing for consumers allergic to hen's egg [ ] . [ ] report that while examining patients allergic to russian thistle (salsola kali) pollen, they found that all of them had ige against sal k , a protein from the pectin methylesterase family. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. epidemiological studies: belleri et al [ ] examined workers from a pectinase production factory and showed that % of them had ige to pectinase. % of those workers had both respiratory symptoms and detectable ige against pectinase. case reports: sen et al [ ] reported cases of asthma and conjunctivitis among workers from a fuit salad processing plant. all three had ige against pectinase. it is interesting to note that in this factory, pectinase was used in a liquid form. animal studies: n/a in-silico work: n/a allergens of a similar class outside of food and feed area: ibarrola et al [ ] examined patients with allergy to platanus acerifolia pollen. % of the patients had rhinoconjunctivitis, % had asthma, % had rhinitis, % had urticaria. % of the patients had ige against pla a , a pectinase. in-silico work: n/a allergens of a similar class: a peroxidase from wheat has been reported to be an allergen, with sera from out of wheat-allergic patients reacting to the purified dot-blotted allergen [ ] . the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. allergens of a similar class: phospholipases are very common allergens of venom from bees and other arthropods [ ] . [ ] followed the fao/who decision tree to assess the safety of transglutaminase and found no concern with regard to its allergenic potential. epidemiological studies: in a cross-sectional study, elms et al [ ] analyzed the serum of bakery workers. % of those workers had ige against xylanase, and there was a significant correlation between specific ige and nasal symptoms. in a retrospective study, sander et al [ ] examined bakers exhibiting symptoms of asthma, rhinitis and conjunctivitis. % of them were found to have specific ige to glucoamylase. vanhanen et al [ ] conducted a cross sectional study on workers in four bakeries, one flour mill, and one crispbread factory. % of the workers presented respiratory symptoms, and % had a positive skin prick test to a mix of enzymes including xylanase. in a similar study in the animal feed industry, the same group found that % of exposed workers were sensitised to enzymes to a mix of enzymes including xylanase. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. [ ] report the case of a bakery worker with rhinoconjunctivitis, cough, and shortness of breath. the patient reacted to an inhalative challenge with ≈ . μg of xylanase. merget et al [ ] describe the case of a baker with occupational asthma (nasal congestion, sneezing, running nose, watering of the eyes and shortness of breath). the patient showed significant levels of ige against xylanase. in-silico work: n/a allergens of a similar class outside of food and feed area: proteases are a very well known class of allergenic enzymes, especially but not exclusively in the context of detergent manufacturing [ ] . in addition to their intrinsic allergenicity, proteases have potential effects on several processes involved in allergic diseases: epithelial integrity and permeability, mast cell degranulation, cytokine release from the respiratory epithelium [ , ] . proteases have been shown to interact with receptors from the lining of the respiratory tract, leading to the activation of immunological pathways that can potentially trigger allergic reactions, for example via stimulating epithelial cells to release metalloproteinase and open up tight junctions promoting allergen penetration into the submucosa [ ] . proteases from pollen [ ] , house dust mites [ ] , cockroaches [ ] and fungi [ ] have been shown to have these effects. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. we have conducted a survey of the existing legislation and regulations regarding enzymes and microorganisms and the basis for occupational exposure limits (oels) and labelling. the integrale version of this survey can be found in appendix . it appears from this survey that oels have been set for enzymes (subtilisins) in the uk and the us, but that these limits are bypassed by the limits set "in-house" by the detergent industry. it is also expected that the european regulations will in the future set some derived no effect level (dnel) or derived minimal effect level (dmel) for macromolecular compounds including enzymes. the appendix also includes a list of proteins as airway allergens together with the corresponding labelling and oel, whenever they exist. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. it appears from our research that there is currently no established model to predict the allergenicity of a molecule. although in-silico models can be useful to predict cross-reactivity between allergens, they only take into account the structure of the allergen, which is only one of the many factors implicated in sensitization, and in our opinion only a minor one. in-silico models fail to take into account the context in which the allergen is presented to the immune system. in-vitro models are currently far from being used for predictive studies. although many cell lines are currently used in the lab, research is still trying to understand the role and mechanisms of the cells in the development of allergies, rather than using them to test the allergenicity of various molecules. although co-culture three-dimensional models might someday achieve that purpose, they remain very far from re-creating the complexity of the immune response in a living organism. among in-vivo inhalation animal models, the mouse is currently the best. however, it suffers from several shortcomings that prevent it from being a reliable predictive model: most models require systemic sensitization by injection, followed by multiple respiratory challenges, which hardly resembles the exposure conditions in humans. additionally, most mouse models are strong "atopic-like" responders that can be sensitized to molecules that are not allergenic in humans. in general, exposure to bacterial species leads to a th -type response in humans, increasing macrophages efficiency and leading to the production of opsonising antibodies, two systems very useful to deal with that type of pathogens. in contrast, th responses to bacterial infections are rare, and can sometimes be indicative of a poor prognosis, such as in the case of leprosy. as an illustration of the apparent lack of role of bacteria in allergic diseases, we have been unable to find an example of regulations such as oel to work with such organisms. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. having possible allergic effects. thus, it is not considered a general feature of bacteria to express sensitising properties. although one study reporting type- reactions to b. subtilis extracts was found in our survey, the overwhelming majority of reports were arguing for a beneficial role of bacteria to allergy symptoms, through the induction of th and/or treg responses. this observation is of course partially biased by the fact that lactic acid bacteria were the most studied microorganisms in our list, and these are actually used for their immunomodulatory properties. it is also worth noting that these properties are strain-dependant: only a few strains of a given species have an immunomodulatory effect, while the others have usually no beneficial effect on allergy symptoms. in addition, one recent study introduced the idea that th -inducing bacteria only might not be sufficient for inducing tolerance, but that treg-induction might also be required [ ] . it seems that the best-documented allergic risk linked to occupational exposure to bacteria is hypersensitivity pneumonitis. from our survey, it seems that hp has been linked to exposure to b. pumilis (machine operator's lung), b. subtilis (wood dust) and a. globiformis (agricultural workers). hp remains, however, a very rare disease, and is associated with very specific conditions and high level of exposure. none of the cases described resulted from an intentional use of the bacteria. the case of fungi is quite different. strong evidence of a link between asthma severity and fungal exposure has been previously established. sensitization to fungal species has been shown to be a significant risk factor for severe asthma [ ] . most likely, fungal sensitization can have a causative role in severe asthma [ ] . furthermore, sensitization and exposure to fungi have been linked to hospital admissions and emergency department visits for asthma, life-threatening asthma, admission to icu, and asthma-related mortality. the role of fungi in severe asthma hospitalizations and mortality has been reviewed in [ ] . indoor fungal exposure is particularly relevant to this report. verhoeff et al reviewed studies on indoor fungi exposure and hypersensitivity. the majority of those studies argued for a link between domestic fungal levels and allergic symptoms [ ] . however, it is worth noting that at least one review disputes the studies linking indoor moulds exposure to upper airway allergy (rhinitis), while agreeing with its role in asthma [ ] . it is interesting to see that, as opposed to bacteria; there are regulations regarding the acceptable levels of exposure to several fungi (moulds). the above-mentioned european classification has been applied to (groups of) fungal species: aspergillus fumigatus, candida albicans, coccidioide sinunitis, cryptococcus neoformans var. neofonnans (filobas idiella neofonnans var. neofonnans), cryptococcus neoformans var. gattii (filobasidiella bacillispora), epidermophyton floccosum, microsporum spp., penicillium marneffei. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. our survey of the litterature unveiled species of fungi associated with type allergies, including several studies demonstrating the link between occupational exposure to fungi and hypersensitivity. cases of ige-mediated hypersensitivity as well as hypersensitivity pneumonitis are well documented and involve several distinct genera. in general it seems that fungi can be sensitizers, although allergic symptoms are associated with high levels of exposure compared to other respiratory sensitizers, such as pollen. in contrast to the situation with bacteria, some of the cases that we have presented result from an intentional use of the microorganism. in particular, the cases of p. roqueforti (cheesemaking), s. cerevisiae and a. oryzae are very relevant for this report, as they are directly linked to food production processes. in addition to compiling scientific litterature on the topic, we also contacted one of the leading companies in the field of industrial cultures of microorganisms, chr. hansen a/s. the company claims no cases of allergy related to microorganisms among employees. the only incident involving respiratory symptoms (likely be caused by a toxic/irritative mechanism) occurred when a customer in brazil sprayed the cultures in an indoor environment. this was a highly unusual process, and very different from the intended use of the product. the lack of documented allergic reactions to microorganisms can of course be tied to the comprehsensive preventive measures applied throughout the production process. few workers are directly exposed to inhalable products, and those who do obey strict rules, including wearing personal protection equipment. although very commendable, it is possible that those measures partly hide the sensitizing potential of these microorganisms. sensitizing properties of enzymes have been a cause of concern for several decades in food and feed industry, albeit at lower levels than in the bakery or detergent industry [ ] . allergies to b. subtilis proteases, for example, have been described since the late s [ ] . the most reported cause of respiratory sensitization by enzymes in the food and feed industry is perhaps the case of alpha-amylase and baker's asthma, but other enzymes such as proteases, cellulase and glucose oxydase are also a cause of concern. to compound the problem, industrial enzyme grades are often poor and contain other enzymes from the same production organism, leading to unexpected sensitization and cross-reactions [ ] . studies conducted in enzyme-producing plants have confirmed that enzymes can induce sensitization and allergy [ ] , regardless of their enzymatic activities, or production organism. although not related to food and feed, and therefore out of the scope of this report, the detergent industry also has an extensive experience with enzymes. while respiratory allergies in consumers proved to be a problem in the early years of enzyme addition to detergents ( s), granulated formulations have reduced type- reactions in the general population to the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. virtually nothing [ ] . the problem is now limited to manufactoring processes, and stringent rules in the modern enzyme-and detergent-producing field have contributed to greatly diminish sensitization in workers. like for microorganisms, hypersensitivity pneumonitis to enzymes has been described, for example to phytase [ ] . cases remain however extremely rare, and seem associated with very high levels of exposure. out of enzymes listed as food or feed additive by the amfep, our survey found that were related to allergies. all but one of these were linked to respiratory sensitization. although our survey seems to indicate that the general trend is towards a decrease in exposure levels and sensitization to enzyme in the industry, it is worth noting that studies in the field suffer from inherent selection biases and therefore probably underestimate the true prevalence of sensitization to enzymes. as we did previously for microorganisms, we completed our survey of the literature with a direct contact with a leading producer of enzyme, novozymes. according to the company, less than cases of symptomatic workers are identified every year, and the symptoms/sensitivity ratio has been decreasing due to reduced exposure in the recent years, pointing at a dose-response effect. interestingly, these numbers seem higher than those observed in the case of microbial cultures, although it is difficult to make a comparison between two different production processes taking place in two different companies. no cases have been reported among customers since . novozymes a/s acknowledges the sensitising role of enzymes and relies on stringent personal protection measures to minimize exposure of workers. the use of granulate formulation has led to a steep decrease in exposure levels, with differences beetween formulations (more effective for detergent than bakery uses). the company research has observed no differences in relative allergenicity between enzymes, which seems to indicate that the physical properties of enzymes (formulation, molecular weight…) might be more important than their function or amino-acid sequences. the role of the production organism in allergenicity is currently being investigated. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. when considering the findings on inhalation allergy risks presented in this report, a number of observations stand out: the enzymatic activity in terms of substrate specificity etc, does not seem to explain whether an enzyme has been incriminated as an inhalation allergen or not. neither does the production organism seem to decide whether or not an enzyme has been reported as an inhalation allergen. while it is possible to device sensitization regimes in animals, in particular in mouse, neither these, nor in-silico or in-vitro systems seem to work well in predicting the allergenicity of enzymes in real life. this would suggest that the inherent properties of an enzyme may not be decisive, as to whether it is going to create inhalation allergies. looking at where and when enzyme allergies have been reported, there seems to be a clear gradient from the producers of enzymes, over manufacturers that formulate their products with inclusion of enzymes, to end users that may be exposed to enzyme-containing products (fig. ) . also -and in spite of an increase in the worldwide use of enzymes -there seems to be fewer recorded cases of allergies and sensitizations compared to the situation in the 'ies and 'ies where much higher levels of airborne enzymes prevailed. it seems reasonable to conclude that this reduction is caused by a reduced exposure, which is likely to be caused by different formulations of the raw enzymes which are now dispensed in liquid or granulated forms rather than the former use of powder of small particle sizes. the exact regimens, i.e. temporal concentration profiles, that cause sensitization and may lead to inhalation allergies are not known, but in spite of the fact that in allergy development doserelationships may not always be linear, it is likely that increasing doses of inhaled enzyme lead to increased risks. also -estimated from the lack of reports of enzymes allergies in the general population -it seems that there are thresholds under which exposure does not cause sensitization and allergy. from this follows that setting demands on the formulation (distribution in non-inhalable forms) and designing of the processes in which enzymecontaining (food or feed) products are used, would be a way to reduce the sensitization and allergy risk. if these measures do not lead to consistently low airborne exposure throughout the life cycle of the product, it is necessary to label products and protect exposed persons by personal equipment such as masks etc. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. it is not known whether sensitization always lead to clinical allergy, but based on industrial hygiene experiences and knowledge from other areas of allergy, it must be assumed that sensitization, i.e. the formation of ige antibodies against an enzyme, would be a very strong risk factor for also developing symptoms upon continued exposure. the different steps in developing enzyme allergy and the potential prevention measures are depicted in fig. . the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. figure : flow of enzymes in society, from the producr to the end-user. the shade of darks represent the number of prevalence of sensitization for each category of person exposed. figure : key steps in developing enzyme allergy. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. propionibacterium shermanii the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. [ ] . here the relevant definition of the criteria that should be used for identifying the inhalation sensitising potential is described, i.e. which criteria that should be applied for evaluation of whether a substance is eligible for being labelled with r : the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. substances and preparations shall be classified as sensitising and assigned the symbol .xn., the indication of danger .harmful. and the risk phrase r in accordance with the criteria given below. if there is evidence that the substance or preparation can induce specific respiratory hypersensitivity, where there are positive results from appropriate animal tests, or if the substance is an isocyanate, unless there is evidence that the specific isocyanate does not cause respiratory hypersensitivity. comments regarding the use of r : human evidence evidence that the substance or preparation can induce specific respiratory hypersensitivity will normally be based on human experience. in this context hypersensitivity is normally seen as asthma, but other hypersensitivity reactions such as rhinitis and alveolitis are also considered. the condition will have the clinical character of an allergic reaction. however, immunological mechanisms do not have to be demonstrated. when considering the evidence from human exposure, it is necessary for a decision on classification to take into account in addition to the evidence from the cases: the size of the population exposed, the extent of exposure. the evidence referred to above could be: the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the results of positive bronchial challenge tests are considered to provide sufficient evidence for classification on their own. it is however recognised that in practice many of the examinations listed above will already have been carried out. substances that elicit symptoms of asthma by irritation only in people with bronchial hyperreactivity should not be assigned r . animal studies enzymes and microorganisms as respiratory sensitizers the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. data from tests which may be indicative of the potential of a substance or preparation to cause sensitization by inhalation in humans may include: ige measurements (e.g. in mice), or specific pulmonary responses in guinea pigs. whole microorganisms are different because they are living cells, i.e. biological agents. the above-mentioned directive covers dangerous substances. according to the new eu regulations on chemicals (reach), the definitions of chemicals (substances and preparations) according to reach are: substance means a chemical element and its compounds in the natural state or obtained by any manufacturing process, including any additive necessary to preserve its stability and any impurity deriving from the process used, but excluding any solvent which may be separated without affecting the stability of the substance or changing its composition. preparation means a mixture or solution composed of two or more substances microorganisms such as a bacterium, a yeast cell or spores of these do not fall under the definition of a substance or a preparation according to the eu regulations. application of the classification criteria and labelling rules to the area of microorganisms would thus be by way of analogy with substances and preparations, since they do not seem to be covered by the regulations mentioned above. eu has, however, covered the occupational hazards of working with biological agents in a directive on protection of workers from risks related to exposure to biological agents at work a definition of "biological agents" and "microorganisms appears in / /eec council directive / /eec of november on the protection of workers from risks related to exposure to biological agents at work: the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. biological agents shall mean microorganisms, including those which have been genetically modified, cell cultures and human endoparasites, which may be able to provoke any infection, allergy or toxicity. microorganism shall mean a microbiological entity, cellular or non-cellular, capable of replication or of transferring genetic material. in the biological agent directive [ ] while there are no bacteria classified as having possible allergic effects this classification has been applied to (groups of) fungal species: the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the above-mentioned list formed the basis of national lists in the uk and germany. these are some years newer, and have been slightly revised for virus (sars), but there has been no changes regarding (the missing) classification of bacteria as allergenic, and no further fungal species defined as allergenic. the list from directive / /ec only include organisms in classes , and , but the german list also mention of class organisms [ ] . unfortunately this list does not provide information on potential sensitization. a recent guideline from germany [ ] the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. konzentration zu einer sensibilisierung bis hin zu schwerwiegenden allergischen erkrankungen. (p. thus it is not considered a general feature of bacteria to express sensitising properties. also as indicated in the introduction the regulation on (chemical) substances and preparations does not cover biological agents, thus it is not surprising that there is no mention of microorganisms in this legal complex. in conclusion, no bacterial species per se have previously been considered as allergenic in the european legislation. on the other hand, individual fungi and yeasts have been recognized as allergenic in accordance with the scientific literature as discussed later. occupational exposure limits (oels) are concentrations of products in the air. health-based oels are set by a risk assessment approach, which includes evaluating all hazards (inborn toxicological properties) linked to a compound and establishing exposure-response relationships for the hazardous effects, followed by risk characterization. concentrations below the oels are considered to protect nearly all occupationally exposed individuals against adverse effects, although it is realized that a minor and especially sensitive part of the population may not be protected [ ] . this subpopulation may include individuals highly asthmatic/allergic to the allergen of interest. a prerequisite for establishing a health-based oel is that adverse reactions are exposure-dependent and that there is a concentration where adverse effects no longer appear, i.e. it is possible to establish a no-observed-(adverse)-effect level (no(a)el) for offending effects [ ] . for allergen exposures, risk assessment may be evaluated by means of the airborne concentration per m , as with oels. appropriate analytical methods are prerequisites for establishing exposure-response relationships. airborne allergens are collected on filters by means of a pump and sampling may be enzymes and microorganisms as respiratory sensitizers the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. in the breathing zone by a person-carried filter cassette or it may be by high volume static samplers. the filter content of allergens is then analyzed by immunochemical methods [ , ] . a risk may also be evaluated by means of the allergen concentration per gram of settled dust [ ] . the idea behind this approach is that settled dust can become airborne and thus the concentration in the settled dust can be used as a proxy for the exposure. this approach has been found useful for evaluation of indoor allergen exposures from house dust mite, dog, cat, cockroach, mouse urinary protein and alternaria alternaria [ ] . for toxicological reactions in general, one no(a)el is considered for the key effect. however, for allergic reactions two phases have to be considered. first, exposure to an allergen may induce "sensitization" that implies production of specific antibodies or activated immune cells. sensitization is not per se a disease. second, "elicitation" of symptoms occurs with further exposures to the allergen at a sufficient dose [ , ] . thus, two limits may be set, one where no sensitization is observed and another one that prevents the elicitation of allergic reactions in already sensitized individuals [ , ] . it may be difficult to establish thresholds for allergen exposures due to inter-individual variations in susceptibility to both sensitization and elicitation. those may be caused by genetic differences (atopy versus non-atopy), heterogeneity of immune properties of allergens, age-dependent effects and lifestyle factors (e.g. smoking). there may also be differences between doses that induce sensitization and those that induce elicitation of symptoms [ , ] . co-exposure to endotoxins may also play a role in development of sensitization and asthma [ ] . nevertheless, "practical" noaels have been proposed for several environmental and occupational allergen exposures [ , ] (table ) . the prerequisite of a clear exposure-response relationship has been demonstrated for several airway allergens [ , , ] . this has been shown for indoor and outdoor allergen exposure to house dust mites, cockroaches, pets, pollen and moulds. the exposure-response relationship often shows a monotonous increase in sensitization and development of allergy with increasing allergen exposure enzymes and microorganisms as respiratory sensitizers the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. [ ] . however, cross-sectional studies have suggested a bell-shaped relationship, like in the case of cat allergens, where high exposure levels may induce tolerance [ ] . a similar relationship has been observed in laboratory animal workers exposed to rat allergens, which may be due to a "modified t helper type (th ) response" where specific igg antibodies plays a role [ ] . this is heavily debated though, as a behavioral effect, i.e. "healthy petkeeper effect" cannot be totally excluded [ ] . the general trend that ige sensitization and ige-mediated allergies increase with exposure levels has been substantiated by studies of exposure to enzymes used in the detergent industry [ , ] , of exposure to natural rubber latex allergens and of exposure to flour dust as discussed below. enzymes and natural rubber latex allergens are used as examples of highly potent allergens, where an oel has been set for the proteolytic enzyme subtilisin. flour dust is used as an example of how to set an oel for a low-potent allergen. the exposure-response relationships for allergens have two important features, the steepness of the relationship and the position of the exposure-response curve. thus, different proteins have different sensitization potencies. for example, sensitization to rat urinary allergens occurs in the pg/m range, sensitization to fungal α-amylase in the ng/m range, whereas sensitization to wheat, pig and cow proteins occurs in the μg/m range [ ] . that different allergens seem to have different potencies is also deduced from environmental allergen exposures as only a limited number of allergens are of major importance in the general population [ ] . thus, the number of allergy cases in a population depends both on the potency of the allergen, the presence of adjuvants and the particle size, as well as on the specific exposure levels. in the german population in , the number of occupational asthma cases caused by various exposures was in the order flour>latex>food and feed [ ] . few oels have been set officially for macromolecular biological compounds. they comprise oels for the enzyme subtilisin and for flour dust. however, industry has often set its own internal the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. oels for enzyme, and oels for enzymes have been proposed in the scientific literature with the purpose to protect against ige-mediated allergies (type i reactions). in addition, more of these agents have an eu labelling or have warnings in oel lists. these warnings are based on recognized hazards, i.e. when an increased incidence of allergy has been observed in an occupationally exposed population. hypersensitivity pneumonitis (hp) is caused by immune reactions of precipitating antibodies and activated immune cells. few cases of enzyme-induced hp have been reported. on the other hand, many cases of type i allergies to enzymes have been documented. therefore, the critical effect is considered to be the type i allergy and thus, measures preventing the type i enzyme allergy are considered also to prevent hp. most hp reactions appear from high exposures to airborne moulds and bacteria, where growth has been promoted by wet and humid conditions. in these cases, risk management should limit the wet and humid conditions, or otherwise limit growth of microorganisms. if not possible appropriate airway protection should be used. the subtilisins are proteolytic enzymes derived from bacillus subtilis or closely related organisms. in the early s, the american conference of governmental industrial hygienists (acgih) established an oel for subtilisins as a ceiling level of ng/m of the % active pure enzyme [ ]; a ceiling level being a concentration that should not be exceeded during any part of the working exposure [ ] . thus, for a well controlled working environment, the exposure concentration has to be considerably lower than the ceiling level in general, which is important to consider when comparing the value, for example, with an -hour time-weighted average oels (table ) . the oel was derived from the experiences in the surfactant industry in the late s and the beginning of the s. the main purpose was to minimize the potential for symptoms as sore throat, nasal congestion, cough, wheezing, allergic respiratory sensitization, and to minimize skin irritation [ ] . this value is one of the lowest oels ever established and it still applies [ ] . no oel has been set for other enzymes by acgih [ ] . no oel has been set for enzymes in enzymes and microorganisms as respiratory sensitizers the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. these values are basically bypassed by the much lower exposure levels presently attended by the detergent industry. also, the lower "in house" oels in industry take into account that other constituents of detergents may have adjuvant effects [ , , ] . in the near future, oels will have to be set for many enzymes to fulfil the requirement of the new eu reach legislation. thus, a value has to be based on a noael approach to establish the "derived no effect level" (dnel) or, where a noael cannot be established, the oel has to be based on an acceptable risk to set a "derived minimal effect level" (dmel) [ ] . setting and using oels in relation to unintended development of microorganisms in naturally occurring constituents would not be possible in most cases due to the limited possibility of identifying the offending microorganisms, the limited knowledge about exposure-response relationships, and the limited possibility to implement such values in relation to risk management strategies. in these cases, the risk management strategy should directly address the appropriate prevention of the offending exposure. in the eu, harmonized classification and labelling are adopted for about , compounds (http://www.reach-compliance.eu/english/legislation/docs/launchers/launch-annex- - - -eec.html), which uses a hazard-based approach. of these compounds, very few are proteins, which are known airway allergens (table a . ). in the eu, hazards are described by symbols, e.g. harmful is indicated by "xn" and an irritant is assigned the symbol "xi", and by risk (r) phrases (http://ec.europa.eu/environment/chemicals/dansub/consolidated_en.htm). thus, specific respiratory enzymes and microorganisms as respiratory sensitizers the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. hypersensitivity (asthma, rhinitis and alveolitis) is indicated by the risk phase r : may cause sensitization by inhalation. although animal studies may be used, the evidences are mainly from human studies, where the size of the population exposed and the extent of exposure are taken into account. for α-amylase, β-glucosidase, cellulase, exo-cellobiohydrolase and laccase, respiratory hypersensitivity was the only hazard identified. for the proteases, chymotrypsin, ficin, papain, pepsin a, rennin, subtilisin and trypsin, additionally hazards were identified as expressed from r : irritating to eyes, r : irritating to respiratory system, r : irritating to skin, and r : risk of serious damage to eyes or the combination of these sentences. the eu classification and labelling indicates different risk management approaches by means of no account shall be taken of substances classified as harmful, corrosive or irritant if they exist as impurities or as additives, if their concentration by weight is less than % and if not otherwise specified (http://www.dehp-facts.com/upload/documents/webpage/document .pdf). hazard identifications from organizations setting occupational exposure limits are limited. in germany, no oel is set for the protein allergens, but hazard-based warnings are added to several proteins (table a . ). thus, respiratory allergens are indicated by "sa"or "sah" if it is also a skin sensitizer. only human exposure effects are accepted. sufficient evidence is accepted if specific hyperreactivity has been observed in relation to exposure in more than one subject in at least two independent testing centres and if there is indication for an immunological mechanism. an allergic effect is also accepted from one single case report of a specific hyperreactivity of the airways or the lungs together with other sensitizating effects, e.g. a close structure-effect relationship with known airway allergens [ ] . other examples of warnings to sensitizers are included in table . the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. read-across to establish allergenic properties within a certain type of enzymes may not be possible. for example, sensitization to phytase derived from trichoderma and peniophora species did not show cross-reactions. however, cross-reactions were observed for different types of enzymes derived from the same microorganism [ ] . overall, warning systems about sensitizing properties have to take into account that different allergens or allergenic systems can show highly different potencies and that exposures of humans need to reach a certain level before clinical effects appear. established warning systems take into account that a certain number of observed allergies have been observed in exposed populations. enzymes and microorganisms as respiratory sensitizers the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. : the short-term exposure limit is indicated by "stel" ( -min reference period) and sensitization by "sen"; baur [ ] : "noael" and "loael" for humans, "sen" indicates sensitization and "a" asthma. for further explanations see text. a warning, including a labelling, is relevant when a group of individuals is at a substantiated risk for developing diseases. to be meaningful, a warning about airway allergy or other immunological airway reactions should be based on the incidence of sensitization or the incidence of disease in exposed populations. the highest incidence is expected where process concentrations are highest, which may be in relation to production or downstream use of pure compounds. health monitoring at such workplaces is therefore especially important for hazard identification. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. in general, oels are set to allow exposure control in relation to production of compounds as well as control of exposures in downstream users. setting an oel requires hazard identification as for labelling, but also a quantitative risk assessment. this requires that the adverse effect is exposuredependent and that a level can be set where a potential risk is so low that it is of no concern. thus, a quantitative relationship about the exposure-response relationship has to be established. appropriate analytical methods must be available for this purpose, but also for the control of compliance with the established oel. as for flour dust, the overall dust concentration in the air may sometimes be used as proxy for the airborne allergen concentration, but in most cases, the analytical methods have to address the air concentration of the allergenic protein itself. at present, very few oels exist for the macromolecular biological compounds. in the near future, it is expected dnels or dmels will be set for several industrial enzymes according the european reach regulation and thus providing a new type of oels. for enzymes, the key effect seems to be ige-mediated allergy. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. • how many workers are exposed to each of those categories? • when did your company start to handle those enzymes or cultures? occupational part: • is your company monitoring allergies among workers and/or customers? • are you aware of allergy cases (within the company or with your customers or endusers) related to some of the enzymes or cultures that your company handles? • have your company developed any special formulations that reduce the risk of exposure to cultures or enzymes? • can you pinpoint any products, processes or unit operations that are related to either particularly high risk for sensitization or particularly low risk for sensitization? • does your company take specific measures for working with some of those cultures or enzymes (e.g.: personal protection equipment) within the company or for custormers or end-users toxicological part: • has your work with enzymes or cultures given you an impression on the doseresponse-relationsships in allergy development? • are you able to rank the allergy risk between individual processes or products? • are your company using any methods for predicting a possible risk of sensitization for any particular product? technical guidance on the studies concerning the safety of use of the additives for users/workers. prepared by the panel on additives and products or substances used in animal feed insights into immunoglobulin e receptor signaling from structurally defined ligands the role of basophils in helminth infection characterisation of pollen allergens chapter cumulative environmental changes, skewed antigen exposure, and 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cheese-packers' disease--respiratory complaints at a cheese-packing dairy symptoms and serum precipitins in workers exposed to dry sausage mould: consequences of exposure to sausage mould penicillium nalgiovense as an occupational and contact allergen cheese worker's hypersensitivity pneumonitis skin prick test reactions to brewer's yeast (saccharomyces cerevisiae) in adult atopic dermatitis patients baker's asthma caused by saccharomyces cerevisiae in dry powder form looking for immunotolerance: a case of allergy to baker's yeast (saccharomyces cerevisiae) occupational ige sensitisation to phytase, a phosphatase derived from aspergillus niger response to a case of occupational asthma due to the enzymes phytase and beta-glucanase ige-mediated allergy to phytase --a new animal feed additive occupational sensitization to fungal enzymes used in animal feed industry report of occupational asthma due to phytase and beta-glucanase exposure-sensitization relationship for alpha-amylase allergens in the baking industry occupational allergic contact urticaria from fungal but not bacterial alpha-amylase two cases of occupational rhinitis caused by biodiastase in hospital and pharmaceutical workers is fungal alpha-amylase in bread an allergen? a dynamic population-based model for the development of work-related respiratory health effects among bakery workers a case of occupational rhinitis caused by porcine pancreatic extract developing into occupational asthma mite amylase from blomia tropicalis (blo t ): differential allergenicity linked to geographical regions diagnosis and management of hypersensitivity reactions related to common cancer chemotherapy agents occupational allergy in a researcher due to ole e , an allergenic , -beta-glucanase from olive pollen biochemical changes and environmental factors in manual and semiautomatic bakeries human serum ige reacts with a metarhizium anisopliae fungal catalase comparative genomics of fungal allergens and epitopes shows widespread distribution of closely related allergen and epitope orthologues prevalence of sensitisation to cellulase and xylanase in bakery workers occupational asthma caused by cellulase and lipase in the detergent industry pharyngeal edema caused by occupational exposure to cellulase enzyme occupational contact urticaria from cellulase enzyme occupational asthma and ige sensitization to cellulase in a textile industry worker enzymes and microorganisms as respiratory sensitizers in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document baker's asthma due to xylanase and cellulase without sensitization to alpha-amylase and only weak sensitization to flour maize food allergy: lipid-transfer proteins, endochitinases, and alpha-zein precursor are relevant maize allergens in double-blind placebo-controlled maize-challenge-positive patients atopic allergens of plant foods a chitinase-like protein in the lung and circulation of patients with severe asthma chitinases and chitinase-like proteins in t(h) inflammation and asthma allergenicity of hev b , a major esterase allergen in natural rubber latex (hevea brasiliensis) allergy, does not only depend on its carbohydrate moiety aerobiological and immunochemical studies on carica papaya l. pollen: an aeroallergen from india allergy to aspergillus-derived enzymes in the baking industry: identification of beta-xylosidase from aspergillus niger as a new allergen glucoamylase: another fungal enzyme associated with baker's asthma occupational protein contact 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protection of workers from risks related to exposure to biological agents at work (seventh individual directive within the meaning of article ( ) of directive / /eec) einstufung von bakterien (bacteria) und archaebakterien (archaea) in risikogruppen. bundesarbeitsblatt - : - . enzymes and microorganisms as respiratory sensitizers in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document trba/trgs : sensibilisierende stoffe für die atemwege background, approaches and recent trends for setting healthbased occupational exposure limits: a minireview exposure to airborne allergens: a review of sampling methods detection of mouse and rat urinary aeroallergens with an improved elisa latex sensitivity: an industrial hygiene perspective exposure to multiple indoor allergens in us homes and its relationship to asthma ige-mediated asthma and rhinitis i: a role of allergen exposure? defining occupational and consumer exposure limits for enzyme protein respiratory allergens under reach are we closer to developing threshold limit values for allergens in the workplace? annals of allergy, asthma & immunology : official publication of the american college of allergy exposure-response in occupational allergy exposure assessment of high molecular weight sensitisers: contribution to occupational epidemiology and disease prevention asthma and indoor air: contrasts in the dose response to cat and dust-mite how do we avoid developing allergy: modifications of the th response from a b-cell perspective pet-keeping in early childhood and airway, nose and skin symptoms later in life biological miracles and misadventures: identification of sensitization and asthma in enzyme detergent workers allergic asthma to <i>bacillus subtilis</i> enzyme: a model for the effects of inhalable proteins this contract/grant was awarded by efsa to:technical university of denmark, national food institute, mørkhøj bygade , dk- søborg, denmark. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the company was founded in , and has been handling dried bacteria cultures for years. they produce tons of cultures per year in factories, and handle about different species in the process. this accounts for about % of the world production. they currently have employees active in the production chain. this company produces enzymes for industrial purposes, both from yeast and bacteria and claim to hold more than % of the global market. it has been operating at an industrial scale since the s and employs workers potentially exposed to the enzymes are in the range, each potentially exposed to several products. novozymes a/s monitors allergies among workers thru screening programs, cap-tests, and self-reporting, and also conducts a significant amount of research in the field of allergy.questions to the companies technical part:• which categories of enzyme and/or microbial cultures does your company produce and/or use? • of these categories, which volume does your company handle on a yearly or monthly basis?the present document has been produced and adopted by the bodies identified above as author(s). in accordance with article of regulation (ec) no / , this task has been carried out exclusively by the author(s) in the context of a grant agreement between the european food safety authority and the author(s). the present document is published complying with the transparency principle to which the european food safety authority is subject. it may not be considered as an output adopted by efsa. efsa reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. key: cord- -tbtlnv b authors: massó sagüés, elena; fernández-carrión, eduardo; sánchez-vizcaíno, jose manuel title: risk of introduction of infectious animal diseases for europe based on the health situation of north africa and the arabian peninsula date: - - journal: front vet sci doi: . /fvets. . sha: doc_id: cord_uid: tbtlnv b the current growth of the human population, the intensification of animal production, climate change or globalization favors an increase in the transmission of infectious diseases. risk analysis is the tool that allows the identification of the factors involved in the introduction and the spread of infectious diseases. the main objective of this work is to evaluate the risk of entry of animal infectious zoonotic and non-zoonotic diseases from north africa and the arabian peninsula to countries of the european union. a probabilistic formulation has been developed to obtain the probabilities of introduction of diseases associated with each possible route of entry in the european union. the results show that, among the infectious diseases analyzed in this study, avian influenza and newcastle disease are the ones with a higher risk of entry in the european union and the wild bird's migration is the route with greater impact. it is confirmed a moderate probability of entry of some vector-borne diseases, bluetongue and epizootic haemorrhagic disease, through wind flow from morocco, algeria and tunisia. due to the absence of live dromedary movement to europe, the more likely way of entry of the middle east respiratory syndrome is through the infected people movement from saudi arabia, kuwait, qatar and oman. this study includes different methodologies. a model of vectors dispersion in wind currents has been established to assess the risk of introduction of vector borne diseases. it is applicable both in animal health and public health. a periodical update would be useful to obtain a periodically updated risk analysis and to allow early detection of potential hazard with an increased risk over the previous years. the current growth of human population, the intensification of animal production, climate change or globalization favors an increase in the transmission of infectious diseases between distant geographical areas ( , ) . these diseases represent a threat for both human and animal health, resulting in higher mortality rates and serious economic losses with the decline in production. the risk analysis is an important tool in epidemiology and it is essential to assess the risk of introduction and possible spread of diseases in areas with little impact (emerging and re-emerging diseases). it enables the design and implementation of control measures for the disease eradication, in order to avoid, prevent and minimize the losses or consequences arising from the transmission of diseases ( ) . the development of a conceptual model is necessary to analyze the risk of introduction of diseases. it has to integrate all the identified elements that are going to intervene in the assessment of the risk. the final risk will be estimated from the evaluation of each one of the elements considered in the conceptual model, called "parameters, " and each parameter will be defined by "variables" ( , ) . the parameters considered in this study are the different routes of entry of diseases coming from north africa and the arabian peninsula into the european union (figure ) . the movement of animals, both domestic and wild, is one of the main routes of entry of diseases. animal products such as meat, milk or eggs, along with the transport of semen for artificial insemination, are possible ways of entry of pathogens ( , ) . wild birds are an important agent of transmission of diseases, hosting diseases such as newcastle disease and avian influenza ( , ) , or carrying vectors like ticks that could transmit pathogens (coxiella, anaplasma, babesia, borrelia, rickettsia, and tickborne encephalitis virus) ( , ) . the wind can be a vehicle of pathogens. many of the epidemiological suspicions are based on the fact that certain vector-borne diseases are introduced to distant regions by the dispersion of vectors in the wind flow. it has been suggested that with a wind speed of between and km/h, with a temperature of between and • c, the vector can be transported up to about km ( ) . rift valley fever, epizootic haemorrhagic diseases, lumpy skin disease and west nile fever are examples of vector-borne diseases, transmitted by insects as culicoides and mosquitoes like aedes. scientific studies prove that culicoides imicola, mainly afro-asiatic dispersion, is spreading through europe and it is expected a rise favored by global warming ( , ) . there are other diseases, such as foot and mouth disease, which can also be transmitted by the wind flow (especially in warm regions) up to km by land and km by sea, without being a vector-borne disease ( ) . the tourism and the high frequency of travels may favor an increase in the risk of transmission of diseases. spain is the route in and out europe. immigrants coming from africa and european citizens of african descent are driving frequently from morocco to europe across the iberian peninsula. we consider this entry route for diseases such as the middle east respiratory syndrome (mers), a zoonotic emerging disease with possible transmission within the human species. the majority of the cases have been related to healthcare centers, but transmission is possible between relatives. in an outbreak on south korea in , only one patient infected more than people during his stay in a hospital emergency room for days ( ). according to the data provided at the who database ( ) another route of entry is the movement of contaminated vehicles, potential source of transmission of diseases and can also vehicle vectors. vehicles transporting animals that have not been correctly disinfected could be an important source of pathogens; therefore the animals that are loaded after have a high probability of infection during transport ( , ) . glanders is a zoonotic infectious disease, endemic in africa. it is important to maintain the vigilance with this disease due to the importance of the horses' social network and the high number of movements for international competitions, and to control the disease to avoid its use as a biological weapon ( ) . there are other possible routes of entry such as bioterrorism, biological leaks, illegal trade; but not referred to this work for not having access to detailed and concrete data for a quantitative analysis of the risk of introduction of diseases by these pathways. the main objective of this work is to evaluate the risk of entry of animal infectious zoonotic and non-zoonotic diseases from north africa and the arabian peninsula to countries of the european union by different pathways. sixteen countries of north africa and the arabian peninsula are included in this study (table ) the possible routes of entry of each selected disease, based on an extensive literature review, are provided ( table ). the identification of the routes of entry has been conceptually based on the possibility of occurrence, regardless of its likelihood and frequency. the necessary data for the analysis is collected from different official databases of free access. the information relating to the movements of live animals and animal product origin between countries, are extracted from the faostat database ( ) and completed with cites trade database ( ) . it provides animal movement and animal product data for imports and exports between countries. the data is obtained in quantities. to obtain information relating to the movement of vehicles and people between countries, eurostat ( ) database is queried. it provides flights data (origin, destiny ( ) it is extracted the data referring to animal population in each selected country, the health status for each disease of the different countries and the annual immediate notifications of each disease. demographic information is obtained from the world health organization database ( ) . after carrying out a literature review of the migratory areas in the world and the more frequented areas by migratory birds, "critical site network tool-species search" ( ) was used to obtain data on the species of birds present in each country and the census, and to visualize its distribution on the global map. for the study of the possible introduction of vector borne diseases through the drag of culicoides and/or mosquitoes by wind currents, simulations (figure ) are made for the estimation of wind and particle's dispersion trajectories that reached the european territory during the year . the program "hysplit-hybrid single particle lagrangian integrated trajectory model" ( ) was used to locate areas and periods of maximum diffusion of particles. this program, facilitated by u.s. national oceanic and atmospheric administration's air resources laboratory (noaa-arl), allows creating simulations in any geographical coordinate of the globe, to different heights and dates, by using a file of gdas (global data assimilation system) climate data. this model has been used in other studies of dispersion of arthropods, especially of culicoides as carriers of bluetongue ( , ) . the program allows the creation of dispersion models for different densities (ρ) and particles size. densities of . g/cc for culicoides and . g/cc for aedes aegypti has been obtained using the formula ρ = m/v, m mass and v volume. the following data has been used for this calculation: culicoides with . mm size and . mg mass ( ) , and aedes aegypti with mm size and mg mass ( ) . our interest is to calculate the probability of occurrence of specific events: the probability of entry of j diseases by different routes from country i; where, henceforth, i∈{ ,. . ., } and j ∈{ ,. . ., }. the probability of occurrence of an event a, p (a), is defined as the quotient between the number of favorable cases to the event a and the number of possible cases ( , ) : each probability is calculated as the probability of a set of independent events (x , x . . . ), being the first event the probability of infection in the country of origin and the last one the likelihood of introduction into the country of destination. the first event is the probability of the country i being affected by the disease j, named as ppa j i , calculated as an average based on the number of outbreaks notified in the last years (from to ). we used a weighted moving average formula that gives a high probability when the country of origin had notified outbreaks in the last few years ( , , . . . ) and a lower probability to those countries that had notified outbreaks in the first years included in the study ( , , . . . ) . therefore, in the extreme cases, when a disease has never been notified or, conversely, it has been endemic in a country during the period of study, the probability will be zero or one, respectively. the formula that is needed for the estimation of the likelihood of a country i being affected by the diseases j is: the b j i, xx represents the binary value / -the value is associated to the occurrence of an outbreak of the disease j on the country i in the year xx. the results are shown in table . the probability of introduction of the j disease into the european union through live animal's trade from the country i (pia i j ) is calculated as the proportion of animals that are annually the formula chosen to estimate the probability that a j disease reaches the european union coming from the country i through transport-vehicles (pit i j ) is a ratio between the vehicles or load transported arriving to the european union from the country i and the total vehicles or load transported to the european union, multiplied by the probability of the country i being affected by the disease j (ppa j i ) and the probability of the vehicles being contaminated by the pathogen responsible for the disease j. the probability of a vehicle being contaminated is obtained from the average survival of the pathogen in surfaces (psa j ) and the average disinfection of vehicles ( . ). it is estimated that one out of every ten vehicles cannot be effectively disinfected allowing the survival of pathogens ( ) . the average survival of pathogens in surfaces (psa j ) is obtained based in the literature review ( ) and available data. glanders, newcastle disease and heartwater infective pathogens have a long average survival, more than days (psa j = ). foot and mouth disease and highly pathogenic avian influenza infective pathogens have an intermediate average survival there is no available information about some diseases in some countries; therefore, no probability has been assigned for those countries. in addition, for the countries that had never notified a disease, it has been assigned a zero probability. the probability of introduction of the j disease into the european union through the importation of animal products from the country i (pis i j ) is the ratio between the proportion of animal products that are annually transported to the european union coming from the country i and the total animal products transported to the european union, multiplied by the probability of the country i being affected by the disease j (ppa the probability of the vector or infective particle remaining viable during the route (psv j i ) is established with a review of the conditions of rainfall and humidity in the countries of study (weather history|weather underground). according to the sources consulted, the foot and mouth disease virus needs specific conditions for the dispersal over long distances by air and its survival is determined by the relative humidity, below per cent the virus is inactivated ( ) . for mosquitos and culicoides, above • c temperature and below • c temperature, their survival is limited ( ). a zero probability is assigned when the weather conditions do not allow the survival of the vector or infective particle, and a probability one is assigned when the conditions are optimal for their survival and dispersion. there is no available information about some diseases in some countries; therefore, no probability has been assigned for those countries. in addition, for the countries that had never notified a disease, it has been assigned a zero probability. there is no available information about some diseases in some countries; therefore, no probability has been assigned for those countries. in addition, for the countries that had never notified a disease, it has been assigned a zero probability. with all the probabilities calculated already, we can calculate the total probability of entry of the disease j from the country i to the european union, taking into account all the routes of entry already evaluated(pi j i ). to do this, we calculate the probability of occurrence of the opposite case, the probability of no introduction of the j disease by any of the routes of entry, using the following formula: there is no available information about some diseases in some countries; therefore, no probability has been assigned for those countries. in addition, for the countries that had never notified a disease, it has been assigned a zero probability. for the validation of the model, historic data has been used to relate the obtained results with the historic events. the information available in the oie database wahis interface ( ) has been used to the extraction of the data referring to the exceptional epidemiological events in the european union countries during the last years ( and ). following the methodology described in the previous section quantitative probabilities were obtained. the probabilities of entry for each route, country and disease are detailed in the following tables: tables - . there are some countries without information available regarding certain diseases, therefore, no probability has been assigned. with all the probabilities already calculated, the total probability of entry of the disease j from the country i into the european union by any combination of routes of entry (pi j i ) is obtained and detailed in table . furthermore, the likelihood of entry of a disease j in the european union is represented in figure . the results show that contagious bovine pleuropneumonia has no risk of entry. this disease is not only absent in europe at the moment, but it is only transmissible by direct contact between cattle, and according to the databases consulted, the movement of cattle among the affected countries and the european union is not made. there is no available information about some diseases in some countries; therefore, no probability has been assigned for those countries. in addition, for the countries that had never notified a disease, it has been assigned a zero probability. on the other hand, highly pathogenic avian influenza and newcastle disease have a major risk of entry into the european union. these disease have a high risk of entry into the european union through wild birds migration, due to the existence of different migratory routes that link the european breeding areas with african wintering areas ( , ) , resulting in a constant flow of wild birds among the affected countries of africa and the arabian peninsula, and the european union. bluetongue, epizootic haemorrhagic disease, foot and mouth disease and rift valle fever resulted to have a moderate risk of entry into the european union. it has been demonstrated the possibility of entry of this diseases into the european union through wind dispersion of vectors for bluetongue, epizootic haemorrhagic disease and rift valley fever; or through wind dispersion of virus for foot and mouth disease. there is no available information about some diseases in some countries; therefore, no probability has been assigned for those countries. in addition, for the countries that had never notified a disease, it has been assigned a zero probability. a low risk of entry has been obtained for the following diseases: middle east respiratory syndrome, peste des petits ruminants, glanders and heartwater. to complete the study, the results obtained are compared with the historic events occurred during the last years, shown in table . in and , european countries had notified outbreaks of bluetongue, highly pathogenic avian influenza, newcastle disease and rift valley fever (among all the diseases included in this study). this work demonstrates the possibility of assessing the risk of entry of different infectious diseases at the same time, and by different routes of entry into a large geographical area. different methods has been used; some of them already deployed in other studies about one single disease, african swine disease ( ), avian influenza ( ), bluetongue ( ), etc. in particular, the dispersion of particles in wind currents model has proved to be a very useful tool for the analysis of risk of spreading of certain vector borne diseases. it is applicable both in animal health and in public health, in the interest to improve the health and well-being through the prevention of risk and mitigation of the effects of emerging diseases that originate from the interface between humans, animals and the natural environment ( ). once the model is complete, an annual review may be useful to update the variables and parameters that are being used and to obtain a periodically updated risk analysis which allows detecting potential hazard with an increased risk over the previous years. in this study we worked with a few parameters and variables that can be subject to improvement. it is possible to increase table | total probability of entry of the j disease from the country i to the european union. disease "j" country "i" the number of diseases studied as well as the routes of entry included in this model. it would be interesting, with the necessary data, to include the illegal trade as another possible way of entry. we have evaluated the risk of introduction, not the risk of spread of disease, it may also be interesting to add the probability of the infectious agent that has been introduced, to come into contact with sensitive populations and disseminate. with the obtained results, the following diseases are not currently presenting a risk of entry through animal trade: heartwater, bovine contagious peripneumonia, middle east respiratory syndrome, and peste des petits ruminants. glanders has no risk of introduction through equine meat, although it has risk of entry through horse trade from bahrain and through people and transport vehicles movement (as fomites). rift valley fever has no risk of entry in the european union through wind dispersion of viruses, although it has a certain risk of entry through animal and dairy products trade, and transport vehicles movement containing vectors of the disease. the probability of entry of infectious diseases through animal and animal products trade is relatively low, due to the shortage of commercial movements between these countries and the european union (see supplementary material) . in the movement of live animals, tunisia represents a greater risk of entry of bluetongue and epizootic haemorrhagic disease for the european union, bahrain of glanders and united arab emirates of foot and mouth disease. there is a lower risk of entry of highly pathogenic avian influenza through the movement of live birds from egypt and saudi arabia. in the movement of animal products, the countries representing a higher risk of entry of diseases are egypt, mauritania, niger and saudi arabia, the three of them with a higher risk of introduction of foot and mouth disease into the european union. the probability of entry of infectious diseases through people movement and transport-vehicles is low, but they are the only likely routes of entry of certain diseases such as heartwater, peste des petits ruminants, and middle east respiratory syndrome (mers). cowdriosis is a vector-borne disease transmitted by ticks (amblyoma genus in the majority of cases). the only likely route of entry of this disease into the european union is through the movement of infected animals or carriers of the tick (with a cero probability obtained) of through the transport of the infected tick in transport-vehicles. the most likely entrance of the middle east respiratory syndrome in the european union is through infected people movement (travelers of immigrants) from countries of the arabian peninsula. outbreaks of this disease had occurred in europe in , the origin was a person coming from saudi arabia ( ) . the peste des petits ruminants, transmissible by direct contact and fomites, represents a risk for europe only by the movement of vehicles contaminated by previous trips in which carrying this type of cattle. the probability of introduction of highly pathogenic avian influenza and newcastle disease through wild bird migration is high due to the existence of different migratory routes that link the european breeding areas with african wintering areas ( , ) , resulting in a constant flow of wild birds among the affected countries of africa and the arabian peninsula, and the european union. this pathway may be responsible for the recent entry of newcastle disease in the european union, in the last years ( and ) outbreaks of newcastle disease has been notified in portugal, france, rumania, bulgaria, sweden, switzerland, check republic and cyprus ( table ). the origin of the outbreaks is not defined but following the results obtained, it may be possible that a percentage of these outbreaks may be caused by wild bird's migration from african and middle east wintering areas. there is a high risk of introduction of bluetongue and epizootic haemorrhagic disease in the european union through dispersion of particles in wind currents from morocco, algeria, tunisia and libya, and a moderate risk of introduction of foot and mouth disease through this pathway. the result of bluetongue is validated by the recent notifications; in the last years ( and ) outbreaks of bluetongue have been notified in greece, italy, and switzerland (table ) . in the other hand, there is no risk of entry of rift valley fever through this pathway. if the north african coast countries, the countries of the study that show in the simulations a certain probability of particles dispersion arriving to the european union, remain free of rift valley fever, there will be no risk of introduction of this disease. to confirm the absence of risk of introduction of rift valley fever through this pathway, the situation of the disease in the countries of the study, which so far are not affected, should be checked periodically. the main limitation of the model is the information available. the majority of information managed in this study is coming from official databases (pubmed, oie , gdas , eurostat , faostat ), although they do not have information from all countries of interest. there is only complete information of five countries: morocco, algeria, tunisia, egypt, and saudi arabia. there is not enough information for certain emerging diseases such as middle east respiratory syndrome, so this study can be updated as the research on the different diseases transmission routes progresses. in the same way, there are countries without information available, for lack of declaration on the current health situation or underreporting of diseases outbreaks. once the model is complete, could conduct an annual or biannual review to update the variables and parameters in use, to obtain an updated risk analysis that allows detecting potential dangerous routes of entry by an increase in the risk from the previous period. this work has made possible to assess the risk of entry of different infectious diseases at the same time, and through different routes of entry into a large geographical area. the use of spread sheets for the development of probabilistic formulation has been of vital importance for the collection and analysis of data, although its validity depends on the confidence and quality of the available information. in this case, there is only complete information of five countries: morocco, algeria, tunisia, egypt, and saudi arabia. it has been established a model for vectors introduction in wind flow that confirms the potential entry by this pathway of some vector-borne diseases, bluetongue and epizootic haemorrhagic disease, from morocco, algeria and tunisia. of all the diseases analyzed in this study, newcastle disease and avian influenza are the ones with a higher risk of entry in the european union. the pathway with more relevance in the risk of entry of these diseases is the wild bird's migration. the diseases with a moderate risk of entry are bluetongue, epizootic haemorrhagic disease and foot and mouth disease. these diseases have in common the possible entry through wind dispersion. in the case of vector-borne diseases it is possible by vectors dispersion in wind currents, and in the case of foot and mouth disease it is possible by virus spreading through wind currents. due to the absence of live dromedary movement to europe, the more likely way of entry of the middle east respiratory syndrome is through infected people movement, from saudi arabia, kuwait, qatar and oman. the contagious bovine pleuropneumonia is the only disease with no risk of introduction in the european union, due to the absence of cattle movement from the countries affected by this disease, chad, niger, mali, and mauritania. publicly available datasets were analyzed in this study. this data can be found here: http://www.fao.org/faostat/en/#data/ ta, http://www.fao.org/faostat/en/#data/tp, https://ec.europa. eu/eurostat/data/database, http://www.oie.int/wahis_ /public/ wahid.php/wahidhome/home, http://apps.who.int/gho/data/ node.country, http://csntool.wingsoverwetlands.org/csn/default. html#state=speciessearch, not available any more and changed into http://critical-sites.wetlands.org/en, http://ready.arl.noaa. gov/hysplit.php. js-v conceived the idea for the work. em and ef-c designed the work. em carried out the data extraction for the risk analysis. em and ef-c developed the probabilistic formulation. em designed the tables with the results for interpretation. ef-c contributed in the interpretation of the program hysplit. em obtained the wind and particle dispersion simulations and estimated trajectories. em obtained data from critical site network tool-species search and studied species distribution in the global map. js-v reviewed the design of the work and the results obtained. em wrote the manuscript. ef-c and js-v provided critical feedback and helped shape the research, analysis, and manuscript. cambio climático en españa y riesgo de enfermedades infecciosas y parasitarias transmitidas por artrópodos y roedores foot-and-mouth disease veterinary epidemiology análisis probabilístico del riesgo potencial de entrada y difusión de la análisis probabilístico del riesgo de introducción de la peste porcina clásica en españa risk assessment applied to spain's prevention strategy against highly pathogenic avian influenza virus h n transport of ixodid ticks and tick-borne pathogens by migratory birds transport of ixodes ricinus infected with borrelia species to norway by northwardmigrating passerine birds bluetongue virus serotype outbreak in the basque country (northern spain) - . data support a primary vector windborne transport using climate data to map the potential distribution of culicoides imicola (diptera: ceratopogonidae range expansion of the bluetongue vector, culicoides imicola middle east respiratory syndrome coronavirus: a comprehensive review middle east respiratory syndrome coronavirus (mers-cov)-emergencies glanders: off to the races with burkholderia mallei statistical databases of the food and agriculture organization of the united nations available online at: https://trade.cites. org/ . eurostat. statistical databases of the european commission available online at air resources laboratory-hybrid single particle lagrangian integrated trajectory model long-distance aerial dispersal modelling of culicoides biting midges: case studies of incursions into australia aedes aegypti (l.), the yellow fever mosquito: its life history, bionomics and structure significance of fomites in the spread of respiratory and enteric viral disease sánchez-vizcaíno f, martínez-lópez b, sánchez-vizcaíno jm. identification of suitable areas for the occurrence of rift valley fever outbreaks in spain using a multiple criteria decision framework manual of procedures for wildlife disease risk analysis. paris: world organisation for animal health the supplementary material for this article can be found online at: https://www.frontiersin.org/articles/ . /fvets. conflict of interest statement: the authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.copyright © massó sagüés, fernández-carrión and sánchez-vizcaíno. this is an open-access article distributed under the terms of the creative commons attribution license (cc by). the use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. no use, distribution or reproduction is permitted which does not comply with these terms. key: cord- -j vylbhy authors: martin, r. title: the role of law in pandemic influenza preparedness in europe date: - - journal: public health doi: . /j.puhe. . . sha: doc_id: cord_uid: j vylbhy the european union (eu) is composed of states with widely varying histories, economies, cultures, legal systems, medical systems and approaches to the balance between public good and private right. the individual nation states within europe are signatories to the international health regulations , but the capacity of states to undertake measures to control communicable disease is constrained by their obligations to comply with eu law. some but not all states are signatories to the schengen agreement that provides further constraints on disease control measures. the porous nature of borders between eu states, and of their borders with other non-eu states, limits the extent to which states are able to protect their populations in a disease pandemic. this paper considers the role that public health laws can play in the control of pandemic disease in europe. the states of the european union (eu) form a political and economic community with supranational and intergovernmental responsibilities, and constitute a single market that seeks to guarantee the freedom of movement of people, goods, services and capital between member states. the emergence or re-emergence of diseases such as severe acute respiratory syndrome (sars) and tuberculosis highlighted the need for eu-level health policy, and led to the community action programme - in the field of public health. this programme is now the cornerstone of community public health strategy, focusing on health information and on the community's capacity to react to health threats. in the context of disease control, the executive arm of the eu, the european commission (ec), has responsibility for the co-ordination of epidemiological surveillance of disease between member states and for regulating matters such as case definitions, disease notification and development of disease networks across europe. the ec is assisted by the european centre for disease control (ecdc), which issues protocols on matters of disease reporting and communication of disease information between states and to the ec. the ec and ecdc can only recommend appropriate disease control measures to states. neither is responsible for the management of disease protection and control in individual states. public health powers in relation to disease lie with national governments. it is member states, not the eu, which are signatories to the revised international health regulations (ihr) , although the ihr recognize the role of 'regional economic integration organizations' such as the eu. a thus, if the world health organization (who) were to recommend under the ihr measures falling within eu legislation, such as restrictions on the movement of goods or the processing of personal data, b the eu would need to act collectively, at the initiative of the ec, as member states would be unable to take unilateral action. otherwise, ihr responsibilities lie with individual states. in , a report on pandemic influenza preparedness in the eu noted that substantial progress had been made in preparing for a possible pandemic influenza, but it remained the case that disease control operated at national level. despite encouragement from the eu towards harmonization of approaches, european national plans vary widely in the strategies they have adopted and the public health powers they propose for implementation of those strategies. harmonization of legislative responses to infectious diseases, based upon sound evidence, will be necessary if collaborative efforts in support of infectious disease control are to be effective. to assist in drawing together national responses to pandemic disease, the phlawflu project c was funded to develop public health law expertise across europe, and to examine the legal underpinning of pandemic disease preparedness across the eu and five further european states. d this paper examines obstacles to european commonality of legal responses to communicable disease. there is no doubt that law is an important tool in containment of communicable and non-communicable disease. in the context of pandemic influenza, it is considered that social measures authorized by law will be at least as important as medical interventions. evidence from the influenza pandemic suggests that compulsory home isolation and quarantine were not particularly effective disease control measures because of the difficulty in diagnosing mild cases. nor were such measures likely to be feasible beyond the initial cases. however, compulsory interventions such as school closures, closure of public places and restriction of mass gatherings, along with disease surveillance and hygiene improvement, have proved effective both in influenza outbreaks and in the sars epidemic. international and eu instruments require states to undertake such measures, many of which will require a legal underpinning. the requirement of compliance with the revised ihr and the globalization of disease information and exchange have prompted many states to revise their public health laws. other states, which had no public health legislation, have now enshrined public health laws in legislative form. these initiatives were long overdue. across europe, as elsewhere, national public health laws tended to be old, based on flawed science, and to predate contemporary understandings and protection of human rights. , the allocation of responsibility for public health practice and the role of the state in regulating private behaviours have very different histories across europe. not surprisingly, states have adopted very different positions on the issue of the extent to which constraints can be placed upon individuals for the public benefit. earlier research on european national public health laws in relation to tuberculosis enabled the identification of four different 'families' of public health legislative models in europe. these were: authoritarian (the enforcement of a high number of compulsory control measures); moderate (the enforcement of predominantly compulsory control measures without recourse to prevention powers such as compulsory vaccination or population screening); preventive (where compulsory provisions were oriented towards preventive measures, including screening, medical examination and/or vaccination, rather than compulsory treatment or detention); and the laissez faire model, where few or no compulsory measures existed. a further complication to a comparison of national legal approaches to disease control is the range of different legal systems in europe. while the majority of european states have a civil law legal system based on the french or german systems, some states operate common law systems, e the former soviet states have vestiges of soviet law, and the legal systems of the scandinavian states recognize civil law overlain with some common law. whereas the definitive public health law of some states can be found in statutory form, other states also include the binding decisions of courts. some national legal systems recognize customary law, local edicts or administrative orders as having legal authority. it cannot be assumed that because a particular power does not lie within public health legislation, that power does not exist. determination of the full range of public health legal powers across european states is a technical and difficult task. following the eu working paper on community influenza pandemic preparedness and response planning in , european states have published national preparedness plans. as with the range of approaches to law, there is a wide range of approaches to pandemic preparedness planning across europe. coker and mounier-jack examined european national plans against a who checklist and found considerable gaps and inconsistencies among preparedness plans, with implications for health in both individual states and for europe as a whole. the authors noted that 'the eu has a critical function in protecting its citizens from public health threats. the role of the eu will be essential to ensure improved sharing of knowledge on pandemic response among eu members, to support the effective provision of services, and to coordinate the response at a community level'. few plans address the extent to which proposed interventions are authorized by their national laws. indeed, few state plans acknowledge the need for legal authorization for their proposed measures, and there is often a lack of clarity about the legality of measures. f while there is some commonality across european states in the measures considered appropriate in an epidemic, the formulation of those measures differs from state to state, reflecting the culture and social priorities of individual states. across europe, states have proposed disease reporting networks, social distancing powers, restriction on travel and trade, closure of premises and facilities, and measures regulating the provision of goods and services. however, the extent and scope of these powers vary widely. while most states contemplate powers of isolation and quarantine, some states also propose quarantining flight crews, and authorize compulsory vaccination, compulsory administration of prophylaxis, and compulsory medical treatment. most states authorize the closure of schools and leisure facilities in a pandemic, but some states would also close diplomatic and consular representation, restrict trade union activity or prohibit visitors to inpatients in hospitals. there is variation in the extent to which states will be prepared to requisition persons and property. many states have passed, or are in the process of passing, new legislation to support their preparedness planning. in england and wales, for example, the health and social care act has introduced into the public health act new powers of isolation outside a hospital, powers of quarantine, powers to require the wearing of protective equipment, powers to require people to attend counselling or disease risk training, and the power to require individuals to provide health information. it also provides for the application of compulsory power orders to groups of persons as well as to individuals, provides new border control measures and imposes new obligations to monitor health risk. states that had taken a liberal approach to intrusion on individual liberties for the benefit of the public health have, in the face of the threat of a pandemic, passed laws providing considerable public health powers. french public health law had previously c the phlawflu project has received funding from the eu in the framework of the public health programme. d croatia, turkey, iceland, liechtenstein and norway. e including the countries of the uk, ireland, malta and cyprus. f the spanish preparedness plan, for example, expresses concern regarding the legality of proposals for isolation, restriction of movement and the proposal to make compulsory the administration of antivirals to staff in contact with patients, noting that 'the legal services of the ministry of health will need to study the legal aspects relative to compulsory vaccination and isolation and the restriction of movement according to the constitutional act / of april of special public health measures in public health, articles and '. focused on preventive measures and provided few compulsory powers for disease control. the new french public health code now authorizes isolation and quarantine, obliges individuals to submit to temperature checks, and provides powers to close facilities such as schools, restrict use of public transport, requisition health personnel including students and retired persons, and regulate distribution of medicines. school buildings will be used as centres for vaccination and for accommodation of vulnerable persons. the new code withdraws employment rights such as the right not to work in a situation of danger. employees and public servants in france currently have the right to withdraw from their workplace if they reasonably believe that their work situation presents a grave and imminent danger to life and health, provided that they have alerted their employer to the danger and provided that their leaving does not create a new risk for others. under the new code, which only applies in the particular case of pandemic influenza, this right of withdrawal will not apply in circumstances where the employer has taken all foreseeable measures to reduce the risk of exposure to disease. the formulation of public health measures across europe reflects cultural values and priorities. french law, for example, proposes the possible closure of schools in an epidemic, but the new french public health code, recognizing the importance of education in france, provides very specific measures to protect the right to education of its children. the code acknowledges the need for school closures because children are more susceptible to the influenza virus than adults. however, the code requires that during a school closure, every effort must be made to continue educational provision via the internet, radio and television, and sets out detailed provisions on ways in which education might be continued throughout the pandemic. the difficulty of predicting what legal powers will be needed to exercise effective disease control has led some states to include in their public health legislation a power to make emergency regulations to provide powers that were not foreseen or which would not be appropriate outside an emergency. the health and social care act for england and wales, for example, proposes that where there is sufficient urgency, a legal instrument may be made without following normal parliamentary procedures. the regulation will then cease to have effect after days, unless it has been ratified by a resolution of each of the houses of parliament. the new french code allows that in the case of a grave threat calling for urgent measures, particularly in the case of an epidemic, the minister responsible for health can, by means of an arête, dictate in the interest of public health measures that are proportionate to the risk and appropriate to the time and place, in order to prevent or to limit the consequences of possible threats to the health of the population. in addition to emergency powers specifically addressed to pandemic disease, many european g and other h states have also introduced or updated separate emergency powers legislation to address unexpected threats, to authorize measures that would not normally be acceptable, or to provide powers as a last resort in the face of emergencies where existing legislation is insufficient. other states have constitutional provisions authorizing emergency powers. i it has until now been the case that for the purpose of legislation, emergencies have been conceptualized as aberrations, normally involving an aspect of violence such as war, rebellion or a violent natural disaster. european emergency powers have generally been limited to a 'state of siege' (france), armed rebellion (hungary), or industrial and natural disasters such as earthquakes or the forest fires in greece. they have not been considered a tool for disease control. in the uk, the civil contingencies act has replaced the emergency powers act j in relation to temporary special legislation to respond to serious emergencies. the emergency powers act had provided power to make emergency regulations, following a royal proclamation of a state of emergency, in case of an interference with the supply or distribution of food, water, fuel, light or the means of locomotion that deprived the community, or part of it, of the 'essentials of life'. the civil contingencies act expands the domain of emergency powers so that an emergency is widely defined to include 'an event or situation which threatens serious damage to human welfare', which could potentially include a public health threat such as a serious disease outbreak. while no regulations have been passed to date, there is clearly scope for a heavy-handed response in the event of a public health threat. the civil contingencies bill in its original form underwent prelegislative scrutiny by a joint committee which noted that the bill: ' in the wrong hands, [the bill] could be used to undermine or even remove legislation underpinning the british constitution and infringe human rights. our democracy and civil liberties could be in danger if the government does not take account of our recommended improvements'. the bill was revised and the government agreed to remove a clause that would have prevented emergency regulations from being subject to judicial review with the consequence that the regulations could not be suspended or struck down by a court if they were challenged on human rights grounds. the committee recommended that certain acts of parliament of major constitutional significance should be exempted from a power to modify or disapply legislation, but this remains in the final legislation. the committee also proposed that those powers set out in part of the bill should be subject to a 'sunset clause' and expire every years from royal assent unless renewed by parliament. this was rejected by the government as inappropriate, because the bill contained enabling powers that were intended to deal with a problem that was 'not short-term'. this suggests that a new approach is being taken to the meaning of 'emergency'. under earlier emergency powers legislation, an emergency was determined by a royal proclamation, but under the civil contingencies act, a state of emergency is to be announced, without initial reference to parliament, by the secretary of state or a senior minister. public health emergency planning in the uk appears to acknowledge that the civil contingencies act will have a more general role in the control of disease, although how these plans relate to new powers under the public health act remains to be seen. a senior spokesperson from the english department of health told the author that the department does not intend to use emergency powers contained in the civil contingencies act for pandemic influenza. powers, contained in the health and social care act should provide all the necessary legal powers to contain and control disease. however, it is clear from government documents on pandemic planning that there is every expectation that civil contingencies act powers will be used should the need arise. , the finnish national preparedness plan for pandemic influenza recommended amending finland's emergency powers act so that a major epidemic can be classified as a state of emergency as defined in the act. previously, an emergency was defined to include an armed attack against finland, a serious violation of the territorial integrity of finland, a threat of war, a serious threat to the livelihood of the population or the economy by interrupted import of indispensable fuels and other energy, or a catastrophe. finland's communicable disease act of already contains quite intrusive powers including the power to administer compulsory mass vaccination by the defence forces, compulsory medical treatment, isolation from the workplace, and disease reporting that discloses personal information. the concern with use of emergency powers for disease control is that disease control ceases to be a matter of health protection, and becomes an issue of foreign and national security, with the risk of being hijacked by the agendas of security policy and politics. ,l this has become even more pronounced with the merging of responses to naturally occurring infectious disease and bioterrorism within emergency powers legislation. the who and european commission have both established committees with responsibility for public health as a security issue. m it is questionable whether the suspension of separation of powers and potentially of civil rights and liberties is justified in the name of public health, and arguable that recognition of human rights is essential for dealing effectively with an epidemic. draconian quarantine measures can be counterproductive, and may even encourage people to avoid seeking medical treatment. emergency powers exercised for public health reasons treat citizens as the enemy, and reinforce the philosophy of original public health legislation which classified diseased persons as a public health nuisance to be removed and excluded from society for the benefit of the well. hong kong, which has had recent experience of epidemic disease, considered but rejected expanding its emergency regulations ordinance to cover pandemic influenza, concluding that public health powers were sufficient and appropriate to disease control even during a pandemic. n hong kong has instead amended its quarantine and prevention of disease ordinance in the light of its sars experience. emergency powers and human rights all council of europe member states are party to the european convention for the protection of human rights and fundamental freedoms, and any person whose convention rights have been violated by a state party can take a case to the european court of human rights. in the uk, the human rights act brings provisions of the convention into uk domestic law and enables human rights actions to be brought in a domestic court. rights with particular relevance to public health powers include article (right to life), article (an absolute right to freedom from torture and inhumane and degrading treatment), article (a qualified right to liberty) and article (a qualified right to private and family life). in enhorn v sweden, a human immunodeficiency virus (hiv)-positive man detained by swedish public health authorities on public health grounds successfully challenged his detention on the grounds that it breached articles and of the convention. the european court of human rights held that any detention must comply with the principle of proportionality, there must be an absence of arbitrariness, detention must be a last resort measure, and any detention must have as its objective not only protection of the healthy but also care of the ill. there has been little judicial challenge in british courts of the exercise of emergency powers. in relation to the english emergency powers act and the emergency powers (defence) act, the courts have played a minimal role, striking down only a handful of emergency measures as ultra vires, usually well after the emergency. more recently, in a case where the british parliament had sanctioned the indefinite detention of any person not a british citizen and certified as a 'suspected terrorist', and where the government had derogated from both the european convention and the international covenant on civil and political rights (iccpr) on the grounds that there was a 'public emergency', the house of lords rejected the government's assertion that the derogation was consistent with the european convention. although the majority of judges declined to question whether there was a public emergency on the grounds that the existence of such an emergency was largely a matter for the government to determine, they concluded that imprisonment of non-citizens alone was neither proportional, given the equal threat from citizens, nor necessary, and questioned the irrationality of singling out a minority (non-citizens) for special burdens, when members of the majority could present an equal risk. lord hoffmann was prepared to consider the notion of an emergency and he found it to be a threat to the 'organised life of the community', which would include not merely a threat to the physical safety of the nation, but also to its fundamental values: 'the real threat to the life of the nation, in the sense of a people living in accordance with its traditional laws and political values, comes not from terrorism but from laws such as these.' the decision suggests a judicial role in overseeing government powers in emergencies. the concept of a 'public emergency' is considered under article of the european convention for the protection of human rights and fundamental freedoms as 'a situation of exceptional and imminent danger or crisis affecting the general public, as distinct from particular groups, and constituting a threat to the organised life of the community which composes the state in question.' o article allows that states might derogate from some of their obligations under the convention 'in time of war or other public emergency threatening the life of the nation', but not from article (right to life) or article (prohibition of torture and inhuman or degrading treatment). the former european commission of human rights, which in defined a public emergency to consist of a 'threat to the organised life of the community', was called upon to determine the criteria of a public emergency threatening the life of the nation in a case in which the greek government sought to justify derogation of rights on grounds of a public emergency. the commission held that the emergency must be actual or imminent; it must affect the whole nation; the continuance of the organised life of the community must be threatened; and the crisis or danger must be exceptional, in that the normal exceptions permitted by the convention for the maintenance of public safety, health and order are inadequate. derogations may only last for as long as, and only be exercised to the extent required by, the demands of the circumstances. they must not limit the subject's rights of access to court protected in article of the convention, nor the right of a remedy protected in article . in circumstances where a state wishes to exercise emergency powers which might contravene human rights, the state is required to make a formal derogation under article of the european convention indicating the rights and the territory to which the derogation applies, and to keep the secretary general of the council of europe informed of the measures taken, the justifications, and the cessation of operation of emergency powers. similar requirements can be found in the iccpr. if european states are to abide by their commitments under the convention, it seems that use of emergency powers will be subject to human rights examination. states will not then be able to exercise their powers in an arbitrary way, and will not be able to respond in a manner that is not proportional to the risk. despite these safeguards, the use of emergency powers legislation for serious ongoing disease outbreaks is questionable. an influenza epidemic could, on a worst case scenario, last for years, which would potentially allow the operation of emergency powers that derogate from human rights protections for a considerable period of time. there is a danger that laws made in the form of emergency regulations might, if in force for long enough, become embedded in the legal system and so constitute a permanent assault on liberties which had previously been achieved, as might be suggested of terrorism legislation in the uk. emergency powers in the context of disease, based as they are on responses to war and catastrophes, tend to operate in such a way that persons affected with disease are characterized as the enemy. they propose that in public health emergencies, there must be a trade-off between the protection of civil rights and effective public health interventions. however, the ideals of democracy, individual rights, legitimacy, accountability and the rule of law suggest that even in times of acute danger, government should be limited in the activities that it can pursue and the powers that it can exercise. as gostin points out in the context of the us model state emergency health powers act, this is not to say that individual rights should always trump public health, but that individual rights should never be infringed 'unnecessarily, arbitrarily or brutally'. nevertheless, there has been significant criticism of the us legislation, and concern that measures proposed in the act are sufficiently dangerous as to 'undermine.constitutional values'. while emergency powers might provide short-term solutions to serious threats, they could also do long-term harm to public trust in public health services, and encourage health behaviours which are counterproductive to the public health. in many countries, including the usa, there are signs that public health and national security are increasingly conflated. the ihr are framed around the assumption that disease is a security issue. however, the danger of subsuming disease control within foreign and national security is that the focus is on security rather than on health. wider national and international interests may not always coincide with public health. global public health may not always coincide with the security concerns of individual states, particularly more powerful states. mcinnes and lee note that policy responses to the sars epidemic elicited a 'garrison mentality' whereby strict border controls and control of movement of persons became central to disease containment, with consequences for the movement of persons, goods and services. it has been widely argued that the promotion and protection of human rights is inextricably linked to the promotion and protection of public health, and that lack of respect for the rights and dignity of persons or groups of persons can increase their vulnerability to disease contagion. , the importance of human rights to health has been acknowledged in the revised ihr article , which requires that the ihr be implemented with full respect for the dignity, human rights and fundamental freedoms of persons. as mann argues, '.the human rights framework is indispensable both for analyzing the central societal issues which must be confronted and for guiding the direction of societal transformation needed to promote and protect health.' states have a significant number of non-medical tools at their disposal in a disease pandemic, and public health law reform has been undertaken with pandemic influenza in mind. public health legislation around the world now authorizes a wide range of social distancing powers and compulsory screening, examination and treatment measures. in addition, much public health legislation provides for the possibility of some limited emergency measures. nevertheless, some european states have proposed the use of emergency powers legislation to provide exceptional powers in the case of a pandemic; powers which will inevitably constrain the rights of individuals. the evidence base for the need for such exceptional powers has yet to be established, and in the absence of such evidence, there is concern that too heavy a hand will result in long-term harm to public trust in the exercise of population-based disease prevention strategies. early responses to public health threats as reflected in th century public health legislation were premised on building fortresses to protect the healthy (and generally wealthy) from those suffering from disease, rather than on care and protection of the population. public health legal powers tend to focus on containment and exclusion, representing 'the community response to social and economic pressures and the wide spread fear of death and disease' rather than on positive public health outcomes. immigrant populations have long been targeted as carriers of disease, and in relation to diseases such as drug-resistant tuberculosis, increasing incidence in the western world is often attributed to persons entering from states with high tuberculosis rates. much contemporary public health policy has rejected the 'fortress' approach to disease control in favour of seeing the public health mandate as imposing duties upon all members of a society or population, or indeed duties of global health protection. the evidence base for border control as a public health, as distinct from a security, measure is limited, especially in a pandemic. in relation to other diseases such as hiv and tuberculosis, border screening has proved to be unreliable and has shown little benefit for the health of the population. compulsory border screening and refusal of entry to affected persons are contemplated by many states in their pandemic influenza preparedness plans, p and the revised ihr contemplate that who might recommend refusal of entry of suspect and affected persons and refusal of entry of unaffected persons to affected areas, subject to the ethical consideration of respect, to the extent possible, for the individual right to freedom of movement. article of the ihr requires all signatory states to establish points of entry with surveillance and border control capacities. a consequence of having no internal eu borders is that the eu needs a strong common external border. under the eu free p for example, bulgaria, denmark, greece and latvia. movement directive, member states may deny entry of eu citizens and their family members if they are considered to be a threat to public health, but only if this is proportionate and meets strict material and procedural safeguards. most eu member states q have signed the schengen convention, eliminating border controls between participating countries and creating an external frontier. the convention called for a common visa policy, harmonization of policies to deter illegal migration, and an automated schengen information system to coordinate actions in relation to individuals who had been denied entry. the amsterdam treaty incorporated the schengen convention into eu treaties, and set out a plan to integrate policies on visas, asylum, immigration and external border controls into community procedures and into the community legal framework. this has resulted in what is for all intents and purposes an eu external border, with much social and economic activity operating at regional rather than national level. however opt-out r and opt-in s possibilities make it difficult to define an administrative space that falls within the frontier, and there is no overarching political control. rather, decisions are made by means of a complexity of intergovernmental and supranational institutions, and there remains considerable sovereign power in relation to many issues of border and public health relevance. the schengen agreement includes consent to share information about people, via the schengen information system. this means that a person cannot 'disappear' simply by moving from one participant country to another. a country is permitted by article . of the schengen agreement to reinstate border controls for a short period if it is deemed to be in the interest of national security. any schengen country can impose temporary or permanent border controls if it believes itself to be unprotected by other members. under this provision, portugal restricted border entry during the european football championship, as did france for the ceremonies marking the th anniversary of d-day, and again shortly after the london terrorism bombings of july . with foot-and-mouth disease having been confirmed in france, the netherlands and britain, norway, in particular, put its border officers on high alert to prevent spread of the disease into the country. other nordic countries have also increased spot checks on entries into the region, irrespective of their new borderless status, in an attempt to contain foot-and-mouth disease. under the schengen borders code, third-country nationals may be refused entry if considered a threat to public health. one issue that arises from the lack of border controls within europe is the disparity in levels of disease preparedness across europe. in , new member states joined the eu, eight of which are former communist countries in central and eastern europe (slovenia, hungary, czech republic, poland, lithuania, latvia and estonia). these states are characterized by a history of underfunding of health and surveillance systems, unreliability of access to drugs, continuing increase in diseases such as drug-resistant tuberculosis and hiv/acquired immunodeficiency syndrome, and inadequate public health responses to disease. since these states have entered into the eu, citizens can cross borders into other, better-resourced states. in the context of a pandemic, this could mean an influx of persons who are possible disease carriers from poor states with a frail public health system and with insufficient medicines, to other eu states, putting citizens at risk and draining health resources in those states. this creates difficult choices for host countries in terms of the assistance they offer. should they fail to offer healthcare services to mobile populations, these populations will put state population health at risk. should they offer healthcare services to mobile populations, this will strain resources and drain services from home populations. in their comparative study of european national preparedness plans, mounier-jack and coker found that eu states intended to take at least one measure to restrict travel to and from the state during a pandemic, and of these states recommended border restrictions on entry and departure. one state proposed drafting new laws to give stronger border control powers. other states, however, conceded that by phase of a pandemic, while there might be political grounds for restrictions on travel, there would be little public health benefit. the possibility of border closure was an issue examined in exercise common ground, a pandemic influenza exercise for the european union, conducted by the uk's health protection agency over a -day period in november . this was the second of two eu exercises commissioned by the ec to evaluate the ability and capabilities of member states to respond to a health-related crisis, in this case an influenza pandemic. concern was expressed when switzerland indicated that it might consider the closure of its borders, given the location of drug manufacturers in switzerland. france's border closure proposals contained exceptions for pharmaceutical and vaccine materials and workers. the feasibility of instituting border controls within europe in a pandemic was then examined at an eu pandemic influenza workshop in august . it was concluded that while border closure might be a useful early containment strategy, at a pandemic stage, it would be impractical to enforce border controls within europe because of the porous nature of european borders and because of the need for cross-border traffic of goods. any prolonged border control would disrupt critical supply chains, and there was a risk that the consequent disruption of border controls within europe would result in greater harm than benefit. screening at borders for diseases such as tuberculosis and hiv is common practice in many states, but has been much criticized on grounds of evidence and ethics. a systematic review looking at the effectiveness of physical interventions such as screening in relation to respiratory viruses concluded that '(g)lobal and highly resource intensive measures such as screening at entry ports.lacked proper evaluation'. there is also limited evidence regarding the efficacy of screening international passengers on departure or arrival in a flu pandemic, except possibly in the early phase. while control and screening measures may have worked in the days of slow travel, it is now the case that travel times are likely to be shorter than incubation periods, such that port screening will be ineffective in disease identification. nevertheless, the mounier-jack and coker study found that eight eu states proposed entry screening in their pandemic preparedness plans. some european states indicated in exercise common ground that they intend to undertake border screening regardless of the evidence base, on the grounds that such measures provide reassurance to the public, and because the surveillance information might prove useful. the ihr authorize states to require information from travellers about their travels, and to undertake a non-invasive medical examination which is the least intrusive to achieve the public health objective. entry may be refused where the traveller refuses to co-operate. article of the ihr provides that invasive medical examination, t vaccination or other prophylaxis shall not be required as a condition of entry except in limited circumstances, such as to determine whether a public health risk exists, or in relation to persons seeking temporary or permanent residence. in these circumstances, if a traveller refuses to comply, entry may be refused or be made subject to the least invasive procedure to achieve the public health objective. article stipulates that such measures be undertaken within the confines of express informed consent and national and international safety guidelines, and article requires that in implementing measures, travellers are to be treated with dignity and respect, and with recognition of gender, sociocultural, ethnic or religious concerns. within the eu, border measures are a matter of community competence that require state co-ordination. where eu member states intend to adopt border measures for the control of communicable diseases, they must inform and, where possible, consult other member states and the commission in advance. the exercise common ground report and the eu pandemic influenza workshop concluded that there was variability in the extent to which member states, european economic area states (including all eu countries plus iceland, norway and liechtenstein) and switzerland have included an international dimension in their pandemic influenza plans. rather, they have focused on national, domestic issues. it is necessary to consider an international dimension because: 'in a community like the eu, free of internal borders and with many common activities and free movement of people and goods, any countermeasures taken in one member state will be bound to affect at least some if not all, other member states'. the reports noted that states also needed to address issues surrounding expatriates, travel restrictions, restriction of emigration, issues of contact persons and the potential for social disorder. there was a lack of clarity around community law on implementation of travel restrictions, and some confusion regarding the extent to which issues of freedom of mobility needed to be handled differently according to an individual's nationality. to be practical and costeffective, border measures would require policy coordination between countries of arrival and departure, and consensus between neighbouring states to avoid disruption. however, as mounier-jack and coker note, few countries address the issue of collaboration with neighbouring states on matters of travel restrictions in their plans: ' there is clearly a need for countries within a european region to be informed and to inform others of their respective strategies in order to ensure that policies are consistent where necessary, or pose as few challenges as possible to public health protection where differences or inconsistencies exist. there may also be a need to ensure that european response mechanisms work together in harmony if public health interventions are to be similar in different countries'. national generic plans in europe have addressed issues of border control rather inadequately. questions have been raised about mobile populations and their implications for healthcare resources, but the issues remain unresolved. there appears to be political reluctance in the context of a united europe to invoke exemptions from internal market rules of free movement of goods and persons on grounds that neighbouring states are failing to address public health threats, and while there is concern about the consequent risk to populations, most states have taken the pragmatic view that any border control should take place at europe's external borders and not within europe. the variation in public health resources and in public health legal powers across eu states, in a context of free borders, is a concern for europe-wide pandemic disease strategies. it is not impossible that states with the strongest national public health powers, which permit, for example, compulsory vaccination or detention, will find some citizens moving states to avoid imposition of these powers. ideally, states within the eu will work together to achieve some commonality of pandemic disease policy and some commonality in their public health legal frameworks. much has been done to develop common policy approaches to preparation for an influenza pandemic across europe. however, article of the european treaty, which states the eu objective of a high level of health protection and requires the european community to work with nation states to deal with health threats, does not allow for a policy of harmonization of state laws. the most that can be hoped for is some convergence of legal powers resulting from discussion and negotiation between states. the differing histories, politics, culture and legal systems of this group of highly divergent states does not bode well for agreement across europe on the appropriate legal response to disease threats. in an attempt to identify the extent to which there is variation in public health legal powers and the consequences of such variation for public health in europe, the phlawflu project is examining the role of national laws in the control of and protection against pandemic human influenza across europe. the objective of the project is to provide an evolving critical study of national laws supporting and constraining defined issues of communicable disease control across europe, and to provide a resource to support public health law reform and public health policy making in europe. the project methodology includes workshops bringing together public health policy makers from european states to examine legal responses to disease scenarios. it is to be hoped that some common legal responses emerge from these exercises, and given the absence of attention paid to public health laws in europe in recent years, that much can be learned by all states on ways in which to use law as a tool in pandemic disease control. meanwhile, europe is in a complex place in relation to its public health approach to pandemic disease. in times of economic strength and freedom from threats of war and disease, the commonality of eu states comes to the fore, and states are ready and willing to engage in joint enterprise. where states are at threat, however, they tend to turn inward on themselves, and political and cultural differences emerge. in times of threat, states which have traditionally been strong on public intervention in private rights are unwilling to accede to the approaches of more liberal states. traditionally, liberal states are reluctant to impose draconian measures, but at the same time may be unwilling to carry the public health burden of citizens from poorer states. the revised ihr have done much to focus public health law reform measures and to ensure some minimum commonality of content, but it is clear that some states, in accordance with their legal culture, are prepared to undertake more intrusive interventions than others. for all these concerns, it is clear that public health laws will be a mainstay of pandemic disease strategies, both in relation to the eu and in relation to nation states within europe. public health laws will be essential in providing powers to enable actions to be taken to control disease spread, but also to constrain states from taking actions that might reassure short-term security concerns but that have potentially harmful long-term public health consequences. of course, such issues are not unique to europe, but the nature of europe as a continent and as a legal entity creates particular complications for the ways in which law might best be used to create a coordinated european pandemic disease strategy. one unexpected benefit of the pandemic threat has been the renewed interest in exploring the role of law as a communicable disease tool, and in the examination of the range of public health legal approaches across europe. globally, a greater understanding of the role of public health law as a tool for managing and minimizing the spread of communicable disease will be a lasting and invaluable legacy of governance efforts in relation to pandemic influenza. none sought. none declared. european centre for disease prevention and control. technical report: pandemic influenza preparedness in the eu. status report as of autumn european public health law network website. available at: www.ephln.org social measures may control pandemic flu better than drugs and vaccine report on the influenza epidemic in nsw in world health organization writing group. nonpharmaceutical interventions for pandemic influenza, national and community measures the exercise of public health powers in an era of human rights: the particular problem of tuberculosis introduction. the importance of law for public health policy and practice public health powers in relation to tuberculosis in england and france: a comparison of approaches public health law and tuberculosis control in europe how prepared is europe for pandemic influenza? an analysis of national plans cabinet office and department of health. pandemic flu, a national framework for responding to an influenza pandemic. london: department of health pandemic flu, influenza pandemic contingency planning: operational guidance for health service planners. london: department of health ministry of social affairs and health. finnish national preparedness plan for pandemic influenza; proposal of the working group on national pandemic preparedness. helsinki: ministry of social affairs and health uk international priorities: a strategy for the fco. cmnd . london: hmso the limits of law in the protection of public health and the role of public health ethics quarantine and prevention of disease ordinance the exercise of public health powers in cases of infectious disease: human rights implications a v secretary of state for the home department a v secretary of state for the home department greek case, yb , opinion of the commission russian federation. ecthr judgment of european commission for democracy through law (venice commission) the interface between public emergency powers and international law the law of exception: a typology of emergency powers drafted by the centre for law and the public's health at georgetown and johns hopkins universities the model state emergency health powers act bioterrorism, public health and civil liberties public-private health law: multiple directions in public health global public health security health, security and foreign policy public health strategies for pandemic influenza: ethics and the law health and human rights: a reader human rights and the new public health human mobility and population health increasing drug resistant tuberculosis in the uk compulsory screening of immigrants for tuberculosis and hiv world health organization. ethical considerations in developing a public health response to pandemic influenza directive / /ec of the european parliament and of the council of mapping schengenland: denaturalizing the border health-care system frailties and public health control of communicable disease on the european union's new eastern border world health organization pandemic phases exercise common ground: a pandemic influenza exercise of the european union department of health and cabinet office. pandemic influenza -sharing of evidence and response policies across the eu. london: department of health workshops and facilitated by dr miguel betancourt cravioto and dr daniel reynders physical interventions to interrupt or reduce the spread of respiratory diseases: a systematic review specifications attached to the invitation to tender document, sanco/c / / , quoted in health protection agency, exercise common ground, a pandemic influenza exercise for the european union, final report. london: health protection agency none declared. key: cord- - gg fm x authors: corradetti, claudio title: reflecting on the eu: the good and the bad times, and those that are yet to come date: - - journal: jus cogens doi: . /s - - -y sha: doc_id: cord_uid: gg fm x nan 'europe will not be made all at once' since its inception, the unification of europe has been conceived as an open-ended project. after the project for a european defence community was abandoned, the eu community rapidly moved from the european coal and steel community (ecsc) with the treaty of paris of to the european economic community (eec) and the european atomic energy community (euratom) under the treaties of rome of . this was followed by an important shift to the common market with the treaty on european union (teu) of the maastricht treaty in . along with the principle of subsidiarity, as a governance rationale for the political and legal aims of the union, maastricht also introduced the european citizenship. finally, following the treaty of amsterdam ( ) and the treaty of nice ( ) , the lisbon treaty amended the two basic treaties of the union (the maastricht treaty and the treaty of rome) and incorporated the eu charter of fundamental rights as binding law. more and more areas and responsibilities have been delegated by states to supranational eu institutions. yet, no predefined design has informed the evolution of the union. whereas, according to a standard vision of federal constitutionalism, the structural openended finality of the european union appears as a weakness, if seen through the eyes of a confederal union, its undefined horizon acquires the peculiar virtue of expressing a capacity of adaptation to an ever evolving will of citizens and states. yet, every fairytale hides its drama and the "drama of all dramas" of the european integration starts with the definition of its constituent power. europe lacks a de facto single demos. is this a sufficient impediment for resorting to radical alternatives? how should we think of the interconnections between different european demoi? how should we combine a republican type of institutional representation with a so-called liberal form, one based on the maximization of state interests? how should we reconcile the peculiarity of the european constituent power with the idea of its open-ended finality? these are questions for which we need to elaborate more accurate answers. a polity of european citizens should be conceived as grounded both on an irreducible plurality of national identities as well as on the projection into the future of a european identity that is never fully attainable. at the beginning it was a pure international organization subjected to the international law principle of pacta sunt servanda. it was only from the s onward, after the case les verts, that treaties started to be regarded as the constitutional charter of the union. for some, this is reminiscent of the biblical story of moses and his people who accepted a constitution without discussing it. problems of legitimacy arouse, but the structure of legal justification had already been changed. in van gend en loos, the then court of justice of the european communities affirmed that community law not only conferred duties on european individuals, but it also conferred rights. the doctrine of direct effects reintroduced the question concerning "in whose name" the community was legitimated to assign rights and duties applicable to the european citizens, once these were no longer traceable back to states. the answer was: well, to the european citizens! yet, the problem remained regarding which overall conceptualization of citizenship to embrace, whether of state citizenship or of abstract cosmopolitanism. additionally, there followed the problem of defining the transmission chain of the constituent wills into the european institutions. the multiplicity of plans for europe is visible today when the multifaceted eu crisis demands a shift from market integration to an integration of core state powers. are we on the right track when deciding on a governance for the current eu agenda? more generally, has the eu governance and its integration process diminished its original liberal constitutional project and contributed, inadvertently, to the rise of the present crisis in all of its most significant dimension? here are four major challenges. first, if one considers only the eu contemporary regulatory governance adopted to contrast the financial crisis, there emerges a clear and progressive stabilization of emergency mechanisms of control which contravene several democratic standards and escape human rights checks. the reference is to the shift from the temporary measures adopted through the second, linked to the former, is the recent populist turn of some eu national constitutional courts who have started replacing standards once based on "common constitutional traditions" with the promotion of a "constitutional identity." the result has been a retreat from a shared european constitutional pattern based on liberal values of freedom and fundamental rights to self-referential legitimations of national autocracies. thirdly, with the outbreak of the refugee crisis, eu governance has tackled problems of relocation, border control, and registration with a fairly modest budget of . million euros for the years - . both schengen and the dublin regulation of have left individual states to decide on asylum requests. national expenditure on the refugee crisis has proved to be inadequate. it followed that relocation plans have remained unrealized and the hope of a distributive burden-sharing for the refugee crisis within the eu states has yet to occur. finally, following the covid- pandemic, the imf has currently foreseen a contraction of the gross world product (gwp) by %. there will not be winners and losers but only more or less damaged economies. within the european area, the loss will be of approximately . %, with the result of further harshening the dichotomy between virtuous economies of the north and struggling economies of the south. if no agreeable common solution is found, each state will act individually. the price for safeguarding the unity of the eu will be unbearable. for now, the european commission has reached a pro tempore compromise deal which combines different limited resources for a total of billion of euros. the instruments under debate by the ministries of finance include, again, a partial resort to the esm, as well as to the european investment bank (eib). they also foresee the use of the commission president ursula von der leyen's sponsored measure sure (support to mitigate unemployment risks in emergency) and a recovery fund to sustain the economies of the most economically burdened states. on this topic see p. genschel and m. jachtenfuchs , - . see g. de búrca , . see o. pollicino february . see p. genschel and m. jachtenfuchs , - . ivi. will these measures be enough to push back the upcoming crisis? almost certainly not, but they will at least prevent a stalemate in national economies. the rest will have to be worked out together. indeed, one of the problems is that for all these types of crises, the eu has relied either too often on community external support as with the recourse to the international monetary fund (imf), the world bank (wb), and the united nations high commissioner for refugees (unhcr) or it has fallen prey of states' nationalisms and populist parties. the governance strategy currently adopted by the eu, while appearing more effective in the short period, has often replaced and slowed down the construction of a community internal process, exposing the eu to external control (an example being the agreement with turkey for the containment of refugees in ). all in all, the current eu crisis has been accompanied by a deficit in the form of shared governance among the eu members. stagnation of political vision at the eu level has resurfaced deeper questions concerning the type of europe we envision either as a more integrated federalist model or as an improved form of quasiconfederal league. in this respect, the commission's "white paper on the future of europe" suggests interesting options to consider. it discusses five possible scenarios concerning what is open to future political choice. these consist of the following: ( ) the basic option of an unreformed union (considering a "carry on" strategy or a contingent tackling of problems), ( ) the abandoning of a political integration and a strengthening of the single market, and ( ) the possibility of higher integration only among those states that ask for it (a sort of coalition of the willing). the final two scenarios include the following: ( ) an increase of efficiency measures on a restricted number of policy areas (higher integration on core state powers) and ( ) a higher extent of integration of a larger number of policy areas pointing to a more federal europe. where will the eu position itself? 'it will be built through concrete achievements which first create a de facto solidarity' solidarity is a concept that we europeans have inherited from the jacobin motto of the french revolution: "liberté, egalité, fraternité." the term was also reaffirmed by marxist philosophy and worker's movements, until its present codification in human rights charters, the maastricht treaty as well as in the clause of the treaty on the functioning of the european union. since the inception of the european project, the preamble of the treaty establishing the european coal and steel community ( ) , echoing schuman's declaration, stated that "europe can be built only through real practical achievements which will first of all create real solidarity […]". similarly, in the treaty of maastricht, the mandate of the eu was said to be one "to organize, in a manner demonstrating consistency and solidarity, relations between the member states and between their peoples (par. , cl. )." ivi. an explicit reference to the concept was made by mirabeau at the french national assembly of october , , when he presented the word "solidarité" as one of the crucial new moral pillars. see h. brunhorst , . treaty establishing the european coal and steel community (ecsc treaty). ivi. these references leave us with the problem of how to accommodate solidarity in the regional context of the eu once this has undergone an extensive elaboration at the national level for over years. the political and philosophical question then turns to how to define the normative principle of solidarity in the eu. how should we frame legitimate relations of mutual cooperation in a multilevel regional polity? a host of candidates promise to fulfill such task, starting from the very popular idea of "constitutional patriotism" (verfassungspatriotismus), a concept originally connected to jaspers and sternberger and, more recently, to habermas. the creation of a de facto solidarity not only embraces a direct relation of mutual support among european citizens. it rather incorporates a scheme of horizontal burden-sharing also involving the relation of states among themselves and in their relation to eu institutions. certainly, the notion of solidarity cannot be left out of the game when attempting to understand the connection between foreign nationals and hosting states (as in case of internal eu migration due to working reasons, etc.). if a sound moral and political principle can be reconstructed from such value, this has to be one matching solidarity with a number of fiscal and economic reforms within the eu by capturing an ideal type of reciprocity. the problem of the eu integration in the wake of the present multiple crises is one which demands new critical understanding. this is not just a matter of choosing whether it is more desirable to go for supra-state cosmopolitanism or for a federative turn. none of these options are desirable per se when conceived in the absence of an overall process of democratic legitimation (currently lacking). the eu is and should remain a distinctive project. one where the political imagination of the peoples of europe will find institutional expression through forms of representation mediated by states, citizens, and european supranational values. in this respect, the eu should progress and expand the consolidation of its double constitution-a supranational and an intergovernmental one-regulating the single market and the monetary union. cosmopolitan ideals and national sentiment is eu supranational governance a challenge to liberal constitutionalism? which european union? europe after the euro crisis from market integration to core state powers: the eurozone, the refugee crisis and integration theory why europe needs a constitution questioning sovereignty: law, state, and nation in the european commonwealth europe's "democratic deficit": the question of standards wirtschaft und verfassung in der europäischen union: beiträge zu recht, theorie und politik der europäischen integration memoirs: the architect and master builder of the european economic community. doubleday and company identity-based-narrative-in-constitutional-adjudication-when-judicial-dominance-matters sangiovanni a ( ) solidarity in the european union treaty establishing the european coal and steel community (ecsc treaty) european neo-constitutionalism: in search of foundations for the european constitutional order the path is still long and winded, but as jean monnet affirmed: "i have always believed that europe would be built through crises and that it would be the sum of their solutions." hopefully solutions, rather than divisions, will appear. key: cord- -a u e o authors: nadeau, s. a.; vaughan, t. g.; scire, j.; huisman, j. s.; stadler, t. title: the origin and early spread of sars-cov- in europe date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: a u e o the investigation of migratory patterns of the sars-cov- pandemic before border closures in europe is a crucial first step towards an in-depth evaluation of border closure policies. here we analyze viral genome sequences using a phylodynamic model with geographic structure to estimate the origin and spread of sars-cov- in europe prior to border closures. based on sars-cov- genomes, we reconstruct a partial transmission tree of the early pandemic, including inferences of the geographic location of ancestral lineages and the number of migration events into and between european regions. we find that the predominant lineage spreading in europe has a most recent common ancestor in italy and was probably seeded by a transmission event in either hubei or germany. we do not find evidence for preferential migration paths from hubei into different european regions or from each european region to the others. sustained local transmission is first evident in italy and then shortly thereafter in the other european regions considered. before the first border closures in europe, we estimate that the rate of occurrence of new cases from within-country transmission was within the bounds of the estimated rate of new cases from migration. in summary, our analysis offers a view on the early state of the epidemic in europe and on migration patterns of the virus before border closures. this information will enable further study of the necessity and timeliness of border closures. in response to the pandemic potential of the sars-cov- virus, many nations closed their borders in order to curb the virus' spread ( ). these closures continue to incur high economic and social costs. to weigh the relative costs and benefits of border closures, it will be important to understand the efficacy of these policies. at the early stages of an outbreak, border closures can delay a pathogen's arrival, thereby giving countries additional time to prepare ( ). however, the success of this strategy depends on timely implementation and a good knowledge of where the pathogen is already circulating. to evaluate the efficacy of border closures in limiting the spread of sars-cov- , it is important to reconstruct the timeline of the early international spread of the virus, before such policies were implemented. in this analysis, we aim to estimate the early patterns of sars-cov- transmission into and across europe. we also address the more specific question of where the predominant sars-cov- lineage circulating in europe originated. we hope that by addressing these questions we can inform further analysis of the efficacy of border closures as a strategy to combat sars-cov- . the sars-cov- virus was identified as the cause of an epidemic in wuhan, china in late ( ) . the epidemic in wuhan was reported to the who on dec. and within one month, sars-cov- was confirmed to have spread to additional countries ( ) . by the end of february , the virus was detected in all who regions ( ) . currently, several lineages of the sars-cov- virus are circulating across the globe. the intermixing of these lineages in different countries and regions suggests that the virus has been transmitted across borders many times ( ) . here we focus on estimating the early introductions of sars-cov- into europe and the virus' migration across european borders. through national surveillance efforts, the first covid- cases in europe were detected in france on jan. and in germany on jan. ( , ) . of the cases detected in europe by feb. , were infected in china, were linked to the initial cases in germany, were linked to the initial cases in france, and were of unknown origin ( ) . in addition to the unknown sources of transmission, some early introductions may not have been detected. this is especially probable given that a significant proportion of infected individuals are likely to be asymptomatic ( ) . in summary, it is difficult to draw firm conclusions about the source, number, and timing of sars-cov- introductions into europe based on confirmed case data alone. viral genomes are an important secondary source of information on outbreak dynamics. if viruses acquire mutations on the same timescale as an outbreak, these mutations can provide information about past transmission events. phylodynamic methods couple a model of viral evolution describing the mutational process to an epidemiological model describing the transmission process. by fitting the combined model to viral genomes sampled from a cohort of infected individuals, we can infer the evolutionary and epidemiological model parameters. here we fit a phylodynamic model with geographic structure to sars-cov- genomes from hubei, china and several european countries before the first borders were closed in these regions. we co-infer the transmission tree linking these sequences, the geographic location of ancestral lineages, migration rates of infected individuals between regions, the effective reproductive number, and the proportion of no-longer infectious cases sequenced in each region. in addition to these inferences, we specifically focus on estimating the geographic origin of the predominant sars-cov- lineage in europe. this lineage is defined by a characteristic amino acid substitution at position in the orf b gene from proline to leucine and was provisionally named the "a a" lineage by the nextstrain team. in the more dynamic, tree-based nomenclature suggested by ( ) , this lineage corresponds to the "b. " lineage described as "a large lineage that roughly corresponds to the large outbreak in italy, and has since seeded many different countries" ( ) . as of apr. , , two-thirds of the sars-cov- sequences collected in europe belonged to this lineage and just % of sequences from the lineage were collected outside europe (data from ( ) , lineages assigned using ( ) ). here, we use the name a a to refer to the group of sars-cov- viruses defined by the orf b:p l mutation. where the a a lineage originated remains unclear. its characteristic orf b mutation was found in some of the earliest confirmed covid- cases in italy, switzerland, germany, finland, mexico, and brazil in late february ( , ) . intriguingly, a late-january sample from a german case linked to all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted june , . . https://doi.org/ . / . . . doi: medrxiv preprint business travel from singapore shares a mutation in the s gene with the a a lineage, but does not have the lineage-defining orf b mutation. this german sample is part of a smaller "a " clade that is basal to the larger clade of a a sequences ( ) . as a result, it was hypothesized that a german transmission cluster may have seeded the larger european outbreak ( ) ( ) ( ) . however, it was quickly pointed out that incomplete and biased sampling must be taken into account before this hypothesis can be rigorously addressed ( , , ) . phylodynamic models with geographic structure aim to account for such biases. firstly, parameter estimates are generated by integrating over a distribution of potential phylogenies, which acknowledges that we cannot reconstruct the true transmission tree with certainty. secondly, sampling parameters are allowed to differ between regions, which acknowledges that testing and sequencing resources vary across regions. here, we fit a phylodynamic model with geographic structure to full-length sars-cov- genomes to (i) estimate the early patterns of sars-cov- spread into and across europe, (ii) weigh genomic evidence for competing hypotheses about the geographic origin of the predominant a a lineage in europe, (iii) report on the epidemiological parameters, and (iv) compare the rate of new cases arising from within-region transmission versus migration during the early epidemic. we fit a simplified version of the multi-type birth-death model described in ( ) . under this model, beginning with a single infected host in a single geographic region (deme), the virus can be transmitted from one host to another (a birth event), die out due to host recovery or death (a death event), be sequenced (a sampling event, assumed to correspond to a death event), or migrate from one deme to another (a migration event). the birth, death, sampling, and migration processes are assumed to occur at deme-specific rates that are constant through time. importantly, this model aims to capture heterogeneity in epidemiological parameters (birth and death rates) and sequencing effort (sampling proportion) among demes. we used a version of the model parameterized in terms of the effective reproductive number, which allows us to additionally infer this epidemiologically relevant quantity for each deme. we analyzed sars-cov- genome sequences from five different demes: hubei province in china, france, germany, italy, and a composite deme of other european countries ("other european"). all sequences were accessed from gisaid ( ) . to represent the pandemic origin, we randomly chose sequences from hubei collected on or before the lockdown of wuhan city on jan. . to investigate the earliest outbreaks in europe, we considered all available sequences collected in france, germany, and italy on or before the lockdown of the lombardy region of italy on mar. . these countries had the first detected (france and germany) and the largest (italy) early outbreaks in europe ( , ) . by limiting sampling to before regional lockdowns and border closures went into effect, we hope to (i) satisfy model assumptions that epidemiological and migration parameters are constant through time, and (ii) get a picture of the early, unimpeded spread of sars-cov- within europe. to represent the pool of sars-cov- circulating in other european countries during this time, we down-sampled sequences from other countries to the cumulative number of confirmed covid- deaths in each country by mar. plus one (table s ). we used this quantity as a proxy value roughly proportional to the outbreak size in each country. table characterizes the sequences analyzed from each deme for the main analysis. as a sensitivity analysis, we repeated the analysis while down-sampling based on confirmed death data from mar. , considering that deaths occur with a delay after transmission. this yielded a slightly larger sequence set for analysis (results in supplement). table . analyzed sequence information. location is the location of sample collection, as recorded in the nextstrain metadata ( ) . date is the date of sample collection, as given on gisaid ( ) (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted june , . we prepared a sequence alignment from publicly available data on gisaid ( ) using the nextstrain pipeline for sars-cov- ( ). short sequences (< , bases), those without fully specified collection dates, and duplicate sequences from the same case were eliminated, as well as sequences from known transmission clusters or with suspicious amounts of nucleotide divergence (as determined by the nextstrain team). we aligned selected sequences to reference genome genbank accession mn . to eliminate suspected sequencing errors, we masked the first and final sites from the alignment, as well as sites , , , , , , and , (following the nextstrain pipeline). we assume that during the time span considered here, the outbreak in hubei and the different european outbreaks were only sources and not sinks for sars-cov- globally. in other words, we assume that (i) once a strain was in europe, the strain could have been transmitted from europe to other global regions, but subsequent re-introductions of this strain did not occur. similarly, we assume (ii) strains were not re-introduced into hubei. these assumptions allow us to ignore sequences from outside of hubei and europe. to justify assumption (ii), we argue there was not sufficient time between the pandemic origin in hubei and jan. , for a significant amount sars-cov- export, transmission outside-hubei, and subsequent re-introduction into hubei. furthermore, confirmed case data shows that hubei province was the epicenter of the sars-cov- pandemic until this time, with comparatively less transmission occurring outside of the province than within it ( ). to justify assumption (i), we tested whether there was evidence for significant migration into european demes by running a separate analysis on a a sars-cov- sampled from all global regions (results in supplement). for inferences, we used the implementation of the multi-type birth-death model in the bdmm package ( , ) in the beast software ( ) . since this is a parameter-rich model, we fixed some parameters to improve the identifiability of others. the values for fixed parameters, priors for estimated parameters, and the rationale behind these decisions are given in table . we ran four mcmc chains to approximate the posterior distribution of the model parameters. the first % of samples from each chain were discarded as burn-in before samples from the chains were pooled. we used tracer ( ) to assess the convergence and confirm that ess was > for all parameters. to weigh the significance of cases from migration versus within-region transmission during the early epidemic, we compare the rate at which new cases migrate into a region (= per-individual migration rate x case count in source region) to the rate at which new cases arise from within-region transmission (= transmission rate x case count in sink region). when signal in the sequence data is low, e.g. for some migration rates, our prior assumptions determine the magnitude of these rates. to assess the sensitivity of our main conclusions to the prior, we additionally analyzed the same sequences using a lower migration rate prior ( figure s b ). we note that the migration and transmission rates are assumed to be constant through time for this analysis. thus, the temporal trends depend only on the confirmed case data, which we take from the johns hopkins center for systems science and engineering ( ). (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted june , . to test our assumption that europe was primarily a source and not a sink of infections before mar. , we analyzed a a sequences collected from different global regions on or before that date. we aggregated sequences into five demes: africa, asia & oceania, europe, north america, and south & central america (table s ) , and then fit the multi-type birth-death model to these data. the most recent common ancestor of the global set of a a sequences was inferred to be in europe with % posterior support ( figure s ). the posterior distributions for the migration rates into europe closely matched the prior, thus the data contains little information on these rates ( figure s ). however, in the analyzed dataset, introduction events were inferred from other parts of the world into europe, while in total migration events were inferred from europe to other parts of the world (table s ) . for our main analysis we focused on estimating patterns of sars-cov- transmission into and across europe. based on the particular set of sequences analyzed, we infer that sars-cov- was introduced from hubei into france, germany, italy and other european countries approximately - times each before mar. (table ). the largest number of estimated introductions was from italy to other european countries. importantly, these estimates reflect only introductions occurring in the transmission history of the analyzed cases, not the full epidemic. in contrast, the inferred migration rate parameters should describe more general patterns of spread between regions. the sequence data were informative for inferring some, but not all, migration rates. we highlight here only the rates for which the data is the most informative; see figure s for a full comparison of posterior and prior distributions. the highest migration rate was inferred to be from italy into other european countries, with a median rate of . /year. the lowest migration rate was from italy to germany, with a median rate of . /year. the maximum clade credibility tree in figure summarizes the posterior sample of transmission trees linking analyzed sequences. the a a lineage sequences form a clear clade with posterior support of . the most recent common ancestor of the analyzed a a sequences is estimated to be in italy with % posterior support. in contrast, the location of the most recent common ancestor between this clade and the basal, singapore-linked german sequence is less certain. this ancestor is inferred to have been in either germany ( % posterior support), hubei ( %), or italy ( %). we find very little support for this ancestor having been in france or another european country ( %). several epidemiologically relevant parameters were co-inferred along with the transmission tree. firstly, we report on the reproductive number in the different demes, which varied from . to . in hubei to . to . in france ( figure s ). secondly, we report on the prevalence of no-longer infectious cases in each deme as of the collection date of the last analyzed sequence. this quantity can be backcalculated from the estimated sampling proportion (prevalence = # sequences analyzed / sampling proportion). we note that both the sampling proportion and prevalence estimates have large credible intervals ( figures s and s ). of the european demes analyzed, the outbreak in germany was estimated to be smaller in early march ( to cumulative cases) than the outbreaks in france ( to , cases) and other european countries ( to , cases), while the outbreak in italy was the largest ( , to , cases). figure compares the rate at which we estimate new cases to arise in each region from migration versus from within-region transmission. the estimated rates of new cases from migration and withinregion transmission are represented here as point estimates days before the date of case confirmation, which assumes a -day delay between infection and onward transmission or migration. beginning with the first day on which we have case data from hubei, we estimate a substantial risk of infected individuals migrating from hubei into european regions. throughout late january to mid-february , cases were sporadically detected in each european region, each of which is associated with a risk of subsequent within-region transmission. sustained within-region transmission is first evident in italy in mid-february. shortly thereafter, sustained within-region transmission occurred in other european countries, in france, and in germany. by mar. , the estimated rate of occurrence of new cases from within-region transmission is within the estimated bounds on the rate of new cases from migration for each region considered ( figure s a) . we obtain the same qualitative result in our sensitivity analysis using a very different prior on the migration rate ( figure s b ). we note that the rates in figure are underestimates of the rates of new cases arising due to migration or transmission due to the underreporting in the confirmed case data. however, assuming that the amount of underreporting is comparable across regions, we can indeed compare the rates. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted june , . . https://doi.org/ . / . . . doi: medrxiv preprint figure . maximum clade credibility tree. the clade of a a sequences analyzed is highlighted with dashed branches. the values above the branches are the posterior clade probabilities and the pale red bars show the % highest posterior density interval for node ages. the pie charts at nodes show posterior support for the ancestor being located in each deme (note that we assumed the root of the tree was in hubei with probability ). the deme for each tip is the deme in which the sequence was collected, irrespective of travel history. tips are annotated with gisaid accession identifier. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted june , . . https://doi.org/ . / . . . doi: medrxiv preprint table . median inferred number of introductions from each source deme to each sink deme along the transmission tree linking analyzed cases. hubei is assumed to be a source only. values in brackets are the upper and lower bound of the % highest posterior density interval for these estimates. source the median of these rates is show in the "migration" row. we also multiplied the number of newly confirmed cases in each sink region by the posterior sample of transmission rates for the region. the median of these rates is shown in the "within-region transmission" row. dates are lagged days to account for a -day delay between infection and migration or onward transmission. case data comes from ( ). we inferred the early spread of the sars-cov- virus into and across europe as well as the geographic origin of the predominant a a lineage spreading in europe. to do this, we applied a previously published phylodynamic model to analyze publicly available viral genome sequences from the epidemic origin in hubei, china and from the earliest detected and largest european outbreaks before mar. . after performing bayesian inference, we (i) report on inferred patterns of sars-cov- spread into and across europe, (ii) compare posterior support for several hypotheses on the origin of the a a lineage, (iii) report on epidemiological parameters, and (iv) compare the timeline of new cases resulting from migration versus within-region transmission in europe before borders were closed. genome sequence data indicates that prior to mar. , sars-cov- was introduced from hubei province into france, germany, italy and other european countries at least - times each (table ) . these estimates, which are based solely on genome sequence data, provide a complementary account of introduction events compared to line-list data ( ) . the introduction events we report here are inferred to have occurred along the transmission tree specific to the analyzed sequence set and are not all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted june , . . https://doi.org/ . / . . . doi: medrxiv preprint attributable to individual cases. in comparison, line-list data ( , ) attributes introduction events to individual cases but cannot reconstruct previous, unobserved introductions. since we analyze only a fraction of all cases, we expect our estimates to be a minimum bound on the true number of introductions. ideally, we want to go beyond counting migration events amongst the analyzed sequences and investigate general dynamics. to do this, we would interpret inferred migration rates as representing more general patterns of sars-cov- spread. however, the sequence data was only informative for inferring some of these rates ( figure s ). in regions with few lineages circulating during the period considered, there is little signal for the amount of outward migration. we observe information about the per-individual migration rate from italy to other european countries ( figure s ). however, we do not find evidence for preferential migration paths from hubei into different european regions or from each european region to the others, although we cannot exclude this possibility. we estimate that the a a viruses spreading in europe by mar. had a common ancestor in italy sometime between mid-january and mid-february (figure ). in contrast, nextstrain places this ancestor in the u.k. with % confidence ( ) . however, the nextstrain result may be an artefact of disproportionately high sequencing effort in the u.k. since biased sampling violates the assumptions of the "mugration" method employed ( ) . we additionally report that the a a lineage was most likely carried from hubei to italy or from hubei to italy via germany. both transmission routes have almost equal posterior support under our model assumptions (figure ). since we only consider a few geographic regions, these migration routes are not necessarily comprehensive. rather than reconstructing a complete transmission chain, we compare model support for different a a transmission routes amongst the analyzed demes and report two equally plausible routes. although it is not the main focus of our analysis, we also report on epidemiological parameters of the early outbreaks considered. estimates for the reproductive number fall roughly within the range of previous estimates ( ) , though we mention a particular caveat with respect to the reproductive number in hubei below. unsurprisingly, prevalence estimates in early march generally exceed confirmed case counts by a factor of - ( figure s ). our inferences of epidemiological parameters do not challenge the idea that the early reproductive number in different outbreaks is difficult to estimate precisely, but not hugely variable, and that there is substantial under-reporting in line-list data ( ) . finally, we estimated the rate of new cases arising from migration compared with the rate of new cases arising from within-region transmission in the regions analyzed. the magnitudes of these rates are quite uncertain due to uncertainty in the inferred migration and transmission rates ( figure s ) and underreporting in case counts, which we implicitly assume to be constant in time and between demes. however, the temporal trends suggested by these data are still compelling and robust towards different prior assumptions. we see that under sustained risk of case migration from hubei, isolated cases were confirmed throughout europe beginning in late january but did not immediately cause large outbreaks. shortly after the first evidence of sustained within-region transmission in italy, outbreaks in the rest of europe also took hold ( figure ). our results based on the multi-type birth-death model take into account phylogenetic uncertainty and sampling biases between demes, which are two major concerns in genomic analyses of sars-cov- ( ) . indeed, wide confidence intervals around internal nodes in the maximum clade credibility tree and low clade support near the tips (figure ) indicate a high degree of phylogenetic uncertainty. therefore, it is important that the parameter estimates we report result from integrating over a distribution of potential phylogenies with different geographic locations assigned to ancestral lineages. in comparison, some initial studies that estimated international sars-cov- spread constructed a median-joining network instead of a phylogeny to account for this uncertainty ( , ) . in this approach, identical sequences are collapsed to single nodes and edges represent mutational differences. this disregards information from relative sampling times and means that ancestor-descendent relationships are highly dependent on the choice of the network root ( , ) . unaccounted-for sampling biases in these analyses may also yield spurious results for the geographic origin of lineages ( , ) . our analysis, which relies on a mechanistic model of migration and between-deme sampling differences, should be robust to such biases. despite the advantages of the multi-type birth-death model just mentioned, there are also several unique caveats to consider. the birth-death model assumes uniform-at-random sampling from the total all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted june , . . https://doi.org/ . / . . . doi: medrxiv preprint infected population in each deme. however, particularly in the early stages of outbreaks, infected individuals were identified by health ministries via contact tracing ( ) . non-random sampling may be one possible explanation for why we infer markedly different transmission rates in china when analyzing cases from within hubei (as in this analysis) as opposed to cases exposed in hubei but sequenced elsewhere (as in our previous analysis ( ) ). furthermore, the multi-type birth-death model assumes that parameters are constant through time and homogenous within demes. as a result, our inferences based on province-, country-, and continent-level demes are only coarse approximations of the true, heterogeneous epidemic dynamics occurring at a local level. in particular, we do not account for a reduction in airline traffic between europe and china beginning in late january, before borders were closed ( ) . due to these limitations, we focus on estimating and interpreting particular events along the transmission tree of the analyzed sequences (e.g. table , figure ) and advise caution when interpreting inferred migration rates ( figure s ). we expect that our results will be useful in parameterizing more specialized models aimed to understand the efficacy of border closures as a means to fight pandemic disease. so far, such analyses have primarily used line-list data and information on travel networks to estimate sars-cov- migration patterns ( ) ( ) ( ) . here we present independent estimates of migration patterns based on genome sequence data. by combining case count data and our estimates for migration and transmission rates, we provide a timeline of early sars-cov- introduction and spread before border closures were implemented. despite migration risk from hubei being on the same order of magnitude as later migration risk from italy, we only observe sustained outbreaks in other european regions after the onset of sustained within-region transmission in italy. finally, before the first border closures in europe, we estimate the risk of new cases arising from within-region transmission to be within the estimated range for the risk of new migration cases. % of world population lives in countries with restricted travel amid covid- updated who recommendations for international traffic in relation to covid- outbreak a new coronavirus associated with human respiratory disease in china who, covid- situation reports coronavirus disease (covid- ) dashboard first cases of coronavirus disease (covid- ) in the who european region beschreibung des bisherigen ausbruchsgeschehens mit dem neuartigen coronavirus sars-cov- in deutschland estimating the asymptomatic proportion of coronavirus disease (covid- ) cases on board the diamond princess cruise ship a dynamic nomenclature proposal for sars-cov- to assist genomic epidemiology. biorxiv hcov- /lineages: resources for calling and describing the circulating lineages of sars-cov- nextstrain build for novel coronavirus (ncov) genomic characterization and phylogenetic analysis of sars-cov- in italy thanks to rapid global data sharing of #sarscov genomic data via https://t.co/tbvb magpy, we can reconstruct large and small scale patterns of #covid spread. this is a thread discussing this intersection of large and sma phylogenetic network analysis of sars-cov- genomes a follow up to yesterday's thread on the possible connection between the bavarian cluster and the italian #covid epidemic regaining perspective on sars-cov- molecular tracing and its implications improved multi-type birthdeath phylodynamic inference in beast phylodynamics with migration: a computational framework to quantify population structure from genomic data beast . : an advanced software platform for bayesian evolutionary analysis covid- ) cumulative cases by day worldwide download today's data on the geographic distribution of covid- cases worldwide early epidemiological analysis of the coronavirus disease outbreak based on crowdsourced data: a population-level observational study inference of transition between discrete characters and 'mugration' models -treetime . . documentation the reproductive number of covid- is higher compared to sars coronavirus repeated seroprevalence of anti-sars-cov- igg antibodies in a population global transmission network of sars-cov- : from outbreak to pandemic. medrxiv medianjoining network analysis of sars-cov- genomes is neither phylogenetic nor evolutionary on the use of median-joining networks in evolutionary biology evolving covid- conundrum and its impact sampling bias and incorrect rooting make phylogenetic network tracing of sars-cov- infections unreliable airlines suspend flights due to coronavirus outbreak -reuters the effect of travel restrictions on the spread of the novel coronavirus (covid- ) outbreak. science ( -. ) outbreak dynamics of covid- in europe and the effect of travel restrictions impact of international travel and border control measures on the global spread of the novel coronavirus outbreak s.n, t.v., j.s., j.h., and t.s. thank eth zürich for funding. all rights reserved. no reuse allowed without permission.(which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity.the copyright holder for this preprint this version posted june , . key: cord- -wfvc l authors: perrin, christophe; cloez, sandrine; dujardin, catherine; ravinetto, raffaella title: europe should lead in coordinated procurement of quality-assured medicines for programmes in low-income and middle-income countries date: - - journal: bmj glob health doi: . /bmjgh- - sha: doc_id: cord_uid: wfvc l nan a secured supply of quality-assured medicines and other medical products is an essential prerequisite for universal health coverage. unfortunately, on average one in medicines do not meet acceptable quality standards in low-income and middle-income countries (lmics). the high prevalence of poor-quality medicines in lmics greatly depends on the globalisation of pharmaceutical production and distribution, combined with the weakness of many national medicines regulatory authorities (nmras). the use of non-quality-assured medicines, often undetected, causes poor case management and unfavourable medical outcomes in individual patients, while at population level, it is translated in poor control of communicable diseases, emergence of resistance to medicines and loss of trust in health systems. - risks are magnified by the covid- pandemic, which triggered disruption of supply chains, stockouts, substandard production, falsification of repurposed medicines and irrational use of medicines. european taxpayers' money is used to fund medical programmes in lmics in the context of humanitarian aid and development. medicines for these programmes are purchased either at international suppliers specialised in the humanitarian sector, or locally in the countries or regions of intervention. these purchases are not exempted from the quality risks that exist in the local and international market. hence, adequate pharmaceutical procurement and quality assurance (qa) policies are needed for three reasons. first, to mitigate the risk of purchasing products of poor quality. second, to assure the same quality standards that would be required for medicines marketed in the donor country. third, these policies are needed to address fundamental moral obligations in terms of equity, transparency and accountability. various european donors play distinct and complementary roles here: the european commission (ec), the national ministries of foreign affairs and the national development cooperation agencies with their aid implementers. the role of donor agencies is particularly crucial. if a donor does not prioritise qa requirements in pharmaceutical procurement policies, and does not foresee a dedicated budget line to secure quality, its aid implementers might choose supply channels that are not fully reliable, or they might purchase medicines that are not subject to stringent regulation even if they are authorised in the recipient country. summary box ► thoughtful procurement policies in humanitarian and development medical programmes can mitigate the risk of purchasing poor-quality medicines, allowing to address fundamental moral obligation to equity, transparency and accountability. ► european donors are aware of the quality problems in the global pharmaceutical market, and some are already translating awareness into explicit procurement and quality assurance policies. however, a joint position and coordinated action is lacking. ► european donors should share existing knowledge and tools, seek the input of recipient countries, and develop a joint position on how the donor community can help to ensure access to affordable and qualityassured health products-also during public health emergencies such as the covid- pandemic. ► applying stringent and harmonised quality assurance requirements, european donors and their implementing organisations can help shaping the global pharmaceutical market towards affordable, quality assured products. a stakeholder survey carried out at the end of by the institute of tropical medicine in antwerp, belgium, shed some preliminary light on the procurement policies adopted by a sample of european donors and implementing actors of the national cooperation programmes. most european donors in the sample directly or indirectly fund the purchase of medicines for development or humanitarian assistance programmes within their official development assistance (oda) ( %). they are aware of the high prevalence of poor-quality medicines in these contexts ( %) and they acknowledge the need for stringent qa requirements in procurement policies. but awareness is not always translated into formal qa policies and guidelines. only a minority ( %) have developed or implemented internal policy briefs, or procurement policies with clear specifications for pharmaceutical qa. there is a lack of structured mechanisms for the monitoring and evaluation (m&e) of pharmaceutical quality in procurement, and no respondents mentioned any provisions for risk management plans. nonetheless, there are also some positive examples of targeted qa policies. in particular, four european donors set the tone. the directorate-general (dg) european civil protection and humanitarian aid operations (echo) of the ec specifically requires that their aid implementers use positive lists of approved procurement entities. to this aim, dg echo has published since a list of humanitarian procurement centres, assessed according to their quality systems, indicating where to procure medical supplies in priority. among eu member states, belgium explicitly requires since that aid implementers ensure the quality of medicines procured for medical programmes in lmics and avoid double-quality standards between the donor and the recipient country. sweden publicly acknowledges its effort to incorporate guiding principles on qa into its contractual requirements with aid implementing partners ; and in june , the uk implemented an internal qa guidance for procurement and supply of medicines inspiring broader guiding principles for donors. there are various reasons for the apparent delay of other european donors. first, securing safe supply chains meets a variety of hurdles, such as the need of complex contractual arrangements with suppliers, as well as the need of adequate tools for m&e, the institutional lack of specific qa expertise at donors and aid implementers' level and the fear that products that have been rigorously assessed for quality would be more expensive. second, some donors may consider that assuring the quality of medicines remains the sole responsibility of aid implementers and/or recipient countries. third, % of donors in our sample explicitly rely on the qa policies of the international actors they support, such as united nations (un) agencies, the global fund to fight aids, tuberculosis and malaria (gfatm) and the gavi alliance-even if they did not mention any specific policy dialogue with these organisations on pharmaceutical quality in procurement. it is also encouraging that in absence of explicit formal qa policies, awareness is translated into a variety of other initiatives that aim to support recipient countries in pharmaceutical qa, and to mitigate the risk of purchasing poor-quality medicines (table ) . these initiatives are either direct, for example, qa trainings for staff and implementers, and/or capacity building projects for national procurement units or nmras; or indirect, through the support to international mechanisms such as the who prequalification programme and the who global surveillance and monitoring system for substandard and falsified products. many european donors also have internal mechanisms to report quality incidents occurring with medicines purchased with their funds; but it is not clear to what extent findings are shared with peers, and used to adapt and improve existing procurement policies or to orient the policy setting agenda. in an ideal world, each country would count on a stringent nmra, able to ensure the quality of medicines manufactured, distributed or imported into their territory. bilateral and multilateral donors can contribute to reinforcing under-resourced nmras, through targeted capacity building programmes, in the frame of health systems strengthening. however, as long as this longterm aim is not achieved and many recipient lmics cannot secure qa in their own procurement, donors can support them by setting explicit and stringent qa policies for procurement of medicines in the programmes they fund. by doing so, they would be accountable about the optimal and ethical use of oda resources, both to recipient countries, and to tax payers and parliaments in their own countries. pharmaceutical qa should become an integral part of donors' risk management plans and policies. adequate qa policies can be direct or indirect. when funds are directly disbursed by a donor, the donor would require its implementers to purchase medicines according to its own qa policy. when funds are indirectly disbursed through channels such as multilateral or bilateral cooperation, humanitarian programmes, non-governmental organisations, investment funds or development banks, the donors would make use of policy dialogue (eg, via their official representation at board meetings) to monitor whether adequate qa standards are applied and evaluated. monitoring and evaluating a (direct or indirect) qa policy requires donors and aid implementers having easy, ongoing access to disaggregated financial data within oda budgets. this allows them to trace funds spent on pharmaceutical purchases and/or qa capacity building and provides access to up-to-date indicators of availability and quality of essential medicines in medical programmes. presently, the qa policies and the mechanisms for accountability and risk management still vary across european development and humanitarian aid programmes, and only a minority of european donors have explicit qa policies in place. harmonisation of such policies across donors would allow setting adequate standards across aid programmes, and to achieve a better protection of individual and public health in recipient countries. efforts to build a common approach across european donors should be encouraged, but are still in their infancy. the existing models and best practices could serve as a basis for other european donors to develop internal policies adapted to their own cooperation strategies, in the frame of a process of european harmonisation. importantly, the input of aid recipient countries should be requested and taken into due account, so as to codesign policies and procedures which respond to existing needs. but european donors could be more ambitious. in line with the resolution developed for the seventy-third world health assembly on the covid- pandemic, they could develop a joint guiding position to affirm how the european donor community should and can collectively ensure equitable access to and availability of qualityassured health products, including medicines. compared with other approaches that focus on developing market opportunities, or that fail to integrate concerns about pharmaceutical quality, european donors can collectively take leadership in promoting the universal right to safe, quality-assured medicines internationally, in partnership with their counterparts from lmics. european donors could also consider proactively sharing the available information on quality of medicines among themselves, and with recipient countries. they could consider adopting mutual recognition of policies and tools that help securing pharmaceutical quality for all. for instance, european donors could agree on positive lists of procurement entities, at international level and in aid recipient countries; they could share reports on qualified manufacturers at international level and in aid recipient countries; and they could share price lists for priority essential medicines in contexts where several european donors intervene. these measures would be particularly helpful for emergency preparedness. during disasters and outbreaks of infectious diseases there are increased, urgent pharmaceutical needs. in a crisis, purchases need to be done rapidly, with no time for in-depth prequalification of products and suppliers. the ongoing covid- pandemics shows that not only lmics, but also high-income countries are confronted with quality problems under such circumstances, for example, for personal protective equipment and diagnostic tests. under these complex circumstances, the resources and know-how of european donors and their aid implementers could contribute to securing a supply of quality-assured health products, by addressing the underlying vulnerabilities in regulations, markets and supply chains. a comprehensive assessment of european initiatives to support recipient countries could help designing and refining shared best practices. this could be the basis for a reliable procurement system for health products, in line with the joint programming scheme where various european donors and their implementing partners aim at maximal complementarity when addressing health needs in the same recipient countries. there may be fears that quality-assured products are costly, and that additional costs would not be compatible with the attainment of universal health coverage. however, the prices of health products do not depend on manufacturing and qa costs only, but also on manufacturing volumes and market opportunities. if all european donors and their aid implementers would apply stringent and harmonised qa requirements in their procurement policies, they could contribute to shaping the market of lmics towards affordable and quality assured products. this would require awareness and political will at (higher) institutional level, enhanced coordination across european donors, and consideration for the hidden-yet high-cost of inaction for individual and public health. twitter raffaella ravinetto @rravinetto world health organization. global surveillance and monitoring system for substandard and falsified medical products. geneva: world health organization world health organization. a study on the public health and socioeconomic impact of substandard and falsified products: executive summary. geneva: world health organization essential medicines and health products. substandard and falsified (sf) medical products. definitions of substandard and falsified (sf) medical products. geneva: world health organization oxford statement signatories. global access to quality-assured medical products: the oxford statement and call to action a link between poor quality antimalarials and malaria drug resistance? medicines quality assurance to fight antimicrobial resistance rpq/reg/isf/alert . . falsified medical products, including in vitro diagnostics signatories from countries. covid- and risks to the supply and quality of tests, drugs, and vaccines the belgian commitment to pharmaceutical quality: a model policy to improve quality assurance of medicines available through humanitarian and development programs a survey of nongovernmental organizations on their use of who's prequalification program analysis of the quality assurance and pharmaceutical procurement policies of a sample of european donors annex iii: principles and procedures applicable to procurement assuring medicines quality in medicines procurement who global benchmarking tool (gbt) for evaluation of national regulatory system of medical products. national regulatory system (rs): indicators and fact sheets. revision vi version . geneva: world health organization covid- response. draft resolution proposed by albania thailand, the african group and its member states, the european union and its member states covid suspicious certificates for ppe -updated / / itm researchers wrote a position paper to guide the use of rapid diagnostic tests to test for covid- infection cooperating internationally to programme development aid and, develop a coordinated, strategic response to key global challenges universal health coverage: drug quality and affordability can go together acknowledgements we are grateful to dg devco and dg echo of the european commission; the ministries of foreign affairs in belgium, denmark, france, germany, italy, ireland, luxembourg, netherlands, spain, sweden and switzerland; and the aid implementers of the national cooperation programmes in belgium, france, germany, italy, ireland, netherlands, norway, spain, sweden, switzerland and the united kingdom. we thank roeland scholtalbers for re-reading and editing key: cord- -lvn hqk authors: rosenkötter, nicole; clemens, timo; sørensen, kristine; brand, helmut title: twentieth anniversary of the european union health mandate: taking stock of perceived achievements, failures and missed opportunities – a qualitative study date: - - journal: bmc public health doi: . / - - - sha: doc_id: cord_uid: lvn hqk background: the european union (eu) health mandate was initially defined in the maastricht treaty in . the twentieth anniversary of the treaty offers a unique opportunity to take stock of eu health actions by giving an overview of influential public health related eu-level policy outputs and a summary of policy outputs or actions perceived as an achievement, a failure or a missed opportunity. methods: semi-structured expert interviews (n = ) were conducted focusing on eu-level actions that were relevant for health. respondents were asked to name eu policies or actions that they perceived as an achievement, a failure or a missed opportunity. a directed content analysis approach was used to identify expert perceptions on achievements, failures and missed opportunities in the interviews. additionally, a nominal group technique was applied to identify influential and public health relevant eu-level policy outputs. results: the ranking of influential policy outputs resulted in top positions of adjudications and legislations, agencies, european commission (ec) programmes and strategies, official networks, cooperative structures and exchange efforts, the work on health determinants and uptake of scientific knowledge. the assessment of eu health policies as being an achievement, a failure or a missed opportunity was often characterized by diverging respondent views. recurring topics that emerged were the directorate general for health and consumers (dg sanco), eu agencies, life style factors, internal market provisions as well as the eu directive on patients’ rights in cross-border healthcare. among these recurring topics, expert perceptions on the establishment of dg sanco, eu public health agencies, and successes in tobacco control were dominated by aspects of achievements. the implementation status of the health in all policy approach was perceived as a missed opportunity. conclusions: when comparing the emerging themes from the interviews conducted with the responsibilities defined in the eu health mandate, one can identify that these responsibilities were only partly fulfilled or acknowledged by the respondents. in general, the eu is a recognized public health player in europe which over the past two decades, has begun to develop competencies in supporting, coordinating and supplementing member state health actions. however, the assurance of health protection in other european policies seems to require further development. the maastricht treaty from marked the beginning of the health mandate of the european union (eu) as enshrined today in article of the lisbon treaty (tfeu, treaty on the functioning of the european union) [ ] . the original eu health mandate focused primarily on stimulating cooperation between member states and supporting national actions (art. ( ), treaty of the european union (teu)) [ ] . it embodied the union with only limited legislative powers on health matters. although this initial mandate was enhanced through subsequent treaties, today article , still gives the eu relatively circumscribed power in areas of public health (art. ( ), tfeu). healthcare continues to remain a national competence and in this regard, the eu "shall respect the responsibilities of the member states for the definition of their health policy and for the organization and delivery of their health services" (art. ( ), tfeu). despite the restricted treaty-based mandate for health, the eu has a relevant role to play in national public health and health systems policies and has expanded its remit in areas beyond the treaty [ ] . areas affected by eu provisions are extensively described in the literature [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . to illustrate the main developments in the area of what can be called "eu health policy" a timeline is illustrated in table . however, because of its limited legal mandate, some eu legal initiatives were highly contested [ , ] . therefore, one can pose the question of what has been achieved over the last twenty years. it may be argued that, despite its narrow legislative scope, the health mandate has triggered important european actions in certain public health areas like tobacco control [ , ] , infectious disease control [ , ] , european guidelines [ ] [ ] [ ] and the development of an eu public health infrastructure [ ] . in recent years, the ec has summarized in annual reports a diverse nature of key public health achievements such as communications and recommendations, health policies, ec co-financed actions and established networks (e.g. high level groups, scientific committees, platforms) [ ] [ ] [ ] . however, stakeholders in the field provide examples indicating that public health relevant eu policies such as single eu policy assessments on the common agriculture policy (cap) [ , ] , pharmaceuticals [ ] , or the health in all policy (hiap) approach [ ] do not always meet the expectations of the public health community. in these papers the authors express concerns about potentially detrimental health effects [ , ] or disappointment about the support of the policies and approaches aimed at improving health in europe [ , ] . also, eu agencies such as the european centre for disease prevention and control (ecdc) are described as agencies with a limited legal mandate, competences and resources for eu public health but, at the same time, with promising prospects to develop as a renowned international player in the field [ ] . in addition, an evaluation of the eu health strategy acknowledges its status as a guiding framework for ec health policies and joint ec and member state actions table timeline of main developments in eu health policy year eu health policy developments before the introduction of a legal eu health mandate treaty of rome: health is not a priority. two aspects are considered: social security of cross-border workers and occupational health. on health but also identifies the missing impact of the strategy on other ec policies as well as on member state health policies and actions [ ] . evaluations of the eu public health programmes, which are one of the ec's financial instruments to implement its strategic health goals, criticize missing prioritization of topics, barriers for participation in projects for some member states and ineffective dissemination of project results [ , ] . hence, the available evidence of the impact of eu health policies, infrastructure, and actions is elusive, and the identification of the value of public health relevant eu-level actions across all policies is lacking. in this paper, we aim to explore and provide an overview of influential public health relevant eu-level policy outputs and a summary of policy outputs or actions perceived as an achievement, a failure or a missed opportunity by interviewing key experts in the field. by this, we intend to establish a qualitative indication of which eu health policies have contributed to the improvement of population health in europe. the study focused on the evolvement of the health mandate since , the year the maastricht treaty was signed. the study was carried out in two consecutive phases: ( ) qualitative interviews, suitable to identify expert perceptions, and ( ) voting on influential and public health relevant eu policy outputs and actions based on nominal group technique. the study adhered to the rats guidelines on qualitative research [ ] . experts were purposely selected to ensure heterogeneity of opinions. the selection was based on their individual profile and professional affiliation. we selected experts that were renowned in the field due to their current or former affiliation to specific eu-level bodies and institutions, research institutes with eu focus, or eu-level nongovernmental organization. selected experts were actively involved in public health research, policy-making, policy advice or advocacy performed at eu level, internally or externally. in addition, snowball sampling was applied until data saturation was reached. data saturation was assumed as soon as no new eu public health policy actions and their perceptions were mentioned during the interviews. the potential participants were contacted between december and march by a short information email to identify whether they were interested to participate in the interview study. of contacted experts, twenty participated in this study, one participant could not confirm participation due to time constraints and another did not respond to the invitation. of those twenty experts nine belonged to the initial purposively selected sample and eleven were identified during the snowball sampling procedure based on recommendations of already interviewed experts. the majority of experts was affiliated to an institution located in brussels (n = ). the composition of the study sample in terms of represented professional affiliations is outlined in table . upon agreement by the participant, an appointment for the interview was made and participants received an informative letter with more in-depth information about the goal of the study and an informed consent form in which the voluntary basis of the participation has been clarified and anonymized data handling was assured. interviews were conducted either face-to-face (n = ) or via telephone, or voice over ip (n = ) in the period between january and march and were held in english, dutch or german by one of the three principal investigators (nr, tc, ks). the interviews lasted from to minutes. all interviews were audio-recorded, transcribed verbatim and anonymized. the interviews were performed using a specifically designed semi-structured interview guide. the guide was developed on the basis of previous desktop research and an internal brainstorming session of an advisory research group consisting of the three principal investigators, four senior researchers, and one junior researcher of the department of international health at maastricht university to identify items relevant for investigating expert perceptions on european public health policy. during the course of this process, a list of public health relevant eu policy outputs, processes or procedures that were regarded as achievements, failures or missed opportunities were first gathered individually and then, following a group discussion, a common list was compiled. this output was used to construct the interview guide containing six guiding themes from which open-ended questions were formulated. the guiding themes included ( ) a description of the individual role of the expert in european public health, ( ) the individual definition of european public health, ( ) the assessment of public health relevant eu-level actions as being an achievement, missed opportunity or failure, ( ) the formulation of five influential european policy outputs, ( ) consequences of european health policy, and ( ) the policy process at the european level. the semi-structured interview guide was used as a framework for the interviews and allowed the interviewers to address other relevant topics that emerged during the interviews. after completion of interviews the three principal investigators initially performed an internal analysis of each separate interview and an analysis across interviews to identify the scope of eu-level actions and experts' perceptions of these actions as achievements, failures or missed opportunities [ ] . afterwards, a directed (or deductive) content analysis approach [ , ] was applied whereby the initially predefined coding scheme with the main categories of interest (achievement, failure, and missed opportunity) was used to summarize the respective topics and the reasoning that appeared during the interviews. topics that did not fit into one of these main categories were added as new codes and were organized into new categories. the analysis was jointly performed by the principal investigators using nvivo (qsr international pty ltd. version ). furthermore, the results of the content analysis on achievements, failures and missed opportunities were grouped according to the major common themes in a table to provide an overview of the perceptions of the key informants. prominent eu-level outputs or actions, which were discussed by almost all respondents, are described in more detail in the results section. where applicable, we used original quotes to illustrate the views and tendencies of experts' assessments. the professional profile and study id of respondents are indicated behind the respective quotes. quotes which were originally given in dutch or german were translated into english. a slightly adapted nominal group technique [ ] was used for triangulation purposes. during the interviews, the participants were asked for five influential policy outputs of european health policy-making. following the finalization of all interviews, all participant nominations were compiled in one list and reoccurring topics were removed. to ensure comparability of policy outputs and actions, we grouped the nominations into categories under the following headlines: (a) secondary legislation and court decisions; (b) soft laws, strategies, and programmes; (c) agencies, centres, organizations; (d) networks, policy platforms, cooperation; and (e) others. participants were asked in an online survey to select three outputs per category which were, according to their opinion, most influential. based on the participants' nominations, a ranking in terms of a frequency distribution of selected influential policy outputs for each category was determined. the online survey was completed by out of participants who took part in the interviews. the design and analysis of the study was guided by applying guba and lincoln's test for trustworthiness [ ] . credibility and dependability have been ensured by enlarging the sample until saturation was reached in terms of the identification of eu policy actions and their perceptions. moreover, three researchers in the primary research group in combination with an internal advisory research group were involved with the aim of reflecting upon the study design and critically questioning the findings. additionally, the primary research group met regularly during the interview period to exchange initial findings and experiences on the interview process. the members of the internal advisory research group were experienced in eu public health policy research or qualitative research methodologies. the confirmability was strengthened by the use of several investigators both in the data collection process and in the analysis phase, combined with the use of triangulation, where interview participants were also asked to participate in the ranking exercise. the medical ethical committee of the university hospital maastricht and university maastricht declared that no ethical approval was required for this type of research. all participants were informed about their role and rights as study participant prior to their interview participation. all participants provided written or audio-recorded informed consent to be interviewed. overall, respondents consistently mentioned that, during the twenty-year history of the eu health mandate, specific initiators induced change in european public health policy. the most important identified initiators included the maastricht treaty with its later amendments, the health-related rulings of the european court of justice, and the health crises such as boviene spongiforme encefalopathie (bse) and severe acute respiratory syndrome (sars). in addition, the internal market provisions with the foreseen free movement of goods, people, services and capital, initiated change with both negative and positive public health impact. additionally, a set of conditions was identified in the interviews that described and advanced the role of eu health policy as a reference point for public health. these conditions under which eu health policy made progress during the past twenty years were (i) the regulatory power at the eu-level, (ii) eu-led facilitation of cooperation and comparisons across member states; along with (iii) increased capacity building on eu issues and on eu-level (e.g. professionalization, development of interest groups, associations). the ranking of influential policy outputs of eu-level health policy-making is provided in table . in the category "secondary legislation and court decisions", the patients' rights decisions made by the european court of justice (n = ) were chosen by most of the respondents as influential policy output, followed by the directive on the application of patients' rights in cross-border healthcare [ ] (n = ) and the directive on advertising and sponsorship of tobacco products [ ] (n = ). in the category entitled "soft laws, strategies, and programmes", the first and second eu public health programmes [ , ] (n = ) were selected most frequently, followed by the - health strategy "together for health" [ ] (n = ). the third rank is shared by three policy outputs: the "framework for action in the field of public health" [ ] (n = ) which is the commission's first proposal setting out eu-level public health after the introduction of the health mandate in the maastricht treaty, the council conclusions "towards modern, responsive and sustainable health systems" [ ] (n = ), and the current over-arching european strategy "europe " [ ] (n = ). in the third category on "agencies, centres and organizations", the european medicines agency (ema, n = ) ranked top, followed by the ecdc (n = ) and the european food safety authority (efsa, n = ). among "networks, policy platforms and collaborations", the european presidencies (n = ) were selected most often by the respondents, followed by the collaboration of the european commission (ec), the world health organization regional office for europe (who-eur) and the organization for economic co-operation and development (oecd) (n = ). moreover, the three entries on the third rank include the eu health policy forum (n = ), the network on epidemiological surveillance and control of infectious diseases (n = ), and the network on health technology assessment (n = ). the fifth category was not topic specific therefore, work on european level health determinants (n = ), the exchange of best practices (n = ), and published scientific reports which influenced eu policy-making (n = ) were ranked on the first three positions. at a glance, the label "achievement" was allocated to the public health mandate as it is laid down in the treaties, the establishment of eu-level agencies dealing with public health topics and successes in smoking prohibition, food safety and infectious disease control. the label "missed opportunity" was allocated to the insufficient degree to which the hiap approach is implemented and the ways in which health promotional aspects of alcohol and nutrition were handled. the label "failure" was less often assigned with the missing integration or link to social policies appearing in some interviews under this heading as well as the strength of the internal market which annulled national protective alcohol legislations in some member states. in table , we provide the full list of eu-level outputs or actions which, based on the content analysis and the identified thematic categories, were mentioned as achievements, missed opportunities or failures by the key informants. due to a broad and divergent spectrum of perceptions, topics almost always shared aspects of achievements, missed opportunities or failures. in the following section, we focus on those eu-level outputs or actions which were mentioned by the majority of respondents during the interviews and allowed us to draw a comprehensive picture on the breadth and the diversity of expert perceptions. an assessment of the general value of eu-level public health actions over the last twenty years resulted in mainly ambivalent judgments. on the one hand, many relevant activities were performed at an eu-level and the existence of a health mandate contributed to an eu social model. on the other hand, its dependence on political will and economic circumstances influenced the development of eu-level public health policy and led to the perception that more should or could have been achieved within and beyond the possibilities of the current health mandate. the establishment of the directorate general for health and consumers (dg sanco) in as an independent, formal structure for eu health policy was generally discussed as an achievement. the formation of dg sanco, and thus, the political decision to separate the health dossier from dg v, the former dg with the responsibility for health policy as well as a focus on employment and social policies, was controversially perceived. the establishment of dg sanco led, on the one hand, to a more mature health policy field. "…the dg v was a big dg and then dg sanco became separate from that. health had its own commissioner, its own opportunity to protect itself and public health benefits." (# , public health advisor/advocate) on the other hand, aspects of failure were mentioned regarding the detachment of health and social policy at eu-level. according to the respondents being separated led to a loss of collaboration for more holistic health policies and actions in health systems and healthcare at eu-level. a following the formation of dg sanco, it was seen as a beneficial way forward for the dg to shift its sectorial policy approach from a focus on specific topics such as cancer, drug dependence, health monitoring, accidents and injuries, or pollution-related diseases, to a horizontal one with the formulation of the first health strategy with three cross-cutting objectives: health information, health threats and health determinants [ ] . "and this was an important moment in time where the sectorial approach to aids, cancer and other issues has been reduced gradually and that more the integral horizontal approach, which was applied at that time already in all member stateshence europe was running behind in that sense, but ultimately was embraced and taken as guideline for the framing of all sorts of public health actions. "(# , eu/national civil servant) since it fostered more visibility of the public health field and closer cooperation by financing projects, joint actions and research across europe, the public health programme of dg sanco was commonly discussed as being supportive to the development of european public health and the mobilization of the public health community. aspects of missed opportunity became relevant when assessing the representation of health in other eu policies. "i don't know what exactly the reason is, but they [dg sanco] are not strong enough to push for health in [the other] dgs. the obvious example is the latest eu strategy, you cannot find reference to health anywhere it's really a disaster, because of [the] weak dg sanco. health is not among the headline targets, it's not among the flagships." (# , public health advisor/advocate) while the cooperation with other dgs was recurrently discussed as problematic, the potential for the "partnership on active and healthy ageing" under the european innovation union appeared as a unique theme and was regarded as an achievement for strengthening health policy on the general eu policy agenda. ecdc profile should cover also non-communicable diseases and sdoh. coordination of the approval of efficacy, safety and quality of drugs. cost-effectiveness of pharmaceuticals is not taken into account. problem of not being able to tackle pharmaceutical pricing. reversal of the approval of already approved drugs not handled on eu-level. control of health claims of food products. efsa mandate should include/be stronger on health promotion aspects of nutrition (e.g. regulation of advertisement of unhealthy food products). food safety directive. health mandate assures that health protection should be guaranteed in all eu policies. hiap and health impact assessment have never been implemented fully (tick box exercise). leads to the discussion of health in other sectors. control; tobacco product-; tobacco advertising directive). the tobacco regulations could have been designed stronger (e.g. more harmonized realisation of smoking prohibition on public places). tobacco regulation has some aspects of failure since a strict, general ban is not reached. food safety measures and regulations on health claims. missing political will to tackle obesity and related life style factors like unhealthy food products. health research programme eu health research budget and outcomes of the programme. missing integration of the research programme and eu health research outcomes in public health. health research budget. the use of structural funds for investments in health ( ) ( ) ( ) ( ) ( ) ( ) ( ) . internal market rules as source for legislation should be more attentive to health concerns. internal market provisions cause problems if member state regulation is more protective regarding health threats than eu regulation. the patients' rights directive in general. negotiations on patients' rights directive failed to include a strong emphasis on the development of common standards. effect on cross-border cooperation. gives legal certainty to policy makers. policy field which starts to recognize health, e.g. in its white paper on the cap after ( / (ini)). unrecognized potential for health of the cap by public health sector. health life years as indicator in the lisbon strategy. missing health information system. lack of morbidity data. different public health topics health inequalities ec communication: solidarity in health: reducing health inequalities in the eu. strengthening of the hta approach in the eu. coordinating cross-country level health technology assessments. coordinated management of rare diseases. existing drug resistance of tuberculosis as indicator for lacking disease management. health of minorities (e.g. roma) as part of the european agenda. social care is hardly seen as eu competence. environmental standards set by the eu. missing follow-up process on the environment and health action plan ( ) ( ) ( ) ( ) ( ) ( ) ( ) . blocking of direct to consumer advertising of prescription-only pharmaceuticals. white paper on governance ( ) increased transparency. more standardisation of methods (evaluation of indicators, outcomes, policies) and common language. increased understanding of the public health community about the impact of eu policies on public health. cooperation with industry influences the health research agenda and policy-making. evidence-based policy-making: the interest of the industry is against public health. and that's a good sign, if we can get more of those sorts of partnerships on specific policies, then i think, we'll get a better understanding."(# , eu/national politician) with regard to assessing the status of dg sanco cooperation with other international policy actors, respondents had mixed perceptions. whereas some argued that dg sanco's collaboration with international organizations like the who-eur or the oecd is improving and therefore, can be considered as an achievement, others asserted that this collaboration was not sufficiently established and can therefore be categorized as a missed opportunity. the establishment and the work of eu public health agencies like the ema, the ecdc, the efsa, and the european monitoring centre for drugs and drug addiction (emcdda) were regarded as an achievement and as an important step forward towards the strengthening of the european dimension in health. the work and the scope of the agency mandates was a recurring topic and subject to diverging perceptions. as an example, in the case of the ecdc, its development was assessed as an achievement whilst its scope was considered a missed opportunity. the bioterrorism attacks on the united states of america in and the sars crisis led to calls for better international coordination of infectious disease surveillance and the establishment of ecdc in [ ] . hence, the setting up of ecdc was commonly perceived as an achievement, since it gave preceding eu actions in infectious disease control a formal structure and maintained actions in the field. also, the close collaboration with the respective national public health agencies during outbreaks and in negotiating and developing common guidelines for infectious disease control were regarded as an important task of the ecdc. however, a number of respondents were critical of the scope of the ecdc mandate and thus, looked at this as a missed opportunity. questions were raised on whether the ecdc's responsibility in surveillance, risk assessment and training are sufficient or if additional responsibilities in risk communication and management were needed to assure full stewardship during and in the prevention of health crises. "i suppose the flu epidemic […] . that should be put on the table not only as a missed opportunity, big failure, having put ecdc at the center of the development, but the ecdc is not authorized to risk communicational management as you know. so, in that sense, it is a failure that member states were not able to coordinate in this very important public health area and use the eu institution, either ecdc or who to do that." (# , public health advisor/advocate) moreover, interview participants reported tensions between member states and eu agencies regarding the transfer of responsibilities from national to eu-level. "and the member states are very reluctant to hand over power regarding public health to the commission, or to brussels. now if you focus on infectious diseases, that is much better because they understand that there is a need, but again it is not easy." (# , eu/national civil servant) since the largest burden of disease in the eu is caused by non-infectious rather than infectious diseases, a call was put forward to further increase the mandate of ecdc to all public health relevant aspects and not focus only on infectious diseases. the hiap approach was generally assessed as an achievement regarding its potential to address health determinants outside the health sector. "[the article on the health mandate] is very important, because thereby a mandate is created that the commissioner for health and consumer affairs […] approaches his colleagues whenever they make new legislation to ensure that the health protection dimension is guaranteed; it gives partly a mandate to break into the policy and law development in sectors which in principle do not have any links with public health. […] this is very difficult. but its potential is very strong." (# , eu/national civil servant) however, in regard to its degree of implementation participants commonly perceived hiap as a missed opportunity. health impact assessment, the implementing tool to hiap, was regarded as a "tick box exercise" (# , # , both public health advisor/advocate) rather than a thorough consideration of health in other policies areas. explanations given during the interviews demonstrated that conditions to achieve hiap seemed not to be established yet and that there seems to be difficulty in bringing dg sanco interests in line with the interests of other dgs without over-emphasizing the health aspect. political assertiveness in convincing other commissioners and dgs about the relevance for intersectoral cooperation was perceived to be lacking, even though an inter-service group on public health with the participation of more than twenty ec departments was established for this purpose. generally, the work regarding tobacco was regarded as an achievement of how european health policy-making effectively addressed a life style risk factor for health. "the progress around tobacco [directive on tobacco advertising, directive on tobacco products, transparency register], the fact that we have a piece of international law on tobacco [who framework convention on tobacco control] is massive and that was european led." (# , public health advisor/advocate). this quote echoed the perception of the majority of respondents who emphasized the leading role of the eu regarding the support and commitment to the who framework convention on tobacco control. moreover, it was argued that the achievements regarding the regulation of tobacco advertising and smoking prohibition in public places would not have been achieved by single member states independently and thus this was a common achievement initiated and supported by european cooperation. nevertheless, aspects of a missed opportunity or even failure were mentioned in this regard since some would have appreciated stronger legislative measures to achieve a more harmonized realization of smoke-free legislation across the eu. it was considered that the achievements recognized in eu tobacco legislation were missed in the regulation of other health-related life style factors such as nutrition and alcohol. whilst regulations in the area of food safety were generally acknowledged as an achievement by preventing food-borne health threats; a potential mandate to address the composition of food and thereby, prevent, inter alia obesity or non-communicable diseases seemed to be neglected and was labeled as a missed opportunity. "…food safety has been majorly put forward over the last twenty years, in the sense that we know that the food will not be contaminated. but then it is a missed opportunity in the sense that beyond food safety there is health promotion and then one wanted the union to have more powers to regulate issues on the content of saturated fat for instance or the percentages of sugar and so on." (# , public health advisor/advocate) with regard to governmental activity on these issues, the eu platform for action on diet, physical activity and health was named as an example of an achievement as well as a failure. "i think the diet platform […] can be seen as a failure and opportunity. […] if we had not created that platform then arguably the issue wouldn't have been tackled at all. and in a way that has been really brought some issues of complexity to the political discussion around issues around marketing of food, around self-regulation, reformulation, some of the initiative like salt in diet has come as a commitment from that platform."(# , public health advisor/ advocate) the failure aspect of the eu platform for action on diet, physical activity and health was related to the perception that a platform is a rather weak policy instrument and that more political will to tackle these issues with stronger eu policy or legal instruments would have had more impact. additionally, the lack of timely cooperation of public health professionals with other sectors such as agriculture was raised as missed opportunity. it was illustrated that agriculture policy has public health relevant links regarding affordability, accessibility, and the availability of food. however, it was also argued that this cooperation has been developed further over the recent years. "and it is correct, that the common agriculture policy has not been taken up health in the beginning, but by now they are doing this very consciously. […] thus, i really see an improvement; i actually do not see a situation anymore in which health was influenced really negatively [by the common agriculture policy]." (# , eu/national civil servant) internal market provisions were perceived as ambivalent by the respondents. the eu is based on internal market rules that also affect eu health policy. "the engine of european health policies is still the market."(# , academia) however, the influence of eu market regulations, for example on alcohol policies, was perceived as a failure when member states had more protective and stricter national legislation as was the case in the nordic countries. respondents claimed that eu internal market regulations that are more attentive to health issues would be appreciated in this case. moreover, the potential given by articles and of the tfeu, which put limitations on the single market, was mentioned and it was perceived as a missed opportunity that this potential had not been fully taken up by public health experts: "[…] the public health aspect, which is written into article [now article , tfeu] on the internal market, you can put limits on the internal market on the grounds of public safety, public morality and public health, is almost never used. what if dg internal market was turned into our greatest weapon?" (# , public health advisor/advocate) this was positively exemplified by the case of tobacco control which applied internal market rules for public health purposes to assure harmonized labeling, packaging, nicotine content, etc. across the eu. however, the application of the health argument to put limitations on the internal market rules was also perceived as being negatively connoted by non-public health experts: "if you just go to the dg internal market and grab the first person you see and ask them what public health means, they will tell you it's the exception member states use to defend local weird monopolies on peculiar alcohol, or something like that. it's an exception to a rule." (# , academia) the recent eu patients' rights directive in cross-border healthcare [ ] was mainly regarded as an important achievement. this assessment was not necessarily driven by satisfaction with the scope of the directive but, instead, because it is the first eu secondary legislation ever enacted specifically on healthcare. "[…] the cross-border directive will turn out to be incredibly important. particularly because it is so symbolic important if you like because it does represent really the first time that the eu has got any concrete in relation to healthcare as opposed to public health. the consequences of this remains to be seen." (# , eu/national civil servant) "therefore, i see the patients' directive as a true success from a legislative perspective" (# , academia) the achievement aspect was supported by perceptions that the directive will lead to more cross-border cooperation and will have an impact on quality of care as well as on priority setting in healthcare and the packaging of healthcare services. thereby, it was expected that the directive will not only influence people who seek healthcare services in other countries but also those who seek services in their home country. in this regard, some expected that the directive would also ultimately empower patients as consumers of healthcare services. "the cross-border directive […] will have consequences of more consumer empowerment, consumer rights, patient rights, more consumer participation and more literacy,…"(# , public health advisor/advocate) however, there were also critical voices that interpreted the directive, as targeting a limited segment of the european population and hence, potentially increasing health inequalities. these respondents also questioned the willingness of the general population to seek healthcare treatment outside their home country. furthermore, respondents were critical of the extent to which more eu involvement in healthcare of member states would lead to quality assurance in general: "it is positive in the way people can be treated where they want, but it is still my point of view that we […] want to have our own level of quality and we don't want others to decide what level it should be. perhaps, because we have a very high quality […] . but of course we don't mind to tell others about it, we don't mind others to come in, we don't mind to help others to get the same standard -that is cooperation, so i always say i love cooperation but i do mind the harmonization.". (# , eu/national politician) this study provides an overview of public health relevant eu-level actions of the past twenty years. we outlined the diverse nature of expert perceptions on key developments in the field and provided a ranking of the most influential achievements. the assessment of outputs or actions being an achievement appeared across and within interviews along with assessments of outputs or actions being a missed opportunity and less often a failure. thereby, it turned out that the eu public health field has significantly developed its organizational structures (dg sanco, supranational agencies dealing with public health) and incorporated public health topics like infectious disease control and tobacco control, whereas the hiap approach still included untapped potential. this finding confirms "the challenge of implementation" [ ] of the hiap concept in the eu [ , ] . given the fact that according to article and article ( ),tfeu [ ], a high level of human health protection should be ensured within all eu policies and actions, it was seen as a weakness that the uptake of health consideration in the general eu policy-making process was low [ ] . ollila described the importance of communication and cooperation strategies for a successful realization of the hiap approach [ ] . the deficiency of these strategies was raised during the interviews which indicated that the performance of eu health players is perceived to be particularly poor in this regard. concordance of interview responses with tasks formulated in the health mandate of the eu interestingly, the study indicated that the treaty-based tasks such as support of cooperation between member states, development of guidelines and indicators, best practice exchange, and periodic monitoring and evaluation on eu-level public health to ensure 'a high level of human health protection' [ ] were only partially perceived as fulfilled or acknowledged by the interviewed experts. thematic discussions on actions or policies related to the development of guidelines and indicators appeared with regard to infectious disease surveillance and management of rare diseases but were not a major theme across interviews. the eu-level task to promote best practice exchange among member states was regarded as influential, which is represented by a top position in one of the rankings presented in this paper. with regard to the task of establishing monitoring and evaluation structures, some respondents perceived the status of the eu health information system rather as a failure. this corresponds to observations in the literature indicating that although ground work such as the development of a common eu health indicator set is acknowledged [ , ] further efforts are needed to implement and maintain health indicators [ ] and to develop a permanent and sustainable eu public health monitoring and reporting infrastructure that supports decision making in public health on eu level [ , ] . respondents agreed that cooperation in the area of public health between member state representatives and experts as well as with other stakeholder groups has increased and has been facilitated by the eu through various projects, networks, forums, and platforms. this trend was mainly positively perceived since it supported eu-level public health policy by accumulating and exchanging knowledge, generating public support and a legitimacy to act on certain fields [ ] . this finding is corroborated by the literature on the potential of new governance instruments for health-and social policy-making at eu-level [ , [ ] [ ] [ ] . however, these new governance instruments can also be regarded as a rather strategic investment of the ec to keep topics on the agenda until a political window of opportunity opens but as an ineffective policy tool to enforce and implement action in due course [ ] . the collaboration of a diverse set of stakeholders as it is the case for example in the eu platform for action on diet, physical activity and health can lead to actions that constitute rather a compromise of various interests. consequently, the results might be disappointing from the viewpoint of public health experts [ , ] . a final judgment on the impact of facilitating collaboration is to be awaited and may only be made in the long term future. it will require different ways of measuring 'impact' compared to the analysis of domestic adaptations when implementing eu hard law [ ] . the assessments of ec tasks for public health policy making have been influenced by characteristics like the subsidiarity principle throughout several interviews. on the one hand some participants were in favor of more eu influence on health policies and their implementation. in their view integration and harmonization of health policy did not reach far enough and hence their perception of actions was dominated by the category 'missed opportunity'. on the other hand some experts were in favor of keeping certain health issues like health care as national responsibility which led to a perception of too much eu involvement and a negative perception of the evolvement of the health mandate. public health has a cross-cutting nature and cooperation across dg's often poses difficulties. therefore, convincing evidence is required to demonstrate the health impact of policies outside the health domain and strong partnerships are needed to counter strong industrial lobbying groups [ , , ] . the ease of cooperation and the potential to achieve policy coherence between dg sanco and dgs with stronger regulatory competences like the internal market (e.g. regarding tobacco, pharmaceuticals) or agriculture policy (regarding food safety, subsidies of unhealthy versus healthy food products) represented another characteristic that influenced the individual perception of eu public health policies. experts who assessed the value of eu health policy actions under the reality of a rather weak health mandate were more likely to perceive eu actions as achievements. this was in contrast to others who strove for more appreciation of social and health matters in eu policies and who perceived a lot of missed opportunities or failures in this regard as the power of the eu was too weak to realize change and to fulfill the objective of the health mandate to ensure human health protection for citizens in the eu. in summary, underlying themes such as cooperation among european public health professionals, increasing institutionalization, and characteristics such as the issue of subsidiarity or the possibilities to cooperate across eu policy domains influenced experts' perceptions throughout the topics presented in this paper. these conditions and characteristics are part of what lamping called the "chaordic dynamics" of european integration in the field of health policies [ ] . as our study demonstrated eu health policy does not demonstrate a clear-cut success since the logic of action in the field can involve diverging interests. nevertheless, the eu public health has quite systematically developed in terms of scope and impact beyond the original mandate. the ranking of influential policy outputs provided indications on important developments in eu public health policy. however, even though we categorized the outputs, they sometimes differed in character and power which might have led to imbalanced judgments. additionally, we received different reasons for labeling eu-level actions or policies as achievements, missed opportunities or failure for public health. some were identified because they increased the strength or value of eu-level public health policy, whereas, others were identified because they impacted the health of the european population. the findings of the study may not be empirically generalizable since they were closely linked to qualitative individual perceptions and the settings that participants belonged to. however, we are confident that the broad range of profiles of the experts has ensured the diversity of perceptions on the topics varying from achievement to missed opportunity and failure. moreover, given that participants were generally active in health policy at eulevel and mainly positive about the eu, this could also have influenced the obtained results to some extent. eu public health policy is subject to divergent perceptions of how successful or unsuccessful specific topics have been tackled and how far european integration in public health policy should go. from the findings, it is unequivocal that the eu has strengthened its role over the past twenty years in supporting, coordinating, and supplementing member states' actions on public health issues as laid down in article ( ), tfeu. the eu is now a recognized player in public health in europe. however, when it comes "to the promotion of a high level of […] protection of human health […]in defining and implementing its policies and activities" (article , tfeu), further work is needed to achieve the full potential of the eu health mandate. endnote a also several eu member states disconnected on national level the ministry of health from social affairs. at the time of writing only seven out of eu member states organized health and social affairs within one ministry (spain, france, sweden, finland, estonia, greece, the netherlands). regulation (eec) / on the application of social security schemes to employed persons and their families moving within the community (accompanied by implementing ec launches the action programme maastricht treaty: the legal basis for undertaking actions in the field of public health is defined in article year eu health policy developments after the introduction of a legal eu health mandate by the maastricht treaty the european agency for the evaluation of medicinal products (emea), now european medicines agency (ema), has been formed in london. treaty of amsterdam: health impact assessment is implemented directorate general for health and consumers (dg sanco) is established lisbon agenda recognizes health protection as a prerequisite for economic growth measured with the indicator healthy life years the european food safety authority (efsa) has been established in parma first programme of community action in the field of public health the tobacco advertising directive / /ec is adopted after the first version has been annulled by the european court of justice commission decision to set up an executive agency for the public health programme. it has the task to manage community action in the field of public health the european centre for disease prevention and control (ecdc) in stockholm is operational white paper: together for health: a strategic approach for the eu decision for a second programme of community action in the field of health directive / /eu on the application of patients' rights in cross-border healthcare has been adopted. abbreviations aspher: association of schools for public health in the european region boviene spongiforme encefalopathie; dg: directorate general; dg connect: directorate general for communications networks, content and technology; dg sanco: directorate general for health and consumers emcdda: european monitoring centre for drugs and drug addiction; eu: european union; gats: general agreement on trade in services; heidi: health in europe: information and data interface; hia: health impact assessment; hiap: health in all policies; hta: health technology assessment; oecd: organization for economic cooperation and development; sars: severe acute respiratory syndrome; sdoh: social determinants of health; tfeu: treaty on the functioning of the european union; who-eur: world health organization-regional office for european union: the maastricht treaty. the treaty on the european union (teu). maastricht: european union european union and health policy: the "chaordic" dynamics of integration the impact of the eu law on health care systems eu law and the social character of health care health law and policy in the european union the politics of european union health politics health systems governance in europe -the role of european union law and policy health law and the european union health governance in europe -issues, challenges and theories european union public health policies -regional and global trends european policymaking on the tobacco advertising ban: the importance of escape routes patient's rights: a lost cause or missed opportunity? in health care and eu law european parliament and council of the european union: directive on the approximation of the laws, regulations and administrative provisions of the member states relating to the advertising and sponsorship of tobacco products ( / /ec) european parliament and council of the european union: directive on the approximation of the laws, regulations and administrative provisions of the member states concerning the manufacture, presentation and sale of tobacco products ( / /ec) european parliament and council of the european union: decision setting up a network for the epidemiological surveillance and control of communicable diseases in the community ( / /ec) european commission: commission decision / /ec on the early warning and response system for the prevention and control of communicable diseases under decision no / /ec european guidelines for quality assurance in colorectal cancer screening and diagnosis european guidelines for quality assurance in cervical cancer screening european guidelines for quality assurance in breast cancer screening and diagnosis eu regulatory agencies and health protection. in health systems governance in europe public health and risk assessment directorate: key achievements public health and risk assessment directorate: key achievements directorate general for health and consumers (dg sanco): health in the eu. what is in there for you? recent achievements cap on health? the impact of the eu common agricultural policy on public health. london: faculty of public health estimating the cardiovascular mortality burden attributable to the european common agricultural policy on dietary saturated fats regulating medicines in europe: the european medicines agency, marketing authorisation, transparency and pharmacovigilance is health recognized in the eu's policy process? an analysis of the european commission's impact assessments the european centre for disease prevention and control: hub or hollow core? j health polit policy law public health evaluation and impact assessment consortium: mid-term evaluation of the eu health strategy - -final report. bologna: public health evaluation and impact assessment consortium european court of auditors: the european union' s public health programme public health evaluation and impact assessment consortium: mid-term evaluation health programme ( - ) -final report. bologna: public health evaluation and impact assessment consortium how to peer review a qualitative manuscript qualitative data analysis: an expanded sourcebook beltz: weinheim three approaches to qualitative content analysis consensus methods for medical and health services research naturalistic inquiry european parliament, council of the european union: directive on the application of patients' rights in cross-border healthcare council of the european union: decision on adopting a programme of community action in the field of public health council of the european union: decision on establishing a second programme of community action in the field of health ( - ) ( / /ec) together for health: a strategic approach for the eu - (com( ) final). brussels: commission of the european communities european commission: commission communication on the framework for action in the field of public health (com( ) final). brussels: commission of the european communities council conclusions: towards modern, responsive and sustainable health systems ( /c / ). off j eur union european commission: communication on europe . a strategy for smart, sustainable and inclusive growth (com( ) final). brussels: european commission european union: communication from the commission to the council, the european parliament, the economic and social committee and the committee of the regions on the health strategy of the european community (com( ) final) european union: regulation of the european parliament and of the council. establishing a european centre for disease prevention and control the state of health in all policies (hiap) in the european union: potential and pitfalls health in all policies: from rhetoric to action european union health information infrastructure and policy. in european union public health policy regional and global trends public health indicators for the eu: the joint action for echim (european community health indicators & monitoring). archives of public health = archives belges de sante publique the weakness of strong policies and the strength of weak policies: law, experimentalist governance, and supporting coalitions in european union health care policy the hard politics of soft law: the case of health. in health systems governance in europe the open method of co-ordination in action: the european employment and social inclusion strategies collaboration and consultation: functional representation in eu stakeholder dialogues europeanization: new research agendas buchner b: nutrition, obesity and eu health policy a european alcohol strategy submit your next manuscript to biomed central and take full advantage of: • convenient online submission • thorough peer review • no space constraints or color figure charges • immediate publication on acceptance • inclusion in pubmed, cas, scopus and google scholar • research which is freely available for redistribution the paper was partly presented at a symposium held on th june in brussels, belgium, the european public health conference on th november in malta, and a conference on the th anniversary of the eu health mandate on nd may in maastricht, the netherlands. we would like to express our sincere thanks to the experts who invested their time and participated in the interviews. their views and perceptions on european public health policy were highly valued. we would like to thank wilco tilburgs and hassan el fartakh for their support in transcribing the interviews and ann borg for her support and helpful recommendations during the final editing process. we also appreciate the support of our colleagues at the department of international health at maastricht university; in particular kasia czabanowska, matt commers, kai michelsen, christoph aluttis and beatrice scholtes gave advice in setting up and designing the study, questioning the results, or reviewing the manuscript. the authors declare that they have no competing interests. all authors were involved in setting up the study. nr and tc coordinated the study. nr, tc and ks carried out the interviews, performed the analysis, and interpreted the results. nr drafted the manuscript. all authors revised the manuscript and approved the final version. key: cord- - syce n authors: domínguez-andrés, jorge; netea, mihai g. title: impact of historic migrations and evolutionary processes on human immunity date: - - journal: trends immunol doi: . /j.it. . . sha: doc_id: cord_uid: syce n the evolution of mankind has constantly been influenced by the pathogens encountered. the various populations of modern humans that ventured out of africa adapted to different environments and faced a large variety of infectious agents, resulting in local adaptations of the immune system for these populations. the functional variation of immune genes as a result of evolution is relevant in the responses against infection, as well as in the emergence of autoimmune and inflammatory diseases observed in modern populations. understanding how host–pathogen interactions have influenced the human immune system from an evolutionary perspective might contribute to unveiling the causes behind different immune-mediated disorders and promote the development of new strategies to detect and control such diseases. the evolution of mankind has constantly been influenced by the pathogens encountered. the various populations of modern humans that ventured out of africa adapted to different environments and faced a large variety of infectious agents, resulting in local adaptations of the immune system for these populations. the functional variation of immune genes as a result of evolution is relevant in the responses against infection, as well as in the emergence of autoimmune and inflammatory diseases observed in modern populations. understanding how host-pathogen interactions have influenced the human immune system from an evolutionary perspective might contribute to unveiling the causes behind different immune-mediated disorders and promote the development of new strategies to detect and control such diseases. infectious diseases are arguably the main source of evolutionary pressure that humanity has ever confronted. the dispersion of different human communities around the globe has exposed each population to different infectious agents, exerting a selective pressure (see glossary) on them; thus, adaptation to the new environment has favored the selection of the most beneficial genetic variants for the host. as a result, infectious agents have caused the expansion of alleles behind the induction of either protection or tolerance to these diseases; heritable variations, that increased the survival to deadly infectious agents, may have been naturally selected before the hosts had the opportunity to reproduce [ ] . natural selection driven by pathogens is probably more remarkable for those infectious agents that have been among us for a longer time, namely the causative agents of well-known diseases such as leprosy, smallpox, malaria, or tuberculosis. the genetic imprint of pathogen-driven selection depends on the length and the virulence of the infections and also the geographical distribution. the human genome presents more than genetic loci with traces of selective pressure [ ] . this group includes more than immune-related genes with functional variations between populations, which are probably behind the variability of responses to immune-related diseases reported nowadays [ , ] . besides natural selection, other evolutionary mechanisms, such as genetic drift, greatly influence the frequencies of the genetic variants found within diverse populations throughout the world [ ] (box and figure ). with the burst of next-generation sequencing and the development of cutting-edge technologies such as transcriptomics, proteomics, and systems biology, we are starting to witness the great impact of evolutionary processes on human immunity and how the interactions between microorganisms and humans that took place millennia ago might play a fundamental role not only in the response against modern pathogenic threats, but also in the emergence of autoimmune and inflammatory diseases observed in modern populations worldwide. in this review we offer a novel perspective on the role of infectious diseases as agents of natural selection and as forces behind the evolutionary pressure encountered by human ancestors and modern humans in their migrations around the globe. specific genetic variants selected throughout different periods of human history may have influenced immune responses of present-day populations against pathogenic microorganisms and may have played a role in the development of certain inflammatory and autoimmune diseases. the majority of experts agree that africa is where our species originated. genetic studies conducted in diverse contemporary populations suggest connections with ancestors that lived on the continent up to years ago [ ] . human evolution has been constantly influenced by pathogens; therefore, a great number of human genes linked to immune functions and immunity-related disorders have evolved along with humans. the heterogeneity in the immune response to infectious diseases across different populations is under genetic control and is the result of evolutionary processes. genetic variants that have been under evolutionary pressure can contribute to explaining the differences in the susceptibility to diseases observed across different populations. the ancestry of individuals from different populations across the globe greatly influences their possibility of developing certain autoimmune diseases and inflammatory disorders. the lifestyle of western societies affects the symbiotic relationships between humans, viruses, and other organisms and might contribute to the rise of certain autoimmune and inflammatory diseases. pathogens have played a central role as agents of natural selection from those very early days. among various infectious diseases, malaria has exerted the highest evolutionary pressure on the communities across the african continent ( figure ) [ ] . populations remaining in sub-saharan africa have been exposed to malaria for such long periods of time that their genetic structures have been shaped by the severity of malaria (plasmodium sp.) infections. in , allison described that sickle cell disease distribution was confined to africa and was associated with the geographical presence of malaria [ ] . this finding led to the more recent description of the existence of mutations in the hemoglobin-b (hbb) gene as a result of natural selection driven by evolutionary pressure for protection against malaria [ ] (table ) . similarly to hbb, some areas of west africa with a high incidence of malaria the gene variations that pass from one generation to the next are often transmitted as a random process known as genetic drift, while selection of advantageous variants tends to be preferentially transmitted. mutations, genetic drift, migration, and environmental selective pressure are among the fundamental processes behind the evolution of humans. the influence of these mechanisms in the diverse communities that were mobilized and then became isolated, as well as severe external factors such as epidemics, caused successive genetic bottlenecks in populations (see figure in main text) [ ] . human evolutionary studies are currently considered under 'modern synthesis', which merges darwin's ideas of natural selection with theoretical population genetics and mendelian principles, stating that evolution occurs via small genetic changes that are regulated by natural selection [ ] . these beneficial adaptations subsequently expand within the members of a population and become evident in the ancestral specificity and the geographical distribution of the advantageous alleles in the genomes of contemporary humans. genetic bottlenecks occur when the number of individuals in a population is reduced drastically due to a catastrophic event such as an earthquake, a flood, a famine, or the outbreak of an infectious disease. these events limit the genetic variation of a population and can lead to genetic drift. as a result, a smaller population, with a correspondingly reduced genetic diversity, remains to transmit genes to future generations through sexual reproduction. even if this reduction in the genetic diversity is temporary, it can lead to long-lasting effects on the genetic variation of the offspring populations. genetic drift: changes in the allele frequencies of a population over generations due to chance. genetic locus: fixed position on a chromosome (e.g., the position of a gene or a genetic marker). histocompatibility complex: region of approximately kb, located on human chromosome , that contains a large number of genes whose products are expressed as proteins on immune cells. of these genes, the best known are hla genes. introgressive hybridization: incorporation of genes from one species into the genetic reserves of another by interspecific hybridization and backcrossing with the parent species. present a high frequency of hemoglobin-c (hbc) in their populations, which is associated with a - % decrease in the possibility of developing the disease [ ] . this is also the case for the duffy antigen receptor gene (darc) in erythrocytes and single nucleotide polymorphisms (snps) in human leukocyte antigen (hla), which have been associated with protection against plasmodium vivax malaria in certain areas of africa where this disease is endemic [ , ] . another example of natural selection driven by evolutionary pressure for protection against malaria are thalassemia (a and b) pathologies, a group of hemoglobin disorders that presents an incidence of up to % among communities of west africa [ ] . the human casp t c snp, expression of which is restricted to the african subcontinent, south america, and certain areas of asia, can modulate immune and inflammatory responses to malaria by antagonizing interleukin (il)- b and nf-kb signaling in innate immune cells; moreover, caspase -deficient mice (casp -/-) exhibit decreased interferon (ifn)-g production and clearance of the parasite, relative to wild type (wt) infected mice [ ] . however, others have questioned these findings, given that caspase -deficient mice also lack caspase expression, so the effects observed might not be specific to caspase [ ] . mycobacterium tuberculosis (mtb) has caused infections in our species and ancestors for at least years [ ] . this long-standing relationship between humans and mtb probably underlies the large variety of immune-related factors that modulate susceptibility to mtb infection, including vitamin d receptor (vdr), natural resistance-associated macrophage protein (slc a ), tir domain containing adaptor protein (tirap), hla, monocyte chemoattractant protein (mcp- ), and cytokines such as il- and ifn-g [ ] ( table ) . patients with african ancestry present a higher frequency of mtb-related genetic variants than individuals from other populations, including variants in the gene encoding for toll-like receptor (tlr ), mediating cellular responses to bacterial malaria is one of the greatest causes of morbidity and mortality in the history of humanity. most human populations with a long history of endemic malaria have evolved genetic adaptations to malaria parasites due to the strong selective pressure that this infection has exerted. since the parasite infects erythrocytes, the evolutionary pressure has selected genetic variants that affect red blood cells and, therefore, the survival of the parasite as well. genetic variants conferring resistance to the disease have spread through human populations over time, including several abnormal hemoglobins that protect against malaria but usually cause erythrocyte-associated diseases in the populations where these adaptations are prevalent. these factors include the t c polymorphism in the caspase gene (casp ); the hemoglobin b (hbb) and hemoglobin c (hbc) variants; mutations in the duffy antigen receptor gene (darc); thalassemias (a and b); sickle cell disease; and polymorphisms in the human leukocyte antigen (hla) loci. dna nucleotide. for example, an snp may replace cytosine (c) with thymine (t) in a certain segment of dna. selective pressure: phenomenon that alters the behavior and fitness of living organisms within a given environment. it is the driving force of evolution and natural selection. thalassemias (a and b): inherited hemoglobinopathies characterized by a failure in the synthesis of the globin alpha or beta chains. toll-like receptors: family of transmembrane pattern recognition receptors expressed by immune and nonimmune cells that recognize conserved pathogenassociated molecular patterns. they play a pivotal role in innate immunity. transgenerational inheritance: transmission of traits from generation to generation. trends in immunology, december , vol. , no. lipoproteins [ , ] . selective pressure has also shaped the mechanisms that modulate the expression of genes implicated in immune responses against lassa virus, such as il- (il ) and the glycosyltransferase-like protein large (large), suggesting that the natural selection exerted by the virus drove the expansion of genetic variants that enhance immunity against lassa fever [ ] . these examples indicate that infectious diseases have contributed to shaping the genetic landscape of african populations and their descendants, and highlight the great impact of pathogens as an evolutionary force in humans. our homo sapiens ancestors were not the only species to venture out of africa, with other homo species performing a similar migration much earlier, such as homo ergaster, homo erectus, or homo heidelbergensis [ ] . from these early migrations, local populations such as the denisovans and neanderthals evolved [ ] . these lineages were not geographically isolated, but lived side by side with modern humans and interbred with them, leaving a genetic footprint in their common progeny. accordingly, - % of the genome of european and asian populations is thought to derive from these now-extinct hominid lineages [ ] . neanderthals spent close to years adapting to their environment and their immune systems were shaped by the infections they faced. by interbreeding with archaic humans, modern humans incorporated these advantageous adaptations in the genome of their descendants. this was highlighted by different studies that showed that the introgression of diverse genes related to immune functions, such as the oas cluster, tlr , or the histocompatibility complex from denisovans and neanderthals shaped the genetic landscape of present-day eurasian, but not african, communities. genomic sequences and expression data from lymphoblastoid cell lines from individuals of european and african ancestry confirmed that the tlr -tlr -tlr genetic loci, presenting signs of local positive selection and repeated introgression from both neanderthal and denisovan genomes [ ] [ ] [ ] , showed a significantly higher expression in individuals carrying archaic-like alleles than in individuals carrying the nonintrogressed modern human alleles [ , , ] . the expression of these genes has shaped human immune responses against different types of pathogens. for example, the gp protein of the hiv- virus has been recently recognized as a tlr ligand [ ] . in this regard, increased tlr expression has been correlated with higher il- production by the macrophage cell line thp- and higher titers of hiv- in the breast milk of hiv- -infected nigerian women relative to controls [ ] . tlr and tlr form dimers with tlr , triggering immune responses against different types of bacteria, fungi, virus, and parasites [ ] . variation in tlr -tlr -tlr is the major genetic determinant of human interindividual differences in tlr / -mediated responses, including cytokine production to a number of clinically relevant pathogens such as staphylococcus aureus and listeria monocytogenes [ ] (table ). this inheritance from archaic humans may have also left some human individuals more prone than others to developing asthma, hay fever, and other allergies (of snps associated with susceptibility to allergic disease, had a neanderthal or denisovan origin) [ ] , although these associations remain to be fully demonstrated [ ] . these reports demonstrate that by interbreeding with archaic humans, modern humans incorporated a group of advantageous adaptations to the genome of their descendants and contributed to shaping immune responses in modern human populations. the migration of our human ancestors out of africa implied the exposure to different types of infectious diseases (box and figure ). one study tested the responsiveness of human macrophages to pathogenic bacteria in vitro, finding that almost % of the genes present in human macrophages infected with the bacteria salmonella typhimurium or l. monocytogenes present different regulatory responses directly linked to the lineage of the donors and, also, that macrophages obtained from individuals of african origin display enhanced bactericidal activity compared with those from individuals of european lineage [ ] . the trend towards lower inflammatory responses in european populations is strengthened by the fixation of a tlr gene variant that results in lower proinflammatory gene expression in populations with a european ancestry compared with those with an african one [ ] . the largest population differences in gene expression between africans and europeans have been found in the macrophage receptor with collagenous structure (marco), a protein implicated in responses against viral infections and tlr-induced dendritic cell activation [ ] , the chemokine receptor cx cr , which mediates effector lymphocyte functions, and also several ifn-stimulated genes [ ] . west eurasian populations present a high frequency of tirap ser leu snps [ ] . tirap is an adaptor protein in tlr and tlr signaling pathways, involved in inflammatory responses and cytokine production. the heterozygous expression of the ser leu snp is protective against invasive pneumococcal disease, bacteremia, malaria, and tuberculosis, as shown in a case-control study of individuals from the uk, vietnam, and several african countries, and it is associated with lower tlr signaling in humans [ ] . this variant is considered to be a consequence of the natural selection that may have taken place in an early period following the migration of modern humans out of africa [ ] . european populations present a selective adaptation of the ifn gene that allows a high production of ifn-g in infectious scenarios due to the positive selection of ifng variants + g and + t; this suggests the existence of strong environmental pressures linked to higher ifn-g concentrations in plasma during mtb infection in european individuals relative to other populations [ , ] . in line with this, a database meta-analysis showed that individuals expressing the + t/a variant of ifng presented higher susceptibility to tuberculosis mtb infection than individuals without it, which might be considered a putative prognostic factor for the development of tuberculosis [ ] , although this remains to be robustly demonstrated. one of the most interesting aspects of humans is their ability to adapt to almost every ecosystem of the planet. the history of mankind is also the history of millions of individuals wandering around the world, looking for a better place to live. a glimpse to the migratory legacy of humanity around the globe reveals the great impact that the massive population movements defined the world as we know it today (see figure in main text). the distances ancient humans travelled are impressive, from the first hominids colonizing africa to the conquest of the americas in a time when the bering strait was not yet under water. the historical exodus of mankind started almost million years ago with the migration of homo erectus from africa through eurasia. from this event on, relatively isolated human populations evolved separately on different continents, leading to the emergence of different human species, such as neanderthals in europe, the denisovans in asia, and, later, modern homo sapiens in africa [ , ] . h. sapiens first colonized large areas of the continent around years ago [ ] , spread towards the middle east at some point between and years ago, and migrated through eurasia, reaching australia within years [ ] . asian human ancestors went through the frozen waters of the bering strait in two distinct waves to colonize the american continent approximately years before the present time [ ] . when humans ventured out of africa, they faced different types of pathogens than the communities that stayed in the african continent. with time, the series of events faced by diverse populations has generated differences in the immune responses to pathogens in the populations with an african or eurasian origin and which have spread throughout the world. the indigenous populations of south america are descendants of migrating populations of north-east asians that crossed the bering strait around years ago [ ] . five centuries ago, european settlers disembarked on the american continent, bringing a large collection of pathogens such as those causing measles, pneumonic plague, and influenza infections, which the indigenous populations had never faced before. these diseases rapidly spread and caused mortality rates above % [ ] . the consequences of these pandemics are still visible in current populations; one report studied dna from the bones of ancient humans from the tsimshian community, living in the british columbia region in canada until the th century, identifying marks of positive selection in a number of immune-related variants [ ] . specifically, the hla-dqa variant was present in almost the totality of tsimshian individuals, but only in one third of present-day humans studied; this suggested that ancient american genomes were evolutionarily selected to respond to local diseases but not to fight against pathogens brought by the europeans [ ] . another study compiled information on infectious diseases that have killed more than individuals among indigenous communities of the amazonia in the past two centuries, showing that the mortality rates and the incidence of infectious diseases rapidly decayed within the time following the first encounter with the pathogen, compatible with genetic adaptation [ ] . european colonizers underwent purifying selection in situations of intense pressure. such scenarios were documented when dutch colonists migrated to surinam and encountered epidemics of yellow and typhoid fever that caused a % mortality rate among settlers [ ] . variations in the frequencies of c , glo, and hla-b genes among the descendants were not likely caused by genetic drift, but rather, it has been proposed that these populations were probably selected through genetic control of survival to the epidemics [ ] (box ). africans and americans with an african ancestry present a much higher number of genetic variants related to robust inflammatory reactions, increased cytokine secretion, and bactericidal activities compared with the other populations [ ] , including more than genes with traces of recent selection, such as il a and il b gene variants [ ] . the degree of african ancestry, analyzed by fine-mapping analysis refined to the duffy-null allele of rs , was correlated with an increased amount of the proinflammatory chemokines ccl and ccl in plasma relative to controls [ ] . a study involving african american and hispanic american women found that the higher values of c-reactive protein (crp) in blood found in these populations compared with european americans were related to a crp-associated variant of triggering receptors expressed by myeloid cells (trem ) [ ] . moreover, comparison of health record data from individuals with connective tissue diseases, including rheumatoid arthritis and systemic lupus erythematosus (sle), as well as atherosclerotic cardiovascular disease from almost african american and european american adults was conducted; the study reported for the latter, a prevalence of atherosclerotic cardiovascular disease in . % african americans (particularly high in young individuals), relative to . % in european americans [ ] . these studies highlight certain genetic links to inflammatory predisposition/manifestation. however, increased proinflammatory activity is a double-edged sword. in the absence of regular pathogen challenges that require maintained modulation of the balance between inflammation and suppression of the immune response, the organism can overreact to inflammatory stimuli and trigger exacerbated responses. for instance, descendants of african populations generally present higher susceptibility to a variety of autoimmune syndromes such as inflammation-associated carcinomas, lupus, asthma, and multiple sclerosis (ms), the overall prevalence of which is up to three times higher in individuals with african ancestry relative to individuals with european ancestry [ , , ] . there are extensive differences in immune cell gene expression between americans with african and european ancestry. the increased proinflammatory responses observed in american individuals relative to other populations might be beneficial to combatting infections, but might also increase the chances of developing inflammatory and autoimmune disorders, which warrants further investigation. our gastrointestinal tract provides residence to both beneficial and potentially pathogenic microorganisms, harboring ten million different microbial genes in the human fecal microbiome [ ] . the microbiome has its own evolutionary scenario across different populations with divergent lifestyles, nutrition, and exposure to environmental agents, generating extraordinary heterogeneity. the ongoing process of 'lifestyle westernization' of different societies has an important impact on the mutualistic relationships between humans and commensal organisms worldwide. african tribes are adopting western subsistence patterns, leading to remarkable changes in the composition of their microbiota [ ] . the comparison of the intestinal flora of the baaka hunter-gatherers and the bantu agriculturalists (both from the central africa republic), with a group of us-born african americans showed a great example of the evolution of the human microbiome [ ] . specifically, the bantu, still engaged in hunting, have a greater bacterial gut diversity than their baaka neighbors, who left the jungle for agriculture, and even more than urbanized westerners (us african-americans) [ ] . this reduced microbiota diversity in western societies has been associated with a higher incidence of the so-called 'diseases of civilization' such as cardiovascular diseases, diabetes, obesity, and autoimmune disorders, which are very unusual in hunter-gatherer societies compared with communities living a western-type lifestyle [ , ] . although viruses are mainly seen as pathogenic agents, they also play a fundamental role in the evolution and maturation of the human immune system [ , ] . approximately % of the human genome is composed of endogenous retroviruses (ervs), sequences derived from past retroviral infections and permanently inserted into different regions of the human genome [ ] . one study showed that ervs played a central role in the induction of ifn-dependent immune responses and that the removal of one or more of these viral dna elements in the hela human cell line severely impaired the recruitment of transcription factors necessary to trigger the expression of ifng against vaccinia virus infection relative to controls [ ] . viruses can also influence the severity of infections caused by other viruses. for example, cytomegalovirus infection in hiv- seropositive humans can potentiate the effects of hiv- infection by expanding the pool of circulating regulatory t cells (immunosuppressive); these were shown to inhibit the proliferation of autologous peripheral blood mononuclear cell the origins of the hiv virus are still a matter of scientific discussion. the most accepted scenario argues that hiv originated in simians and was transmitted to humans in west africa in the s, likely due to local ingestion of ape meat infected with the simian immunodeficiency virus. around , the virus reached wide parts of the continent and finally spread overseas thanks to a group of haitian professors coming back from africa. in the following decades, the virus spread worldwide and generated the pandemic we now know. today, there are approximately million people worldwide living with hiv- /aids [ ] . ccr is a receptor of chemokines that plays a fundamental role in hiv- pathogenesis and it is also one of the most promising targets to restrict the infection, since mutations in this receptor turn individuals resistant [ ] . the ccr -d mutation results in a deletion that eliminates the hiv- co-receptor on lymphocytes, providing robust protection against hiv- and, therefore, aids [ ] . ccr -d allele frequencies reach % in northern europe populations, whereas it is not present in populations with different ancestry, such as east asian, native american, or african groups [ ] . this regional distribution of ccr -d variants is most likely related to a naturally selective episode that struck european populations around years ago and involved a strong infectious agent that also employed ccr [ ]. a mathematical model studying the changes in the european populations in the middle ages suggested yersinia pestis (bubonic plague) as the most probable infectious agent behind the pressure that selected this particular genetic variant [ ] . this is in agreement with the finding that european rroma populations, but not northwestern indian populations that inhabit the area where the rroma originally lived, present signatures of positive selection in tlr -tlr -tlr , which influence cytokine responses in y. pestis infections [ ] . (pbmc) in response to cytomegalovirus infection in vitro [ ] . in one study, patients with chronic hepatitis c virus (hcv) infection and hepatitis a virus (hav) superinfection presented lower titers of hcv rna than patients harboring only hcv, suggesting that hcv replication might be potentially suppressed during hav infection [ ] , although this will still require further investigation. the relationships between humans and pathogenic or nonpathogenic organisms are extraordinarily complex and include tripartite evolutionary interactions between humans and microbes competing with each other. this is the case of parasites that infect other parasites, such as bacteriophage viruses, that can influence the outcome of bacterial infections. for example, in a cohort of individuals with chronic wounds, a report showed that the phage pf, which coexists with pseudomonas aeruginosa in infected wounds, triggered the production of type i ifn, the inhibition of tumor necrosis factor (tnf) production, and the suppression of phagocytosis in human primary monocytes and mouse bone marrow-derived macrophages, dampening the antibacterial response and promoting the bacterial infection [ ] . however, bacteriophages can also provide protection to the human host by directly attacking pathogenic bacteria and by upregulating in human pbmcs the expression of proinflammatory genes such as il a, il b, il , tnfa, cxcl , and cxcl , as shown for several s. aureus and p. aeruginosa phages, including pnm, luz , - , and ge-vb_pae-kakheti [ ] . cooperative relationships between organisms are evolutionary processes themselves. the way microorganisms and their hosts associate can lead to interactions of mutualism, in which the interplay may be so intimate as to provide benefit for each party and influence immune responses against different types of pathogens. a great number of humans live far away from the original settlements of their ancestors and are subject to radically different environmental conditions. between two and three million people with european genealogy suffer from autoimmune diseases, the prevalence of which is also increasing in other populations across the globe [ ]. there is rising evidence that the emergence of autoimmune diseases is associated with the presence of a number of immune-related alleles that have been selected via evolutionary processes; and, furthermore, that the contrasting differences in the prevalence of autoimmune diseases between populations may be a result of different selective pressures [ ] . alleles associated with inflammatory diseases that present marks of modern positive selection include the risk allele fut at rs for crohn's disease (cd) or the risk variant sh b at rs for celiac disease [ ] ; such variants have been linked to the development of several human autoimmune diseases, such as type diabetes, ms, and celiac disease [ , ] . the analysis of the integrated haplotype score [ ] of loci associated with sle that might provide protection against infections, such as tnip , itgam, ptpn , tnfsf , uhrf bp , tet -dguok, and blk, has suggested that these loci exhibit robust signs of positive selection [ ] (figure , key figure and table ). african and asian human populations exposed to trypanosoma brucei or plasmodium sp. have presented positive selection of snps in the apol and fcgr b genes; indeed, by enhancing human macrophage-mediated phagocytosis of infected erythrocytes, despite their association with an sle predisposition, these snps have been associated with protective roles against sleeping sickness and malaria, respectively [ , ] . an analysis of human loci associated with inflammatory bowel disease (ibd) concluded that the majority of the loci associated with cd are also linked with a higher risk of developing ulcerative colitis [ ] ( table ). many of these loci were also associated with the development of other autoimmune diseases, namely psoriasis and ankylosing spondylitis [ ] . pbmcs from individuals carrying the sh b variant rs *a (associated with a risk for developing cd) [ ] presented higher production of il- and il- b after stimulation with lipopolysaccharide or muramyl dipeptide due to enhanced activity of the nod inflammasome pathway relative to controls [ ] ; this has suggested an immune-related role for sh b in the context of bacterial infections, which might help explain the positive selection of sh b approximately years ago [ ] . others found an association between genetic variants in nod , cd , and increased susceptibility to cd [ ] , in line with previous results showing that mutations in nod and tlr /cd are related to an increased risk of developing ibd [ ] . from another angle, changes in hygiene patterns seen in the past two centuries brought vast improvements in sanitation, drinking water, and garbage collection, which greatly reduced the exposure to many infectious diseases. however, these conditions may have reduced the exposure to viral and microbial agents that help the immune system to develop tolerance during childhood. the hygiene hypothesis proposes that the lack of exposure to microbial agents in the early stages of life is related to a higher risk of developing hypersensitivity reactions, based on the fact that children that are exposed to higher amounts of microbial stimuli (e.g., by growing on farms) are less prone to develop allergies and asthma [ , ] . moreover, reduced exposure to infectious agents can have a much wider effect than initially believed. lack of exposure to microbes in childhood can cause aberrant responses to infection and potentiate the effects of etv -runx mutations in the pathogenesis of acute lymphoblastic leukemia [ ] . by contrast, a meta-analysis of six observational studies, including participants, showed a correlation between low helicobacter pylori infection and ms, suggesting that low h. pylori prevalence might be a putative protective factor in ms, although this remains to be experimentally validated [ ] . one study also reported that antibodies against toxoplasma gondii were detected less often in patients with ms compared with healthy controls [ ] . however, these findings warrant further and robust investigation. overall, it is clear that evolutionary processes can drive the fixation of genetic variations that increase (or decrease) our defense against infections upon sensing microbial ligands, but can also lead to a greater risk of developing certain autoimmune diseases in which endogenous ligands can cause tissue damage and inflammation. a growing number of reports suggest that inheritance is not always governed by classical darwinian evolutionary processes. exposure to certain environmental stimuli can cause effects in the progeny of an exposed individual, even though the stimuli are no longer present. this type of transgenerational inheritance might be explained through the effects of epigenetic processes, which are hypothesized to be transmitted through the germline and passed on to the offspring [ ] . for example, the worm caenorhabditis elegans can transmit improved resistance to infections to pathogenic bacteria to their offspring through alterations of the histone landscape [ ] . indian meal moths exposed to low doses of virus are subsequently less susceptible to viral challenge, a protection offered to their offspring as well [ ] . transgenerational inheritance of diverse traits has also been observed in mice, in which the variation of the color of the coat is passed on the next generation [ , ] . offspring of male rats subjected to a high-fat diet present glucose intolerance and reduced insulin secretion, linked to reduced methylation at the il ra gene relative to controls [ ] ; and mice fed scorpions are more resistant to a challenge with scorpion venom than mice on a normal diet [ ] . since infections are one of the strongest factors impacting survival, it is conceivable that transgenerational transmission of traits in mammals, including humans, evolved to improve host defense. the number of studies of the potential role of epigenetic inheritance in shaping the human immune system is still scarce. however, different experiences undergone by certain communities indicate that these mechanisms might be important. for example, the babies of pregnant women who suffered during the early stages of pregnancy (the effects of the dutch hunger winter in ), years later showed reduced dna methylation marks in several genes that control metabolism and cell differentiation during development, such as igf , pim , txnip, abcg , pfkfb , and mettl , compared with their siblings [ , ] . this was related with higher rates of obesity, heart disease, cancer, and depression in individuals whose pregnant mothers suffered the effects of the famine [ , ] . some of these effects seemed to be present in the progeny of this group, that is, in the grandchildren of those who had passed the famine during pregnancy [ ] . the rapid growth in the number of reports covering the impact of epigenetic mechanisms in different human processes warrants further and robust studies on the impact of epigenetic inheritance in shaping the evolution of the human immune system. human immune responses have been shaped by the evolutionary pressure exerted by microorganisms and viruses throughout history. generating a broad range of genetic variations and immune functions in different populations favors the adaptation to new environments and increases the chance of survival of the human species against potential pandemics. much remains to be learned in this exciting field over the coming years in order to identify the main regulatory forces and the time window necessary for the fixation of an advantageous genetic trait in a population (see outstanding questions). the combination of the selective pressure caused by infectious diseases with other evolutionarily relevant processes, such as genetic drift, migratory events, bottlenecks, and introgressive hybridization, contribute to driving the expansion, fixation, or elimination of characteristic immune response-related traits in different populations around the world. these specific genetic variants are able to boost the host response against pathogens by improving the sensing of microbial ligands but can also lead to the development of autoimmune diseases, in which the immune system responds to endogenous ligands and induces abnormal responses targeted against the host's own tissues. of note, it is very difficult to assign certain variants, a specific role in the protection or induction of autoimmunity. to assign changes in the genetic landscape of human populations to certain diseases is an extraordinary challenge. moreover, our species is in constant evolutionary interaction with various microorganisms and viruses. populations of bacteria and their viruses (phages) undergo, under natural conditions, reciprocal evolution in terms of resistance and infection; this, in turn, also affects the evolutionary traits of our immune system. thus, an extraordinarily complex scenario exists in which organisms of different phyla interact, compete, and coevolve, to ensure their own survival. as novel tools, the development and refinement of methods that study large sections of the human genome, epigenome, and microbiota, will help to obtain genome-wide data in diverse human populations, allowing us to follow the evolutionary trails left from the encounters with different organisms, further unveiling the roots of human immunity. high-throughput biotechnology and an expanding computational capacity can enable the study of global population genomics and might contribute to decoding the origin and consequences of functional changes in adaptive alleles down to the single cell level. however, these methods also have limitations associated with the difficulty in linking gene variations to clinical phenotypes and disease, the generation of false positives, or the high number of samples necessary to reach reliable conclusions. expanding the heterogeneity of populations studied for immune gene association studies relevant to disease will be key, as generally, a large focus is placed on certain european or american communities, thus generating results that are difficult to extrapolate to other populations. the knowledge of the evolutionary and genetic basis of human immune traits and their impact on diverse pathologies (e.g., autoimmunity, infections, inflammatory diseases, cancer) increasingly suggests that the genetic basis of disease may be derived from a large number of rare variants of modest effect. the mechanisms described here acquire special importance in the current scenario of world globalization, in which the migration fluxes and the admixture of different populations are reaching unprecedented levels, allowing faster expansion of advantageous alleles. however, these processes may also accelerate the spread of new epidemics, as seen in the cases of hiv infection, or more recently, sars-cov, ebola, and chikungunya viruses; as well as the emergence of multiresistant bacteria and fungi, such as methicillin-resistant s. aureus or candida auris. this is just the starting point to unveil the evolutionary history of the relationships between pathogens, the immune system, and humans. further investigation of the functional adaptations of human populations is warranted to provide a broad picture of the functional consequences of evolution in human immunity. acknowledgments m.g.n. is supported by a spinoza grant of the netherlands organization for scientific research and an erc advanced grant (# ). the funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication. we thank laszlo a. groh for which are the strongest evolutionary forces that drive the evolution of the human innate immune system? how long does the genome of a given population take to adapt to a new infectious threat? are the mechanisms of resistance to infection transmitted only via genetic modifications, or can epigenetic adaptations to resistance also be transmitted to the progeny, and under what circumstances? the development of single-cell sequencing technologies has opened new fields of study. how does genetic variation of the expression of a specific variant vary between different cell subsets and how does it affect the overall phenotype of an individual within a population? are there specific immune processes that are preferentially impacted by evolutionary pressures? in western societies we enjoy a life expectancy vastly superior to that of our predecessors, but at the same time, we suffer diseases that they did not suffer. can some of the reasons for these changes lie among some of those bacteria that we have somehow lost in our microbiome? identifying these bacteria and understanding their effects on the human body might be the first step to developing putative therapies based on bacterial restoration. since many of the variants causing autoimmune diseases are linked to an enhanced responsiveness to pathogens that is no longer needed in developed countries, could these genes and their related pathways be employed as targets for new putative therapeutic approaches against inflammatory/ autoimmune syndromes? are other newly described genetic regulatory pathways, such as interfering rna or long noncoding rnas, also influenced by pathogen-driven evolutionary processes in different populations globally? proofreading and correction of the manuscript. we additionally thank srinivas agra for providing the figure icon 'bacteria' as deposited on https://thenounproject.com. natural selection and infectious disease in human populations genomic signatures of selective pressures and introgression from archaic hominins at human innate immunity genes genome-wide identification of regulatory sequences undergoing accelerated evolution in the human genome divergent selection of 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severe malaria full-exon resequencing reveals toll-like receptor variants contribute to human susceptibility to tuberculosis disease multiple common variants for celiac disease influencing immune gene expression epitope selection for hla-dq presentation: implications for celiac disease and viral defense card /nod mutational analysis and genotype-phenotype correlation in patients with inflammatory bowel disease association of nod leucine-rich repeat variants with susceptibility to crohn's disease nod and nod : beyond peptidoglycan sensing global, regional and countrylevel - - estimates for : assessing progress towards the targe key: cord- -cli mpev authors: schaller, karl title: neurosurgeons in the corona crisis: striving for remedy and redemption. a message from the president of the eans date: - - journal: acta neurochir (wien) doi: . /s - - - sha: doc_id: cord_uid: cli mpev nan it is difficult these days to keep a clear and unbiased mind: real experts, self-declared experts, and politicians are issuing apodictic statements and restrictive orders with a big impact on our daily lives. many of those statements are contradictory, and the various countries are reacting in equally different ways. even within the same country many different public directives may be put in place by the regional governments, due to their federal political organization (i.e. germany). as a result, i happen to know that in some neurosurgical departments not a single or had been shut down, and not a single covid- -positiv patient had been admitted to their icu, whereas in my own, for example, we are not allowed to do elective surgeries anymore, and more than icu beds are taken by intubated and ventilated corona-patients already (as of march , ). in other countries with more centralized health care, specialists proposed some sort of a herd immunization of the whole population (i.e. netherlands). but, with increasing pressure through one or another interest group and/ or so-called public opinion, such major decisions may be revised from one day to the other (also netherlands). inagain other countries (for example the usa), the danger related to that virus has been denied to a large extent ("hoax by the democrats"). now, from one day to the other, and a °-turn later, entire us states are preparing for lockdown, and large navy-ship-based mobile hospital units are being moved around and prepared to anchor along their coastlines. from other mainly eastern countries (russia, kazakhstan etc.,) we don't hear anything through our usual news channels. furthermore, it seems that the profound and special european spirit of a common departure, which reigned in the beginning of the s is being jeopardized by specific political interests, and by the lack of solidarity across european borders: supply chains for surgical masks are interrupted for the reason of national egoisms. tools, and mechanical or electronic components, which are desperately needed to build for example ventilators are no longer delivered straight from one country to the other. all together that will be put on a very big bill, once all will be over, and it will take a long time to reinstall confidence in our political leadership, in our idea of a europe without limits in humanity, as a model for the world, and as the place to be. there is a ghostly feeling when looking outside. it is as if somehow we have become figures inside real-world edward hopper paintings. all that leaves the strange feeling behind that something big may be going on. but there is no clear and consistent answer, neither from the experts, nor from political leadership, as to how to address this exceptional situation. important decisions may become triggered by public opinion, and thus the overall situation may entirely spin out of evidence-based control. it seems that the hour of countless experts, of epidemiologists and lab-scientists has arrived. the hour of those who were hidden behind large screens and next to super-computers for big data management and pcr analysis, in remote corners and basements of our hospitals and faculty buildings, and that these people seem to have been prepared for their moment to arrive. that's what is happening now: we, who are used to treating patients on an individual basis have become subordinates of crisis units and eminent telegenic virologists. that may the right thing for the time being, if there was coherent advice. but i wish these experts would be more consistent in their analysis of the situation and in their recommendations, because even for us as doctors it is difficult to understand what's right and what's wrong. how should we expect the public to understand such a situation and to behave accordingly, if we don't know ourselves what's going on based on facts? having tried to drink a beer with one of my colleagues at the lakeside this week, we were driven away by patrolling police. that made me recall dystopic scenarios from stories and movies of the past, depicting totalitarian states, where entirely new professions were generated, such as stalkers, who used to guide people and take them around to forbidden zones [ ] (https://www. youtube.com/watch?v=xb jvtut -g). the current situation is also unmasking in a painful way that there is also an important economic inequality across all european countries. there is a significant lack of doctors and of nursing staff, and in terms of access to icu-beds with ventilators in some countries, whereas in others, the situation seems to be less precarious. for example, how is it that in the uk, home to some of the finest educational and research institutions and to a good proportion of the richest people in the world, only approximately icu beds exist -for a population of approximately million people? other countries on the other hand, were deprived of their medical staff due to more precarious general economic situations, with doctors and nurses following the westward stream toward politically more transparent and wealthier countries. medical education is expensive, and there is global competition for well-trained and mobile doctors and nurses. poor political planning and cultural differences from an all-inclusive state responsibility to more social darwinist principles may be the reason, and reflect the different approaches across european countries. in times of crisis, nobody wants to be left behind however, and that's why we are hearing so many outcries from various corners for more social fairness at this time. undoubtedly, there is a clear need for guidance and clarity. it is clear, that a day will come when we will understand what happened. that may become redemption day for some, and it may become pay day for others; beyond the giant socioeconomic impact of this crisis, there will be an accounting of medical and surgical collateral damage, which has been caused by the currently imposed restrictions and postponing of otherwise indicated medical and neuro/surgical care. in our field alone thousands of patients, with degenerative spinal disease, low grade gliomas, cavernomas, epilepsy and other pathologies are put on ever-increasing waiting lists. meanwhile, mild paresis may become moderate, tumors may grow to more complexity, cavernomas may bleed, and repeated seizures may have deleterious consequences. all that will happen, and it will have to be accounted for, when there will come a time to do so, and after the whole mountain of delayed surgeries will be worked up. truth will come, and it won't be comfortable for those, who may be riding on hightide right now. this unclarity, radiating from the highest levels of political leadership and from contradictory expert panels, this lack of consistent leadership continues to induce a feeling of insecurity among people. seeing photos of old people, who were overrun by hysterical crowds during the opening of supermarket doors, or of overworked and tired nurses, who find themselves in front of emptied grocery shelves, and of strong men defending the towers of toilet paper, which they had just bought; all these images, that materialized prove that the taint of civilization is so thin that it's being blown away after only a couple of complicated weeks, appalls me and it renders it difficult not to lose faith in our species. and it confirms my long-standing notion that sometimes the seemingly biggest and strongest among us seem to have the least compassion in those moments, when one should stay calm and cool and caring. then, i have to focus my mind on the enlightening idea that not all people are like that, that the majority isn't in fact. and that's why we have to continue to strive to deliver the best possible service to our patients and to have confidence in our species. but how will things develop if the situation is not resolved in a few weeks' time? the current image of humanity leaves a stale feeling. and to relaunch a system with such inertia after weeks and months of deprivation, if many companies no longer exist, will take tremendous effort? doctors will just go back to work, as usual. but to reboot whole economies and have a labour force with less pragmatic backgrounds returning to their jobs may be less evident. all of us are somewhat trapped in our own inertial systems. we have to adapt to the current state-of-mind and to our regional rules and play the game. it is up to us to stay calm and to transmit an anti-hysterical mindset to our peers, be them professional, nurses, secretaries, cleaners, it specialists, and all kind of intermediate hospital staff, or patients and the general public. we have to live up to the current situation, whether the whole hype is justified or not. we have to provide an example to show how to stand together, across all national borders, and with the optimism of pragmatic and generous people, who we should always strive to be. that's why most of us became neurosurgeons in first place. that spirit was generated by european neurosurgical leaders many years ago, not long after the european carnage of world war ii. that spirit has formed the foundations of the eans. even during the iciest times of the cold war, european neurosurgeons were connected without boundaries. the first european training courses started in the s, and the course sites are illustrative for the visionary european thinking of our predecessors. i refer you to our website to see that list and encourage you to upload your own material related to these events, should you happen to have something of interest. otherwise, be referred to the photography books of john garfield, who documented that particular spirit from the beginning with his camera. eans wants to provide a trans-national and inspiring base for us neurosurgeons. we would like to improve our common platforms for the exchange of thoughts, and we shall stand in for each other. let's stay in contact more intensively via social media, telephone calls or video conferences, respectively. i would like to see more networking and information passed on through our own, eans-based communication channels. we have installed an easy-to-use format to upload your comments, your thoughts, data, and scientific material related to the corona crisis now. all that may serve us some day to understand better what's happening currently, and to analyze potential collateral damage associated with it, and to be prepared better should such a situation recur in the future. here, the links to our website: https://www.eans.org/default.aspx https://www.eans.org/news/ /eans-presidents-message-on-corona-virus-feedback.htm please feel encouraged to contact the eans team at any time. in every crisis lies the seed of opportunity. i am sure we will grow from that, and we shall meet and celebrate under liberating circumstances in the near future. and, should it turn out that chloroquine may contribute to ameliorate the course of corona-disease indeed, then be referred to john garfield again [ , ] . neurosurgeons have always been frontrunners, when it came to visionary treatments. take good care of yourself, of those next to you, and of your patients. we will retake the drivers' seats in our natural habitat, in hospitals and in neurosciences, and we shall meet soon again, hopefully in belgrade in october ! with my warmest regards, karl schaller. but a neurosurgeon: after the third gin and tonic roadside picnic. . sf masterworks publisher's note springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations key: cord- -o zspgrk authors: ippolito, g.; fusco, f.m.; caro, a. di; nisii, c.; pompa, m.g.; thinus, g.; pletschette, m.; capobianchi, m.r. title: facing the threat of highly infectious diseases in europe: the need for a networking approach date: - - journal: clin microbiol infect doi: . /j. - . . .x sha: doc_id: cord_uid: o zspgrk in recent years emerging and re-emerging infections, as well as the risk of bioterrorist events, have attracted increasing attention from health authorities because of the epidemic potential that renders some of them a real public health challenge. these highly infectious diseases (hids) are occurring more and more frequently in europe, and despite the many initiatives in place to face them, many unsolved problems remain, and coordinated efforts for dealing with hids appear mandatory. whereas uncoordinated measures would lead to only partial and poor responses to these emerging threats, networking represents a valuable approach to these diseases, in order to: (i) ensure a rapid and effective response; (ii) stimulate complementarity and prevent duplication; (iii) promote international cooperation, exchange of experience, good practice and protocols; and (iv) support the less prepared countries in the european community. despite hopes to the contrary, infectious diseases appear far from being defeated and continue to claim the attention of public health authorities. particularly in recent years, yet to be fully understood changes in the environment, increased movement of goods and persons, and the local influence of global warming and other phenomena concerning vectors and hosts, seem to have promoted and accelerated changes in the presentation of old infectious diseases and the development of new ones [ ] [ ] [ ] . the relevance of the 'emerging and re-emerging' infectious diseases, usually defined as 'infections that have newly appeared in a population or have existed previously but are rapidly increasing in incidence or geographic range', is further noted by the who in its recent world health report [ ] . the who stressed that infectious diseases are spreading faster and emerging more quickly than ever before. some emerging and re-emerging diseases represent a real challenge because of their epidemic potential. recently, many global alarms involving infectious diseases-such as the anthrax crisis in the usa, the emergence of sars, the pandemic threat posed by the highly pathogenic avian influenza a (h n ), and the cases of imported or autochthonous viral haemorrhagic fever (vhfs) in europe-have highlighted the need to improve preparedness for these highly infectious diseases (hids), also in order to increase certain aspects of what is perceived in many areas as an issue of collective and national security [ ] . emerging hids are of particular concern because they usually hit relatively unprepared public health systems, and appropriate diagnostic tests, vaccines, drugs, containment and mitigation measures are frequently not available or not immediately so. a similar situation could occur if a pandemic strain of influenza virus emerges: several surveys conducted in european countries and in the usa have revealed many gaps in their preparedness plans, in particular in terms of making such plans truly operational, in stepping up prevention measures against seasonal influenza, in ensuring essential services, in enhancing collaboration with adjacent countries, and in extending and better directing influenza research [ ] [ ] [ ] . research on emerging infectious diseases has been funded since the inception of the european union (eu) framework programmes (fp) for research in . in the eu developed recommendations for early diagnosis and management of bioterrorism-related infections, with the aim of providing member states with a common basis for dealing with these diseases [ ] . among activities/projects covered in the fp - , more than half are focused on various aspects of influenza, which makes the commission's fps arguably the single largest funding source for influenza research in europe. the other topics covered include: vhfs, sars, transmissible spongiform encephalopathies, food-and water-borne diseases, other zoonoses, as well as issues such as preparedness and capacity building for different diseases in a more generic fashion. in total, both influenza research and research on other emerging infectious diseases have received more than € million of eu funding each since . a complete searchable list and short descriptions of all projects, grouped into different categories (as well as a downloadable pdf version) is available online [ ] . concurrent with the increasing awareness of the threat of a new influenza pandemic, the current fp - introduces for the first time a specific area dedicated to 'potentially new and re-emerging epidemics', specifying that its 'focus will be on confronting emerging pathogens with pandemic potential including zoonoses'. the term 'potentially new and re-emerging epidemics', which is uncommon in the scientific literature, refers mainly to those emerging viral diseases of current or future relevance for europe. this new mandate to cover research systematically in the area of emerging epidemics establishes a focal point within the fp from which calls for proposals in this area can be strategically planned. the past calls in the area of emerging and re-emerging infectious diseases were frequently published ad hoc, in response to specific threats, and the lack of a dedicated area was responsible for the limited coordination and long-term planning. the new mandate in fp specifically dedicated to diseases should overcome these problems. although research on influenza will continue to receive support in view of the magnitude and likelihood of an influenza pandemic, future calls will increasingly build a strategic european research capacity for other emerging and re-emerging hids. a definition of hids and the agents/diseases included are summarized in table . several cases of these diseases have been reported in europe since : cases of sars were imported in eight countries, and approximately imported confirmed or suspected cases of vhfs have been reported, mainly lassa fever [ ] [ ] [ ] [ ] . very recently, two isolated cases of lassa fever have been diagnosed in london in travellers who returned to the uk from nigeria and mali [ , ] , and several cases of autochthonous crimean-congo haemorrhagic fevers have been reported in the european region (in turkey and in some states in the balkans) and in some countries within the eu (bulgaria and greece) [ , ] . no human cases of highly pathogenic influenza a (h n ) virus have occurred in europe, but two suspected cases were managed in the netherlands and belgium, and public health authorities in greece faced a pseudo-outbreak [ ] [ ] [ ] . moreover, several recent cases of cowpox infections have been reported recently in europe: confirmed cases in germany, one suspected case in the netherlands, five confirmed and seven suspected cases in france. although human cowpoxvirus infections are not classified as hid, these cases are worth mentioning here as an example of how an unexpected agent can disseminate rapidly. some of the cases described above were proven to be caused by the same virus, indicating exposure to a common source of infection related to an international trade in pet rats by a czech rat breeder [ ] . two cases of human infection with an orthopoxvirus, similar to but distinct from cowpox, have been identified in north-eastern italy in two veterinary doctors who had been exposed to infected cats [ ] . this finding, and the fact that the two infections occurred independently of one another, underscore the need to enhance awareness of zoonotic poxvirus transmission (possibly endemic) also in regions where this problem has not been addressed so far, e.g. the southern alps. almost all of the cases requiring isolation were first admitted to a general hospital without adequate isolation capabilities, and later transferred to a high-level isolation unit. despite the fact that no outbreaks occurred in europe, these experiences exposed weaknesses in terms of recognition, public health response, and diagnostic and clinical management. indeed, despite the wide availability of national and international plans and guidelines, their application in 'real-life' scenarios remains poor. not surprisingly, public health policies and diagnostic and clinical approaches to hids differ widely among european countries, and a common platform that would enable scientists to respond in a quick and powerful manner is still lacking. hids require multidisciplinary expertise. experts in microbiology (especially virology), public health, epidemiology, infectious diseases, and communication need to work together to respond to such incidents. for hids in particular, because of the rarity of their occurrence, strong collaboration and exchange of data, and attention to lessons learned from previous episodes, are advisable. for these reasons, creating new networks and enhancing those functioning well should be strongly promoted, in order to: ensure a rapid and effective response to health threats deriving from natural infection by or deliberate release of hid agents; stimulate complementarity and prevent duplication; promote international cooperation, exchange of experience, good practice and protocols; support the less prepared countries in the european community. a continuous effort is necessary for sustaining and promoting research on hids, whether basic or translational, in order to promote the increase of general knowledge concerning on these issues and to support the development of new tools for facing them in an effective manner. the development and refinement of new diagnostic tests, new therapeutics and innovative vaccines is mandatory as never before. the use of networking and international partnership could represent the successful strategy in this context. the traditional boundaries between basic science and clinical medicine should be dropped, and through effective networks the few hid events that occur worldwide should be studied thoroughly. a network of top-quality scientists and clinicians will provide the complementarity required for the development of these new approaches. moreover, improved funding for research on hids could come from the involvement of networks in the private sector, which could be encouraged to invest in this area because of the epidemic potential and the possible large-scale economic consequences. early recognition and prompt reaction to hids rest upon adequate preparedness, which should include the availability of adequate infrastructures and specific training for healthcare workers. several differences exist among european countries, due to government policies, as well as to pre-existing conditions, and these may result in delayed and dissimilar public health interventions and non-standardized training programmes. a better coordination of public health approaches to hids may lead to standardization of interventions and protocols, such as the prompt isolation within structures with adequate technical and logistic features, to the development of a common core-curriculum, and substantial improvement in the application of international health regulations. moreover, from a practical perspective, a network involving the main public health institutes may play a key role in the management of: a returning hid patient travelling through more than one country (e.g. in the case of one or more connecting flights), in order to coordinate public health interventions; an hid patient admitted in a country without adequate healthcare settings for isolation (in this case, a cross-border transport by ground or air may be the most appropriate solution); multi-country outbreaks. due to the current perceived international security threats, several eu member states are considering establishing biosafety level (bsl)- diagnostic facilities. to improve and sustain the existing initiatives and networks aimed at promoting collaboration among the existing bsl- laboratories appears mandatory, as well as to provide assistance, through these networks, to other european countries not equipped with such sophisticated and costly facilities [ ] . moreover, among the critical points identified in the context of the laboratory diagnosis of hid agents are the scarcity of biological samples to validate the diagnostic methods and the fact that few commercial diagnostic tests are available for these pathogens. thus, a well-functioning network is essential for: the sharing of diagnostic and research experience of the currently operating bsl- european laboratories, as well as diagnostic protocols, samples, reagents, and personnel for training; the review of current laboratory diagnostic capability for hid agents; the development of new hazard-free diagnostic tests suitable to be transferred to other non-bsl- laboratories; the standardization of procedures for biosafety and biosecurity. intra-hospital procedures for clinical assistance and infection control for hid cases represent 'the core' of managing these diseases, and represent effective measures for hid containment. on the other hand, hospitals may play an important role in the amplification of an outbreak if infection control measures are inconsistently applied. consequently, common protocols for infection control and biosafety during the clinical and diagnostic management of hids patients, based on the available evidence and on 'reallife' experiences, are strongly advisable [ ] [ ] [ ] [ ] . moreover, in order to offer to these patients the best available standards of care, a set of specific skills is required, and thus, given the scarcity of these events, a functioning network for expert consultation, second opinion, and scientific support is needed. as more and more persons, animals and goods move within europe, the need for improved and coordinated responses to hids continues to grow. furthermore, it is increasingly recognized that hids can pose a significant threat to each country's national security. innovative research and coordinated efforts, through the establishment of well-functioning networks, are the only way to deal with these issues, in order to improve preparedness and to react quickly: in short, to be 'prepared for the unknown'. a key role may be played by the european centre for disease prevention and control, whose mission is, among others, to 'coordinate the european networking of bodies operating in the fields within the centre's mission' [ ] . uncoordinated measures can lead to only partial and poor responses, and different approaches to similar health threats in various eu countries are likely to negatively affect the compliance by health professionals, and the perception of the population. although well-functioning networks are already in place, many gaps still exist, as well as opportunities for future collaboration. fortunately, new scientific developments, and new perceptions of these health threats, make this field one of the most stimulating research areas with a direct impact on the health of millions of people. global trends in emerging infectious diseases climate change and infectious disease: a dangerous liaison? european lab network prepares for high-risk pathogen threat world health organisation: geneva infectious diseases and national security how prepared is europe for pandemic influenza? analysis of national plans challenges remain in preparedness european centre for disease prevention and control gouvras g task force on biological and chemical agent threats, public health directorate, european commission, luxembourg. bichat clinical guidelines for bioterrorist agents european commission quality assurance for the diagnostics of viral diseases to enhance the emergency preparedness in europe no authors listed e-alert july: case of lassa fever imported into germany from sierra leone marburg hemorrhagic fever -the netherlands ex uganda a fatal case of lassa fever in london the first case of lassa fever imported from mali to the united kingdom crimean-congo hemorrhagic fever in greece: a public health perspective probable cases of crimean-congohaemorrhagic fever in bulgaria: a preliminary report management of potential human cases of influenza a/h n : lessons from belgium a dutch case of atypical pneumonia after culling of h n positive ducks in bavaria was found infected with chlamydophila psittaci a pseudo-outbreak of human a/h n infections in greece and its public health implications european centre for disease prevention and control. cowpox in germany and france related to rodent pets cat-to-human orthopoxvirus transmission, northeastern italy networking for infectious-disease emergencies in europe risk management of febrile respiratory illness in emergency departments the initial hospital response to an epidemic framework for the design and operation of highlevel isolation units: consensus of the european network of infectious diseases infection control in the management of highly pathogenic infectious diseases: consensus statement from the european network for infectious diseases the authors wish to thank c. schmaltz (european commission, research directorate general, brussels) for the data about eu-funded research projects, and for his invaluable suggestions and critical reading of the manuscript. all authors declare no dual or conflicting interests. key: cord- -cwhmm f authors: nan title: challenges to the european exception: what can s&t do? date: journal: a new deal for an effective european research policy doi: . / - - - - _ sha: doc_id: cord_uid: cwhmm f nan a quick review of the available evidence shows, however, that, while great strides have been made over the past few decades towards the achievement of these goals, europe is facing significant challenges in most if not all of these areas. economic growth is slow. europe's competitive position is feeble. there are not enough jobs, and not enough of them are high-level. europe is still characterised by significant poverty and regional inequality. an important demographic challenge is emerging. europeans' health is affected by serious lifestyle and contagious diseases. and the environment is being degraded. this is undermining what europeans are most proud of and turning europe into a negative exception at global level. the term "european exception" is most often used to refer to a european country not acting in accordance with what most other european countries are doing, whatever the field. sometimes, however, the vocabulary is also used to refer to how europe behaves differently from other advanced world economies. usually, reference to the european exception has a positive tone to it. europeans are proud of their commonly held values, their social model based on egalitarianism and solidarity, their high level of environmental awareness and protection and so on. however, europe appears to be the only advanced economy suffering from chronic low growth and high unemployment, and an unceasing lack of dynamism. its levels of poverty and of individual and regional income inequality are not that far removed from us levels. and this makes europeans feel anxious, and unsure of themselves, their future and further european integration. significant change has characterised the world economy over the past few decades. world trade has been liberalised as both formal and informal trade barriers have been reduced significantly, or disappeared altogether. capital roams the planet freely in search of the best investment opportunities as barriers to capital mobility have been eliminated. global communication and transportation networks have become denser and better integrated through a combination of technological and organisational innovation. the speed of technological change has accelerated while technologies are standardised more rapidly and use is made of modular production systems. as the combination of these factors has made it possible to locate the production of goods and services anywhere on the planet and still serve global markets, the global production system is in the process of being reconfigured. the new international division of labour not only provides both developing and developed countries with ample opportunities, it also has shady sides. on the one hand, low-, medium-and to an increasing extent high-technology manufacturing and services industries are under threat from delocalisation or so-called off-shoring and outsourcing, resulting in at least short-term disruption and unemployment. employment is also under threat from rapid process innovation leading to productivity increases. on the other hand, rapid product innovation provides developed countries with opportunities to improve competitiveness and serve global markets by fleeing forward as it were. the race to upgrade the economy is never-ending, however, and innovation-based advantages are fleeting and unsustainable as rapid standardisation and modular production techniques quickly allow the production process to move partially or completely to developing countries. as reflected in its lacklustre economic growth performance, europe has not yet adapted to the rules of this new game. in the first half of the post-war period, the european economy grew as fast as the world economy ( fig. . ) . in the second half of the post-war period, however, the decline in economic growth was more pronounced in europe than in the united states, japan and other oecd economies (figs . and . ) . in the last years or so, europe has done worse than the united states, while japan has once again started to outperform europe, and the large bric (brazil, russia, india, china) economies and smaller east asian economies continue to grow rapidly. the growth of output amounted to . per cent in the euro area in , substantially lower than the . per cent in the united states and the . per cent in japan, and the . per cent at world level. output is projected to grow by a higher . per cent in the euro area in , still economic growth in the euro area has been lagging that of the best performing oecd countries since the mid- s. it should be acknowledged, however, that some eu countries have performed rather well economically in the past decade. this group includes the member states formerly classified as cohesion countries (especially ireland), as well as finland, the netherlands and the uk. year cumulative growth gap fig. . . slow european economic growth in the second half of the post-war period compared to other industrialised countries (cumulative economic growth gap between the eu and the other industrialised countries (current prices and current ppps)) source: dg research data: oecd note: for both the eu- and the non-eu- oecd countries, gdp at current prices and current ppps (billions of dollars) was taken as . for all following years, gdp growth in percentages relative to the amount was calculated. then the series for the non-eu oecd countries (australia, canada, iceland, japan, korea, mexico, new zealand, norway, switzerland, turkey, us) was set to and the difference with the series for the eu- calculated. significantly lower than the . per cent in the united states and the . per cent in japan, and the . per cent at world level. whenever europe has been able to increase productivity in the past it has suffered in the field of employment, and vice versa, pointing to the existence of structural barriers to growth. underlying europe's lacklustre economic growth performance is its weak competitive position. the most common definition of competitiveness refers to the overall capacity to improve standards of living in a sustainable way. states during the s and s. but since the s, european standards of living have not increased relative to the united states ( fig. . ) . labour productivity is another common measure of competitiveness. though, except for a few countries, the productivity gap was never closed in the end, for most of the post-war period the eu somehow caught up on average with the united states. this catch-up has now stopped and is even being reversed. since , for the first time in three decades, growth in us labour productivity has outstripped that of the union (fig. . ) . this eu productivity downturn is of a structural nature and mainly due to an outdated and inflexible industrial structure slow to adapt to the intensifying pressures of globalisation and rapid technological change. deindustrialisation is often taken as a further sign of europe's deteriorating competitiveness. the fear is that slow labour productivity growth, high labour costs, year gdp per capita (us= ) eu euro area fig. and short and inflexible working hours drive entire industries to low-cost, hightech countries in eastern europe and asia. the evidence for deindustrialisation is not clear-cut. some analyses point out that industry still accounts for the same important share of gross domestic product in terms of volume as in the past, while the declining share in terms of value added and employment is due simply to decreasing prices because of productivity gains and exposure to competition higher than that for services. should it occur, the impact of deindustrialisation would indeed be worrying: the existence of many services depends on the presence of industry; industry pays better wages than services, even for low-skilled jobs; industry accounts for most innovations and technological revolutions; and industry has an important strategic role. europe's feeble competitive position is also clear from its weak trade performance, especially that at the high-tech end. europe's most dynamic export products are generally not those one would closely associate with the knowledge-based economy. the top three products with the fastest growing market share are floor coverings, pork and poultry fat, and hemp. on the other hand, if one looks at products for which market share is in major decline (> per cent loss in market share), the eu has many more ( product groups) than the united states ( ) or japan ( ). what is more, in europe many technological products are among them (e.g. air launchers, turbines, insulating glazing, drugs containing alkaloids or hormones, telephones, photographic film). high-tech manufacturing exports represent a much smaller proportion of total manufacturing exports in europe than in the united states or japan (in , . per cent vs. . per cent and . per cent respectively). europe's share of global high-tech manufacturing exports, though increasing, is lower than that of the united states (in , . per cent vs. . per cent respectively). and europe runs a structural deficit in high-tech manufacturing trade, whereas the united states and japan run surpluses. the european employment input is significantly lower than that in the united states. first, though apparently catching-up, the european employment rate is still substantially lower than that of the united states ( fig. . ). in , the eu- employment rate was . per cent and the eu- one . per cent, so to percentage points below the target under the lisbon agenda, compared to . per cent in the united states. this is mainly due to the limited participation of women, the young, and the elderly in the labour force. at . per cent and . per cent, the female and older people's employment rates were about and percentage points below the lisbon targets for . second, europe also scores lower than the united states in terms of the number of hours worked annually per employee ( fig. . ). for a long time, the low employment rate and number of hours worked annually per employee were explained with reference to the european emphasis on work-life balance. a growing number of authors draw attention to the existence of disincentives to work, however, the main one being the lack of employment opportunities. this lack of employment opportunities is clear from the high unemployment rates. in , about . million europeans were out of work. this equalled . per cent of the labour force, some percentage points higher than the rates in the united states and japan ( fig. . ) . the proportion of high-level jobs is also considerably lower in europe than in the united states. though europe likes to pride itself on its superior social model, poverty rates are rather high, and regional inequality is substantial. in , the at-risk-of-poverty year eu- the gini coefficienta number between and used to express the degree of income inequality, where corresponds to perfect income equality and corresponds to perfect income inequality -was . in both the eu- and the eu- ( fig. . ) . the share of children living in households with income below the poverty line ranges from per cent in slovenia and per cent in denmark to per cent in slovakia. the eu is also marked by substantial inequality in income levels. in , gross domestic product (gdp) per capita was below per cent of the eu- average in out of nuts regions examined in the eu- . the highest eurostat. at the beginning of the s, eurostat set up the 'nomenclature of statistical territorial units' (nuts) as a single, coherent system for dividing up the european union's territory in order to produce regional statistics for the community. nuts subdivides each member state into a whole number of regions at nuts level. each of these is then subdivided into regions at nuts level , and these in turn into regions at nuts level . leaving aside regional gross domestic product per capita (inner london -united kingdom) was about times the lowest one (lubelskie -poland). enlargement, for the european union, is at one and the same time a challenge and an achievement, a "raison d'être" and a "façon d'être". it is a continuation of the historical process that started over years ago with the communities' inception, developed through several steps (in , , , , ) , and reached a high point -albeit not an end-point -with the enlargement of the european union to countries of eastern and southern europe on may . preparation for that enlargement took several years and by the time they joined, the eu- had successfully transformed their economies from centrally planned to functioning free market ones. compliance with the copenhagen criteria for accession served as a powerful catalyst for change. this assessment is detailed in a recent stock-taking exercise in which the commission services have provided strong evidence and analyses indicating that the enlargement constitutes an economic success for the "old" and the "new" member states alike. it has to be noted that enlargement has been a dynamic process rather than a discrete event and that its effects will become visible over time. figure . shows that convergence and catching up in real income have been at work throughout the period since the late s. per-capita incomes are now much closer to eu- levels than they were in , the year in which enlargement prospects crystallized in the commission's agenda . after the output collapse in the early years of transition, growth rates in the eu- have been higher than in the eu- , but also more volatile. the key contributors to actual and potential economic growth in the eu- have been capital accumulation and technical progress (the so-called total factor productivity, tfp), while the contribution of labour has been mostly negative (that being a reflection of weak employment growth and, to a lesser extent, of an ongoing decline in hours worked per employee). in general, and consistent with the convergence hypothesis, member states with lower initial ( ) per capita income tended to grow faster in the intervening years. birth rates continue to be low in europe. everywhere, the fertility rate is below the threshold needed to renew the population (around . children per woman), the local level (municipalities), the internal administrative structure of the member states is generally based on two of these three main regional levels. this existing national administrative structure may be, for example, at nuts and nuts levels (respectively the länder and kreise in germany, or at nuts and nuts (régions and départements in france, comunidades autónomas and provincias in spain). of aids deaths was estimated at . million. in europe, the number of newly reported hiv infections is increasing, while that of newly diagnosed aids cases is decreasing. in the eu countries with data available for and for both hiv infections and aids cases, the number of newly reported hiv infections increased by almost per cent (from to , ) while the number of newly diagnosed aids cases fell by over per cent (from to ). europe is also affected by other communicable diseases including sars and avian influenza. one of the most worrying challenges for europe, and indeed for the whole world, concerns the deterioration of the environment. european citizens overwhelmingly agree that the state of the environment influences their quality of life ( per cent), that policy-makers should consider the environment to be just as important as economic and social policies ( per cent), and that policy-makers should take into account environmental concerns when deciding policy in other areas such as the economy and employment. "a high level of protection and improvement of the quality of the environment" is a european community objective (see above). europe has been implementing environmental action plans and pursuing sustainable development strategies at both national and european level for quite some time now. it plays a leading role in the fight against global warming. and it occupies a strong position in the field of environmental technologies. yet, because of population growth; consumption patterns; market, policy and political failures; features of existing technologies; and world views and values, europe and the world at large are still far removed from a development trajectory that is truly sustainable, that is, which satisfies the current needs of society (growth, competitiveness, employment, etc.) without compromising the needs of future generations. european citizens worry most about water pollution (of seas, rivers, lakes, underground sources, etc.) ( per cent); man-made disasters (major oil spills, industrial accidents, etc.) ( per cent); climate change ( per cent); and air pollution ( per cent). the sixth environment action programme of the european community - ( th eap) identifies four priority areas for urgent action: ( ) climate change; ( ) nature and biodiversity; ( ) environment and health and quality of life; and ( ) resources and waste. the environmental objectives of the eu sustainable development strategy include: ( ) addressing climate change; ( ) better management of natural resources; and ( ) making transport more sustainable. a review of nine recent comprehensive analyses of global environmental problems (table . ) showed near-unanimous agreement that the three problems posing the greatest threats to the global environment and continuing economic development include: ( ) water quality and access; ( ) climate change; and ( ) loss of biodiversity. climate change forecasts indicate that, if the level of emissions is not curbed, the temperature level will rise and risks such as water shortage, malaria and hunger will increase and affect millions of people by ( fig. . ). addressing such environmental problems is highly complex. one of the premises of sustainable development is that environmental problems interact with each other, as well as with economic and social issues. climate change affects agriculture, forestry, water availability, marine systems, terrestrial ecosystems, health and, last but not least, the economy. forests and oceans act as climate regulators but also harbour a wide diversity of species. decisively tackling the issue of biodiversity will require i.a. making forestry sustainable, addressing pollution, and dealing with climate change. pollution negatively affects health, from allergies and infertility to cancer and premature death. in the mid- s damage costs to the eu caused by air pollution originated in the eu (see table . ) were calculated to be around per cent of eu gdp (ranging from . to . per cent) and damages to eu and non eu countries caused by air pollution originated within the eu were estimated to be . per cent of eu gdp (with ranges between . and . per cent), with health damages accounting for the largest share. an animal and human health problem like aviary flu also constitutes a threat to biodiversity. environmental degradation contributes to the increase recorded in the number of disasters and, in relation to this, to a heightened sense of vulnerability (see fig. . in the last section of this chapter). disasters can be man-made or natural and include wildland fires, earthquakes, volcanic eruptions, landslides/debris flows, floods, extreme weather, tropical cyclones, sea and lake ice, coastal hazards including tsunamis, pollution events, and so on. during the period - , disasters killed , people and caused billion dollars of damage. throughout history, the relation between science and society has been marked by both continuity and change. the continuity is situated in the tension between the c h a p t e r philosophical and intellectual pursuit of and search for knowledge on the one hand, and the desire of researchers and their supporters to make scientific knowledge useful and apply it on the other hand. this tension was first recognised by the ancient greek philosophers, and has been reflected in recurring calls from philosophers and scientists throughout history, including today, for more "research for its own sake". within the context of this tension, the change has been located in what has constituted or better what has been considered useful knowledge in each age, in other words in "the changing social expectation of science": "what counts as useful knowledge differed from patron to patron and society to society, so that cosimo de medici and the united states department of energy looked for quite different 'products' to be created by their clients, but both traded support for the potential of utility". from century to century, societal expectations of s&t have not just changed. they have also increased. in the era of the ancient greek philosophers, societal expectations of s&t were rather low. s&t was a highly controlled activity carried out by a small elite group of people for philosophical or religious objectives. at present, however, it is considered a powerful tool for political, economic, and social change. in between, s&t helped exploit worldwide resources as trade empires and colonies expanded ( th century); helped expanding and consolidating trade empires and colonies, and turn their natural resources into wealth, or make up for the lack of trade empires and colonies ( th century); helped fight wars (first world war and second world war); and helped producing consumer goods, consumer medicines, exploring space, addressing environmental challenges, exploring the human genome, and so on (post-war period). it is no exaggeration to say that as a result today societal expectations of s&t have never been higher in industrial countries. in the united states, the carnegie commission on science, technology, and government listed in no less than major societal goals to which s&t can contribute (table . ). and a national academies report noted in that "the nation increasingly looks to the scientific and engineering communities for solutions to some of its most intractable problems, from chronic disease to missile defence, to transportation woes, to energy security, to ensuring clean air and clean water. expectations for s&t are perhaps higher than at any other time in our history and are placing unprecedented demands on leadership". needs of society as they change over time, or in other words, to become a 'science and technology for society' ". things are no different in europe. in , the european commission remarked that "expectations of science and technology are getting higher and higher, and there are few problems facing european society where science and technology are not called upon, one way or another, to provide solutions". out of the challenges europe is facing, and recommendations have been made on how to address them. time and again the same wide range of urgently to be addressed challenges is identified. the reports are also near-unanimous in the key role assigned to s&t in this respect, as will be seen in chapter . in other words, great expectations are held of s&t as regards the tackling of the multitude of challenges europe is facing. this will be developed in chapter as part of the new policy context that enabled the genesis of the lisbon strategy as well as of the th framework programme. the role that s&t can play in addressing all these challenges is expected to be substantial. this section will show that s&t indeed has the potential to contribute to a range of economic, social and environmental challenges: it can improve economic performance, promote employment, improve public health, tackle demographic, cohesion and environmental challenges, and so on. modern mainstream economic theory -whether neoclassical, endogenous or evolutionary -has recognised for quite some time now that technological progress and innovation are the main engines of economic growth. according to baumol, innovation explains much of the extraordinary economic growth record under capitalism. the reason is that in important parts of the economy, competition is based on innovation rather than price. firms are therefore forced by market pressure to support innovative activity systematically and substantially. according to romer, productivity growth is driven by innovation resulting in the creation of new though not necessarily improved product varieties. and under the schumpeterian paradigm, growth results from "quality improving innovations that render old products obsolete, and hence involves the force that schumpeter called 'creative destruction' ". even basic research generates several direct economic benefits. it is a source of useful new information; it creates new instrumentation and methodologies. those engaged in basic research develop skills which yield economic benefits when individuals move from basic research carrying codified and tacit knowledge. through participation in basic research, access is granted to networks of experts and information. those there is also empirical support for the contribution of s&t to economic performance (see tables and sources in annex). estimates of private returns to firms' own investment in r&d still produce varying figures, but there is an emerging consensus that gross returns between and per cent are common and plausible (table . ) . microeconomic studies confirm the existence of significant spillovers of knowledge from the firms that perform the r&d to other firms and industries. taking account of measured spillovers typically raises the estimated gross rate of return on business investment into the range of to per cent (tables . - . ) . macroeconomic studies, which by definition cover all sectors of the economy, also find significantly higher returns to r&d in oecd countries, with estimates ranging from per cent to over per cent. a recent austrian report found that the rise of corporate spending on r&d from . per cent to . per cent of gross domestic product in the second half of the s produced a boost of three tenths of a per cent in growth. both microeconomic and macroeconomic studies find that an important source of productivity growth in all oecd countries comes from the international diffusion of technology. a country's ability to absorb those foreign technologies is enhanced by investment in education and by investment in own r&d. the economic literature is not conclusive on the employment effects of innovation, since process innovation (the introduction of labour-saving technologies) is likely to have a negative effect on employment, assuming all other factors remain constant, while product innovation creates new markets and employment opportunities. but empirical evidence suggests that technological change promotes employment. such evidence includes a recent study of the directorate-general employment which found that the rate of growth of total factor productivity (due to improvements in the efficiency of production or to pure technological progress) has a positive impact on the employment rate, with a one-year lag, and that both in the short-and long-term, countries with higher than average total factor productivity growth tend also to have higher than average growth in employment. clear evidence exists that more computerised or r&d-intensive industries increased their demand for college-educated workers at a faster rate in the s. such high-skilled workers also command higher wages, as the consensus is that the increase in the schooling wage premium and the rise in wage inequality are driven by technological change. support also comes from the observation that all member states saw employment levels in the high technology sector rise between and , leading to an increase of almost million for the union as a whole, with employment in high-tech services accounting for . million of this total (fig. . ) . through its contribution to product, process and service innovation, productivity growth, and the creation of more and higher paid jobs, research and innovation can also help meet the challenges of ageing and cohesion. higher employment rates and levels of productivity -to which s&t can contribute -would allow for maintaining or increasing living standards, and for the absorption of increasing medical and pension costs. doubling the growth of productivity over the next few decades would allow for maintaining current levels of industrial production and average per capita income with some million elderly in the eu. the best solution to poverty is investing in education. for instance, in general the lower the illiteracy rate, the higher per capita income. higher levels of educational attainment enhance the chance of finding work and enjoying a decent standard of living. however, education is not yet accessible for everyone and often only to those who can afford it. improving access to educations takes time and effort. education is, therefore, in its own right not powerful enough to solve the poverty problem. in the meantime, contributions to a solution to poverty can also be expected from science and technology. besides investing in education and developing skills, this means dedicating research programmes to find ways to fight inner-city poverty, to relieve the effects of urbanisation, to diminish the impacts of ever increasing mobility on our environment, and to improve the quality of life of the vulnerable groups in society, such as the handicapped and the ill, the elderly and the young. in developing countries this can take the form of helping to improve the productivity of natural and physical assets, for example, by protecting farmland against erosion and desertification, preserving an area's natural resources, building easy-tomaintain water storage facilities and de-salinisation installations, and strengthening farmers' diagnostic capabilities in relation to livestock diseases, to name a few. that these advances have important impacts on farmers' income levels has been repeatedly demonstrated by the different targeted activities across the framework programmes. science and technology can also make a large contribution to the improvement of public health. it can assist in prevention (e.g. through the development of vaccinations), it can play an important role in the quicker and more reliable diagnosis of diseases (e.g. through the further development of medical imaging), and it can find treatments for diseases or, in the absence of treatments, it can help finding ways to control them (e.g. hiv/aids retroviral drugs). s&t can also help to lessen the impact of disease. furthermore, s&t can help to find new ways to deliver treatment (e.g. ambulant rather than hospital treatment) and can provide better tools for health care system management. a good illustration of the way in which science and technology can make a positive contribution to public health is the article edctp initiative referred to in chapter . it is also useful to take a step back here. globalisation in this regards also means the globalisation of infection transmission. as travel of people (and goods) intensifies, communicable diseases constitute challenges which it is increasingly difficult to confine. interconnectedness is a defining feature of our modernity. as a case in point, healthcare systems are indeed organised as systems -which can lead to catastrophic failures such as the consequences of hiv-infected blood supplies that took a particular prominence in france but did in fact strike many countries. ours is a vulnerable society. while that vulnerability is most strikingly epitomized by ebola-type viruses, with diverse profiles of outbreaks, it is also revealed through world bank, world development report / . the international s&t cooperation with third countries (inco) is one of those programmes which have been developed around the idea that poverty can be overcome by successfully developing human and institutional resources. european and developing countries clinical trials partnership. the rise of nosocomial infections (i.e. ills originating in the very places which are devised to heal). these further illustrate the flipside -or paradoxical unanticipated consequences -of healthcare as interconnected systems. yet, while avian flu and sars together with the above examples represent the globalisation of infection transmission, they also point to the globalisation of the means to tackle public health challenges. the relative containment of avian flu and sars, and even more so the eradication of smallpox (the variola virus), constitute inspirational successes in that regard. there is no doubt that the solution to the environmental challenge has to come first and foremost from elsewhere than from new technological development. available technological best practices should first of all be disseminated as widely as possible. a change of mentality is also required leading to less consumption of more carefully selected resources and increased reuse and recycling within the limits of the current technological frontier. yet it does not seem unjustified to expect a contribution from new technological development. technology is already used in a variety of ways when it comes to the environment, and everywhere there is great scope for improvement. technology in the form of satellites is used to monitor the global environmental situation and change therein. technology in the form of super computers is used to develop climate models and make predictions. technological development has made industrial production less resource intensive. it has also reduced the energy consumption of machinery (e.g. cars). s&t has been successful at developing alternatives for harmful substances (e.g. within the context of fighting ozone depletion). technological development has increased the extent to which a larger variety of goods can be recycled. the production of green energy is wholly dependent on technological development. and s&t is needed to mitigate the impacts of environmental degradation. this need for a joint undertaking -combining existing technologies, technological innovations, as well as political innovations -is illustrated in fig. . in the case of climate change (the fight to curb greenhouse gas emissions, that is). as the next chapter will further examine, s&t is not only an indispensable source for the evidence base on challenges such as environmental degradation, they are also taken to be one of the causes of such predicaments. one can undoubtedly point to the lack of societal controls on the use of s&t, to environmentally harmful production and consumption patterns, and to other types of failures in this regard. nonetheless, the outlook can change fundamentally if one can conceive of s&t as part of the solution rather than the problem. the "precautionary principle" is a useful notion to mark that double perspective. it can first be taken as stifling innovation in the name of environmental protection; but more interestingly, it can be understood as promoting innovations that take account of social and environmental difficulties, taking account of risks as well as benefits, taking account of less tractable, longer-term consequences. its emphasiseven with its origin in german environmental legislation in the s -was as much on environmental protection as on gaining a competitive advantage through innovations on the backdrop of environmental regulation. indeed, although this remains a fiercely debated question, a recent survey of the literature indicates that a transparent and non-discriminatory regulatory framework, coupled with high environmental standards, is an engine for innovation and business opportunities. this engine functions notably through the creation of lead markets. the story of the catalytic converters provides a compelling example of such r&d-based win-win. a first step in that perspective consists in acknowledging the need to sever the link between economic growth and environmental degradation. the endeavour of a duly responsible polity -with a concern for the quality of life of present and future generations -is then to optimise the effects of its economic activity, that is to minimise adverse externalities without sacrificing part of its material well-being or endangering economic growth. a second step consists not in ignoring the above "limits to growth" understanding, but in researching other links between development and sustainability. this move is at the heart of the role of s&t in relation to the environment -and is indeed at the heart of the lisbon strategy as underscored in the conclusions of the göteborg summit. the potential of technology to create synergies between environmental protection and economic growth was emphasised by the october european council. that well-established premise is taken to its most fruitful operational conclusions in the environmental technologies action plan. more recently, the benefits of s&t for the economy and environment alike were further examined in the "towards a more sustainable eu" report for the dutch presidency and indeed in the kok report of november . in fact europe occupies a strong position in the field of environmental technologies. of course this also relates to the fragile but powerful synergies, introduced above, between environmental promotion/protection, s&t, and growth and competitiveness. these potential benefits can also be of great importance for developing countries. with appropriate technology transfer they can provide these countries with affordable solutions for reconciling their desire for strong economic growth with the need to do so without increasing the pressure on the local -or the globalenvironment. this north-south dimension highlights the sustainable development predicament as differentiated yet common. the question of sustainable development can be posed along two main lines: a question of adapting -or otherwise innovating -appropriate "clean" technologies, and a question of redefining needs and lifestyles. now it is interesting to re-consider the climate change issue in the light of the above remarks. the european union has taken a leading role in the international process to tackle global warming so as to promote environmentally responsible choices by all actors. the eu has ratified the kyoto protocol early on, joined by almost all of its international counterparts on this course -most recently russia. its successes are also the planet's successes. the eu is committed to meet its kyoto emissions reduction targets and continues to show leadership on this issue. the role of s&t is set to become even more central in the post-kyoto (post- ) regime, for which negotiations are starting now. the need for new and cleaner technologies as an indispensable means to tackle energy demands and co emissions was the main message of the latest yearly report of the more widely, s&t plays an important part in the eu's capacity to shape -and implement -international agreements. by way of conclusion, it is worthy of note that the answers which science and technology can bring to environmental problems are increasingly judged with reference to the changes they bring in society. they demand choices of policies and governance, the impact of which on economic and social groups must be measured in terms of effectiveness and efficiency, the spread of costs and benefits, and social or regional equity. this is only possible if research also seeks to develop the knowledge-base and methodologies needed by such analyses. the ultimate answer? the ultimate challenge? as the previous discussion of the contribution of s&t to employment or environmental challenges has shown, it is not always clear-cut where problems start and where solutions end. or to put these tangled matters even more simply in this case: the role that s&t can play is manifold. and nowhere is this manifoldness better encapsulated than in the predicament of the "knowledge society". here the challenges, the expectations, and indeed the role of s&t in eliciting and addressing them, are brought together in ways that it is most illuminating to examine. first, this section probes the mutual shaping of science and culture. second, it foregrounds some collateral features of the knowledge society, and in particular the vulnerability that accompanies its emergence. this will lead up, in chapter , to a discussion of our modernity -or modernities -as characterised by a distribution of goods but also of ills or risks, and of knowledge or claims thereon. indeed, in this subsequent chapter, the problematic and ambivalent relations between s&t and the public at large will be considered in the perspective of the weaknesses of european s&t. but firstly we must examine the crucial place of s&t within our knowledge society in the making. the mutual shaping of culture and s&t the examples in this chapter have already shown how profoundly our culture is marked by s&t developments. at the same time as s&t shapes our society, they are themselves produced, taken up, reconfigured, shaped by society. that is one (double) way in which culture is decidedly scientific culture, and thus in which s&t is at the heart of this nearly eponymic "knowledge society". but to allow all sections of society to benefit from those advances -as well as to take part in that shaping process -individuals need to be provided with the appropriate equipment, in terms of education, skills, awareness, and appreciation for the stakes in s&t endeavours. vital for a democratic society in this day and age, such demands point towards another crucial sense for scientific culture, also exposing the acute need for it to be developed. actions to foster a thorough public grasp of what is science and how it contributes to society are thus sine qua non to a full-fledged democratic society. importantly, s&t developments accompany and affect lifestyle changes in societies. in this respect the taking up of mobile phones or gsm provides interesting illustrations. the gsm has strikingly changed the way people communicate with their loved ones, organize their work and outings, and live everyday. as regards research, innovation, and competitiveness, the rise of the gsm standard provides an inspiring example of european leadership. in effect, new information and communication technologies open up opportunities for new lifestyles and new ways of working. remote working or online trading decouples economic activity from a particular geographic location (be it the office, capital cities or structurally favoured regions). moreover, such technologies can facilitate access to employment -and other forms of social inclusion/participation -among sections of society (people with physical disabilities, the elderly) who may otherwise be excluded. key to achieving those benefits is ensuring that people are equipped with the necessary skills to get involved. much information society literature also hypothesises that "ework" (remote working) may contribute to environmental sustainability as, in addition to other dematerialisations, travelling to work is reduced. on the other hand, transport technologies themselves -from the wheel through to the airplanecontinue to have a central role in society, for example in enabling communication. the quality of human life is made up of many more components than the ones already mentioned: greater access to knowledge, better nutrition and health services, more secure livelihoods, clean air to breathe, security against crime and physical violence, satisfying leisure hours, political and cultural freedoms and sense of participation in community activities. s&t can contribute to improvements and bring lasting solutions in each of these areas. for example, investment in research and new technologies to achieve sustainable transport solutions generates desirable impacts on the quality of life worldwide: less energy consumption; fewer air pollution; less respiratory diseases; lower noise levels; increased space and security for pedestrians and cyclists resulting in more friendly cities for children and older people; less congestion; fewer road accidents; and so on. besides, it is s&t which makes possible the novel lifestyles -and indeed the novel societydiscussed above. it may be that, in solving some age-old problems, s&t has created the possibility for new problems to emerge. yet even to address these new problems we can hardly do without s&t. but we can -and rightfully do -concern ourselves with the consequences of the solutions we devise. the vulnerable society and the knowledge society s&t has brought a mix of benefits and risks. in the modern world heightened wellbeing and security are accompanied by increased vulnerability and insecurity. this vulnerability can take many forms, from loneliness or travelling accidents to industrial disasters or the twisting of human rights in a totalitarian state. fig. . provides an illustration of the rising challenge represented by disasters. here "disasters" include both technological and natural events. the dramatic increase shown on the graph may be due not only to the consequences of concentrated urbanisation, climate change, and so on, but also to a heightened sense of vulnerability and risk, together with a better ability to measure disasters. hence the emerging knowledge society will have its problems too. besides, it will not depend solely on s&t but also on governance and on the citizens who will make up our society -and shape it. yet it is characterized by an increasingly pivotal role for s&t. the knowledge society requires a revolution in our understanding of knowledge: not only with regard to s&t researchers, but also concerning a democratisation or broadening of knowledge production. this has profound implications for decision-making, for the lay-expert divide, for the handling of risks and uncertainties, and indeed for the relations between citizens and institutions of governance, as every individual should be recognized as -and given the means to be -a person of knowledge. europe finds itself in a peculiar situation in this regard, and the following chapter will unpack the paradoxical relations between s&t and its citizen. this chapter has explored in greater detail some important economic, social and environmental challenges europe is facing, the expectations held of s&t in addressing these challenges, and the role that s&t could potentially play. the th framework programme was designed against the background of europeans feeling anxious because the continent is experiencing a number of important economic, social and environmental challenges -or indeed against the background of a europe turning from a positive into a negative "exception" at global level. economic growth is slow. europe's competitive position is feeble. there are not enough jobs, and not enough of them are high-level. europe is still characterised by significant poverty and regional inequality. an important demographic challenge is emerging. europeans' health is affected by serious lifestyle and contagious diseases. and the environment is being degraded. as will be further examined at the end of chapter and in chapter , expectations of s&t have never been higher than they are now. such expectations held of s&t are partially justified. s&t can indeed play an important role in addressing societal economic, social and environmental challenges. s&t is the engine of economic growth and competitiveness. the employment effects of s&t are positive. s&t can play a major role in addressing the consequences of ageing, and the cohesion and public health challenges. s&t can play a key role in addressing environmental challenges. s&t is part and parcel of our lives, be they framed in a knowledge society or otherwise, and they are the linchpin of the latter's emergence. however, as will be seen in the next chapter, for s&t to be able to realise its potential, some serious s&t weaknesses will have to be addressed. united nations research institute for social development, information and communication technologies and social development in senegal: an overview/les technologies de l'information et de la communication groupe spécial mobile" hosted by the european conference of postal and telecommunications administrations, and its specifications where defined by the european telecommunications standards institute in the late s. commercial operation began -and the world's first gsm phone call was made european foundation for the improvement of living and working conditions & prest european commission, dg jrc -institute for prospective technological studies impact of ict on sustainable development key: cord- -lqzgz p authors: gallo, juan e.; ochoa, juan e.; warren, helen r.; misas, elizabeth; correa, monica m.; gallo-villegas, jaime a.; bedoya, gabriel; aristizábal, dagnóvar; mcewen, juan g.; caulfield, mark j.; parati, gianfranco; clay, oliver k. title: hypertension and the roles of the p . risk locus: classic findings and new association data date: - - journal: nan doi: . /j.ijchy. . sha: doc_id: cord_uid: lqzgz p background the band p . contains an established genomic risk zone for cardiovascular disease (cvd). since the initial wellcome trust case control consortium study (wtccc), the increased cvd risk associated with p . has been confirmed by multiple studies in different continents. however, many years later there was still no confirmed report of a corresponding association of p . with hypertension, a major cv risk factor, nor with blood pressure (bp). theory in this contribution, we review the bipartite haplotype structure of the p . risk locus: one block is devoid of protein-coding genes but contains the lead cvd risk snps, while the other block contains the first exon and regulatory dna of the gene for the cell cycle inhibitor p . we consider how findings from molecular biology offer possibilities of an involvement of p in hypertension etiology, with expression of the p gene modulated by genetic variation from within the p . risk locus. results we present original results from a colombian study revealing moderate but persistent association signals for bp and hypertension within the classic p . cvd risk locus. these snps are mostly confined to a ‘hypertension island’ that spans less than kb and coincides with the p haplotype block. we find confirmation in data originating from much larger, recent european bp studies, albeit with opposite effect directions. conclusion although more work will be needed to elucidate possible mechanisms, previous findings and new data prompt reconsidering the question of how variation in p . might influence hypertension components of cardiovascular risk. legend to graphical abstract: schematic depiction of the main observation presented and discussed in this article. two adjacent haplotype blocks characterize the p . cardiovascular risk locus: left, the block or island containing the first part of the p gene and its wellcharacterized promoter, in which we observed clearly elevated associations (red) with blood pressure (dbp, sbp) and/or hypertension in a colombian and a european study sample, and right, the block hypertension and bp association 'hypertension island' * (haplotype block < kb) lead cvd risk snps (haplotype block < kb) significance of association strongest cvd association current threshold for genome-wide reporting ( p = x - ) fair cvd association contains start and promoter of p gene contains no protein-coding genes it had long been a widely held belief that common genetic variation in the established cardiovascular risk locus of cytogenetic band p . , as discovered and delimited via corresponding genetic associations in [ , , ] , is not reproducibly associated with high blood pressure or hypertension, a prime risk factor for cardiovascular disease [ , ] . the pleiotropic nature of the ≈ -kb classic risk locus of p . , the strongest known single contributor in the human genome to genetic risk of cardiovascular disease [ , , ] , viewed together with the complexity of the genetic and molecular regulation of hypertension, prompted us to reopen the question if this region might, after all, be associated with a contribution of hypertension to the increased cardiovascular risk that characterizes the locus. in the theory section, we briefly depict the classic two-block structure of the region in the light of current knowledge, and review some findings and hypotheses that could admit a step of a hypertension etiology being modulated at this locus. in the results we first present results from an original, small colombian association study focusing on p . variation. we then present largely corroborating results from much larger, recently published european studies, which we identified in a second step. taken together, the evidence accumulating so far suggests that common genetic variation in a well-described block (< kb) of the p . risk region -and, more specifically, in the regulatory dna of the p gene it harbors -may play a role in promoting hypertension, for example via vascular modifications in resistance arteries (arterioles). demographic and clinical information, including selected cardiovascular risk factors, was collected for all participants in a study conducted in medellín, colombia (see supplementary data). systolic (s) and diastolic (d) blood pressure (bp) levels were defined by the average of two conventional auscultatory bp measurements through a mercury manometer according to an approach in line with current european guidelines [ ] . presence of hypertension was identified based on physician's diagnosis, prescription of antihypertensive drugs, average office sbp ≥ and/or dbp ≥ mmhg, or any combination of these possibilities. a total of participants were genotyped. in view of this small sample size and in line with traditional case-control methodology [ ] , [ , chapter ] , the highest and lowest dbp tertiles of the population were slightly overrepresented in the genotyped individuals. dna was extracted from white blood cells following standard salting-out procedures and genotyped by lgc group, uk with kasp tm technology (https://www. biosearchtech.com). all snps genotyped were biallelic in our study sample. snp rs is triallelic in some other kg populations (not clm), its rare third allele frequency being . in amr, . in eur and . in afr. statistical analyses were performed considering bp either as a continuous or a categorical variable (above or below the current [ ] threshold for hypertension diagnosis). effect sizes were defined as differences of mean values for continuous variables (e.g., sbp, dbp), and as odds ratios for dichotomous variables (e.g., presence or absence of hypertension). variables considered as possible covariates in the statistical analyses included gender, age, smoking and body mass index (bmi). to calculate association p-values and effect sizes between snp states and phenotypic variables we used the r package snpassoc [ , ] . models fitted included codominant, dominant, recessive, overdominant and log-additive models. participants signed an informed consent formulated for this genotype-phenotype association study. the study was approved by the ethical committee of the corporación para investigaciones biológicas, medellín, colombia. all procedures were performed in agreement with the declaration of helsinki. additional material and methods are given in supplementary material s . methods of european studies were as described in the publications presenting those studies [ , , ] . the originally described risk locus of p . has a bipartite or two-segment structure, in which each of the two mutually adjacent segments is dominated by a distinct haplotype block. although the haplotypes (i.e., states) of the two blocks are correlated, via a linkage disequilibrium originally estimated as around . (for average |d |) [ ] , their frequencies are different. the minor allele frequency (maf) plots shown in figure depict individual snp-and haplotype-level variation in the locus. two of the superpopulations that were sequenced and genotyped by the genomes project ( kg; [ ]) are represented: amr (populations from the americas having a native american ancestry component) and eur (populations of european ancestry). the 'left-hand' block of the risk region, colored green in figure , is composed of essentially three haplotype classes, giving one low and two higher, similar minor allele frequencies (separated by a green horizontal bar). the 'right-hand' block is dominated by two haplotype classes and has fairly constant allele frequencies close to %; the lead cvd risk snps of the region are at its rightmost fringe, denoted by ochre triangles. the left-hand block includes the upstream regulatory dna and start of the protein-coding gene encoding p (transcribed leftward). the figure shows a big difference between amr and eur in this block. the figure also shows a matrix, in / haplotype (schema) coding, for snps that are representative of variation within this block, as obtained by genomes' sampling of individuals (i.e., chromosomes) from clm (medellín, colombia, an amr population). where three or more major haplotype classes dominate a region, individual bialellic snps can provide only aggregated resolution of what is happening at the haplotype level [ ] . here, snp classes s (blue identifiers), s (red) or s provide information on, respectively, the contrast of haplotype class c , c or c versus the remaining two haplotype classes of the haplotype block. the cvd risk locus in p . is a confirmed risk locus not only for cardiovascular disease but also for other diseases, including cancers. the pleiotropies appear to be partly antagonistic, in that the risk allele for one disease can be the protective allele for another disease. for example, the nhgri-ebi gwas catalog (www.ebi.ac.uk/gwas) reports snps of the left block where the protective allele for an age-related condition (e.g., cvd, glaucoma) would be the risk allele for a cancer-related condition (e.g., breast cancer, glioma, pediatric brain tumor, endometriosis; see supplementary material s . ). antagonistic pleiotropies have been noted or proposed before for this locus ( [ ] ; see also [ ] ), with data suggesting a corresponding positive selection in some populations having native american ancestry [ ] . recent literature on the risk locus of p . and its associations has emphasized the enigmatic long noncoding rna (lncrna) gene anril, which has regulatory roles that are still only partly understood, but that can affect expression of proteincoding genes in the risk locus [ ] . earlier molecular biology studies had focused on two protein-coding genes for cyclin-dependent kinase inhibitors, which when activated can arrest or inhibit the cell cycle at the g stage: the gene cdkn a encodes both p arf (p ) and p ink a (p ), and cdkn b encodes p ink b (p ). a third protein-coding gene, mtap, which is located outside the risk locus, is also relevant for understanding the region. mtap encodes a key enzyme in the biosynthetic pathway of polyamines (which plays an important role in the stabilization of atherosclerotic plaques); there is also evidence that mtap may be partly regulated from within the p . cvd risk locus despite the intervening distance [ , , ] . . . the p gene some common genetic variation in the p haplotype block is in regulatory dna, where it could in principle modulate transcription of the p gene. thus, the common snp rs , in an established promoter of the p gene, borders a tetranucleotide that was critical for c/ebpβ binding and transcription of the p gene in experiments in vitro ( figure ) [ , , ] ; its common variation might also modulate a predicted stemloop (see supplementary material s . ). the functionally strategic position of snp rs was also noted before, but in the apparently antagonistic context of glioma risk [ ] . the well-studied promoter of the p gene [ ] is a 'battlefield' in which the proto-oncogene c-myc and tgfβ/smad struggle to control the p gene. indeed, control of this gene can be important: where expression of the cell cycle inhibitor and tumor suppressor p is suppressed, cancer risk can rise [ ] . tgfβ is a main j o u r n a l p r e -p r o o f regulator of blood vessel development and maintenance, but it acts through several alternative 'arms', or pathways, e.g., via crosstalk. we ask when and where the arm of tgfβ's activities that utilizes p , as part of the tgfβ/smad pathway, might be employed under physiological conditions in a context that would be relevant for the genesis of hypertension. figure shows a rough sketch of elements of three pathways in which tgfβ can play a role and/or modulate hypertension etiologies: the tgfβ/smad pathway with target gene p , in the central lane, and two other pathways that are trans-activatable by 'crosstalk'' with tgfβ, respectively mediated by the type angiotensin-ii receptor (at r / classic ras pathway, left) and the epidermal growth factor receptor (egfr, right). the central and rightmost of these three pathways and their crosstalk, for which roles in hypertension etiologies remain to be elucidated in more detail, are examined in this context in supplementary material s . ; refs. [ , , , ] describe experimental results that may be relevant. in addition to such cues from experiments, an independent line of evidence supporting a likely role for the central pathway comes from large-scale wholegenome association studies and metastudies that can now identify whole networks showing evidence of collective association with blood pressure [ , ] . the results indicate that the tgfβ/smad pathway plays an important role in bp regulation, as it is enriched for bp-associated genes [ ] . it then seems a plausible hypothesis that p , as an important tgfβ/smad-responsive target, could be involved in the modulating of blood pressure. although the complexity and possible sensitivity of such a network can render difficult the prediction of how a genetic change at a given locus will affect bp or the risk of developing hypertension, already figure hints that the p context puts us in a locus and a scenario where key players, such as cell cycle inhibitors, are only a step away from etiologically familiar routes to pathogenesis of hypertension or its prevention. we conducted a local, small study in the city of medellín, colombia (n= individuals without overt cvd) to explore possible associations between hypertension and p . ( common snps spanning > kb). one of the motivations for the study was that we knew of no p . studies of a population living in latin america. it was immediately clear from the sample size that even for minor allele frequencies (maf) above % we would find at best nominally significant evidence for associations from this study alone ( . > p ≥ × − ; [ ] ). still, we wished to see what could be achieved by collecting and analyzing local data of modest size and integrating them into the global knowledge base [ , p. ] , [ ] . the screening of p . for associations in the colombian study sample revealed an 'island' spanning consecutive snps ( . kb), in which sbp and dbp levels as well as the presence of hypertension gave consistent association signals, and the island coincided almost exactly with the 'left' haplotype block of the p . risk locus (figures and ; see theory). effect sizes for the snps in the island were approximately allele-additive. snps in the immediately flanking regions gave only sporadic or no association signals (supplementary material s . ). for instance, despite the small sample size, snp rs in the island achieved p values of × − for hypertension, and × − / × − for sbp/dbp increase, with a per-allele odds ratio of . for hypertension and a mean difference (effect size) of . / . mmhg for sbp/dbp levels, after correcting for sex and age. comparable results were obtained when including smoking and/or bmi as covariates (supplementary material s . and supplementary data). we compared the inferred risk and protector alleles for hypertension or blood pressure at our genotyped snps with the risk and protector alleles of disease phenotypes that had been previously reported at those same snps. risk alleles were defined as those for which mean difference or beta was positive (continuous variables), for which effect size was greater than one (dichotomous variables), or that were explicitly labeled as 'risk allele'. previously published associations listed in the nhgri-ebi gwas database suggested two mutually 'antagonistic' groups (see section . , supplementary material s . , or [ , supplementary information]), the risk alleles for the cvd-or aging-related conditions (e.g., coronary heart disease, or intracranial aneurism) being the protector alleles for cancerrelated conditions (e.g., glioma). in our colombian study and in the p . island where we observed the best association signals, the inferred risk alleles for hypertension or bp (e.g., a, the major allele, for rs ) were also the previ- j o u r n a l p r e -p r o o f ously reported risk alleles for cvd-or aging-related conditions at all snps that we genotyped. although our study focused on blood pressure variables and hypertension, we did consider, and subjected to the same basic phenotype-genotype analysis, several dozen (not all independent) alternative candidates for the phenotypic/outcome variable of primary interest, selected on the basis of their hemodynamic or other relevance to the development of hypertension (listed in supplementary data). however, the bp and hypertension variables consistently gave by far the best support for an association at the snps we genotyped (see supplementary data for the results obtained for rs and rs , representing the two equivalence classes of snps s and s , which are characterized by high linkage disequilibrium). a review of the data from recently published large european studies [ , , ] with detailed look-ups of results in this region of interest, again showed higher dbp association signals in the same island that we had delimited in the colombian study than in its flanking dna, with p-values that almost reached genome-wide significance. thus, in the studies by warren et al. [ ] and by evangelou et al. [ ] (n > , ), the two lowest p-values for dbp, . × − and . × − , were found, respectively, at the 'left' fringe of the island's haplotype block (rs ) and at a snp in the interior of the island that we had not genotyped in the colombian study (rs ). furthermore, in the european blood pressure studies [ , ] genome-wide significance of dbp associations was attained, outside of the classic p . cvd risk locus and its flanking regions, in the next gene mtap (see figure and theory), with a lowest p-value of . × − for the sentinel snp rs (red asterisk and red horizontal bar at left in figure ; see also the locuszoom plot in supplementary material s . ) . the bp association with the mtap gene had not been noted in previous studies, and indeed the region had not been sufficiently covered in earlier imputation panels. in the earlier study by ehret et al ( [ ] , n > , ), the best p-values observed were within the island, and again for dbp, although they did not reach values below . (see supplementary material, subsection s . ). we were even able to detect concordant results as far back as , in a study screening close to . million snps for bp associations [ ] . the snps in the island again had much lower p-values than those in the lead cvd risk zone and regions flanking the island. thus, the snp that had the lowest p-value for hypertension in our colombian study sample, rs , had similarly the lowest p-value for dbp in the island in ref. [ ] , namely . as its genome-wide meta-analysis p-value corrected for genomic control. a notable difference with respect to the colombian results was the effect direction. indeed, directions of the bp effects in the european studies [ , ] were consistently opposite to those observed in the colombian study at the same snps. in other words, the bp effects were also opposite to the established cvd effects for those snps in the literature. the now established cvd effect directions within the island are the same in european populations, deduced via direct genotyping of island snps, as they are in populations with a latin american or native american ancestry component (e.g., [ , ] ), where they can be inferred from genotyped snps in linkage disequilibrium located close to the island. thus, in the european study samples, but not in the colombian study sample, the inferred protector alleles for hypertension (i.e., earlier in disease etiology) are the known, constant risk alleles for cardiovascular disease or events (i.e., later in the etiology). three other european reports we found, in ref. [ ] and in the phenoscanner and roslin geneatlas association databases, had varying significance and independence, but the reported effect directions appear compatible with our findings (supplementary material s . ). j o u r n a l p r e -p r o o f the results from the trans-ethnic investigations we describe here are compatible with the persisting presence, in a south american and in a european population, of associations between bp levels/hypertension and genetic variation specifically in a previously identified haplotype block or 'island' nested within the cardiovascular risk locus of band p . . in the european population, dense coverage of the association screening also allowed the recognition of an association peak for bp in a neighboring gene, mtap, which is located at about kb from the risk locus. according to the data collected so far, the hypertension or bp effect within the cvd risk locus p . would have the opposite direction in the south american compared to the european population, whereas the cvd effect appears to be the same in both populations, according to published reports [ , ] . to our knowledge, however, our study is the first screen of the p . 'island' for hypertension or bp associations in a south american population, and we also know of no dedicated cvd association study with dense coverage in this part of the p . risk zone. it is therefore too early to generalize or extrapolate from this one small study, and we must await confirmation from independent genotypephenotype screening in future, if possible also in south american populations having other ancestry proportions, such as those of peru or bolivia (see, for example, [ ] ). the larger networks in which the p gene is embedded (e.g., the pathways of figure ) are likely to be complex and resilient, and not just genetic background but also environment (e.g., diet) and local epidemiology (e.g., disease prevalences) could influence routing or rebalancing of the networks and thus affect associations. according to theoretical arguments, genotype-phenotype associations that change direction of effect between geographic, population or other environments (e.g., in the form of crossing reaction norms [ , ] ) could actually be favored, because a variant that is inferior in all environments would be rendered unstable and would eventually be eliminated from the gene pool [ , ] , [ , section ] . in line with this view, trans-ethnically significant bp associations can exhibit sizable directional inconsistency rates. for example, among snps genome-wide that were associated with bp traits (sbp, dbp, or pulse pressure) at a high significance of p < − in a european sample (n = , ) in ref. [ , supplementary table ] , of the snps having p < . also in a south asian population, and of the snps having p < . in an african population, had effect directions that were opposite to those in the european population. as a final note, a wider view of physical or regulatory gene interactions may help understand potential roles of the cvd risk locus in modulating blood pressure or hypertension. one line of investigation that could be relevant is represented by hi-c data that capture interactions between different chromosomal regions. thus, hi-c data reported in ref. [ , supplementary table ] show, at least in mesenchymal stem cells, an interaction cluster that includes the classic risk region and other regions from almost all of cytogenetic band p . ( . / . mb; for details see supplementary material s . ). the clustering regions include (or immediately flank) the p . interferon gene group, relevant in inflammation, and two genes (gadd g and focad) that have been considered of interest in cardiovascular as well as cancer contexts. of particular note, the cluster also includes the mtap and dmrta genes, which have been recently reported to be associated with dbp [ ] and sbp [ ] , respectively, and are located at a distance on either side of the classic risk region. such findings suggest that a full understanding of the potential role of p and the classic cvd risk locus in modulating hypertension and cvd might require a wider view that takes into account the regulatory interactions among genes distributed across, and possibly beyond, the . -mb cytogenetic band p . . additional (extended) materials & methods, theory, and results are included in supplementary material sections s , s , and s , respectively. de-identified genotype data and detailed association results of the medellín study are given in a supplementary data file (excel workbook) and described in supplementary material section s . j o u r n a l p r e -p r o o f pomethylation that is disrupted in some cancers [ ] ) illustrate the functional importance of the region around snp rs . bottom: functional p promoter binding sites (boxes), critical subsequences (beige) and attempted experimental replacements [ , ] that compromised normal transcription of p (bottom subsequences). legend to figure : schematic diagram sketching three well-characterized pathways ( vertical lanes) in which tgfβ plays a role via signaling and/or transactivation/crosstalk and which may act to promote or prevent hypertension. at the top of each lane, preparation steps needed for a master product that is essential for the pathway's activation are summarized. asterisks and daggers indicate gene products in which common snps in or in the gene's vicinity have been reported to be associated with blood pressure or hypertension in ref. [ , ] (*; (*) for pathway) or have been reviewed as being associated with bp or hypertension in ref. [ ] ( †). dashed horizontal arrows indicate experimentally observed or hypothesized crosstalk/transactivation between pathways (not just receptors). not shown, for simplicity, is another potentially relevant system, descending alongside the classic ras pathway at left, namely the 'nonclassical' ras, composed primarily of the angii/ang iii-at r pathway and the ace -ang-( - )-at r axis [ ] , which generally counteracts the effects of a stimulated classic ang ii-at r axis as shown in the figure rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs kg-clm,% kg-gbr,% class s s s s s s s s s s s s s j o u r n a l p r e -p r o o f rs genome-wide association study of , cases of seven common diseases and , shared controls genomewide association analysis of coronary artery disease a common allele on chromosome associated with coronary heart disease chromosome p . locus for coronary artery disease: how little we know alu elements in anril non-coding rna at chromosome p modulate atherogenic cell functions through trans-regulation of gene networks recent studies of the human chromosome p locus, which is associated with atherosclerosis in human populations. arterioscler from genotype to phenotype in human atherosclerosis -recent findings genomic risk prediction of coronary artery disease in , adults: implications for primary prevention esc/esh guidelines for the management of arterial hypertension haplotypes of the beta- adrenergic receptor associate with high diastolic blood pressure in the caerphilly prospective study principles of statistical inference snpassoc: an r package to perform whole genome association studies basic statistical analysis in genetic case-control studies the genetics of blood pressure regulation and its target organs from association studies in , individuals genomewide association analysis identifies novel blood pressure loci and offers biological insights into cardiovascular risk genetic analysis of over million people identifies new loci associated with blood pressure traits genomes project consortium. a global reference for human genetic variation toward multiple snp motif analyses of loci associated with phenotypic traits antagonistic pleiotropy and mutation accumulation influence human senescence and disease chromosome p snps associated with multiple disease phenotypes correlate with anril expression genetic loci associated with coronary artery disease harbor evidence of selection and antagonistic pleiotropy long noncoding rna anril: lnc-ing genetic variation at the chromosome p locus to molecular mechanisms of atherosclerosis expression of chr p genes cdkn b (p ink b ), cdkn a (p ink a , p arf ) and mt ap in human atherosclerotic plaque cytoplasmic mtap expression loss detected by immunohistochemistry correlates with p homozygous deletion detected by fish in pleural effusion cytology of mesothelioma tgfβ influences myc, miz- and smad to control the cdk inhibitor p ink b c/ebpβ at the core of the tgfβ cytostatic response and its evasion in metastatic breast cancer cells tgf-β family signaling in tumor suppression and cancer progression variants in the cdkn b and rtel regions are associated with high-grade glioma susceptibility battle at the p promoter tgf-beta regulation by emilin : new links in the etiology of hypertension loss of emilin- enhances arteriolar myogenic tone through tgf-β (transforming growth factor-β)-dependent transactivation of egfr (epidermal growth factor receptor) and is relevant for hypertension in mice and humans dissecting the role of epidermal growth factor receptor catalytic activity during liver regeneration and hepatocarcinogenesis p < × − has emerged as a standard of statistical significance for genome-wide association studies promoting cardiovascular health in the developing world: a critical challenge to achieve global health the missing diversity in human genetic studies genetic variants in novel pathways influence blood pressure and cardiovascular disease risk susceptibility locus for clinical and subclinical coronary artery disease at chromosome p in the multi-ethnic advance study the effect of chromosome p variants on cardiovascular disease may be modified by dietary intake: evidence from a case/control and a prospective study association of a chromosome locus p . cdkn b-as variant rs with hypertension: the tam-risk study natural selection on genes related to cardiovascular health in high-altitude adapted andeans the evolution of life histories introduction to genetic analysis trade-offs in life-history evolution the causes of evolution selective variegated methylation of the p cpg island in acute myeloid leukemia elastin microfibril interface-located protein , transforming growth factor beta, and implications on cardiovascular complications nonclassical renin-angiotensin system and renal function abstract: schematic depiction of the main observation from association data presented or discussed in this article. two adjacent haplotype blocks characterize the p . cardiovascular risk locus: left, the block or island containing the first part of the p gene and its well-studied promoter, characterized by elevated associations (red) with blood pressure (dbp, sbp) and/or hypertension in colombian and european studies, and right, the block containing lead cardiovascular risk kg-eur and kg-amr, representing mainly european origin and mainly or partly native american origin, respectively. color-shaded matrix shows -snp haplotype motifs (rows of red 's/major allele and 's/minor allele: master motifs, yellow shading: one-snp mutants) of the kg-amr individuals ( haplotypes) from medellín, kg-clm, which correspond in ancestry and admixture to the individuals m -clm studied here from the same city (snp allele frequencies are shown below matrix). the major haplotype classes are shaded light green (c ), light blue (c ) and sand (c ); by inclusion/exclusion they define the snp classes s (blue rs identifiers), s (red rs identifiers) and s . relative haplotype frequencies are shown at right for kg-clm, corresponding to our association results from medellín, and for the british population this figure illustrates a nested-plot overview of genotyped p . snps, the association signals for hypertension in the medellín study, and elements of a causal hypothesis involving a snp in the p promoter. colored bars and asterisks (top track) show extent and sentinel/lead snp of regions of strongest bp (red) or cardiovascular risk (blue) association in the studies by warren we thank professors i. king jordan and greg gibson (georgia institute of technology) for suggestions and references early in this study, drs. edwin garcía and juan ramón gonzález for advice on association analysis, and richard jacobs and jane mcdougall (lgc genomics uk) for genotyping assistance. we also thank two anonymous referees of a earlier version of this paper for suggestions. this rs rs rs rs rs rs rs rs rs rs rs rs rs * rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs rs key: cord- - hf qw authors: romette, j. l.; prat, c. m.; gould, e. a.; de lamballerie, x.; charrel, r.; coutard, b.; fooks, a. r.; bardsley, m.; carroll, m.; drosten, c.; drexler, j. f.; günther, s.; klempa, b.; pinschewer, d.; klimkait, t.; avsic-zupanc, t.; capobianchi, m. r.; dicaro, a.; ippolito, g.; nitsche, a.; koopmans, m.; reusken, c.; gorbalenya, a.; raoul, h.; bourhy, h.; mettenleiter, t.; reiche, s.; batten, c.; sabeta, c.; paweska, j. t.; eropkin, m.; zverev, v.; hu, z.; mac cullough, s.; mirazimi, a.; pradel, f.; lieutaud, p. title: the european virus archive goes global: a growing resource for research date: - - journal: antiviral research doi: . /j.antiviral. . . sha: doc_id: cord_uid: hf qw abstract the european virus archive (eva) was created in with funding from the fp -eu infrastructure programme, in response to the need for a coordinated and readily accessible collection of viruses that could be made available to academia, public health organisations and industry. within three years, it developed from a consortium of nine european laboratories to encompass associated partners in africa, russia, china, turkey, germany and italy. in , the h research and innovation framework programme (infras projects) provided support for the transformation of the eva from a european to a global organization (evag). the evag now operates as a non-profit consortium, with partners and associated partners from eu and non-eu countries. in this paper, we outline the structure, management and goals of the evag, to bring to the attention of researchers the wealth of products it can provide and to illustrate how end-users can gain access to these resources. organisations or individuals who would like to be considered as contributors are invited to contact the evag coordinator, jean-louis romette, at jean-louis.romette@univmed.fr. the european virus archive (eva) was created in with funding from the fp -eu infrastructure programme, in response to the need for a coordinated and readily accessible collection of viruses that could be made available to academia, public health organisations and industry (gould et al., ) . within three years, it developed from a consortium of nine european laboratories to encompass associated partners in africa, russia, china, turkey, germany and italy. following the h , call for research and innovation framework programme (infras projects) the concept of developing the eva into a global organization was a natural progression and the project was funded for a four-year period. in april , the evag started to operate as a non-profit consortium including partners and associated partners from eu and non-eu countries. based on excellent progress during the first two operational years, we have every reason to believe that the consortium will continue to expand and will ultimately constitute the world's largest virus collection. in this paper, we outline the structure, management and goals of the evag, to bring to the attention of researchers the wealth of products it can provide and to illustrate how end-users can gain access to these resources. laboratory isolation and characterisation of viral pathogens, such as vaccinia virus, poliovirus, foot and mouth disease virus, yellow fever virus and others, began at the turn of the twentieth century. this represented a pioneering era in the development of medical and veterinary virology. over the years, diagnostic and research centres were established worldwide to investigate viruses causing a wide range of diseases including those that had hitherto escaped all attempts to be identified and cultured under laboratory conditions. in the st century, ever-growing numbers of mammalian viruses continue to be isolated. they may represent only the tip of the iceberg, as more viruses are being discovered and reported using rapidly developing molecular technologies. as a result of these advances, the isolation, identification, characterisation and preservation of growing virus collections has inevitably generated new challenges. many of the known viruses are in collections that may be lost or discarded when the virologist in charge retires or when a laboratory closes or is reorganized. other collections may not be available to the wider virology community, because scientists are not always willing or in a position to share their viral isolates. they may lack the infrastructure, time (manpower), facilities and experience to characterize and present their collection in an accessible catalogue and further be able to ship their collected strains to foreign countries due to the restraints imposed by increasingly complex transportation regulations. furthermore, many governments have introduced additional security measures that specifically regulate the transfer and/or exchange, between countries, of high-risk-group pathogens that may attract the interest of terrorist organisations. additionally, laboratories in russia, china, india, southeast asia and south america hold their own virus collections, many of which, to date, have been relatively inaccessible to european laboratories. viruses from many taxonomic families have also been identified on the african continent and some well-developed laboratories in africa have made their collections readily accessible to researchers in europe and the americas. currently, however, except for a few countries, including senegal, south africa and kenya, authenticated virus collections from other regions of africa are virtually nonexistent. one objective of the evag therefore has been to establish trust among these widely disparate institutes and to associate the scientists in these countries with partners in the consortium under carefully monitored conditions that protect these developing laboratories, to the mutual benefit of everyone. the current project aims at the development of virus collections in low-income countries where, in some cases, there has been an understandable reluctance to exchange viruses for fear of losing their resource to institutes from more highly developed countries. the evag infrastructure provides assured access to modern capacities, thereby assisting these institutes to train their staff in developing their own resources and expanding and maintaining these capacities locally. end-user requests for viruses and reagents from these developing laboratories will be entirely managed from within, but supported by the quality management and distribution systems of the consortium. in common with all other members of the consortium, these developing laboratories will automatically retain full ownership of their collections. elsewhere, research laboratories often accumulate collections of viruses that are primarily dependent on the speciality of the laboratory scientists. for example, the american type culture collection (atcc) and the us centres for disease control and prevention (cdc) maintain large collections of mammalian viral pathogens, including risk group (rg ) agents. laboratories in russia, china, india, southeast asia, south africa, australia and new zealand hold collections that have been isolated specifically in their countries. relatively large collections of arboviruses are currently held in the usa (galveston national laboratory, texas), france, the czech republic, slovakia and also in some countries of scandinavia. european collections of specified animal pathogens, particularly lyssaviruses, pestiviruses, arboviruses and influenza viruses are maintained at the animal and plant health agency and at the pirbright institute in the united kingdom; in italy at the istituto zooprofilattico sperimentale delle venezie (izsve) hosting the world organization for animal health/food and agriculture organization (oie/fao) and eu reference laboratory for avian influenza; and in france at the institut pasteur. rg viruses such as ebola and nipah are maintained in specialised facilities in france, germany russia, the usa, china, south africa and the uk. diverse viruses of medical or veterinary importance (coronaviruses, herpesviruses, retroviruses, adenoviruses, enteroviruses, etc.) are held in laboratories around the world. as the evag continues to develop and to expand its boundaries, access to a greater proportion of these viruses will be enabled where possible. recently, in meeting the challenges of emerging viruses, the consortium has voluntarily oriented a part of its activities to the support of the world health organization (who) during emerging health crises by dedicating a new work-package entitled "response to emergence". moreover, the evag is currently an associated partner of the global outbreak alert and response network (goarn). current processes of coordinating virus standardisation, characterisation, preservation and distribution are inevitably relatively arbitrary, and are largely dependent on the speciality of each laboratory. in fact, besides the evag, we are unaware of any non-profit organization that is concerned with facilitating reliable access globally to viruses and associated reagents from individual virus collections for research and/or diagnostic laboratories, teaching centres or industries involved in the production of diagnostic reagents, pharmaceuticals and vaccines solely for the benefit of science, in a safe and carefully regulated manner. it would be virtually impossible to establish a single laboratory to maintain supplies of all recognised pathogenic mammalian viruses. based on the knowledge and experience gained from the former eva project, we have addressed these problems by using web-based tools to centralise access to quality-controlled, standardised preparations of viruses held in laboratories worldwide. the underlying concept is simple, but because virologists are naturally protective of their collections, its success has required a paradigm shift in the mind-set of the partners or associated partners in the individual laboratories. the h -infraia- - framework programme of the european union (eu) placed a call for funding applications under the heading "research and innovation action," with sub-headings, one of which was "research infrastructures." accordingly, the concept of the evag was created and a funding proposal was submitted. in preparing the proposal, it was emphasised that it would be virtually impossible to generate a wide-ranging virus archive in a single laboratory complex. thus, although many high-calibre virology laboratories operate in europe, it was decided to retain the "virology club tradition" by developing the principle of an integrated international infrastructure via consortia, operating through validated networks. as far as we are aware, this concept had not previously been encompassed by virologists in the context of producing and providing access to viruses. the evag was therefore conceived to fill the gaps in accessing viruses through identification of qualified international laboratories that could become partners of the consortium. it was agreed that by integrating their collections or resources and devising appropriate validated protocols and effective dissemination procedures, the partner laboratories would be able to achieve common high standards in producing and supplying authenticated viruses to the scientific community, both within and outside europe. the evag is unique. it was conceived to move beyond the current state of the art and provide access to the largest collection of mammalian viruses and associated reagents in the world to the global scientific community, to government health and environmental departments, to higher education institutes and to industry, through appropriate information systems. the new approach was, in the first instance, to integrate non-eu partners as beneficiaries of the project. this was possible because the rule of reciprocal benefit was agreed: the non-eu partner provision activity of material to an eu entity (research institute, health organization, animal health-care structure, etc.) was eligible for financial compensation from the consortium eu budget. consequently, highcalibre european and non-european partner laboratories agreed to become the primary members of the evag consortium. these are listed in table partner laboratories that form the core of the evag, listed in chronological order of their integration into the project. both medical and veterinarian institutes are included to cover viral zoonotic diseases. it also illustrates the wide geographic coverage already attained and demonstrates that bsl laboratories are major contributors to this consortium. the choice of institutes also reflects the specific available skills and know-how of scientists in institutes closely associated with the most well-known contemporary emerging viruses. we have also begun to integrate network organisations such as the global approach to biological research, infectious diseases and epidemics in low-income countries (gabriel) network, of the fondation merieux and the international pasteur institutes. future plans include expansion of partnerships to countries not currently represented, such as malaysia, vietnam, cambodia, indonesia, south america, and additional african countries. in addition, associate partners who share the same interest in the dissemination of viruses and reagents, but are not beneficiaries of the eu grant, have now been included (see table ). this association was formalized by the signature of a memorandum of understanding. the archive is founded on the principle that members and associate members of the consortium retain ownership of the viruses that they disseminate via the centralised distribution centre. the virologist or institution that supplies the viruses to the customer retains the ownership and enters into agreements with the customer via material transfer agreements (mtas). all orders for viruses and associated reagents (cell cultures, recombinant proteins, rna, dna, recombinant viruses, plasmids, monoclonal antibodies), are placed via the website https://www. european-virus-archive.com/which is managed centrally in marseille by the management team (partner# , aix-marseille university). during the early stages of the existence of the original eva, emphasis was placed on establishing the collections of viruses and associated reagents and ensuring that quality control was managed centrally. this ensured that all laboratories complied with identical standards. this principle has been further developed in the project. a manager was appointed to head up a management quality committee with the responsibility of establishing and maintaining appropriate standards through close monitoring of procedures, protocols and record maintenance. to ensure uniformity throughout the consortium, the quality managers representing each partner form the body of the committee. previously, non-eu participants were given the status of associate partners, i.e. they were associated with the consortium, but were not full members. the objective was to evaluate the feasibility of this approach concerning both the collaborative exchanges related to development of a common infrastructure and the operational effectiveness of resource distribution. from the point of view of scientific collaboration, the process was established rapidly and functioned efficiently. however, standardisation of the procedure for the distribution of materials to end-users proved to be challenging, and it is still being developed and improved within the current consortium. the reason for this slow progress is almost entirely due to the complexity of national regulations. whilst the european regulations retain the principle of being national and are enforced primarily to control the circulation of pathogens, they have been developed to maintain uniformity throughout the eu countries. thus, the administrative pathway to obtaining authorisation for an exchange of resources between, for example, france and germany or the netherlands and italy are very similar and are principally based on agreements provided by the national biosafety agencies. in contrast, the process of exchanging resources between the eu and russia or between the eu and the usa involves many more constraints: the intervention of different administrations in processing authorisation for delivery of hazardous goods including, biosafety agencies, customs administration, commerce and trade administration, ministry of health, etc. (also, the law in russia does not permit the import or export of pathogens.) nevertheless, our expertise in overcoming these hurdles is exemplified by the report of studies on the rg virus crimean-congo haemorrhagic fever virus in which the virus was supplied by a european partner to an end-user in the usa (haddock et al., ) . not surprisingly, when zika virus emerged in south america and was found to be responsible for microcephaly, we received numerous requests for zika virus, and many papers have been published with recognition of the evag as the source of virus. five examples encompassing, i) simultaneous transmission with chikungunya virus, ii) diagnosis, iii) signal processing pathways, iv) pathogenesis in relation to foetal infections and v) virus structure, are cited here (goertz et al., ; chan et al., ; zhang et al., ; miner et al., ; sirohi et al., ) . these anticipated problems of transporting pathogenic viruses between countries, particularly outside europe, were considered during the preparation of the project. each potential barrier to efficient exchanges between different countries was extensively discussed and product dissemination objectives for each non-eu partner were defined depending on their specific status. once established and operating satisfactorily, the management team continued the process of contacting high-calibre laboratories outside europe, organising meetings and conferences with these laboratories and subsequently inviting them to become associate partners. currently associate partnerships have been negotiated and agreed with laboratories in africa, russia, china, turkey, australia, japan, germany, italy, hungary, south korea, south east asia and south america ( table ). the initiation and early progress of the evag are already tremendous achievements, and subject to its continued success and expansion, it is anticipated that it will become the largest global network of laboratories contributing to the sharing of resources and promoting advances in virology. currently agreed associate partners in the evag. the integration of institutes that share the same interest in research and in the provision of high quality products to the scientific community is made through an association formalized by the signature of a memorandum of understanding. the choice of the associate partners has been established to fill gaps or to extend the list of viruses currently offered in the catalogue, and to have evag present in geographical areas concerned by emerging virus outbreaks. dissemination of resources including know-how and material to the scientific community is one of the major deliverable of the project. it has been addressed by the creation of web-based catalogue accessible on the evag portal. quality of the offer is a key factor to differentiate evag from the other infrastructures proposing similar products to the end-user. the gold standard products listed on the catalogue have been clearly at the origin of the fast growing dissemination activity of the consortium and of its robust reputation as reliable provider of key components for the detection of emerging viruses acting under the umbrella of international health organisations. the evag infrastructure provides wide-ranging and efficient access to virus collections held in laboratories worldwide, with the potential for access to recently isolated viruses from clinical, veterinary and field samples. the collection will continually expand, as the number of contributing laboratories increases and will provide participants with access to materials and capacities, including: • freeze-dried viruses for long-term storage; • viruses validated by sequencing, including the "rescue" and characterisation of archived material; • reagents and kits, including primer sequences and appropriate protocols for virus identification; • standardised diagnostic kits and recombinant proteins representing genes of the viruses, or hyperimmune antisera with defined specificities; • custom-prepared kits, proteins and antibodies; • high-quality associated research laboratories accessible to perform experiments and theoretical and practical training. relevant information can be obtained by placing an enquiry on the website (www.european-virus-archive.com/evag-portal). at the time of writing the catalogue shows more than products consisting of viruses and associated reagents. the list which is constantly increasing includes human, animal, insect, fish and plant viruses of medical, veterinary, academic, industrial and economic importance in addition, a range of arthropod tissue culture cell lines, monoclonal antibodies, recombinant protein expression systems, purified viral rna, specialised kits for diagnosis and custom-prepared proteins, can also be obtained through the website. the approach to quality management is directed by the project's own quality standard, based upon organization for economic cooperation and development (oecd) guidelines, and critically focusses on the acquisition, characterisation and storage of virus products. each partner institute seeks to comply with this way of working to harmonise laboratory processes across the consortium. the quality manager conducts "best practice" audits to quantify improvement progress. all facilities are inspected and regulated by their own national bio safety authorities, and employ staff trained in procedures compliant with the requirements of the international air transport association (iata) for the safe dispatch of potentially hazardous materials. trans-national access (tna) ensures free of charge access to european research infrastructures and the costs of the research, products and associated activities. this opportunity is open to all european researchers and to some extent to researchers from non-eu countries. tna dispatches include regulatory documentation such as import/export permits, and a certificate of analysis (coa) provided by the supplying partner. the evag project has established a grading system that defines virus product quality ranked from to , in which grade is the basic level, described as partially sequenced to confirm identity; not checked for mycoplasma contamination; unknown infectivity; and stored frozen. the "gold standard" is a fully sequenced virus, mycoplasma-free, with defined infectivity and lyophilised for long-term storage. derived products such as proteins, nucleic acids or antigens are also prepared using standardised protocols and checked for quality before storage or dispatch. security at the consortium institutes is maintained using cctv, pincoded passes and at remote sites, -h manned security/vehicle identification. entry into bio containment laboratories is strictly controlled. supply of products to the applicants operates via the tna approach defined above, whereby research groups apply through the web portal by submission of a completed request form. a successful application must be justified by an explanation of scientific objectives, techniques to be utilised/developed, list of named researchers, list of relevant publications and confirmation that appropriate expertise and facilities to handle the virus are in place. a review of these criteria is carried out by the project quality manager and a selection panel that includes external experts. a team of scientists and administrators is dedicated to the management of the evag, to ensure continuity and to maximize efficiency. whilst the individual laboratories operate through the web-based catalogue, they supply to the end-user only the viruses and products that are held in their collections. the website is the main communication tool with which to inform the end-user and to disseminate the products through the dedicated portal. the portal is conceived as a user-friendly marketplace that lists the viruses and associated reagents of each partner. this web-based catalogue constitutes a single entry point to the archived viruses and reagents. it provides access for end-users to place any enquiry about products of interest through a commonly used e-shop format. any end-user could be entitled to free access to the virus or associated reagent(s). the provider of the requested item(s) receives financial compensation from the eu budget. eu support for free-ofcharge access provided to the end-user has as its main objective to facilitate access of the scientific community to high quality resources. however, this is conditional on the excellence of the project which the end-user aims to develop using the resource. an international panel, consisting of independent experts, reviews the request for free access and subject to approval the customer will receive the item without charge, other than covering the cost of transportation. in the case of a charge being required to supply the virus or reagent, the evag only recovers the cost of production and transportation to the customer. free-of-charge access to end-users employed outside eu-member or euassociate states is limited to % of the total accesses provided in the grant. in addition to core partners, the project also integrates associate partners as an efficient means of enlarging the collection of viruses and reagents in the web-catalogue. easy and efficient access to the viruses and reagents and to the service offered to the scientific community is attracting increasing numbers of requests for the products in the catalogue. in addition, the quality of the products distributed and the assistance provided by the management team to answer specific questions from scientists are establishing the evag as a key biological resource centre worldwide. it is now a recognised entity acting under the umbrella of medical and veterinarian international health organisations which include the who, goarn, oie and the major centres for disease control and prevention around the world which underpin the control of emerging viral diseases. to place an enquiry and receive a quotation for a product, open a web browser and type in "evag." then click on login/register to create an account. the online catalogue can then be accessed via the portal link on the home page. products in the catalogue can be found by typing in a keyword/and/or the appropriate taxonomic name. the filters on the right hand side of the portal can also be used to browse through the products by category. the product(s) of interest should then simply be added to an enquiry cart, and the enquiry form should be completed. a material transfer agreement (mta) and a quotation for the products and shipping costs will then be sent to the end-user. the enquiry then becomes an order when the quote is accepted and the mta is signed. all enquiries are evaluated in terms of biosafety, and an official (signed and stamped) end-user qualification form of the laboratory to receive and handle the products of interest is requested. the emergence of sars at the end of has clearly demonstrated the lack of preparedness of countries to face emerging virus outbreaks. when the concept of eva was discussed in , this aspect of the consortium activity was clearly stated in the project submitted to the european commission. this was reinforced in the second project: evag, submitted in , with a contingency fund dedicated to support this activity. the emergence of the novel mers-cov in humans in saudi arabia in provided an opportunity for consortium partners and members of a european epidemic response network to develop a precise and rapid mers cov diagnostic tool, suitable for field diagnosis under emergency conditions (corman et al., ) . during december , the who recommended pcr assays as the method to detect the virus in blood samples, and identified the evag as a reliable source for the delivery of positive control reagents (who, ) . to date more than kits have been distributed worldwide, to laboratories in countries. this achievement demonstrated the capacity of the consortium to mobilize high-calibre scientists capable of rapidly developing and supplying a technical solution to a high-level emergency situation involving human pathogenic agents. another key feature of this achievement was the efficiency of our logistics platform to distribute the material worldwide under the demanding conditions prevailing at the time. zika virus emerged in french polynesia and spread rapidly to south america and the caribbean, causing at least , clinical cases, within the first year according to paho/who estimates. on the st of february , who declared a public health emergency of international concern (pheic) (see ref ). on the th of february, the us centre for disease control (uscdc) elevated its response to "level activation" i.e. the highest level. prior to may , there were two strains of zika virus in the evag online catalogue. by the first trimester of , at the peak of international concern, partners had added eight more products of direct relevance to the emergency situation. since november, , the evag has received enquiries for more than products to be distributed in countries. a total of products were provided in february, alone, with the highest demands in the usa, singapore, netherlands, germany, france and china. in its interim guidance for zika virus laboratory testing (march , ), who identified the evag as the provider for pcr quality-control material (corman et al., ) . this emergency situation and the resulting need for a coordinated distribution logistic was very challenging. consequently, the management team introduced procedures to alleviate the burden on partners and to improve product access. this was achieved by establishing additional distribution hubs for zika virus products obtainable from: the centre for molecular diagnostics and therapy (crie), moscow, russia; wuhan institute of virology (wiv), wuhan, southern china; national institute of infectious diseases (niid), tokyo, japan. the management team also negotiated a common material transfer agreement (mta), with industry on behalf of all consortium members and a centralised exchange logistics platform was established. this experience underlines the reality that anticipation and preparedness are key components for an effective response to public health emergencies. from december, through may, brazil reported its largest yellow fever (yf) outbreak in decades, with a total of suspected cases and deaths (fischer et al., ) , prompting widespread yellow fever virus (yfv) vaccination campaigns and the need to distinguish between vaccine-and wild-type yfv-associated disease. novel multiplex real-time reverse transcription pcrs that differentiate between vaccine and american wild-type yfv were developed by consortium partners in response to this outbreak and validated under field conditions (fischer et al., ) . centres for disease control (cdc) in countries play an important role for the control of virus outbreaks by the collecting data and samples during the identification phase of the pathogens. however the academic research combined with material collections appears to be an unavoidable actor for the preparedness and the control of an emerging virus disease, each time a variant form or a new form of a pathogen is responsible of it. combining, in the same organization, cdc and high calibre research laboratories was foreseen as the best solution to reach both an efficient control of the virus propagation and a preparedness for the next outbreak. at the same time, to cover the all range of pathogens from rg to rg was only made possible by the constitution of a group of high containment facilities within the organization. sixteen registered biosafety level- (bsl- ) laboratories, with the resources to handle the highest risk hg pathogens and five cdcs are partners or associate partners in the consortium. each cdc contributes directly to the management and control of emerging virus diseases, under the umbrella of the who. the evag is also associated with the global outbreak alert and response network (goarn) and is recognised by goarn as a supplier of high quality viruses and virusrelated reagents and resources. during the - ebola virus epidemics in west africa, all of the bsl- institutes were directly involved in helping local authorities to contain the epidemic. they all gained important experience in the management of such health crises, collecting field samples and isolated relevant virus strains many of which are now characterised and stored in the facilities of the evag bsl- partner at the bernard nocht institute in hamburg, germany. in addition, they addressed specific issues involving management of the exchange of high risk pathogens and activation of platforms dedicated to offering services to appropriate endusers. the consortium is now in its third year of existence under the h framework programme. its ultimate objective is to become a permanent archive that provides access to a very wide range of viruses and reagents globally. this will be achieved firstly, through extension of the funding arrangements, secondly through further increasing the range of contributors to the collection, and thirdly by bridging large european or international infrastructures having activities related to those of evag, including: it would be naive to suggest that the long-term survival of the consortium will be straightforward. for example, its perpetuation will require a continuous and increasingly large funding stream to meet the almost restrictive costs of research, laboratory development and upkeep in the face of newly emerging highly pathogenic viruses. sustainability is a key issue for such an infrastructure and the consortium plan will be to remain a non-profit organization. the obvious consequence of this fundamental choice is to have access to diversified sources of funding. the nations to which partners belong will have to support the collection activities including infrastructure management, as well as research concerning the development of new tools necessary to keep the archiving process up to date. the financial support for all the actions involved in preparedness for emerging viral diseases e.g., distribution of bio-resources to supply reagents to detect infected patients at local level will have to come from government health departments but at the international level, from the european commission, charitable international health funding agencies such as the wellcome trust foundation, the bill and melinda gates foundation, the who, the oie. future financial planning also includes the creation of a specific interface between the industrial sector and the consortium. in the first instance, this interface will explore bi-directional opportunities to organize scientific exchanges, priority access to bio-resources including reference material development of diagnostic services, validation of diagnostic assays and commercialization by industrial partners to the benefit of both parties. signed contracts of collaboration will be facilitated, and procedures harmonized. as noted above, the concept of the evag is unique. as far as we are aware, there is no equivalent viral archive, nor does any other collection provide the accessibility, reagent backup, sequence data, provenance, quality control, and capacity to advertise, inform, negotiate and conduct the entire transaction to the end-user via the web and internationally registered secure transportation companies the reputations, high quality, experience and knowledge of the consortium partners, combined with the integration of the european commission infrastructure, will provide endusers with opportunities to approach new fields of research in structural viral genomics, evolutionary biology, control of infectious diseases, antiviral drug design, fundamental research, public and environmental health, pathogenesis, immunology and a wide variety of associated disciplines. subsequently, subject to continuation of funding and the appropriate justification, the evag aims to extend the diversity of its disciplines to encompass fish, plant, bacterial, fungal, protist and other currently unrecognised viruses. this will be achieved first through expansion within the european community, followed by integration of specialist laboratories in the americas, russia, asia, southeast asia, australia and africa. the recently created african cdc will provide a unique opportunity to establish strong links with public health institutes in africa. the substance of this relationship will be based on trust and mutual benefit. for instance, training workshops and staff exchanges will be organized with the objective of developing appropriate infrastructures that can be directly integrated into the evag format and the levels of scientific expertise and quality and range of products that will justify their integration into evag as full partners within the consortium. the evag provides a range of viruses and relevant reagents to endusers engaged in a wide variety of established infrastructures. for instance, the newly created eu infrastructure, infravec (european infrastructures for insect disease vector research and control -https:// infravec .eu/) supplies infectious vectors (e.g. mosquitoes and ticks) to their end-users. the evag is closely associated with this activity, via the provision of viruses that can infect the vectors. erinha (http://www. erinha.eu/), a european strategy forum on research infrastructures (esfri www.esfri.eu/), offering access to high containment facilities to industrial end-users for their preclinical trials including bsl pathogens, recommends evag as the provider of the viruses necessary for their trials. bbmri, another esfri, plans to create a service unit dedicated to the provision of viruses and derived material from viruses. evag is identified as the recommended supplier. the evag contributes to the functioning of other eu infrastructures involved in related research topics including zoonotic diseases, drug discovery, vaccine development, viral epidemiology and emerging virus diseases. these eu-funded projects include prepare (platform for european preparedness against (re-) emerging epidemics -https://cordis. europa.eu/project/rcn/ _en.html), compare (collaborative management platform for detection and analyses of (re-) emerging and foodborne outbreaks in europe -http://www.compare-europe.eu/about), emerge (efficient response to highly dangerous and emerging pathogens at eu level -https://www.emerge.rki.eu/), zikalliance (a multinational and multidisciplinary research consortium coordinated by insermhttps://zikalliance.tghn.org/about/). as a future objective, one can imagine the generation of links among all of these organisations to constitute a "life science infrastructure cooperative" dedicated to virology-related topics, in which the evag could play a central role. for the long-term future, we are planning to establish closer collaborative links with other compatible scientific infrastructures by initiating teaching and training programmes amongst younger scientists to ensure that people of the right calibre will be able to undertake the roles currently occupied by experienced but "maturing" scientists. indeed, one could envisage e-learning programmes being built into diploma courses in universities. as we continue our ambitious mission to build a scientific infrastructure that can benefit scientists operating in all areas of the viral world, we recognise that other agencies and research teams can provide valuable datasets, and analytical expertise from which disease intervention strategies might be developed. indeed, other compatible agencies are approaching the consortium to share their resources and to promote their activities through the consortium website. this is yet another avenue that is being explored, based on the complementarities of these parties with the existing group of partners. at the time of writing, we are witnessing the massive impact of nextgeneration sequencing technology that will undoubtedly continue to evolve and provide even more discoveries, any one of which might then be exploitable by a future larger and more comprehensive form of agency. since the first recognition of viruses and our obsession with trying to understand and control them, it has been an exciting time for virologists. long may the evag and its successors ensure that this is always the case! this publication was supported by the european virus archive global (evag) project that has received funding from the european union's horizon research and innovation programme under grant agreement no . improved detection of zika virus rna in human and animal specimens by a novel, highly sensitive and specific real-time rt-pcr assay targeting the '-untranslated region of zika virus detection of a novel human coronavirus by real-time reverse-transcription polymerase chain reaction assay optimization for molecular detection of zika virus lineage-specific real-time rt-pcr for yellow fever virus outbreak surveillance mosquito co-infection with zika and chikungunya virus allows simultaneous transmission without affecting vector competence of aedes aegypti the european virus archive: a new resource for virology research a cynomolgus macaque model for crimean-congo haemorrhagic fever zika virus infection during pregnancy in mice causes placental damage and fetal demise the . a resolution cryo-em structure of zika virus laboratory testing for middle east respiratory syncytial coronavirus: interim guidance a crispr screen defines a signal peptide processing pathway required by flaviviruses we wish to thank the following scientists for their expertise and contributions during the successful establishment and development of supplementary data related to this article can be found at https:// doi.org/ . /j.antiviral. . . . key: cord- -hw aijwf authors: banyard, ashley c.; johnson, n.; voller, k.; hicks, d.; nunez, a.; hartley, m.; fooks, a. r. title: repeated detection of european bat lyssavirus type in dead bats found at a single roost site in the uk date: - - journal: arch virol doi: . /s - - - sha: doc_id: cord_uid: hw aijwf in august , european bat lyssavirus type (eblv- ) was isolated from a daubenton’s bat found at stokesay castle. in september , another bat from the same vicinity of stokesay castle also tested positive for eblv- . this is the first occurrence of repeated detection of eblv- from a single site. here, we report the detection of low levels of viral rna in various bat organs by qrt-pcr and detection of viral antigen by immunohistochemistry. we also report sequence data from both cases and compare data with those derived from other eblv- isolations in the uk. cases of european bat lyssaviruses type- and - (eblv) continue to occur across europe. eblv- seems to be restricted to the infection of serotine bats (eptesicus serotinus) [ ] , although it has been reported in 'spill-over' events into incidental hosts [ , , ] . in comparison to eblv- , eblv- has been reported on fewer occasions, having been isolated from both daubenton's bats (myotis daubentonii) and pond bats (myotis dasycneme) [ ] . the discovery of eblv- in a daubenton's bat in june in newhaven, east sussex, prompted concerns that bat rabies may be present within the uk and, furthermore, that the threat of rabies entering the uk via migratory bats was realistic [ ] . indeed, bat rabies cases in european bats in the mid s indicated a possible spread of the virus, especially in denmark and the netherlands. as well as having been isolated in the uk, eblv- isolates have also been reported in switzerland, holland and finland [ , ] . eblv- infection has also been observed in daubenton's bats in germany [ , ] . since , eblv- has been identified from several locations across the uk, suggesting that eblv- is endemic at a low level in british bats (table ) , and in august , a further uk isolate was identified in an adult female daubenton's bat found by a member of the public at stokesay castle, shropshire [ ] . the bat carcass was submitted to the veterinary laboratories agency (vla) for laboratory testing to confirm the presence of eblv- and was shown to be positive by the fluorescent antibody test (fat). confirmatory diagnosis was achieved by pcr using a hemi-nested rt-pcr that detected viral rna in brain, salivary gland and tongue samples (fig. a) . quantitative rt-pcr (qrt-pcr) was undertaken using previously described methods [ ] to determine levels of viral rna present within different organs of the infected animal ( fig. b) . high levels of viral rna were detected in the brain, with lower levels in the salivary glands, stomach and tongue, with rna also being detected in the intestine and heart (fig. c) . immunohistochemical analyses were restricted to sections of the spinal cord and detected virus antigen in dorsal root ganglia (fig. a) . the original observation of an eblv- positive bat at this site prompted measures to be taken to minimise the risk to members of the public encountering other bats from the roost. twice-daily checks were implemented to ensure that no injured, sick or dead bats were present in areas of the castle open to public access. importantly, control measures were implemented, including the wearing of protective gloves so that staff could handle a bat with minimal risk during the regular checks before members of the public entered the tower of the castle [ ] . signs were also erected at the entrance of the tower warning members of the public not to handle bats. in september , a dead bat was discovered on the top floor of the south tower of stokesay castle during one such check. the bat was a juvenile male daubenton's bat and was submitted to the vla for routine testing. this bat was also shown to be positive for infection with eblv- by rt-pcr, a -base-pair (bp) fragment of the nucleocapsid (n) gene being successfully amplified. unfortunately, the standard fat could not be undertaken, as the brain material had decomposed during storage and transit. the remainder of the carcass of this bat was, however, submitted for histopathological examination. several sections were taken, and despite advanced autolysis, specific labelling was found throughout the spinal cord in neurons, dorsal root ganglia (fig. b ) and peripheral nerves. the n-gene pcr product was sequenced in its entirety and found to be % identical to the isolate across the -bp region analysed. a phylogenetic analysis of the eblv- isolates across the uk to date was generated using the neighbour-joining method, using mega software (fig. d) . unfortunately, the carcass of this bat was severely decomposed, and further molecular tests, such as comparative qrt-pcr on a range of organs, could not be performed. the identification of eblv- positive bats from the same site more than year apart raises several questions regarding the basic transmission and biology of this virus within bat roosts. recent attempts to undertake scientific studies with bat species regarding the transmission and maintenance of these viruses between bats have resulted in limited conclusions as to how the virus is maintained within colonies. bite transmission seems the most likely route of transmission, although no direct evidence for this in captive bats infected with eblvs has been observed [ , , ] . the detection of high levels of virus antigen in the brain of infected bats is typical for these viruses, and generally, where eblv- has been detected in british bats, live virus has been isolated from brain material where samples have not decomposed. here, we have reported the detection of virus in other tissue types. presence of virus in these regions is likely to be linked to the degree of innervation, principally by the autonomic nervous system, although quantitation of neuronal involvement within different organs and tissue types to establish a basis for this hypothesis has not been undertaken [ , ] . however, studies with eblv- infection in the natural host, eptesicus serotinus, showed no substantial pattern of virus distribution in different non-neuronal organs in bats that developed disease [ ] . mechanisms of virus transmission within roosts remain an enigma. for genotype lyssaviruses it has been established that infection of the salivary glands may lead to the secretion of virus in the saliva for several days before the onset of clinical signs. whilst this is widely documented for larger species, low levels of viable virus or viral rna detected in saliva swabs tested during experimental studies with different bat lyssaviruses highlight the difficulty in determining the importance of this route of transmission for virus dissemination within a roost [ , , ] . aerosol spread of virus within a roost would seem feasible, as bats live in very close proximity. however, to date, transmission studies have only been reported with genotype rabies viruses, experimental attempts to transmit eblv- via this route proving unsuccessful [ ] . for the stokesay castle case, the detection of eblv- rna in tongue lends support to the transmission through bite or grooming, although again, this has not been conclusively shown. in , advanced autolysis prevented thorough histological examination or molecular assessment of the tongue. granular immunolabelling was, however, seen in nerves at the base of the tongue and along the jaw. unfortunately, no taste buds or epithelial tissue was available for further testing. the mechanisms of maintenance of eblvs within bat roosts and transmission between individuals remains unknown, although the 'small vector hypothesis' remains plausible [ ] . the restriction of eblv infection to certain species of bat as well as mechanisms by which individuals are able to survive infection, or at least exposure to the virus, are key questions that remain [ ] . viral load, bite/ exposure site and immunological competence of the exposed animal may all affect the outcome of infection. factors such as seasonal variation, pregnancy, nutritional status and immune status must also play an important part in dictating whether or not bats succumb to infection [ ] , but currently, our understanding of bat biology and immunology is low. the detection of a large number of viruses within different bat species has highlighted this lack of knowledge [ ] . isolation of a number of zoonotic viruses in bat species, including coronaviruses, astroviruses, henipaviruses and other lyssaviruses, will surely drive further scientific investigation [ , , , ] . clearly, (d) fig. a hemi-nested second-round pcr results for bat / . firstround pcr (primers jw and ) produced a negative result for all three bat samples tested. however, the second-round pcr (primers jw and ) produced a positive result for the brain and salivary gland [ ] ( brain, salivary gland, tongue, negative control mouse brain, rt positive control, rt negative control, pcr positive control, pcr negative control, pcr negative control for second-round reaction. with eblv- having now been identified on two separate occasions from the same roost, virus is being maintained and transmitted from bat-to-bat by some as yet undefined mechanism. the status of daubenton's bats, and indeed all bats across the uk, as protected species makes it difficult to undertake investigative studies at such sites. however, serosurveillance of bats are planned at the stokesay castle site that will allow determination of the level of seroconversion within this roost and highlight possible transmission mechanisms that will help understand the transmission biology of these elusive viruses and may identify those parameters needed to enhance strategies to combat neuroinvasion and subsequent disease development. bats: important reservoir hosts of emerging viruses european bat lyssavirus type : human exposure in england european bat lyssaviruses: an emerging zoonosis detection of high levels of european bat lyssavirus type- viral rna in the thyroid gland of experimentally infected eptesicus fuscus bats susceptibility of north american big brown bats (eptesicus fuscus) to infection with european bat lyssavirus type first isolation of eblv- in germany experimental infection of serotine bats (eptesicus serotinus) with european bat lyssavirus type a (eblv- a) isolation of european bat lyssavirus type from a daubenton's bat (myotis daubentonii) in shropshire antibodies against lagos bat virus in megachiroptera from west africa heminested pcr assay for detection of six genotypes of rabies and rabies-related viruses experimental infection of big brown bats (eptesicus fuscus) with eurasian bat lyssaviruses aravan, khujand, and irkut virus isolation of a european bat lyssavirus type from a daubenton's bat in the united kingdom airbourne transmission of lyssaviruses experimental study of european bat lyssavirus type- infection in daubenton's bats (myotis daubentonii) experimental infection of big brown bats (eptesicus fuscus) with west caucasian bat virus (wcbv) human rabies of bat origin in europe emerging pattern of rabies deaths and increased viral infectivity spill-over of european bat lyssavirus type into a stone marten (martes foina) in germany epidemiology of bat rabies in germany bat lyssaviruses in europe studies on the pathogenesis of rabies in insectivorous bats. iii. influence of the gravid state rabies in individual countries-denmark prevalence and genetic diversity of coronaviruses in bats from china natural and experimental infection of sheep with european bat lyssavirus type- of danish bat origin european bat lyssaviruses european bat lyssaviruses-an ecological enigma development of a real-time, differential rt-pcr taqman assay for lyssavirus genotypes , and first isolation of a rabies-related virus from a daubenton's bat in the united kingdom detection of diverse astroviruses from bats in china acknowledgments we wish to acknowledge denise marston for technical assistance. this work was supported by defra grants se and sev . key: cord- - bo s hz authors: lezotre, pierre-louis title: part i state of play and review of major cooperation initiatives date: - - journal: international cooperation, convergence and harmonization of pharmaceutical regulations doi: . /b - - - - . - sha: doc_id: cord_uid: bo s hz abstract the basic principle of international cooperation is to establish bilateral and multilateral efforts to leverage the human, scientific and financial resources and the knowledge and experience of other key regulatory authorities to avoid duplication of efforts, to make activities more efficient and to allow the focussing of limited resources on higher-risk areas of concern. this increased cooperation between worldwide regulators has necessitated proactive deliberate efforts towards convergence/harmonisation of regulation, practices and requirements to eliminate or reduce differences. cooperation and harmonisation of standards in the pharmaceutical domain are already a reality and have become increasingly important during recent decades, with a high level of commitment to these activities by all stakeholders. the worldwide drug regulatory authorities (dras) have been working to end an isolationist attitude that cannot resolve current worldwide issues and challenges caused by an ever increasing globalisation. as a result, many cooperation and harmonisation initiatives have been established at the bilateral, regional and global levels as a response to the changing geo-economic-political situation. the spectrum of collaboration varies from simple informal technical cooperation to full integration of systems and regulations. indeed, all these initiatives can be very different in scope (some are part of a broader harmonisation initiative), level of harmonisation (depending on the political support/commitment), organisation (well-structured versus simple discussion) or advancement (established process vs. pilot projects), but they all work towards convergence of requirements and/or practices. all these multiple worldwide cooperation and harmonisation programmes have evolved rapidly over the past decades. this book section provides the current status of this complex and broad phenomenon of cooperation, convergence and harmonisation in the pharmaceutical sector. it reviews all major global, regional and bilateral cooperation initiatives. many aspects of increased globalization also have profound implications on pharmaceutical regulation worldwide. in general, globalization of the economy (with increased travel of people and exchange of goods, finance, and information), and also globalization of the pharmaceutical market (including development, manufacture, and distribution activities), requires increased cooperation and harmonization of pharmaceutical standards and regulation. pharmaceutical industries have asked for better harmonization of requirements for the development and manufacture of pharmaceutical products to avoid duplication of work that ultimately creates delays in drug availability [ ] . in this context, harmonization of pharmaceutical regulations has naturally become an important topic of discussion among worldwide drug regulatory authorities (dras). over the past several decades, they have been working to end an isolationist attitude that cannot resolve current worldwide issues and challenges. as a result, many cooperative initiatives (bilateral, regional, and global) were established, and harmonization efforts have been enhanced. all these initiatives can be very different in scope (some are part of a broader harmonization initiative), level of harmonization (depending on the political support/commitment), organization (well structured versus simple discussion), or advancement (established process versus pilot projects), but they all work towards harmonization of requirements and/or practices. increased exchange of information on a regular basis (e.g., more than countries and international organizations from australia to vietnam now have agreements to share information with the united states food and drug administration [us fda]) [ ] also contributes to the natural convergence of requirements and practices. harmonization models can be distinguished by their scope and objectives. indeed, the spectrum of collaborations varies from simple technical cooperation to full integration of systems and regulations: ▸ integration model: in this type of agreement, most of the time driven by political decision, deeper harmonization of regulation is achieved with the creation of supranational central authorities in order to support integration and/or creation of a single market (e.g., eu, the association of southeast asian nations [asean] ). in this case, harmonization of standards and regulations is critical in reducing trade barriers. in this model, countries give up some of their autonomy on certain matters by transferring the power to make decisions to the common supranational authority or by automatically recognizing decisions from the other party (via mutual agreement mechanisms). the african medicines registration harmonization (amrh) initiative has defined five identifiable levels of harmonization ( figure ). to facilitate cooperation, a mutual recognition agreement (or arrangement) (mra) can be signed by one or more parties to mutually recognize or accept some or all aspects of one another's requirements. they can be concluded at the technical level (e.g., the status and future plans," november . confidentiality arrangements between the us fda and european medicines agency [ema] , or the mra between eu and australia) or at the government level (e.g., european treaty). these multilateral initiatives are major projects as they involve multiple organizations and countries and represent the highest degree of harmonization. the objective of this technical and scientific intergovernmental cooperation is to globally discuss scientific issues that support the decisions made by individual governments and international regulatory bodies in order to achieve global scientific consensus. the goal is to facilitate the development of new medicines and to make them available to the maximum number of people worldwide. there is no intent of full integration of systems and regulations. the main difficulty faced by these initiatives is the complexity and management of the structure due to the important number of participants (e.g., the world health organization [who] has member states) and the diversity of needs, challenges, and level of development of its members. the world health organization (who) was established in as a specialized agency of the united nations (un) [ ] . it is accountable to its member states and works closely with other entities of the un system. this agency has a very broad scope of responsibilities as it is the directing and coordinating authority for international health matters and public health within the un system. who is well known for some of its work (e.g., the coordination of influenza surveillance and monitoring activities, emergency assistance to people affected by disasters, mass immunization campaigns or actions against human immunodeficiency virus/acquired immunodeficiency syndrome [hiv/aids], tuberculosis, and malaria). however, who undertakes many more activities because it is responsible for providing leadership on global health matters, shaping the health research agenda, setting norms and standards, articulating evidence-based policy options, providing technical support to countries, and monitoring and assessing health trends. most of these core functions, as further defined in its " th general programme of work," [ ] rely on cooperation and harmonization of standards. this focus on regional and global collaboration, and especially aid from developed countries to developing countries, is aligned with the un millennium development goals (mdgs). a a the united nations millennium development goals (mdgs) are eight international goals that un member states (and international organizations) have agreed to achieve by the year . they are derived from the united nations millennium declaration, signed in september , which endorsed a framework for development and commits world leaders to combat poverty, hunger, disease, illiteracy, environmental degradation, and discrimination against women. these mdgs are interdependent and several relate either directly or indirectly to health. who is therefore very involved in this process and works with countries to achieve the health-related mdgs. indeed, the objective of these mdgs is that countries and development partners work together to improve the global situation and resolve major issues. a number of specific targets and indicators have been identified to monitor progress towards the goals. goal ("develop a global partnership for development") specifically recognizes the role of developed nations and addresses global cooperation and partnerships. who has worked in the area of pharmaceuticals since its creation approximately years ago. during this time, many products and services have been created that are widely recognized as core functions of who. the role of who in pharmaceutical regulations is based on its constitutional mandate and various world health assembly (wha) resolutions. this support is twofold. one aspect relates to the development of internationally recognized norms, standards, and guidelines. the second relates to providing guidance, technical assistance, and training in order to enable countries to implement global guidelines to meet their specific medicines regulatory environment and needs [ ] . all countries that are members of the un may become members of who by accepting its constitution. other countries may be admitted as members when their application has been approved by a simple majority vote of the world health assembly (wha). territories that are not responsible for the conduct of their international relations may be admitted as associate members upon application made on their behalf by the member or other authority responsible for their international relations. members of who are grouped according to regional distribution. who's strength lies in its neutral status and nearly universal membership. today, it represents countries and two associate members (puerto rico and tokelau). one country is an observer (vatican) [ , ] . the organization is headed by the director-general, b but the wha is the supreme decisionmaking body for who. it generally meets in geneva, switzerland in may of each year, and is attended by delegations from all member states. its main function is to determine the policies of the organization. the health assembly also appoints the director-general (on the nomination of the executive board), supervises the financial policies of the organization, and reviews and approves the proposed budget. the work of the assembly is supported by the executive board, which it elects. this executive arm of the assembly is composed of members technically qualified in the health field. members are elected for three-year terms. the main board meeting, at which the agenda for the forthcoming health assembly is agreed upon and resolutions for forwarding to the health assembly are adopted, is held in january. a second shorter meeting in may, immediately after the health assembly, is held to address more administrative matters. the primary functions of the board are to give effect to the decisions and policies of the health assembly, to advise it, and generally to facilitate its work. under the leadership of the director-general, c more than , people from more than countries work for who. this who staff includes health professionals (including medical doctors, public health specialists, epidemiologists, and scientists) as well as managers, economists, administrators, and other professionals. they are located in country offices, six regional offices, and at the headquarters in geneva, switzerland [ ] . one of the unique aspects of who is its decentralized structure. who's work is a great combination of actions at the country, regional, and global levels. these efforts to decentralize its structure are aimed at getting closer to the ground (field) where decisions made can be more responsive to actual needs. indeed, this decentralized and regionalized structure provides who with multiple opportunities for engaging with countries. who's global headquarters is located in geneva, switzerland. the team based at the global headquarters supports and builds on all of the regional and local efforts. it sets global policies and standards, facilitates technical support to regions and countries, monitors and publicizes progress, and helps mobilize political and financial support. at the who headquarters, medicine activities are conducted within the cluster of health systems and services (hss) and are coordinated by the department of essential medicines and health products (emp). this department (which employs about staff members [ ] ) is involved in the harmonization of pharmaceutical regulations because it coordinates various activities in the areas of quality assurance (e.g., the international pharmacopoeia, international nonproprietary names [inn] , prequalification of medicines, counterfeit medicines), regulation and legislation (e.g., international conference of drug regulatory authorities [icdras]), and safety and efficacy (e.g., drug alerts). these activities comprise guideline development, workshops, and training courses, coordination and promotion of pharmacovigilance for global medicine safety, regulatory and other information exchange, and review of narcotic and psychotropic substances. who member states are grouped into six regions, each of them having a regional office: ▸ who regional office for africa in brazzaville, republic of congo. ▸ who regional office for europe in copenhagen, denmark. ▸ who regional office for southeast asia in new delhi, india. ▸ who regional office for the americas/pan american health organization (paho) in washington dc, united states. ▸ who regional office for the eastern mediterranean in cairo, egypt. ▸ who regional office for the western pacific in manila, the philippines. each of who's regional offices are the first point of contact for country offices that need extra technical or financial help. these regional offices also give special attention to adapting global policies to fit specific needs in their regions. indeed, the regional level is important in the who organization as it links the global strategy and plan with the country's reality and needs. they play a key role in the implementation of who norms and standards by ensuring that: ▸ country and regional needs are taken into consideration when who norms and standards are developed ▸ global guidelines and internationally recognized norms and standards are appropriately implemented in their regions (in the context of their own specific regulatory environment and challenges) by providing guidance, technical assistance, and training in addition to global activities coordinated from who headquarters, who regional and country offices can also carry out a variety of medicine-related activities specific to their regions. in addition to the regional and headquarters offices, who has country offices that cover member states. d there are also two field offices (the who humanitarian assistance office in pristina, kosovo and the west bank and gaza office) and offices covering two different areas, the us-mexican border field office in el paso, texas (us), and the office of caribbean program coordination in barbados. who has also established "who liaison offices" in key locations (e.g., at the european union in brussels, belgium, at the african union and the economic commission for africa in addis ababa, ethiopia, in washington dc, us, and at the un in new york city) and more than "technical offices" (e.g., the european observatory on health systems and policies in berlin, germany) [ ] . d some countries that do not have a physical who country office are served by the who representative of another country (for instance, the who representative to malaysia covers not just malaysia, but also brunei, darussalam, and singapore) . approximately % of who country offices are either owned or supported by the government and ministries of health. some of these who country offices are located in independent premises either rented or owned by who, while others are located within ministries of health or un common premises. these country offices are led by the head of who office (hwo), who are designated by the director-general and by the respective regional directors. the hwo manages who core functions at the country level and provides leadership in the following key functional areas: ▸ advocacy, partnership, and representation ▸ support for policy development and technical cooperation ▸ administration and management it is important to note that who is focused on needs of countries and emphasizes in particular the decentralization process that is aimed at increasing who's impact on health and development at the country level. this country focus tailors who's technical collaboration to the needs and capacities of each member state, with a special emphasis on the poorest countries and most fragile contexts. the key principles guiding who cooperation in countries are [ ] : ▸ ownership of the development process and projects by the country ▸ alignment with national priorities and strengthening national systems ▸ harmonization with the work of sister un agencies and other partners in the country towards better aid effectiveness ▸ collaboration as a two-way process that fosters member states' contributions to the global health agenda who's country presence is the platform for effective cooperation with countries for advancing the global agenda, contributing to national health strategies and planning, and bringing country realities and perspectives into global policies and priorities. according to the above principles and its structure, who is indeed able to focus on countries' needs and better define its priorities to actively support the development, implementation, monitoring, and assessment of national health policies, strategies, and plans. but it also allows for better monitoring implementation of global agreements such as the millennium development goals (mdgs) and the international health regulations (ihr [ ] ). these activities in countries are governed by the country cooperation strategy (ccs), which is who's key instrument to guide its work in countries. it is a medium-term vision (generally covering four to six years) for its technical cooperation with a given member state, in support of the country's national health policy, strategy, or plan. it is an organization-wide reference that guides partnership, planning, budgeting, and resource allocation. who also established the department of country focus (cco) to support and advocate for who country offices, develop the capacity of who country teams for effective engagement in partnership platforms, and facilitate and monitor who's engagement in the aid effectiveness agenda at the country level. for example, cco provides support for the development, dissemination, and use of the country cooperation strategy. ▸ expert committees: expert committees have an important role in who activities. they are defined in the who constitution. e in addition to the constitution, regulations for expert advisory panels and committees are also included in the who document entitled "regulations for expert advisory panels and committees." f an expert committee is the highest official advisory body to the director-general of who as well as to all the organization's member states. it is established by the wha or by an executive board decision. there are various types of who expert committees. for example, the who expert committee on specifications for pharmaceutical preparations (ecspp) has been providing, for more than years, recommendations and tools to assure the quality of medicines from their development phase to their final distribution to patients. there is also the expert committee on biological standardization (ecbs), which is as old as the ecspp. in addition to its structured organization, the who has been supported since its creation by its "collaborating centers." these are institutions such as research institutes and parts of universities or academies that are designated by the director-general to carry out activities in support of who programs. currently there are over who collaborating centers in over member states working with who in several areas (one of them being "pharmaceuticals"). several collaborating centers may exist for the same topic (e.g., international classifications or traditional medicines) and form a specific network to help who regarding this specific topic. of causes of death. who also started to publish its bulletin, which is today an international peer-reviewed monthly journal of public health with a special focus on developing countries. j in its early years, who's priority was the prevention and control of specific diseases (e.g., malaria, tuberculosis, smallpox, yaws, onchocerciasis, and venereal disease), some of which are still a problem today. they also focused on women's and children's health and nutrition, and environmental sanitation. who's work has since grown to cover other (sometimes new) health problems (including polio, hiv/aids, and severe acute respiratory syndrome [sars] ), but it also works to control tobacco and alcohol use and to promote diet and physical activity to prevent the four main noncommunicable diseases (cardiovascular disease, cancer, chronic lung diseases, and diabetes) [ ] . who has also been increasingly involved in the global regulation and control of medicines. in , the first essential medicines list was released two years after the wha introduced the concepts of "essential drugs" and "national drug policy." one hundred and fifty-six countries today have a national list of essential medicines. who has also funded many projects over the years to facilitate global cooperation and harmonization of standards. the purpose of all these activities in the pharmaceutical domain is aimed at increasing global and equitable access to safe, effective medicines of assured quality. this specific goal is derived from the overall objective of who to improve and maintain global public health. this objective has been regularly reiterated in several wha resolutions and during other events such as the icdras. in , the international conference on primary health care (alma-ata, kazakhstan) set the historic goal of "health for all," to which who continues to aspire. more recently, the un mdgs have further clarified the objectives and priorities of global cooperation derived from the un millennium declaration signed in september . one of who's mandates is "to develop, establish and promote international standards with respect to food, biological, pharmaceutical and similar products" [ ] . who member states (especially developing countries) rely on who for expertise and guidance in regulation, safety, and quality assurance of medicines through development and promotion of international norms, standards, guidelines, and nomenclature. to achieve this goal, who relies on cooperation and uses its decentralized organization to facilitate implementation of projects and agreed-upon standards. the harmonization activities are initiated according to the who's medicines strategy. trigger actions to initiate a new project or development of a standard are given at different levels and bodies (i.e., the wha, executive board resolutions, icdras, or who programs and j since it was first published in , the bulletin has become one of the world's leading public health journals. as the flagship periodical of who, the bulletin draws on both who experts (as editorial advisors, reviewers, and authors) and external collaborators. clusters). these projects and standards are then developed through a vast global consultation process involving who member states, national and regional authorities, international agencies, and with specialists from industry, national institutions, nongovernmental organizations, etc. project updates and approved standards become publically available through the extensive list of who publications to support national, regional, and global health strategies. k because the global dissemination and exchange of information is important, who secures the broad international distribution of its publications and encourages their translation. l this ensures the widest possible availability of authoritative information and guidance on health matters. the department of emp, based at the who global headquarters in geneva, works closely with expert committees, other regulators, and relevant who collaborating centers to develop and implement these harmonization activities. this department coordinates these activities globally with the support of who's regional advisors and country project staff in each of the regional offices and many country offices. each of the regional offices has two to five professionals coordinating the medicines strategy, and who country offices have full-time pharmaceutical policy experts [ ] . it is worth mentioning that in addition to its normative activities and harmonization projects, who also assists countries in capacity building by assessing regulatory systems. it does this by facilitating cooperation and information exchange between countries and by providing technical support. it is very important to involve all countries (whatever their development level), and to facilitate the implementation of norms and standards. finally, who has developed relationships with a lot of nongovernmental and civil society organizations on a global basis via the civil society initiative (csi) , and also at regional and national levels. the objectives of who's relations with nongovernmental organizations (ngos) are to promote the policies, strategies, and activities of who to facilitate their implementation. who has a large repertoire of global normative work relevant for all levels of development. in the area of medicines, a lot of standards, norms, and classifications have been developed, and forums/networks have been created to enhance global cooperation. important initiatives are presented below. k who publishes practical manuals, handbooks, and training material; internationally applicable guidelines and standards; reviews and analyses of health policies, programs, and research; and state-of-the-art consensus reports that offer technical advice and recommendations for decision makers. also, the who technical report series makes available the findings of various international groups of experts that provide who with the latest scientific and technical advice on a broad range of medical and public health subjects. l in , the world health assembly turned multilingualism into a who policy by establishing six official languages (arabic, chinese, english, french, russian, and spanish) . since the adoption of a resolution, all governing bodies' documents and corporate materials have been made available online in all official languages. the international conference of drug regulatory authorities (icdras) provides drug regulatory authorities of who member states with a forum to meet and discuss ways to strengthen collaboration and harmonization of pharmaceutical regulations. this is a key accomplishment of who that has been instrumental in guiding dras, who, and interested stakeholders to develop national, regional, and international medicines regulation, and it continues to be a cornerstone of international harmonization of medicines regulation. these conferences have been held since , and they have involved both developed and developing countries. the th icdras, held in singapore from november to december , , involved participants from over agencies. the th icdras, which took place in tallinn, estonia from october to , , was attended by over participants from countries. the aim of these conferences is to promote the exchange of information and collaborative approaches to issues of common concern. topics discussed include quality issues, herbal medicines, homeopathy, regulatory reform, medicine safety, counterfeiting, regulation of clinical trials, harmonization, new technologies, and e-commerce. recommendations are proposed for actions to take among agencies, who, and related institutions. it is worth mentioning that the idea to create ich began to formulate after background discussions between the us, the european union (eu), and japan during the th icdras conference in paris, france in [ ] . as a platform was established to develop international consensus, the icdras continues to be an important tool for who and dras in their efforts to harmonize regulation and improve the safety, efficacy, and quality of medicines on a worldwide basis. the who constitution mandates the production of international classifications on health. these internationally endorsed classifications, developed through the who network m are very important as they facilitate the storage, retrieval, analysis, interpretation, and comparison of data. they support global cooperation and harmonization by providing a consensual framework that governments, healthcare providers, and consumers can use as a common language. they also permit the comparison of data not only within populations over time, but also between populations. who reference classifications are the international classification of diseases (icd), the international classification of functioning, disability and health (icf), and the international classification of health interventions (ichi). in addition, related and derived classifications (based on the reference classifications) have also been developed (e.g., the anatomical therapeutic chemical classification with defined daily doses (atc/ddd) that classifies m who has designated a number of collaborating centers to work with it in the development, dissemination, maintenance, and use of the who international classifications. therapeutic drugs according to the organ/system on which they act, and their chemical, pharmacological, and therapeutic properties). the who international clinical trials registry platform (ictrp) is a global initiative that aims to make information about all worldwide clinical trials involving humans publicly available. this activity was launched during the th wha in n following discussions and recommendations from a ministerial summit on health research in mexico city, mexico in november . the ictrp is not itself a clinical trials registry, but a central repository that can be searched using the who search portal (http://apps.who.int/trialsearch/). all items in the trials registration data set are copied from individual registries onto the who central repository, and data is updated regularly. indeed, details on clinical trials come directly from one of the primary registries o in the who registry network (e.g., the european clinical trials register that became a member of the network in september p ). by consolidating clinical trials information from several worldwide sources using standardized data set format/criteria, and by implementing unambiguous identification (i.e., a universal trial number [utn] ), the ictrp not only facilitates the exchange of information, but also promotes harmonization of this information. harmonization is also further achieved because who proactively supports countries/regions in establishing who-compliant clinical trials registries or policies on trial registration. quality assurance is a wide-ranging concept covering all matters that individually or collectively influence the quality of a product. this is a major public health challenge, particularly in light of growing cross-border health issues and the growing international dimensions of trade. the quality of pharmaceuticals has been a concern of who since its inception. the development of norms, standards, and guidelines to promote quality assurance is an integral part of who's constitution, and has been endorsed and supported through numerous wha resolutions. more recently, the who medium-term strategic plan for - requested that the organization develop international standards, recommendations, and instruments to assure the quality of medicines, whether produced and traded nationally or internationally. n resolution wha . called on the global scientific community, international partners, the private sector, civil society, and other relevant stakeholders to "establish a voluntary platform to link clinical trials registers in order to ensure a single point of access and the unambiguous identification of trials with a view to enhancing access to information by patients, families, patient groups and others." o a primary registry in the who registry network is a clinical trial registry with at least a national remit that meets who registry criteria for content, quality and validity, accessibility, unique identification, technical capacity, and governance and administration. p the european clinical trials register provides public access to information extracted from the eu clinical trial database ("eudract"). the who medicines quality assurance program, which is part of the emp department, produces norms, standards, and guidelines on the quality assurance of pharmaceuticals. these regulatory tools are prepared through a vast global consultative process, and are ultimately approved by the who ecspp, q which meets annually. the report of each meeting (technical report series) includes newly adopted guidelines in its annexes. when adopted, the norms, standards, and guidelines become international harmonized standards intended for use by national dras, manufacturers, and other interested parties. many important international standards and projects have been developed in this area: ▸ good manufacturing practice (gmp) ▸ guidelines for regulatory approval (e.g., the guidelines on stability testing or on registration requirements to establish the interchangeability of multisource generic pharmaceutical products and the proposal to waive in vivo bioequivalence requirements) ▸ prequalification of medicines, laboratories, and supply agencies ▸ model certificates for quality assurance-related activities ▸ quality control testing ▸ new specifications for inclusion in the basic tests series and the international pharmacopoeia ▸ international chemical reference substances (icrs) r ▸ the inn program some of these international guidelines and projects are further developed below. ▸ good manufacturing practice: good manufacturing practice (gmp) is the part of quality assurance that ensures products are consistently produced and controlled to the quality standards appropriate to their intended use and as required by the marketing authorization. gmp is aimed primarily at diminishing the risks involved in any pharmaceutical production that cannot be eliminated through testing of the final product. s gmp covers all aspects of production: from the starting materials, premises, and equipment, to the training and personal hygiene of staff. detailed, written procedures are essential for each process that could affect the quality of the finished product. panel on the international pharmacopoeia and pharmaceutical preparations. r icrs are used by laboratories to test pharmaceuticals for the purpose of quality control. these substances are mainly used for validating the results from specific tests, and as primary standards for calibrating secondary standards. who's collection of icrs is now maintained by the council of europe's european directorate for quality of medicines and healthcare (edqm) , which also distributes the substances worldwide. edqm is responsible for obtaining candidate material, testing it to ensure its purity and suitability, and reporting results with recommendations to who. s the main risks are the following: unexpected contamination of products causing damage to health or even death; incorrect labels on containers, which could mean that patients receive the wrong medicine; and insufficient or too much active ingredient resulting in ineffective treatment or adverse effects. recognizing the importance of gmp in international commerce of pharmaceutical products, who developed requirements early on. the first who draft text on gmp was prepared in by a group of consultants at the request of the th wha [ ] . it was subsequently submitted to the st wha under the title "draft requirements for good manufacturing practice in the manufacture and quality control of medicines and pharmaceutical specialties" and was accepted. in , the revised text was discussed by the who ecspp and published as an annex to its nd report. the text was then reproduced, with some revisions, in in the supplement to the nd edition of the international pharmacopoeia (ph. int.). since then, who has further defined its general principles and requirements regarding gmp [ ] , and it has also established several detailed guidelines covering specific needs for gmp of active pharmaceutical ingredients [ ] , pharmaceutical excipients [ ] , sterile pharmaceutical products [ ] , biological products [ ] , blood establishments [ ] , pharmaceutical products containing hazardous substances [ ] , investigational pharmaceutical products for clinical trials in humans [ ] , herbal medicinal products [ ] , radiopharmaceutical products [ ] , and water for pharmaceutical use [ ] . finally, it also developed guidelines of a more general scope such as validation [ ] , risk analysis [ ] , technology transfer [ ] , and inspection [ ] , and has created appropriate training materials for countries. many countries have formulated their own requirements for gmp based on the who gmp. the international pharmacopoeia (ph. int.) comprises a collection of quality specifications for pharmaceutical substances (i.e., active ingredients and excipients) and dosage forms together with supporting general methods of analysis. it is intended to serve as source material for reference or adaptation by any who member state. clearly defined steps are followed in the development of new monographs. the ph. int. is published by who with the goal of achieving a wide global harmonization of quality specifications for selected pharmaceutical products, excipients, and dosage forms. the ph. int., or any part of it, has legal status whenever a national or regional authority expressly introduces it into appropriate legislation. the history of the ph. int. dates back to when the need to standardize terminology and to specify dosages and composition of drugs led to attempts to produce an international pharmacopoeia compendium. the first conference, called by the belgian government and held in brussels in , resulted in an agreement for the unification of the formulae of potent drugs, which was ratified in by countries. the outcome considerably influenced the subsequent publication of national pharmacopoeias. in , the interim commission of the who took over the work on pharmacopoeias previously undertaken by the health organization of the league of nations. the rd wha, held in may , formally approved the publication of the "pharmacopoea internationalis" and recommended, in accordance with article of the who constitution, "the eventual inclusion of its provisions by the authorities responsible for the pharmacopoeias." it was thus recommended that the "pharmacopoea internationalis" not be used as a legal pharmacopoeia in any country unless adopted by the pharmacopoeial authority of that country. this first edition, published with the aim of creating a worldwide, unified pharmacopoeia, relied on collaboration with national pharmacopoeia commissions for its preparation. in , the purpose of the ph. int. was reconsidered. it was decided that the publication should focus more on the needs of developing countries (because developed countries had established their own pharmacopoeias), and recommended only simple, classical chemical techniques that had been shown to be sound. since , the drugs appearing in the ph. int. have therefore been selected from the list of essential drugs based on the first report of the who expert committee on the selection of essential drugs. also, whenever possible, classical procedures are used in the analytical methods so that the use of expensive equipment is minimized in the application of the ph. int. to facilitate its implementation by developing countries. the work on the ph. int. is carried out by the who ecspp in collaboration with members of the who expert advisory panel on the international pharmacopoeia and pharmaceutical preparations and other specialists [ ] . the process involves consultation with, and input from, who member states and dras, who collaborating centers and national drug quality control laboratories in all six who regions, standard-setting organizations and parties including regional and national pharmacopoeias, and manufacturers around the world. in , the wha adopted a resolution [ ] to create the international nonproprietary names (inn) program in order to identify pharmaceutical substances unambiguously on a worldwide basis, and to provide a universal, unique, nonproprietary name to be used in pharmacopoeia monographs. it began operating in when the first list of inns for pharmaceutical substances was published. today, this program is coordinated by the who emp department. the selection of a new inn relies on a strict procedure [ , ] . this process is supported by the expert advisory panel on the international pharmacopoeia and pharmaceutical preparations, which provides advice on proposed names following an application made by the manufacturer or inventor. the procedure also involves the who secretariat, which examines the suggested names for conformity with the general rules, similarities with published inns, and potential conflicts with existing names. after a time period for objections has lapsed, the name will obtain the status of a recommended inn and will be published as such in "who drug information" if no objection has been raised. to make inns universally available, they are formally placed by who in the public domain, hence their designation as "nonproprietary" names (also known as "generic names"). the existence of this international nomenclature for pharmaceutical substances is important for the clear identification, safe prescription, and dispensing of medicines to patients, but also for communication and exchange of information among health professionals and scientists and regulators worldwide. it provides them with a unique and universally available designated name to identify each pharmaceutical substance. today, inn names are widely used and globally recognized. at present, more than , inns have been published, and this number is growing every year. the majority of pharmaceutical substances used in medical practice are designated by an inn, and their use is already common in research and clinical documentation. nonproprietary names are intended for use in pharmacopoeias, labeling, product information, advertising and other promotional material, drug regulation and scientific literature, and as a basis for product names (e.g., for generics). also inn collaborates closely with numerous national drug nomenclature bodies. the use of inn names is normally required by national authorities and also by the european community. as a result of ongoing collaboration, national names such as british approved names (ban), dénominations communes françaises (dcf), japanese adopted names (jan), and united states adopted names (usan) are nowadays, with rare exceptions, identical to the inn. in addition to the quality standards, who also developed norms and standards for pharmacovigilance, and promotes information exchange on medicine safety. the aim is to assure the safety of medicines by ensuring reliable and timely exchange of information on drug safety issues, promoting pharmacovigilance activities on an international basis, and encouraging participation in the who program for international drug monitoring [ ]. in , who established its program for international drug monitoring in response to the thalidomide disaster in . at the end of , countries were part of the who pharmacovigilance program. an international system for monitoring adverse drug reactions (adrs) using information derived from member states was established in . this allows who to issue a rapid drug alert whenever a serious problem in the safety of any medicinal product arises. who headquarters in geneva is responsible for policy issues, while the operational responsibility for the program rests with the who collaborating centre for international drug monitoring, uppsala monitoring centre in sweden. a common reporting form was developed, agreedupon guidelines for entering information were formulated, common terminologies and classifications were prepared, and compatible systems for transmitting, storing and retrieving, and disseminating data were created. the adrs database in uppsala currently contains over three million reports of suspected adrs. in , a who advisory committee on safety of medicinal products (acsomp) was established to guide who on general and specific issues related to pharmacovigilance. additionally, a network of "information officers" was established in to allow a direct relationship between who and all national dras in member states. each national information officer is charged with providing information to who on the safety and efficacy of pharmaceutical preparations, and with securing prompt transmission to national health authorities regarding new information on serious adverse effects. this certification scheme was initially adopted by the nd wha in [ ], but since then it has been amended. it is an administrative instrument that requires each participating member state, upon application by a commercially interested party, to attest to the competent authority of another participating member state whereby: ▸ a specific product is authorized for placement on the market within its jurisdiction, or if it is not authorized, the reason why that authorization has not been accorded. ▸ the manufacturing plant in which it is produced is subject to inspections at suitable intervals to establish that the manufacturer conforms to gmp as recommended by who. ▸ all submitted product information, including labeling, is currently authorized in the certifying country. the primary document delivered under this scheme is the certificate of pharmaceutical product (cpp), but two other documents can be requested within the scope of the scheme. the first is a statement of licensing status of pharmaceutical product(s), and the second is a batch certificate of a pharmaceutical product (this document is rarely applied other than to vaccines, sera, and biologicals). these documents are used by dras of importing countries in their decision to approve, renew, extend, or vary a license. who created models for these confidential documents and listed the information that such certificates need to include. obligations that certifying authorities need to fulfill in order to be able to deliver a certificate have also been defined [ ]: ▸ possess an effective national licensing system, not only for pharmaceutical products, but also for responsible manufacturers and distributors. ▸ have gmp requirements, in agreement with those recommended by who, to which all manufacturers of finished pharmaceutical products are required to conform. ▸ effective controls must be in place to monitor the quality of pharmaceutical products registered or manufactured within its country, including access to an independent quality control laboratory. ▸ have a national pharmaceuticals inspectorate, operating as an arm of the national dra, and having the technical competence, experience, and resources to assess whether gmp and other controls are being effectively implemented, and the legal power to conduct appropriate investigations to ensure that manufacturers conform to these requirements by, for example, examining premises and records and taking samples. ▸ support administrative capacity to issue the required certificates, to institute inquiries in the case of complaint, and to notify expeditiously both who and the competent authority in any member state known to have imported a specific product that is subsequently associated with a potentially serious quality defect or other hazard. gmp standards provide the basis for the who certification scheme that relies on the capacity, experience, and expertise of the certifying authority of the exporting country. this scheme is a great example of cooperation between countries and is an important tool to support a regulatory system in developing countries that do not have enough capacity, resources, or expertise. biological medicinal products, such as vaccines, blood and blood products, diagnostics, gene therapy, biotechnology products, cytokines and growth factors, and cell and tissue products, rely heavily on international standardization to ensure their quality and their equivalence across manufacturers. this is especially true due to the increasing globalization in the production and distribution of these biological medicines. over the past years, who has worked to standardize these biological materials by establishing international biological reference materials t as well as developing international guidelines and recommendations on the production and control of biological products and technologies. guidelines provide more general information on a range of topics of interest to national dras and manufacturers (e.g., "guidelines on evaluation of similar biotherapeutic products, sbps"), whereas recommendations establish the technical specifications for manufacturing and quality control of specific products (e.g., "recommendations to assure the quality, safety and efficacy of bcg vaccines"). who has also released many other documents on general topics (such as "regulation and licensing of biological products in countries with newly developing regulatory authorities" [ ] and "good manufacturing practices for biological products" [ ]) or on a specific type of product (e.g., blood products and related biologicals, cells and tissues, cytokines, or vaccines) to facilitate control of these biological products on a worldwide basis. these norms and standards have been developed to assist who member states in ensuring the quality and safety of biological medicines and related in vitro biological diagnostic tests worldwide. by adopting these guidance documents in their pharmacopoeias or equivalent legislation, each country ensures that the products produced and used in their country conform to current international standards. by advising national dras and manufacturers on the control of biological products, regulatory guidance documents also establish a harmonized regulatory framework for products in international markets. who accomplishes its biological program through the who collaborating centers and the who ecbs. members of the ecbs are scientists from national control agencies, academia, research institutes, public health bodies, and the pharmaceutical industry acting as individual experts and not as representatives of their respective organizations or employers. its work is based on scientific consensus achieved through this international consultation and collaboration. this committee, which directly reports to the executive board, has met on an annual additionally, who has been particularly active in the specific field of blood products and related biologicals. it has provided technical guidance and quality assurance tools to dras, national control laboratories, and manufacturers to support implementation of quality and safety systems for the production and control of blood products and related in vitro diagnostic devices worldwide. indeed, many countries have significant difficulties in fulfilling their responsibilities in this field because processing blood (with inherent variability due to the nature of the source materials) is a highly specialized process that requires a high degree of expertise. this development of who international reference materials and guidelines supports the technical capacity of national dras and assures the compliance of manufacturers to quality and safety measures globally in order to prevent transmission of diseases via blood products. it also contributes to technology transfer, global cooperation, and harmonization of regulations via the blood regulators network (brn). finally, the who has been very involved in the development of standards and guidelines regarding vaccines due to the importance of these products in public health. v moreover, who established the "prequalification of vaccines" (regarding the acceptability, in principle, of vaccines from different sources for supply) to help the united nations children's fund (unicef) and other un agencies that purchase vaccines. finally, through its regulatory pathways initiative it also helps to address the challenges faced by developing countries that are targets for clinical trials or introduction of new vaccines not registered in the country of manufacture. the objective is to support the establishment of regulatory mechanisms for the licensing of products in those countries that have not yet fully developed the expertise for the review of technical applications. this is achieved via workshops and technical assistance in collaboration with the european medicines agency (ema) through its article scientific opinion procedure, w the us fda, and other national dras in developed countries. a developing countries' vaccine regulators network (dcvrn) was created in september , and regional initiatives were also established. in many countries (developed and undeveloped), there is recognition of the significant need for research and development of medicines specifically for pediatric use (or data from pediatric studies using medicines that have been developed for adults). this lack of pediatric data became an important problem despite many initiatives from different regions or countries. the lack of suitable pediatric medicines, paired with inconsistent regulatory frameworks, poses significant risks to a particularly vulnerable patient population. the overall aim of the pmrn x is to promote availability of quality medicines (including biological medicines and vaccines) for children by facilitating communication, collaboration, and regulatory harmonization across manufacturing, licensing, and research [ ] . more specifically, among several objectives, this network tries to: ▸ provide a forum for discussion between worldwide dras to build awareness of pediatric medicines regulatory considerations ▸ facilitate the collaboration, discussion, and work towards consensus on regulatory standards for pediatric medicines (i.e., the development of international recommendations and common standards for clinical trials and registration of medicines for children based on the existing ich, ema, and us fda guidelines) ▸ strengthen licensing (approval) systems for pediatric medicines by increasing regulatory cooperation, information sharing, and training traditional medicines y have been used in many countries throughout the world over many centuries. today, these medicines still represent an important part of healthcare in some countries. z for example, more than countries have regulations for herbal medicines, but practices of traditional medicine vary greatly from country to country and from region to region, as they are influenced by factors such as culture, history, personal attitudes, and philosophy. however, while it is often necessary to tailor legislation and delivery to reflect the needs and traditions of the individual countries, a number of themes and issues are common, such as the importance of practitioner training, the issues related to safety, the need to enhance research into both products and practices, and the importance of labeling. also, the use of traditional medicines has expanded globally and has gained popularity in the last few decades. specifically, these practices have not only continued to be used for primary healthcare of the poor in developing countries, but have also been used in other countries where conventional medicines are predominant in the national healthcare system. aa with this tremendous expansion in the use of traditional medicines worldwide, safety and efficacy as well as quality control of herbal medicines and traditional procedure-based therapies have become important concerns for many of these countries. for this reason, who has been increasingly involved in developing international standards and technical guidelines for these types of medicines, and also in increasing communication and cooperation between countries [ ] . the challenge now is to ensure that traditional medicines are used properly, and to determine how research and the evaluation of traditional medicines should be carried out. supported by several wha and executive board resolutions, who has developed and issued a series of technical guidelines (e.g., guidelines for the assessment of herbal medicines, research guidelines for evaluating the safety and efficacy of herbal medicines, and guidelines for clinical acupuncture research). in , who developed draft guidelines for "methodology on research and evaluation of traditional medicine" that was finally approved in april [ ] . the purpose of this document is to promote the proper development, registration, and use of traditional medicines and to harmonize the use of certain terms in traditional medicine. moreover, in , who established a global network (called the international regulatory cooperation for herbal medicines [irch]) to allow communication and exchange between worldwide regulatory authorities responsible for the regulation of herbal medicines. the mission of this program is "to make quality priority medicines available for the benefit of those in need." this is achieved through evaluation and inspection activities, and in cooperation with national dras and partner organizations. the list of prequalified medicinal products (updated regularly) is used principally by un agencies (including unicef and the joint united nations programme on hiv/aids [unaids]) to guide their procurement decisions. but, the list has also become a vital tool for any agency or organization involved in bulk purchasing of medicines, as demonstrated by the global fund to fight aids, tuberculosis and malaria. the strategy is to apply unified standards of acceptable quality, safety, and efficacy and to build the capacity of staff from national dras, quality control laboratories, and from manufacturers or other private companies, to ensure quality medicines. technical assistance, training, and capacity building are an important part of the program [ ] . when a product is included on the who list, the relevant product dossier has been evaluated and the manufacturing sites inspected by who-appointed assessors and inspectors and found to comply with who standards. who also recognizes the evaluation of products by some major dras that apply stringent standards for quality, including, but not limited to, the us fda, ema, and health canada. bb however, it is important to note that the inclusion of a product (or a laboratory) on this list does not imply any approval by who because it is the sole prerogative of national authorities. who inspections are done by a team of inspectors, including: ▸ an inspector/expert from one of the pharmaceutical inspection co-operation scheme (pic/s) countries ▸ a who representative (inspector/expert) ▸ an inspector (or inspectors) as an observer from the national dra of the country in which the laboratory is located at the end of , the who list of prequalified medicines included products (manufactured in countries); a total of quality control laboratories had been prequalified (covering all who regions). the program had also prequalified its first active pharmaceutical ingredients (apis) [ ]. the above projects are specifically related to the harmonization of pharmaceutical regulations and regulatory standards related to medicinal products. however, it is important to note that several other who projects not directly related to the harmonization of pharmaceutical bb when a product is listed with a reference to us fda or ema, the alternative listing procedure was used, and the products have been added to the list relying on the assessment and inspections conducted by the us fda or ema. regulations cc have been or are also very important because they facilitate implementation of common systems, agreements on terminology, and the establishment of a forum for exchange of not only information, but also expertise and experience. these other who projects ultimately facilitate overall dialogue, cooperation, convergence, and harmonization between countries and regions. moreover, other more general projects can also promote regional and subregional collaboration and harmonization of the regulation. for example, one of the principles of the general ec-acp-who partnership established in dd was to "strengthen existing collaborative arrangements (e.g. pooled procurement in the caribbean) and catalyse the creation of new ones, which can work together to achieve pooled procurement, common policies and harmonization of legislation." in addition, who publishes many documents regarding pharmaceuticals and regulations (i.e., newsletters, periodicals, reports status, or special publications such as the who blue book [ ] ) that allow the diffusion and exchange of information and data everywhere in the world. for example, "who drug information" is a quarterly journal, launched in , which provides an overview of topics relating to medicine development and regulation that is targeted to a wide audience of health professionals and policymakers. it communicates the latest international news and trends. finally, some other specific who projects are also very important in facilitating the implementation of the international standards. these following projects need to be reviewed even though they are not directly related to the harmonization of pharmaceutical regulations because they demonstrate the key role of who in the global regulatory system, and therefore show how this organization has the legitimacy to further coordinate global harmonization. ▸ who review of drug regulatory systems: to ensure that public health is appropriately supported, national regulatory capacity needs to be regularly assessed, areas of weakness need to be identified, and necessary measures need to be taken. the objectives of this review are to strengthen national regulatory and control capacity through the identification of specific needs and the provision of appropriate technical support and training. this is done via the evaluation of existing legal framework, regulations, and control activities in order to assess the national regulatory capacity against a set of predefined parameters. who can then provide technical input if gaps are identified. this activity is very important, especially in developing countries, to ensure that international standards can be appropriately implemented at the national level. it is also an important tool to have a clear status of national regulatory systems to evaluate appropriate needs from developing countries and therefore necessary support from regional and international organizations. the who multicountry study (involving only countries) also showed that such assessments represent significant opportunities to learn more about the strengths and weaknesses of dras and the different strategies used to improve drug regulation performance [ ] . the international health regulations (ihr), first adopted by the health assembly in and then significantly revised in in consideration of the growth in international travel and trade and the emergence or reemergence of international disease threats and other public health risks [ ] , were finally adopted by the th wha on may , and entered into force on june , . the ihr is an international legal instrument that is binding on all the who member states. these global rules were developed and implemented to enhance national, regional, and global public health security. its aim is to help the international community prevent and respond to acute public health risks that have the potential to cross borders and threaten people worldwide. the stated purpose and scope of the ihr are "to prevent, protect against, control and provide a public health response to the international spread of disease in ways that are commensurate with and restricted to public health risks, and which avoid unnecessary interference with international traffic and trade." the ihr has been used for the h n pandemic crisis [ ] . the revised ihr requires countries to strengthen their core surveillance and response capacities so that they can report certain disease outbreaks and public health events to who. building on the unique experience of who in global disease surveillance, alert, and response, the ihr defines the rights and obligations of countries to report public health events, and establishes a number of procedures that who must follow in its work to uphold global public health security. as mentioned above, this document was not specifically developed for pharmaceutical products, but is an important global tool that enhances cooperation between all countries in the world. indeed, even if this agreement does not specifically relate to the harmonization of pharmaceutical regulations, it is very interesting for many reasons. first, this project helps strengthen worldwide capacities for public health and global cooperation in general, which is important for the implementation of harmonized global standards. more importantly, this is one of the first agreements that manages public health as a truly global issue and proposes further action using an integrated international approach and network. it shows that further integrated global cooperation in the area of health (with who being at the center of this cooperation to coordinate this effort) is possible and beneficial [ ]. the mission of who's program on medicines and pharmaceutical policies is to support the achievement of the health-related mdgs by assisting governments and organizations to ensure global and equitable access to safe, effective medicines of assured quality. goal ee and target e ff are particularly applicable to who harmonization activities in the pharmaceutical domain. many of who's activities in the pharmaceutical domain support the achievement of these mdgs because they globalize the resolution of major public health issues (that cannot be resolved at the national/local level), they promote collaboration between countries and regions, and they provide tools and standards to allow such international collaboration. since its creation, who has indeed played a significant role in the global harmonization of pharmaceutical regulations. as per its mandate and the responsibilities defined in its constitution, it has developed and maintained numerous international standards, norms, guidelines, classifications, and nomenclatures through a rigorous, international, and independent scientific consultative process. in addition to this normative role, who has also established important networks to facilitate global cooperation. for example, icdras has been an important player in global regulatory harmonization. it launched many projects that have facilitated and promoted harmonization and cooperation between countries [ ]. cooperation projects have also been pioneered over the years with a specific interest in essential medicines. gg the who prequalification program has been an important step since it demonstrated that cooperation in the domain of medicine evaluation is possible and beneficial. indeed, this program has been very positive and its scope has continually been extended since its creation in . it has clearly accelerated the access of essential medicines worldwide (especially in low and middle income countries) [ ] . this model should be used to further develop regional and global collaboration for medicine evaluation. the example of the / pilot who/east african community (eac) collaborative procedure initiated to facilitate registration of prequalified medicines in the eac [ ] was positive. the overall aim was to identify a framework for who/eac, for joint evaluation and approval of dossiers and inspections of medicine manufacturing sites, and to ensure that these assessments are integrated into national regulatory decision making. two assessors each from three eac countries (kenya, tanzania, and uganda) and six who assessors jointly assessed two product dossiers submitted by a single manufacturer. the dossiers were submitted in parallel, and with identical content, to each participating eac country and to who. the products were both prequalified. the principal benefit of this joint assessment was that once the products had been jointly assessed and approved by who/eac, they were granted immediate access to the markets of each of the countries that had participated in the joint assessment. also, such joint assessment contributes to harmonization of regulatory requirements at the regional level. this pilot who/eac project also exemplifies the role of who in providing technical assistance to countries and supporting local capacity building. indeed, by acknowledging the important role of adequate systems to implement sound and effective pharmaceutical regulation, who has supported developing countries in addressing their deficiencies or capacity problems through various types of training, assessment of regulatory capacity, and the recommendation of institutional development plans. these activities have been very beneficial in the past, but work needs to continue and grow in this domain, as problems still exist. indeed, the extent of implementation of standards varies from one region to another. there are a number of factors that explain observed weaknesses of medicine regulation, and these differ from country to country and depend also on the individual health systems. countries may vary regarding their registration system, and not all of them can implement a comprehensive medicine evaluation and registration system. also, who encourages regional and international collaboration among national dras in order to promote the harmonization of requirements and practices, and to strengthen professional competence [ ] . however, as recognized in its medicine strategy plan, cooperation with regional harmonization initiatives and organizations should be further enhanced [ ] . closer cooperation and coordination should also be sought with other global initiatives such as ich. further assistance to countries and cooperation with other regional and global initiatives are indeed possible and can be facilitated by who's regionalized structure. this specific threelevel organization provides multiple opportunities for engaging with countries. the headquarters focus on initiation, development, and global coordination of harmonization projects, while regional offices focus on technical support and building national capacities to support implementation. who's presence in countries also allows a close relationship with ministries of health and its partners inside and outside of government. this work at the regional and country levels is critical in ensuring that local and regional needs and challenges are taken into consideration when international standards and projects are developed. to conclude, although some improvements may address current challenges, who has been very successful and beneficial for all member states (developing and also developed countries). it has promoted evidence-based debate, analysis, and recommendations for health through its own work and that of the numerous formal and informal networks and collaborating centers around the world. these networks facilitate lively cooperation between scientists across nations and allow governments to jointly tackle global health problems. development and promotion of global norms and standards in medicine is one of who's efforts that is widely perceived as being in an area in which who has a comparative advantage. this advantage is due to the recognition of who as the global leader and coordinating authority on global public health. the achievement of the mdgs and the renewal of primary healthcare are indeed unthinkable without who's norms and standards, policy guidelines, and technical cooperation. this is why the development and promotion of global norms and standards are an area of continued focus for who [ ] . it is indeed critical that who continue its work towards better harmonization and cooperation in the pharmaceutical domain. acknowledging the unique neutral and independent role of who, its numerous successes in the past, and its nearly universal membership, it would be appropriate to further extend the leadership of who in this domain. this increased responsibility in the coordination of medicines would also further fulfill its mandate "to act as the directing and co-ordinating authority on international health work." [ ] the international conference on harmonization of technical requirements for registration of pharmaceuticals for human use (ich) is a -year-old program. this unique initiative was established with the objective to bring together the dras of europe, japan, and the united states and experts from the pharmaceutical industry in these three regions to discuss scientific and technical aspects of pharmaceutical product registration. the drug regulatory systems in all three regions share the same fundamental concerns for the safety, efficacy, and quality of medicines. however, many time-consuming and expensive experiments have been repeated in all three regions to meet specific regional requirements. the goal of ich has been to increase harmonization of technical requirements to ensure that safe, effective, and high-quality medicines are developed and registered in the most efficient and cost-effective manner in order to be delivered to the maximum number of patients in the world without delay. these activities have been undertaken to promote public health, prevent unnecessary duplication of clinical trials in humans, and minimize the use of animal testing without compromising safety and effectiveness. by making recommendations on ways to achieve greater harmonization of technical requirements for product registration, the objective is indeed to reduce or obviate the need to duplicate the testing carried out during the research and development of a new product. since its inception in , ich has evolved, through its global cooperation group (gcg), to respond to the increasingly global face of drug development, so that the benefits of international harmonization for better global health can be realized worldwide. this ich mission is embodied in its current terms of reference: ▸ to maintain a forum for a constructive dialogue between regulatory authorities and the pharmaceutical industry on the real and perceived differences in the technical requirements for product registration in the eu, us, and japan in order to ensure a more timely introduction of new medicinal products, and their availability to patients; ▸ to contribute to the protection of public health from an international perspective (added upon revision in ); ▸ to monitor and update harmonized technical requirements leading to a greater mutual acceptance of research and development data; ▸ to avoid divergent future requirements through harmonization of selected topics needed as a result of therapeutic advances and the development of new technologies for the production of medicinal products; ▸ to facilitate the adoption of new or improved technical research and development approaches which update or replace current practices, where these permit a more economical use of human, animal, and material resources, without compromising safety; ▸ to facilitate the dissemination and communication of information on harmonized guidelines and their use such as to encourage the implementation and integration of common standards. ich is comprised of representatives from six parties (the founding members of ich) that represent the regulatory bodies and research-based industry in the eu, japan, and the us: since , when ich was initiated, members have been added: ▸ the international federation of pharmaceutical manufacturers & associations (ifpma), the global nonprofit, nongovernmental organization, founded in to represent the research-based pharmaceutical, biotech, and vaccine sectors. its members are comprised of over leading international companies and over national and regional industry associations covering both developed and developing countries. ifpma is very involved in all subjects related to the improvement of global health. it has been closely associated with ich since its inception to ensure contact with the research-based industry (especially outside the ich regions). ifpma provides the ich secretariat. this important group of nonvoting members was established as a link between ich and non-ich countries and regions. the ich organization consists of the ich steering committee, ich coordinators, ich secretariat, and ich working groups. the ich global cooperation group (gcg) and the ich medical dictionary for regulatory activities (meddra) management board are subcommittees of the ich steering committee. the steering committee is the body that governs the ich, determines the policies and procedures, selects topics for harmonization, and monitors the progress of harmonization initiatives. this committee, established at the first ich meeting in april , has met at least twice a year since, with the location rotating between three regions (eu, japan, and us). during these committee meetings, new topics are considered for adoption, reports are received on the progress of existing topics, and maintenance and implementation of the guidelines are discussed. each of the six ich parties has two seats on the ich steering committee. each of the observers nominates nonvoting participants to attend the ich steering committee meetings. ifpma also participates as a nonvoting member. meetings of the ich meddra management board, ich gcg, and the regulators forum also occur during the same week as the steering committee meeting. ich working groups are the key players of the ich harmonization process. they are responsible for the development, implementation, or maintenance of ich guidelines. each of the six ich parties is represented in every working group. the official membership of an expert working group/implementation working group (ewg/iwg) is usually limited to two officials per party (one topic leader and one deputy topic leader). one of these topic leaders is nominated rapporteur (and sometimes a second is nominated co-rapporteur) by the steering committee. ich observers and interested parties hh can also nominate one representative. the pharmacopoeial authorities and representatives from the self-medication industry and the generic industry were invited to participate in the various working groups. finally, the three regulatory parties of the steering committee officially designate a regulatory chair when a new ich topic is formally adopted. the regulatory chair, designated among the three regulatory parties, regularly presents reports to the steering committee and ensures, in close collaboration with the rapporteur, timely execution of the ich process and adherence to the concept paper and business plan, including scope and timelines. depending on the type of harmonization activity required, the steering committee will endorse the establishment of one of three types of working groups: ▸ expert working group (ewg): these working groups are appointed by the steering committee when new topics are accepted for harmonization. the objective of each ewg is to review the differences in requirements between the three regions and develop scientific consensus required to reconcile those differences. it is charged with developing a harmonized guideline that meets the objectives defined in the concept paper and business plan. ▸ implementation working group (iwg): an iwg's task is to develop questions and answers (q&a) to facilitate implementation of existing guidelines. ▸ informal ewg/iwg: these working groups are formed prior to any official ich harmonization activity. their objective is to develop a concept paper and business plan. working groups meet in the same week as the steering committee and report on their progress to the committee. these one-week meetings are key for the ich organization as they allow for a regular review of efforts and achievements and adjust them if necessary. ich discussion groups are established to discuss specific scientific considerations or views (e.g., gene therapy discussion group) to facilitate the exchange of information on a specific topic, and ultimately the harmonization of the requirements. the coordinators are fundamental to the smooth running of the ich and are nominated by each of the six parties. an ich coordinator acts as the main contact with the ich secretariat and ensures that ich documents are distributed to the appropriate persons within the area of their responsibility. each party has also established a contact network of experts within their own organization or region in order to ensure that, in the discussions, they reflect the views and policies of the cosponsor they represent. the way this network operates differs according to the administrative structure of the party concerned. due to structural differences within the eu and mhlw, ich technical coordinators are also designated from the ema and pmda, respectively. they support the ich coordinator and facilitate every action of the steering committee members in the region, mainly by applying their scientific knowledge. their roles include acting as a contact point between the experts within the ema and pmda and the ich coordinator at the main regulatory body, and as a contact point with the ich secretariat. the ich secretariat operates from the ifpma offices in geneva (switzerland), and provides support to the ich steering committee. the secretariat is primarily concerned with preparations for, and documentation of, meetings of the steering committee, as well as coordination of preparations for working group (ewg, iwg, and informal wg) and discussion group meetings. the secretariat also provides administrative support for the gcg and the meddra management board, and maintains the ich website. the meddra management board, appointed by the ich steering committee, has overall responsibility for direction of meddra, an ich standardized dictionary of medical terminology. the board oversees the activities of the meddra maintenance and support services organization (msso), which serves as the repository, maintainer, developer, and distributor of med-dra. the management board is composed of the six ich parties, the medicines and healthcare products regulatory agency (mhra) of the uk, health canada, and who (as observer). the ifpma acts as a nonvoting observer on the management board and also chairs the board. as stated in its mission statement adopted by all parties in may , this group "promotes a mutual understanding of regional harmonization initiatives in order to facilitate the harmonization process related to ich guidelines regionally and globally, and to facilitate the capacity of drug regulatory authorities and industry to utilize them." this group ensures that the benefits of ich harmonization extend beyond the three ich regions (japan, eu, and us). the role of the gcg has changed over time as the focus on collaboration with the non-ich regions increased. from its creation to today, three phases can be differentiated: ▸ first phase ( to : information sharing outside ich: during these first three years, the gcg mandate was to share information outside ich (via preparation of brochures, presentations at international meetings, etc.). the objectives were to make available to any country or pharmaceutical company that requested it information on ich, ich activities, and ich guidelines. to this end, the group created a series of brochures intended to guide its activities as it answers requests for information and responds to non-ich regulators and industry: • ▸ second phase ( phase ( to : integration and collaboration with rhis: on november , , the ich gcg released their terms of reference in which they extended their action to act as the primary representative of the ich steering committee outside the ich regions, and equally as such as a conduit for non-ich parties to the ich steering committee. to do so, the group developed a privileged relationship with other non-ich harmonization initiatives. this key activity of the gcg had three advantages: • to share ich discussions and actions with the non-ich regions (allowing, when possible, harmonization and implementation of ich activities on a worldwide basis) • to promote and organize the involvement of the non-ich regions experts in ich discussions (via expert meetings, comments on step guidelines, and training on guidelines) • to facilitate interregional collaboration in order to promote transparency, better understanding of challenges and potential solutions to harmonization issues, leverage collective experience and knowledge (allowing easier standardization and development of good harmonization practice) when, in , the gcg decided to include representatives from the non-ich regions, the relationship with the non-ich regions became more collaborative and proactive, and the focus shifted from information sharing to a two-way dialogue to set up training and work on implementation. the results of these collaborations allowed the organization of workshops in the regions (e.g., apec workshops on clinical research inspections in and in thailand, the sadc quality guideline workshop in in zambia, and the apec quality guideline workshop in in china). as an example, the gcg also endorsed the apec life sciences innovation forum (lsif) sponsored workshop on ich quality guidelines (q , q and q ), held in september in seoul, south korea. this workshop was a great success for the spread of ich concepts and recommendations in this region as it was attended by more than participants (i.e., regulators, policymakers, academia, and industry) from countries. this type of workshop allows for practical explanation of ich guidelines, but also opens up discussion and exchange on the anticipated challenges and opportunities associated with the implementation of ich guidelines in order to better prepare implementation. the participation of these individual countries is distinct and complementary to the participation of official rhi representatives. in june , the inaugural meeting of the expanded gcg occurred. today, the key focus of the gcg continues to be the implementation of ich guidelines via the organization of training that began in . this training is indeed an important means for the promotion of better understanding of ich and ich guidelines beyond the ich regions. it developed a framework and mechanism for policy [ - ], a procedure for selection and prioritization, a template for training requests, definitions of roles and responsibilities for the organization and coordination of training activities, and a clearinghouse of training events for public access. these training activities (most of the time coordinated with the respective rhis) involve ich experts. during the meeting in october in yokohama, japan, the ich steering committee also decided to complement the gcg with the regulators forum. the ich regulators forum is the latest idea implemented by ich to increase communications and sow relationships between worldwide dras in order to ensure adoption and implementation of ich guidelines. following a proposal from the us fda in , the first meeting occurred in portland, oregon, us in june . this is a good complement to the gcg activities and includes authorities from the three ich regions, the observers, the rhis, and individual dras such as australia, brazil, china, chinese taipei, india, korea, russia, and singapore. this ich regulators forum allows frank discussion and the sharing of expertise among dras regarding best practices and challenges related to the implementation of ich guidelines and their impact on regulatory systems. this discussion assists in identifying training and capacity needs for action by the gcg. more importantly, it also builds mutual understanding, relationships, and trust. in the s, many varied efforts of harmonization of pharmaceutical regulatory requirements were conducted. first, the european community, who was developing a single market for pharmaceuticals, had shown that harmonization among different countries (with different medical cultures/practices and regulatory systems) was possible. at the same time, bilateral discussions between europe, japan, and the us on the possibility for harmonization were ongoing. the concretization of these ad hoc discussions happened during the world health organization (who) international conference of drug regulatory authorities (icdras) in paris in , where specific plans were agreed to. following this meeting, the three authorities approached ifpma to discuss a joint regulatory-industry initiative on international harmonization. the spirit and concept of ich was then agreed on between the different parties. in april , ich was officially created at its inaugural steering committee meeting, hosted by the efpia in brussels, belgium. representatives of the regulatory agencies and industry associations of europe, japan, and the us met primarily to plan an international conference, but at the meeting the wider implications and terms of reference of ich were also discussed. during this first meeting, the structure (including a steering committee and expert working groups) and the focus of ich activities (harmonization of safety, efficacy, and quality guidelines for human drugs and biological products) were agreed on. eleven topics were selected for discussion at the first conference. finally, it was agreed to expand the membership of the steering committee to include representatives from who, efta, and canada as observers because the harmonized guidelines could be useful to other non-ich regions. additionally, agreement was reached on the full name of ich. this name was chosen because one of the objectives of this group was to organize international conferences on harmonization. today, this name is associated with the overall initiative. the ich members officially confirmed their commitment to ich in a statement following the nd steering committee meeting: the parties cosponsoring this conference, represented at the nd steering committee meeting in tokyo, - october re-affirmed their commitment to increased international harmonization, aimed at ensuring that good quality, safe, and effective medicines are developed and registered in the most efficient and cost-effective manner. these activities are pursued in the interest of the consumer and public health, to prevent unnecessary duplication of clinical trials in humans and to minimise the use of animal testing without compromising the regulatory obligations of safety and effectiveness. this conference will provide a unique opportunity for regulators and industry to reach consensus on the steps needed to achieve this objective through greater harmonization of technical requirements and to set out practical and realistic targets for harmonising requirements where significant obstacles to drug development and the regulatory process have been identified. recognising the substantial progress which has already been made in achieving harmonization within europe and through bilateral contacts between europe, japan, us, and other regions, the conference will seek to make further progress through a trilateral approach, with clearly defined priorities, methods of work and recommendations to both industry and regulatory authorities. whilst the conference will be an important step forward, it is not seen as an end in itself, but as a stage in a developing process, at a high level, between regulators and industry. the conference, its preparations, and follow-up activities will be conducted in an open and transparent manner and the presence of observers from other regulatory authorities and who is welcomed as a means of ensuring that the benefits of progress towards harmonization can be utilized world-wide. the conference will not only look at existing issues but will, based on past experience, seek to minimise future divergence of new registration requirements, as a consequence of technical progress. this initial ich statement is important because it provides the spirit of ich that has been followed and implemented in all subsequent ich activities since. from its creation in to , the initial focus of ich was to promote technical and scientific exchanges and discussions in order to find consensus on divergent technical requirements for registration of medicinal products in the ich regions. the goal was indeed to remove redundancy and duplication in the development and review process, such that a single data set could be generated to demonstrate the quality, safety, and efficacy of new products. during this first phase of its activities, the ich structure and process were defined, a lot of harmonization activities started, and several guidelines/standards developed. these first harmonization discussions were directed to both technical scientific content (related to quality, safety, or efficacy topics) and to format and communication tools (development of e and the start of meddra, electronic standards for transmission of regulatory information (estri) and common technical document (ctd) projects). during these first years, there was a growing interest in ich products beyond ich countries, and ich recognized early that harmonization within the ich regions would not suffice. however, during these first years, discussions and activities focused mainly on harmonization among ich parties (even though ich agreed to include observers as a link to the non-ich regions) because it was important to start the process with a limited number of committed parties. in november , the th international conference on harmonization (ich ) in san diego, california, us marked the end of the first years of ich activity. this conference provided an opportunity to evaluate results and to identify future needs in the area of international harmonization. at the conference, results were presented of a survey on utilization of ich guidelines confirming the positive contribution of ich in improving the international drug regulatory approval process, thus speeding the availability of new medicines to the public. in its statement titled "the future of ich" released at ich , the steering committee emphasized its intentions to focus the second phase of ich on implementing and maintaining existing guidelines, preventing disharmony, encouraging scientific dialogue and harmonization in new areas, and undertaking efforts towards global cooperation with non-ich regions and countries. during its second phase, ich continues to develop and implement tripartite guidance on specific technical requirements, and also increase its effort on the implementation of harmonized regulatory communication tools (i.e. meddra, ctd, estri, etc.) between authorities and industry. indeed, one of the areas of focus of this second phase is to ensure adequate implementation and maintenance of all the guidelines developed since . today, new guidelines continue to be developed, but less frequently. these new guidelines cover important technical subjects related to pharmacovigilance (i.e., guidelines e d, e e, and e f) or improvement of quality systems (i.e., guidelines q , q , and q ). new emerging topics (such as gene therapy) have also been discussed. however, the main challenge of ich is now to maintain and update the collection of guidelines already developed (i.e., follow the evolution of science, the experience gained, etc.). the second focus and priority of this ich phase has been, and continues to be, the extension of relationships with non-ich regions. it began with the creation of the gcg as a subcommittee of the ich steering committee in . since this time, ich has developed its relationship with non-ich regions and tried to facilitate the implementation of its standards and guidelines on a broader territory via collaboration and training. even if some relationships existed before, the gcg has been key for this geographical extension, and its role increased over time by moving from information sharing (via preparation of brochures, presentations at international meetings, etc.) to a collaborative and proactive dialogue (via the incorporation of non-ich regions and countries in the group). further evolution of the ich structure and the gcg's terms of reference are expected to continue to promote greater involvement of global regulators [ - , - ]. the first activity of ich was to organize the ich conference in , one year after its creation, in order to exchange points of view and discuss divergences among different parties involved. since ich , five additional conferences have been organized (table ) . these regular, well-attended conferences helped communicate the results of the harmonization activities to the largest possible audience. they were designed as an open forum (in breakout sessions) to gather additional public comments and provide updates on ich's scientific activities. these six conferences were well attended (e.g., , participants representing industry and authorities of over countries for ich and , participants representing industry and authorities of over countries for ich ). the early ich conferences were very important in increasing visibility on the process of harmonization and for ensuring that the process was carried out in a transparent manner. ich focused primarily on the finalization and completion of the ctd guideline. the last ich conference organized, ich , focused on areas such as new technologies and global cooperation with regulatory harmonization initiatives outside the ich regions. the three satellite sessions (related to "partnerships in harmonization," "gene therapy," and "meddra users' group") also confirmed the priorities of the meeting. during this conference, opportunities and new challenges for regulatory harmonization were discussed. the practical implementation of the ctd was also reviewed. after ich , no additional international conferences were scheduled. ich was planned to have taken place in europe in , but it was then canceled. instead, in may , the ich steering committee decided to replace these large international ich conferences with smaller and more frequent regional public meetings at the time of the ich steering committee meetings in the region (in order to benefit from the presence of steering committee members and ich experts). now that the ich process is well recognized, these smaller regional meetings allow for a better focus on regional issues and challenges. it also provides everyone the opportunity to meet with regulators and industry experts involved in ich activities, to be regularly informed on recent developments, and to exchange information on different hot topics. following this decision, regional meetings have been organized: ▸ in europe, the first eu regional public meeting took place in brussels, belgium in november . ▸ in north america, the first regional public meeting took place in washington, dc, us in october . ▸ in asia, the first regional public meeting took place in tokyo, japan in november . the ich process was first drawn up at the steering committee meeting in washington, dc in march , and amended in tokyo, japan in september . since then, the ich procedures have been revised several times . moreover, the new principles of governance, agreed to at the ich steering committee meeting in june , have revised the role of regulator and suggestions for new harmonization initiatives may arise in a number of forums (ich regional guideline workshops; regional and international conferences, workshops, and symposia dealing with research and development (r&d) and regulatory affairs; recognized associations; testing and registration of medicines, etc.). from the suggestion of a new harmonization action to the development of a new guideline (or modification of an existing guideline), there are three sequential steps: • submission of a concept paper to the ich steering committee by an ich party or an observer • endorsement by the steering committee • establishment of a working group the concept paper is the start of all ich harmonization activities. this document provides a short summary of the proposal (maximum two pages) and provides the information indicated below: • type of harmonization action proposed: for example, a new harmonized tripartite guideline and recommendation, or a revision of an existing guideline (indicating the category of procedure). • statement of the perceived problem: brief description with an indication of the magnitude of the problem currently caused by a lack of harmonization, or anticipated if harmonization action is not taken. • issues to be resolved: a summary of the main technical and scientific issues that require harmonization. • background to the proposal: further relevant information (e.g., the origin of the proposal, references to publications, and discussions in other forums). • type of expert working group: recommendation on whether the ewg should be a six-party group (for topics related to the r&d of a new drug substance and product) or an extended ewg (e.g., gmp). if necessary, further documentation and reports may be added to the concept paper. depending on the category of harmonization activity, a business plan may also be required. the business plan outlines the costs and benefits of harmonizing the topic proposed by the concept paper. only when the ich steering committee endorses a concept paper, and where appropriate a business plan, can the harmonization activity be initiated. a preliminary determination will be made on whether the topic is of sufficient interest to all parties and can be accommodated within the ich work schedule. the steering committee takes the following points into account when discussing a concept paper: • objectives and expected outcome of the harmonization action • categories of the ich process • composition of the ewg or iwg appointed to discuss the technical issues • setting a timetable and action plan for the ewg/iwg the concept paper may need to be revised and updated to reflect the steering committee discussions and conclusions. if the steering committee agrees that a topic may warrant further consideration and a business plan needs to be developed, an informal ewg/iwg will be formed and the group will work through e-mail, teleconference, and rarely, face-to-face meetings. the first tasks of the informal ewg/iwg will be to finalize a concept paper and develop a business plan. the revised concept paper and business plan will be sent prior to, and presented at, the next steering committee meeting. at its meeting in yokohama, japan (in june ), the ich steering committee agreed to have the final versions of the concept papers and business plans available on the ich website, for public information. depending on the type of harmonization activity proposed, the ich steering committee will endorse the establishment of either an ewg or an iwg. ich harmonization activities fall into four categories. as presented in table , these four categories cover the creation and development (stepwise progression), implementation, revision, and maintenance of guidelines. no procedure is in place for the withdrawal of existing ich guidelines because it happens very rarely. when guideline q f (stability data package for registration applications in climatic zones iii and iv) was withdrawn, an explanatory note was released following the endorsement of the withdrawal by the ich steering committee at its meeting in yokohama, japan in june . withdrawal notifications were also released by the ema, mhlw, and us fda. ▸ the formal ich procedure: the formal ich procedure follows a stepwise approach consisting of five steps with "decision points" at step and step that enable the steering committee to monitor the progress of the harmonization topics. this procedure is followed for the harmonization of all new ich topics. a streamlined procedure is also available when necessary. the procedure is initiated with the endorsement, by the steering committee, of a concept paper and business plan. an ewg with membership as specified by the concept paper is subsequently established. the ewg works to develop a draft guideline and bring it through the various steps of the procedure that culminate in step and the implementation in the ich regions of a harmonized tripartite guideline. • step : consensus building when the steering committee adopts a concept paper as a new topic, then the process of consensus building begins. the ewg prepares an initial consensus technical document, based on the objectives set out in the concept paper and in consultation with experts designated to the ewg. the initial draft and successive revisions are circulated for comments within the ewg, providing fixed deadlines for receipt of those comments. work is conducted via e-mail, teleconferences, and web conferences. if endorsed by the steering committee, the ewg will also meet face-to-face at the biannual steering committee meetings. interim reports on the progress of the draft are made to the committee on a regular basis. when consensus is reached among all ewg members, the ewg signs the step experts signoff sheet. the experts document with ewg signatures is submitted to the steering committee to request adoption under step a of the ich process. step a is reached when the steering committee agrees, based on the report of the ewg, that there is sufficient scientific consensus on the technical issues for the technical document or recommendation to proceed to the next stage of regulatory consultation.this technical document is made public on the ich website. on the basis of the technical document, the three ich regulatory parties take the actions they deem necessary to develop the "draft guideline." the consensus text approved by the three regulatory ich parties is signed off by the three regulatory ich parties as the step b draft guideline. • step : regulatory consultation and discussion regional regulatory consultation: at this stage, the guideline embodying the scientific consensus leaves the ich process and becomes the subject of normal wide-ranging regulatory consultation in the three regions. in the eu it is published as a draft chmp guideline, in japan it is translated and issued by the mhlw for internal and external consultation, and in the us it is published as draft guidance in the federal register. step guidelines released for consultation are also available on the ich website. dras and industry associations in non-ich regions may also comment on the draft consultation documents by providing their comments to the ich secretariat. after obtaining all regulatory consultation results, the ewg that organized the discussion for consensus building will be resumed. the same procedure described in step is used to address the consultation results into the step final document. the draft document to be generated as a result of step is called the step draft guideline. the step document with regulatory ewg signatures is submitted to the steering committee to request adoption as step of the ich process. step is reached when the steering committee agrees, on the basis of the report from the regulatory chair and the regulatory rapporteur of the ewg, that there is sufficient scientific consensus on the draft guideline. this endorsement is based on the signatures from the three regulatory parties to ich affirming that the guideline is recommended for adoption by the regulatory bodies of the three regions. in the event that one or more parties representing industry have strong objections to the adoption of the guideline on the grounds that the revised draft departs substantially from the original consensus, or introduces new issues, the regulatory parties may agree that a revised document should be submitted for further consultation. in this case, the ewg discussion may be resumed. the step final document is signed off on by the steering committee signatories for the regulatory parties of ich as an ich harmonized tripartite guideline at step of the ich process. • step : implementation having reached step , the harmonized tripartite guideline moves immediately to regulatory implementation, the final step of the process. this step is carried out according to the same national or regional procedures that apply to other regional regulatory guidelines and requirements in the eu, japan, and the us. information on the regulatory action taken and implementation dates are reported back to the steering committee and published by the ich secretariat on the ich website. in the eu, ich guidelines are submitted to the chmp for endorsement and the timeframe for implementation is established (usually six months). ich guidelines are available on the ema website. in japan, ich texts are translated into japanese and subsequent pharmaceutical and medical safety bureau notification for the promulgation of guidelines written in japanese is issued with an implementation date. the notifications on guidelines in japanese and also english attachments (ich texts) are available on the pmda website. in the us, the us fda publishes a notice with the full text of the guidance in the federal register. step guidance is available for use on the date it is published in the federal register. they are available on the us fda website. ▸ the q&a procedure: the q&a procedure is followed when additional guidance is considered necessary to aid in the interpretation of certain ich harmonized tripartite guidelines and ensure a smooth and consistent implementation in the ich regions and beyond. the q&a procedure is initiated with the endorsement of the steering committee of a concept paper. in the case of major implementation activities, the steering committee may also consider the need for a business plan. an iwg with membership as specified by the concept paper is subsequently established. the development and adoption of the q&a follow an established process. questions received from stakeholders are collected, analyzed, reformulated, and ultimately used as model questions for which standard answers are developed and posted on the ich website. the incoming questions are not answered individually, rather they serve to highlight areas that need additional clarification and are then used to develop a model question that will be answered in the q&a document. based on the level of guidance given by the answers, the iwg will assess whether the q&a document should be a step b document and published for comments or a step document and published as final. the document should be step b if, based on the answers provided, it sets forth substantial new interpretations of the guideline(s). the document should be a step if, based on the answers provided, it sets forth existing practices or minor changes in the interpretation or policy of the guideline(s). the document then follows the normal path of a step b/step document as per the formal ich procedure. the revision procedure applies when an existing adopted guideline needs to be revised or modified. it is almost identical to the formal ich procedure (i.e., five ich steps). the only difference is that the final outcome is a revised version of an existing guideline rather than a new guideline. the revision of a guideline is designated by the letter r after the usual denomination of the guideline. when a guideline is revised more than once, the document will be named r , r , r , and so on with each new revision. the maintenance procedure is used to add standards to exist ing guidelines and/or recommendations or to provide an update based on new information. this procedure has been used to amend the addendum of guideline s (r ), "detection of toxicity to reproduction for medicinal products & toxicity to male fertility," and guideline m (r ), "maintenance of the ich guideline on non-clinical safety studies for the conduct of human clinical trials for pharmaceuticals," on november , . it is currently applicable for changes to the q c guideline on residual solvents, the q b annexes, and m recommendations. in each case, the procedure is used when there is new information to be added or when the scientific/technical content is out-of-date or no longer valid. for the q c guideline, this maintenance procedure is used to revise the permitted daily exposure (pde) as new toxicological data for solvents become available. since its creation, and pursuant to its main goal, ich has released a number of guidances, each harmonizing technical requirements for registration of medicinal products. for each technical topic, the relevant ewg discussed the important question of whether there is scientific justification for the different regional requirements, and whether it would be possible to develop a mutually acceptable guidance. the objective of this scientific discussion is to reach a scientific consensus whatever the time and effort it requires [ ] (and not a "compromise" that would be an unacceptable decrease of certain regional requirements without scientific justification/ basis). ich has also worked on broader projects (e.g., meddra and ctd), which have been critical for the international exchange of information. the ultimate goal of ich activities is to remove redundancy and duplication in the development and review process such that a single set of data could be generated to demonstrate the quality, safety, and efficacy of a new medicinal product. the steering committee has given priority to harmonizing the technical content of the sections of the reporting data. the first ich guideline to deal with harmonizing the format of reporting data was e , "content and format of clinical study reports." this guideline describes a single format for reporting the core clinical studies that make up the clinical section of a registration dossier. the goal of developing a harmonized format has led to the creation of the ich guideline m , "the common technical document" (ctd), further described below. at the first ich steering committee meeting it was decided that the topics selected for harmonization would be divided into safety, quality, and efficacy in order to reflect the three criteria that are the basis for approving and authorizing new medicinal products. since then, ich has created a fourth category called multidisciplinary, which covers crosscutting topics that do not fit uniquely into one category or another. therefore, today ich topics are divided into four categories (quality, safety, efficacy, and multidisciplinary) and ich topic codes are assigned according to these categories. a summary of harmonized topics is provided below. an updated list of these guidances (including their status) can also be downloaded from the ich website (and also from the us fda, pmda, and ema websites). the guidelines under this category provide harmonization of information related to the development, manufacturing, and testing of medicines. they specifically cover stability testing (q ), validation of analytical procedures (q ), impurities testing (q ), pharmacopoeial text harmonization and interchangeability (q ), quality information on biotechnological products (q ), specifications (test procedures and acceptance criteria) (q ), gmp (q ), pharmaceutical development (q ), quality risk management (q ), and pharmaceutical quality systems (q ). in addition, the ich steering committee endorsed on april , the development of a new q guidance related to the development and manufacture of drug substances (chemical entities and biotechnological/biological entities). the guidelines under this category provide harmonization of information related to in vitro and in vivo preclinical studies. they cover all preclinical studies performed during the development of new pharmaceutical products, such as carcinogenicity studies (s ), genotoxicity studies (s ), toxicokinetics and pharmacokinetics studies (s ), toxicity studies (s ), reproductive toxicology studies (s ), pharmacology studies (s ), and immunotoxicology studies (s ). guideline s specifically addresses preclinical safety evaluation for the biotechnological products. the ich steering committee also endorsed on may , the development of a new s guideline that provides preclinical guidelines on oncology therapeutic development. finally, the photosafety evaluation of pharmaceuticals was endorsed as a new topic (s ) by the ich steering committee in june . the guidelines under this category provide harmonization of information pertaining to the clinical evaluation of pharmaceutical products. most of these guidelines relate to the assessment and management of safety data (e and e guidelines). these guidelines cover: • all the above efficacy guidelines can be applied to all therapeutic classes of drugs. until now, ich has focused the discussion on general (i.e., nontherapeutic class-specific) guidances. however, there are, in some therapeutic classes, individual drug evaluation guidelines among the three regions. differences between guidelines can result in obstacles to the mutual use and acceptance of clinical data. at the steering committee meeting in september , it was agreed that this should be adopted as a new area of work for ich, with the first such guideline being undertaken as a "pilot study" to assess the feasibility of extending work in this area. it was agreed to develop the first therapeutic class-specific guideline for antihypertensive drugs. no other guideline for clinical evaluation of a specific therapeutic category has been developed since this guideline (e ). this category was created to include guidelines covering topics that do not fit uniquely into one of the above three categories. in addition to the technical guidelines described in previous sections, ich set up ewgs to harmonize medical terminology (m : meddra), drug dictionaries (m ), and the format and organization of data in regulatory applications (m : ctd) in order to ease the exchange of information. the creation of electronic standards (m : estri) was also critical for the quick exchange of common, agreed-upon data. in november , the ich steering committee endorsed the establishment of an ewg for the electronic common technical document (ectd) and assigned the topic code "m " (even though work in relation to the ectd had previously been undertaken by the m ewg). all these harmonization initiatives are critical achievements that required a lot of effort from their respective working groups. they are important activities that greatly contributed to the international harmonization of pharmaceutical regulations because they harmonized and facilitated the exchange of information between regulators and pharmaceutical companies. due to the importance of these initiatives, each of them is detailed in the specific subsections below. guideline m covers a specific topic relating to both safety and efficacy issues. for this reason, it has been classified as a "multidisciplinary topic." this joint safety and efficacy guideline provides principles for nonclinical strategies (i.e., scope, timing, and duration of nonclinical safety studies) in relation to the conduct of clinical trials. it helps to reduce the differences between the ich regions and it also provides recommendations to reduce animal use during research and development (e.g., inclusion of any in vivo evaluations as additions to general toxicity studies instead of performing separate studies). this guideline is definitively aligned with the overall ich objectives, as its purpose is to facilitate the timely conduct of clinical trials, reduce the use of animals in accordance with the rs (reduce/refine/replace) principles, and reduce the use of other drug development resources. it clearly promotes the safe and ethical development and availability of a new pharmaceutical as quickly as possible. finally, the ich steering committee endorsed (in june ) the "assessment and control of dna reactive (mutagenic) impurities in pharmaceuticals to limit potential carcinogenic risk" as a new topic (m ). meddra was developed by an ich ewg in the early s. it was designed to support the classification, retrieval, presentation, and communication of medical information internationally and throughout the product regulatory cycle. prior to meddra, different medical dictionaries (and also different versions of these dictionaries) were used, such as the world health organization adverse reaction terminology (who-art), the coding symbols for a thesaurus of adverse reaction terms (costart) from the us fda, and the japanese adverse reaction terminology (j-art) from the mhlw. at that time, several worldwide authorities were looking for a more cost-and time-efficient way of processing suspected adverse reaction reports (e.g., the united kingdom medicines control agency [uk mca] were developing a new system of coding called adroit). it became obvious that this activity should fall under the auspices of ich. the goal of ich in developing meddra was to have an internationally recognized standard, and medically rigorous and well-maintained terminology to facilitate communication. it is indeed one of the most important ich projects for ensuring the global exchange of clinical information. this international medical terminology is particularly important in the electronic transmission of adverse event reporting (both in the pre-and post-marketing areas), as well as in the coding of clinical trials data. the meddra dictionary is a multi-axial terminology that provides a set of terms that consistently categorizes medical information. it includes terminology for symptoms, signs, diseases and diagnoses, and therapeutic indications. it is organized by system organ class (soc), divided into high-level group terms (hlgt), high-level terms (hlt), preferred terms (pt), and finally into lower-level terms (llt). the meddra dictionary has been translated into many languages. as the terminology itself does not contain specific guidelines for its use, an ich working group has been charged to develop two guides: ▸ "meddra term selection: points to consider": this document was created to achieve consistency in the way users assign particular terms to particular symptoms, signs, diseases, etc. ▸ "meddra data retrieval and presentation: points to consider": this document provides guidance on retrieval and on sorting and presenting data in the most understandable and reproducible way for the benefit of drug development, pharmacovigilance, and risk management. these two documents provide a best practice approach for the use of meddra. they are revised for each new meddra version release. in addition, the meddra dictionary includes standardized meddra queries (smqs) that were developed (in collaboration with cioms) to facilitate the retrieval of meddra-coded data as a first step in investigating drug safety issues in pharmacovigilance and clinical development. smqs are groupings of terms from one or more meddra system organ classes (socs) that relate to a defined medical condition or area of interest. they are intended to aid in case identification. because the terminology requires constant updating and maintenance, it was agreed that a maintenance and support services organization (msso) would be needed to carry out this task and to distribute the terminology, on license, to users in industry and regulatory agencies. the msso, contracted by ich with technical and financial oversight by the meddra management board, is tasked to maintain, develop, and distribute med-dra. since the release of version . in , meddra has become the accepted international standard for all worldwide regulatory activities (meddra is not yet mandatory in the us). as a single global, standardized medical terminology, meddra speeds the exchange of clinical information, facilitating research and safety monitoring, and making the regulatory approval process more efficient and responsive. different translations of meddra have been released. in march/april , meddra was also implemented in the who vigibase, providing a global repository of meddra-coded safety data that can be used as a substantial tool for pharmacovigilance. during a meeting on october - , in yokohama, japan, the meddra management board announced fee reductions for lower revenue subscribers. these reductions are in keeping with the meddra management board's goal of facilitating the use of med-dra for all users. since january , access to meddra has been free for academic organizations, hospitals, healthcare providers, and other users involved in noncommercial activities. the objective of the electronic standards for transmission of regulatory information (estri) project was to facilitate international electronic communication. to this end, an ich multidisciplinary expert working group (called m ewg) was established during the ich meeting of in brussels, belgium. the m ewg was to evaluate and recommend estri that meet the requirements of the pharmaceutical companies and dras from the three ich regions. since , the m ewg has developed the technological framework and recommended solutions for international information exchange. this was obtained by gathering requirements, assigning specific tasks, evaluating international standards and products, and recommending a functional architecture. this project included the verification of procedures for consistent, accurate information transfer, and the evaluation of encryption technologies and key certification procedures for the transfer of regulatory information. the working group has undertaken test projects to define logical electronic communication standards in order to ensure the integrity of information and data exchange between pharmaceutical companies and authorities. to allow flexible change, some of the activities of the ewg result in recommendations that do not follow the formal ich step process. they are agreed upon in the ewg, signed by all parties of the ewg, and are endorsed by the ich steering committee at its different meetings. these recommendations, which have been modified and improved over time, provide various open international standards that allow for the international transmission of information regardless of the technical infrastructure. to facilitate the use of these recommendations, the m ewg has also developed a glossary for the technical terms. today, six m recommendations are available. they cover and standardize general aspects, but also the choice of file format and information transfer as described in table . recommendations were also prepared for the choice of physical media (i.e., floppy disks, cd-r, and dvd-ram). because these physical media are not relevant anymore, these recommendations were retired in june . in addition to the recommendations, the m ewg also developed several specifications with regard to the electronic exchange of information: ▸ the first specification developed by the m ewg was related to the electronic transfer of the individual case safety report (icsr) presented in ich guideline e b (data elements for transmission of individual case safety reports). following the development of the e b guideline, it became necessary to work on an electronic specification to guide the pharmaceutical companies on how to provide the information required by the e b guideline. indeed, successful electronic transmission of icsr relies on the definition of common data elements (provided in the e b guideline), but also a standard electronic transmission procedure. the first version of this specification was approved by the steering committee under step in . since then this specification has been modified because its implementation and use had to be aligned with the evolution of the ich e b and m (meddra) guidelines. as a result of this activity, adverse event (ae) data can be extracted, populated, and electronically transmitted in the manner specified by the ich icsr message from safety and surveillance databases. even if it has required a lot of work, the implementation of electronic reporting of icsrs based on the ich e b, m , and m standards progressed very rapidly across the ich regions. thanks to these standards, pharmaceutical companies can now exchange case reports electronically via gateway with some dras (such as the us fda or ema). ▸ the second specification developed by the m ewg was the electronic common technical document (ectd) created as the electronic message for the common technical document (ctd) detailed in ich guideline m . this specification has since been maintained by the ectd iwg. the ectd specification, based on xml (extensible markup language) technology, allows for the electronic submission of the ctd from applicant to regulator, taking into consideration the facilitation of the creation, review, lifecycle management, and archiving of electronic submissions. while the table of contents is consistent with the harmonized ctd, the ectd also provides a harmonized technical solution to implementing the ctd electronically. this ectd specification is applicable to all modules of initial registration applications and for other submissions of information throughout the lifecycle of the product, such as variations and amendments. the backbone has been developed to handle both the regional and common parts of submissions. implementation of ectd has begun across the ich partner and observer regions. for example, since january , , all electronic submissions to the us fda are required to be in ectd format. ▸ in , the m ewg published the first version of the study tagging file (stf) specification, which is supplemental to the ectd. this specification has since been modified several times. for each study included in modules and of an ectd submission, the stf includes information allowing for the identification of all the files associated with this specific study. this is additional information to the ectd backbone files that already include many items, but do not contain enough information on the subject matter of several documents (e.g., study report documents) to support efficient processing and review of applications. the common technical document (ctd) is one of the major and most well-known achievements of ich, and like all other big harmonization projects of ich, required much effort. it provides a harmonized structure and format for regulatory applications. the objective is to reduce the time and resources needed to compile applications for registration of medicines in the different ich regions. additionally, this new common format allows dras to have more consistent reviews, helping them to perform analysis across applications and to exchange information among them. before the development of the ctd, each region had its own requirements for the organization of technical reports in the submission and for the preparation of the summaries and tables. in japan, applicants had to prepare the gaiyo, which organized and presented a summary of the technical information. in europe, expert reports and tabulated summaries were required, and written summaries were recommended. the us fda had specific guidelines regarding the format and content of the new drug application (nda). in , the ich industry representatives proposed assembling the information generated during the development of a product in the same order. this proposal followed an industry survey in may that assessed the time and resources needed to convert an eu marketing authorization application (maa) into a us nda (and the reverse). this survey showed that an average of three to four months and to people were required for the conversion from one format to the other. with the acceptance in all three regions, the ctd now avoids the need to generate and compile different regional versions of most of the registration dossier sections. the ctd was adopted as an ich topic at the steering committee meeting that took place just before the ich meeting (july ). the ctd specifications reached step of the ich process at the steering committee meeting in july . after public consultation, step was achieved at the ich conference in san diego, california in november . on september , (at the washington, dc meeting), numbering and section headers were then edited for consistency and use in the ectd. the ctd consists of five modules (module is region specific, and modules , , , and are intended to be common for all regions): ▸ module includes administrative information (i.e., application form) and proposed prescribing information. ▸ module summarizes data included in modules , , and and is organized in seven subsections: • ctd the ctd is defined by a general ich guideline (m ) and three specific technical guidelines (m q, m s, and m e, which cover the quality, safety, and efficacy parts of the ctd, respectively). a q&a document is associated with each of these four guidelines to facilitate implementation of the ctd. the ich parties agreed to implement this harmonized format in the three regions by july . it is indeed used today in the three ich regions: it is mandatory in the eu and japan, and "highly recommended" in the us (the current legislation does not allow the us fda to make it mandatory). moreover, this format is also used in other countries (e.g., australia, canada, turkey, etc.), and derivatives of the ctd have been developed in other regions (e.g., the actd developed by the asean countries). this harmonized format is indeed one of the great successes of the ich process. while the realization of the ctd took many years, there is now a common format for the regulatory submissions across the three ich regions (europe, japan, and the us) and beyond. this facilitates pharmaceutical companies in making simultaneous filings in the ich regions as it eliminates the extensive work previously required to convert from one format to another. however, the ctd is not a "global dossier." it remains only a harmonization of format instead of a harmonization of content. this initial misunderstanding, certainly created by the desire of many people to accelerate the harmonization of technical requirements, led to a lot of criticism against this new format. however, the ctd cannot be a truly global identical dossier (including the same information/data/level of detail) if all technical requirements are not fully harmonized. moreover, the submission's content may also be different for several reasons, such as different individual regulations, legal status, or requirements, and different manufacturing situations for the three regions. indeed, although the ctd provides a common format for regulatory applications, the actual content must still meet local regulations, laws, and statutes. as a result, despite being presented in the same order, the required content of modules to may vary by region. for example, the integrated summary of efficacy/integrated summary of safety (ise/ iss) that were requested by the us fda before the implementation of the ctd are still needed. because these integrated summaries are unique to the us, the table of contents of the ctd does not specifically include them. a specific us fda guidance was released in june to help pharmaceutical companies decide where to place these us-specific ise/iss documents within the structure of the ctd. to conclude, even if the ctd is "only" an agreed-upon common format for the modular presentation of summaries, reports, and data, it provides obvious advantages. the ctd allows companies and dras to harmonize the terms and way of communication [ ] . having the same "language" will certainly help the harmonization of content, and ultimately the harmonization of technical requirements. indeed, regulatory reviews and communication with the applicant will be facilitated by a standard document of common elements. in addition, exchange of regulatory information between dras will be simplified. this increase of communication between authorities and between authorities and pharmaceutical companies will obviously facilitate expertise and opinion sharing (related to the safety, efficacy, and quality of the development product) in a timely manner that will ultimately provide benefits to patients by providing quality medicines more quickly on the market. like meddra, the objective of this project was indeed to support all aspects of pre-and post-approval pharmacovigilance activities as well as communication of regulatory information. for example, meddra and the harmonization of drug dictionaries are critical in the transmission of the icsr presented in ich guideline e b (data elements for transmission of individual case safety reports). the transmission of structured data (especially electronically) does imply the use of controlled vocabularies. before the ich initiative, there was no harmonized standard to document information and data on medicinal products. regulators in the different regions had established their own standards, which differed in data format, content, language, and applied standard terminology (e.g., terminology used for substances, routes of administration, pharmaceutical forms, etc.). the who drug dictionary, or a modified version of this product, was sometimes used. this lack of internationally harmonized standards related to core sets of medicinal product information and medicinal product terminology made the scientific evaluation, comparison, and exchange of drug data (especially in the area of pharmacovigilance) very difficult. the activity on the m guideline only began in . following the example and success of meddra, the ich steering committee at its meeting in november agreed to launch this new harmonization initiative and to develop a new tripartite guideline that defines the data elements and standards for drug dictionaries. during the ich meeting in tokyo, japan in february , who presented a white paper regarding the concepts of a global drug-coding dictionary. during this meeting, the steering committee agreed to convene an informal discussion group in brussels, belgium during the ich meeting in july to allow for a discussion of this proposal. an informal working group was then established to develop a concept paper and prepare a business plan. the m guideline was released for consultation at step of the ich process on may , , along with controlled vocabulary lists for routes of administration and units of measurement. this guideline was subsequently submitted to the iso for development under this process. step guideline was updated based upon feedback received during consultation in , as well as additional considerations following its submission to iso for development as an international standard. key parts of this updated guideline will be incorporated into the ich "implementation guide for identification of medicinal products message specification," which is currently undergoing development as an iso standard. ▸ achievements so far: for two decades, the ich process has achieved much success and benefited both dras [ - ] and pharmaceutical industries . more importantly, this harmonization has been pursued in the interest of patients and public health to prevent unnecessary duplication of clinical trials in humans and to minimize the use of animal testing without compromising the regulatory obligations of safety and effectiveness. to achieve this objective, the goal of ich has been to promote international harmonization by bringing together representatives from the eu, japan, and us to discuss and establish common guidelines and standards. through the ich process, considerable harmonization has been achieved in the technical requirements for the registration of pharmaceuticals for human use. this is now a mature harmonization initiative. since its creation, over harmonized guidelines have been developed in the areas of quality, safety, and efficacy in order to eliminate duplication in the development and registration process. moreover, common harmonized tools for regulatory communication (meddra, ctd, estri) have also been made available. this represents an extraordinary contribution to the global harmonization of pharmaceutical regulations. these guidelines already form a solid basis for harmonized application of technical requirements during the registration process. while the technical output of the ich process has been very positive, the importance of the unique way in which ich operates should also be noted. indeed, in addition to the practical harmonization of specific technical items, one of the major outcomes of ich has been to create a forum that allows experts from different countries and with different backgrounds to communicate, exchange, discuss, and share their experience and information in a structured manner. this is of course an essential first step to any harmonization. finally, another important achievement of ich is to be well recognized on a worldwide basis. ich guidelines have been adopted and are now followed outside the ich regions (e.g., switzerland, canada, and australia, and also many rhis). although ich's initial focus was the development of guidelines for use in the ich regions, increased globalization drastically modified the international cooperation environment. in response to a growing interest from beyond the ich regions in the use of ich guidelines, the ich steering committee took the first step in march of establishing the ich gcg. in november , new terms of reference and rules were endorsed for the gcg with the aim of establishing partnerships beyond the ich regions to promote a better understanding of ich guidelines globally. since then, rhis from across the globe, but also representatives from dras and departments of health (doh) that are either a major source of api or clinical trials data have been invited to participate in the gcg meetings and listen to technical topics at the level of the steering committee (at the biannual ich meetings). in addition, as per a decision of the ich steering committee in november , invited rhis and dras/doh may now also nominate technical experts as active members of ich ewgs. the implementation of ich recommendations and standards outside the three ich regions is indeed very important as it allows industry to better develop medicinal products for the global market. as a consequence of this expansion to non-ich regions, training and capacity building have become a key focus of the ich gcg. in , the gcg implemented a strategy for addressing training and capacity needs to help ensure the most effective use of resources, opportunities, and the realization of desired outcomes. over the past few years, the gcg has responded to numerous requests for training, providing ich expertise both for the development of training programs and for the delivery of the training itself. today, the gcg and the ich steering committee continue to implement new tools to promote a better understanding and use of ich guidelines and recommendations ] . one of the drivers of this success is in the fact that this harmonization process is based on scientific consensus developed between industry and dra experts. before ich, the industry and regulators never sat at the same table in an international forum to discuss the science of drug development in order to develop best practices across different regions. this joint effort allows not only for the involvement of the best experts (from both the authorities and pharmaceutical industries) in specific technical discussions, but also for ensuring that discussions take into account both the regional legislations and the practical impact on the development of pharmaceutical products. this inclusion of both industry and regulators increases commitments to the common goal (i.e., implementation of the ich tripartite, harmonized guidelines, and recommendations) that has obviously been a key factor in the success of ich. the results of a survey on the impact of ich, presented during the ich conference in osaka, japan, showed a high degree of satisfaction by both dras and industry with the completed ich guidelines, and continuous support from both sides for ich activities. the second driver of ich's success is linked to its well-defined structure and process. in the beginning years of ich, the steering committee organized its structure around the working groups, which included world-recognized experts. this decision was critical because it allowed ich to have very robust scientific and technical recommendations, most of the time accepted and implemented without fundamental criticism. the steering committee has of course also been key as the governing body that gives direction, selects the topics for harmonization, and ensures completion of projects in a timely manner (not always easy when one's goal is consensus). in addition to the structure, the steering committee has also been able to define a process that supported this incredible harmonization task in a structured and organized way, supported by different players such as the ich secretariat and coordinators. indeed, the stepwise approach that has been put in place for the development of guidelines (the defined five-step process with decision points at step and step ) has been very important. this approach allowed for the creation of comprehensive drafts by a small number of experts (the best environment for facilitating focused discussion and development of consensus) and public review before implementation (which promotes transparency, and avoids surprises and post-approval issues). the creation of concept papers and business plans that the steering committee put in place at a later stage are also fundamental to ( ) define clear goals, and ( ) help to monitor progress towards the predefined goals. finally, the review of progress during regular meetings also ensures commitment, follow-up, and therefore the seriousness of this initiative. finally, the extension of ich beyond the ich regions was possible because the steering committee understood early on that its activity could not be restricted to the ich regions with the increasing globalization of drug development and manufacture. indeed, research and manufacture of new products is not confined to the three ich regions any longer. clinical trials are carried out throughout the world and many non-ich countries are involved in the development and manufacture of pharmaceutical products. to increase transparency and promote collaboration outside ich regions, the steering committee accepted observers (e.g., canada), worked with other international organizations (efta and who), and involved other regions/countries in this process via the ich gcg, which evolved over time. all these actions allowed the ich work to be expanded to most of the regions/countries in the world, and its harmonization benefits to be available worldwide. the collaboration with non-ich regions is today one of the priorities of ich in order to increase commitment of these regions and facilitate worldwide implementation of ich recommendations. ▸ limitations and challenges for the future: as mentioned above, ich has been an incredible contributor to the international harmonization of pharmaceutical regulations. ich has been successful in achieving harmonization (initially of technical guidelines and then on the format and content of registration applications), and has positively impacted the global development of new drugs. all parties agree that there is a need to maintain this harmonization in the interest of the patient and public health. now that the process and networks are in place, it seems indeed obvious that ich needs to continue its activities as one of the major players in the international harmonization of pharmaceutical regulations. further harmonization activities should be continued in a focused manner. however, in an evolving international environment, some aspects of this initiative need to be reviewed as new approaches may be needed. indeed, some aspects of this initiative may be optimized in order to better handle new and future challenges. the first challenge of ich, which the steering committee has already acknowledged, is the implementation and maintenance of already developed guidelines. the current magnitude of successful harmonization actions and the need for these to remain current in a rapidly changing environment calls for focusing more effort on the implementation and monitoring of ich commitments. development of iwgs or task forces to manage this challenge will be key to its success. this focus on implementation and maintenance should not, however, impact the work on new harmonization topics that still need to be discussed. these new topics for harmonization need to be rigorously assessed for need (i.e., scientific merit/emerging science) and feasibility (i.e., expected outcome, timeline, and resource requirements). another major challenge for ich is to confirm its worldwide expansion and to continue to develop and strengthen its collaboration and partnership outside the ich regions in order to better integrate these regions into the ich process. at the time of ich establishment, it was agreed that its scope would be confined to registration of new drugs and medicines in western europe, japan, and the us because the vast majority of the new drugs were developed and manufactured in these three regions. however, since then, there has been strong involvement of other parts of the world. canada and australia are key markets for pharmaceuticals, and are often involved in global clinical studies. more recently, the emergence of other countries has been recognized in all areas, including the pharmaceutical industry. as already recognized by the ich steering committee, the success of ich in the ich regions only will not be relevant any longer. the modification of the landscape obliges ich to review and broaden its objectives. the current organization (with the gcg) that initially responded to this increased globalization may not be the most appropriate solution for future stages of development. the ich organization and systems need to be reviewed and revised to better serve these broader objectives. in (during ich ), the ich steering committee reviewed its structure and concluded that this structure continues to be appropriate. however, in order to increase transparency, they welcomed appropriate participation of other interested parties in a flexible and ad hoc manner on topics that also affected them. a decade later, the new evolving environment requires a bigger revision of its structure and process. the ich steering committee understands this urgent need and has declared that a new ich organizational structure will be adopted. the steering committee will set the framework for new rules on governance, decision making, and membership [ - ]. finally, ich has to become more proactive in new emerging topics to prevent future disharmony. the gene therapy topic is an interesting example that demonstrates the previous lack of commitment of ich to "proactive harmonization." in september , the ich steering committee established a gene therapy discussion group (gtdg) in recognition of the rapidly evolving area of gene therapy medicines. the gtdg developed several ich consideration documents in this area. despite this first positive step/outcome, the development of these consideration papers and the activities towards the development of a new multidisciplinary guideline (guideline m ) was discontinued in september because "currently the ich regions do not have the resources to support the development of further ich consideration documents" in this domain . recently, the ich steering committee started to define a new proactive approach to identify and creatively pursue advancements in science . if ich succeeds in these challenges, it will certainly become a real international organization/forum (vs. a multiregional initiative) where proactive discussion on all past and new technical requirements for registration of pharmaceuticals for human use will be discussed. however, some of these challenges are not new. ich acknowledged these challenges years ago and has already tried to resolve them without succeeding (e.g., proactivity), confirming the difficulties of this task. to face these challenges, ich needs to revise its structure and engage a new phase in order to address the evolution of regulations and the globalization of drug development and manufacturing, and to promote better proactivity in harmonization. the ongoing ich reform is obviously an important milestone toward resolution of current limitations. europe was the first major regional bloc established after world war ii. following this, there have been many regional harmonization activities throughout the world, especially over the past years. countries in different regions of the globe have organized themselves into closer economic and political entities. these movements have transformed the world, both economically and politically, as they create new opportunities and also new challenges (e.g., the management of regulations and standards disharmony). these regional harmonization initiatives include members with closer interests and needs, compared to global initiatives, allowing further harmonization and cooperation. this level of harmonization is also essential for developing countries that may not have access to all global harmonization discussions due to sparse resources or lack of expertise. regional cooperation can represent their interests and challenges and allow them to be heard at the global level. ii this level of cooperation is also essential for establishing region-wide pooled procurement systems. very diverse initiatives (each with a different scope, objective, structure, and working model) were established due to different cultural, historical, and political contexts. they range from a simple technical and scientific intergovernmental cooperation model to an advanced integration model. ii although all countries are part of who, many countries are not represented at ich where global standards are developed. however, most of the major regional harmonization initiatives are today represented via the ich gcg group. the political and economic development of each region, and sometimes subregions, has indeed shaped the level of harmonization in the pharmaceutical area: ▸ scenario -pharmaceutical harmonization in the context of an economical and political integration: in certain regions, economic integration among countries implies integration of pharmaceutical regulations and the harmonization of technical standards. this degree of integration varies from one region to another (and sometimes from one subregion/country to another), but the harmonization of regulations and policies and standards are very important to create a consistent regional legislative framework and a common certification system for products across regions. europe is the best example in terms of advanced harmonization and integration with the development of a centralized system, institutions, and procedures for the registration of medicines to be marketed in the single market. jj ▸ scenario -pharmaceutical harmonization in the context of a general political agreement: other initiatives follow a general political agreement, mostly signed to avoid conflicts or wars in certain areas in the world or to facilitate economic growth and trade within a region (e.g., asia-pacific economic cooperation [apec]), without an integration goal. the output of this harmonization initiative is variable, but most of the time does not produce a deep harmonization of pharmaceutical regulations because it is not the primary objective of the agreement and therefore the resources and efforts from the countries for this pharmaceutical regulation harmonization are variable. ▸ scenario -pharmaceutical harmonization based on a specific intergovernmental agreement: in other regions, a simple technical and scientific intergovernmental cooperation has been established, focusing solely on the harmonization of pharmaceutical regulations. this is the case of the pandhr initiative in the americas where regional integration has not been the objective because countries continue to present very different systems and degrees of development, and there are no political commitments to create a single market. countries only cooperate to promote harmonization without creating common legislation and procedure. this is a scenario that produces good harmonization of pharmaceutical regulations because this is the focus of the initiative, compared to scenario above, which is a derivative of a broader political agreement. however, the risk and difficulty of this scenario is its implementation. because there is not an ultimate economic and political goal (e.g., developing a single market as in scenario ), implementation of the agreed-upon recommendations in the national law is somewhat difficult. its success clearly depends on the commitment of each country. it is important to understand that the scenarios discussed above can also be considered as steps. harmonization is a moving process and harmonization initiatives evolve over time. for example: jj this central system is supported by the national dras that also continue to operate their own registration systems for products limited to national markets. • the european model was initiated to stop war between its countries (scenario ), but has in the time since evolved to an integration model to create further economic and political bonds ( scenario ). • asean is another evolving initiative that may follow the european model. today, it is between scenario and . this evolution to a more integrated model is obviously easier when the members are somewhat limited in number and share common geographical, historical, and cultural roots. it is indeed very difficult to imagine that apec or pandrh will evolve towards integration models such as europe or asean. the european community was created after world war ii in order to develop a more peaceful europe by promoting cooperative projects. since then, it has rapidly evolved to become a unique partnership between european countries. the main goal of the community is the progressive integration of member states' economic and political systems, and the establishment of a single european market based on the free movement of goods, people, money, and services. the european union (eu) is not a federation like the united states of america (us), nor is it simply an organization for cooperation between governments like the united nations. it is, in fact, unique in that the countries that make up the eu (its "member states") remain independent sovereign nations, but pool their sovereignty in order to gain a strength and world influence that none could have on their own. kk with approximately million people (representing % of the world's population), the eu is today the world's third largest population after china and india, representing a huge single market. the eu's gross domestic product (gdp) is now bigger than that of the us, and it is the world's biggest exporter and importer [ ] . diversity is an important characteristic of the eu as symbolized by its motto, "united in diversity," with many differences existing among its member states. this diversity is a positive attribute of the union. however, considering the official languages and the major historic, social, cultural, and economic differences between member states, its development has not been easy. its diversity has also influenced its organization and the way the harmonization process has been structured. it is therefore very important to understand the history and organization of the eu in order to understand how the european pharmaceutical regulation has been structured over time. effectively alone and where cooperative action at the community level is indispensable. these include major health threats and issues with a cross border or international impact, such as pandemics and bioterrorism, as well as issues relating to free movement of goods, services, and people. acknowledging that all countries share common values (i.e., ensure high standards of public health and equity in access to quality healthcare), it is therefore logical that the eu has developed common standards for medicines. moreover, the implementation of a single market requires harmonization of the pharmaceutical market. the ability to travel freely, or to live and work anywhere in the eu, only makes sense if eu citizens can be sure to obtain the same level of healthcare wherever they go. therefore, a number of european community rules have been adopted to ensure the highest possible degree of protection of public health while promoting the free movement of medicines in an internal market without barriers. the european commission (ec)'s role is not to mirror or duplicate national activities, but to coordinate them. work on healthcare at the community level adds value to member states' actions, particularly in the area of illness prevention, including activity on the safety and efficacy of medicines [ ] . today, the european pharmaceutical system is well developed and the vast majority of requirements have been harmonized. this successful european cooperation in pharmaceuticals is also recognized on a worldwide basis due to its major contribution to the global harmonization of pharmaceutical regulations (via its active involvement in international initiatives such as ich and who). today the eu is composed of member states, but the size of the eu has changed over time as it has continually expanded since european integration first began in with only six countries ( table ). the final three enlargements (in , , and ) expanded the eu member states from to , and were rooted in the collapse of communism. it was a historic advancement that offered an unexpected and unprecedented opportunity to extend the union into central and eastern europe. today, the landmass of the eu covers million km ll and can rightly claim to represent a continent (plate ). stretching from the atlantic ocean to the black sea, it reunites western and eastern europe for the first time since they were separated by the cold war. in the future, the eu will continue to grow as an increasing number of countries express interest in membership. the treaty on european union sets out the conditions for such accession (articles and ): any european country which respects the principles of liberty, democracy, respect for human rights and fundamental freedoms, and the rule of law may apply to become a member of the union. the applicant country must meet a core of criteria (e.g., having stable institutions and a functioning market economy) in order to ensure that eu principles will be respected and that eu rules and procedures will be effectively implemented. this is a long and rigorous process that starts when the country submits an application to the council. today, iceland, the former yugoslav republic of macedonia, montenegro, turkey, albania, bosnia and herzegovina, kosovo, and serbia are candidates to join the eu, some of these countries being in more advanced stages of negotiation with the eu than others. membership is only granted when the necessary requirements are met and when candidate countries have demonstrated that they will be able to fulfill their part as members. in the eu regulatory network. for example, the ipa program supported the participation of nominated representatives of the concerned countries in selected meetings and training courses as observers. the program also supported the organization of conferences to prepare the countries for integration into the european regulatory network for medicines. these activities helped identify areas where additional action might be needed to ensure the smooth transposition of the eu "acquis communautaire" mm into the national legislation of these future eu member states. ▸ the specific case of iceland, liechtenstein, and norway: in july , iceland submitted its application for eu membership and the accession negotiations have now been opened. norway, despite two failed attempts by referendum to enter the european community in and the eu in , remains undecided whether or not it will apply once again for eu membership. presently, however, neither norway nor liechtenstein are candidates for eu membership. however, even if these three countries are currently not part of the eu, it is important to note that they have a specific strong relationship with the union through the european economic area (eea) agreement that entered into force on january , . this agreement allows these three eea european free trade association (efta) states nn to participate in the eu internal market on the basis of their application of internal market relevant acquis. oo all new relevant community legislation is dynamically incorporated into the agreement and thus applies throughout the eea, ensuring the homogeneity of the eu internal market. also, the eea agreement allows for eea-efta states to participate in the internal market's relevant community programs and agencies, albeit with no right to vote. in the pharmaceutical sector, norway, iceland, and liechtenstein have adopted the complete community acquis on medicines, and are consequently parties to the european procedures. in the case of the centralized procedure, the representatives from these three countries do not vote, but their position is stated separately in the opinion, where relevant, in the minutes of the committee and in the case of divergent opinions appended to the committee's opinion. their position is not counted in reaching the committee's opinion [ ] . according to decision no. / of the eea joint committee (which entered into force on january , ), when decisions on approval of medicinal products are accepted by the community, these three countries will accept corresponding decisions on the basis of the relevant acts. the liechtenstein authorities have transposed into their national legislation a provision that makes commission decisions automatically applicable on their territory. however, legally mm "acquis communautaire" is a french term referring to the cumulative body of eu laws, comprising the ec's objectives, substantive rules, policies, and in particular, the primary and secondary legislation and case law -all of which form part of the legal order of the eu. nn the european free trade association (efta) is an intergovernmental organization set up for the promotion of free trade and economic integration to the benefit of its four member states: iceland, liechtenstein, norway, and switzerland. although switzerland has many agreements with the eu, it is today not part of the eea agreement due to the rejection of accession by the swiss people. oo the eea agreement is concerned principally with the four fundamental pillars of the internal market, "the four freedoms" (i.e., freedom of movement of goods, persons, services, and capital). binding acts from the community (e.g., commission decisions) do not directly confer rights and obligations in norway and iceland, but first have to be transposed into legally binding acts in these states [ ] . since the end of world war ii, the eu has steadily become more established and organized. the unique european model (not a federation but a more integrated than simple cooperation between governments) requires a complex organization that not only protects the independent sovereignty of the member states, but also allows for the delegation of some of decision-making powers to shared supranational institutions. today, the structure in place was specifically designed to represent the interests of the community, the member states, and the european citizens. within this overall european structure and context, many special domains have been harmonized and organized to support the functioning of the single market. a number of institutions, committees, and technical bodies ( table ) play a significant role in the european pharmaceutical system. the roles and characteristics of these are briefly described in the following sections. ▸ the european parliament is the directly elected eu institution that represents the interests of the eu's citizens. its members are elected once every five years. its origins go back to the s and the founding treaties, but the lisbon treaty significantly increased its role in the decision-making process and budget approval. its legislative powers were reinforced by the extension of the co-decision procedure. today the european parliament is firmly established as a co-legislator, has budgetary powers, and exercises democratic control over all the european institutions. its work is organized through a system of specialized committees that review and prepare legislative proposals and reports to be presented at the plenary assembly. the committee on the environment, public health and food safety is responsible for the legislation covering pharmaceutical products and the ema. the european parliament has three working locations: brussels (belgium), luxembourg, and strasbourg (france). luxembourg is home to the administrative offices of the general secretariat. meetings of the entire parliament, known as "plenary sessions," take place in strasbourg and sometimes in brussels. committee meetings are also held in brussels. ▸ the council of the european union represents the individual member states. it meets in different configurations and is attended by one minister from each of the eu's national governments (depending on the agenda). health-related discussions are handled by the employment, social policy, health and consumer affairs council (epsco). as with the european parliament, the council was set up by the founding treaties in the s. it is a key decision-making body that, among other responsibilities (e.g., coordination of the eu's economic policies and foreign and security policy) shares lawmaking and budgetary powers with the european parliament. its work is facilitated by the committee of permanent representatives (coreper), which is responsible for preparing the work of the council of the european union (all issues must pass through coreper before they can be included in the agenda for an eu council meeting). this committee consists of the member states' ambassadors to the eu. these permanent national representatives and their team are located in brussels, belgium, and protect national interests at the eu level. ▸ the european commission (ec) is independent of national governments as it represents and upholds the interests of the eu as a whole. it acts as the "guardian of the treaties" but remains politically accountable to the parliament. like the parliament and council, the ec was set up in the s under the eu's founding treaties. a new commission, which is formed by a president (designated by the member states and approved by the parliament) and the "commissioners" (each of them responsible for a specific policy area), is appointed every five years. its role is to draft proposals for new european laws (which are presented to the european parliament and the council for adoption). it is also the eu's executive arm because it is responsible for implementing the decisions of the parliament and the council. this means managing the day-to-day business of the eu: implementing its policies, running its programs, allocating its funds, and representing the eu in international negotiations. the day-to-day running of the commission is done by its administrative officials, technical experts (via its various committees and groups), translators, interpreters, and secretarial staff (which represent more than , people). this staff is organized in departments, known as directorates-general (dg), and "services" (such as the legal service). the overall coordination is provided by the secretariat-general. each dg is responsible for a particular policy area and is headed by a director-general who is answerable to one of the commissioners. the regulation of medicinal products was previously under the dg enterprise and industry, but this policy area has been transferred to the dg health and consumers (sanco) as of march , . the commission is based in brussels (belgium), but it also has offices in luxembourg, representation in all eu countries, and delegations in many capital cities around the world. this "institutional triangle" produces the policies and laws (such as european pharmaceutical legislation) that apply throughout the eu. the court of justice upholds the rule of these european laws and makes sure that this eu legislation is interpreted and applied in the same way in all eu countries. the other institutions of the eu (the european council and the court of auditors) are critical for the functioning of the eu, but are not directly involved with the development and harmonization of pharmaceutical legislation. the eu institutions are supported by a number of other bodies (e.g., the european central bank, the european ombudsman, etc.). specialized agencies (e.g., the ema, the european centre for disease prevention and control, and the executive agency for health and consumers) have also been established to handle certain technical, scientific, or management tasks. this agency is headed by an executive director (who is its legal representative responsible for all operational and staffing matters) and has a staff of about full-time members [ ] . the management board is the supervisory body responsible for setting the agency's budget, approving the annual work program, and ensuring that the agency works effectively and cooperates successfully with partner organizations across the eu and beyond. in addition to its staff, the ema is composed of seven committees that conduct the main scientific work of the agency. these committees and their characteristics are reviewed below: • human use. the chmp plays a vital role in the eu marketing procedures as it is responsible for: -conducting the initial scientific assessment and issuing opinions on an maa for medicines registered via the centralized procedure (these opinions are used by the ec as a basis for its legally binding decisions) -coordinating post-marketing activities for medicines registered via the centralized procedure -arbitrating disagreements between member states during mutual recognition and decentralized procedures (arbitration procedure) -acting in referral cases, initiated when there are concerns relating to the protection of public health or where other community interests are at stake (community referral procedure) this committee (and its working parties) also provides assistance to companies during development, prepares scientific and regulatory guidelines, and cooperates with international partners on the harmonization of regulatory requirements for medicines. • the committee for orphan medicinal products (comp), established by regulation (ec) no / , is charged with reviewing applications from companies seeking "orphan medicinal product designation" for products they intend to develop for the diagnosis, prevention, or treatment of rare diseases (so-called "orphan drugs"). this committee is also responsible for advising the european commission on the establishment and development of a policy on orphan medicinal products in the eu, and assists the commission in drawing up detailed guidelines and liaising internationally on matters relating to orphan medicinal products. • submitted by pharmaceutical companies, and to adopt opinions on these plans. this includes assessing applications for full or partial waivers and assessing applications for deferrals of pediatric studies. this committee also assesses data generated in accordance with the agreed-upon pips, provides opinions on the quality, safety, or efficacy of a medicine for use in the pediatric population (at the request of the chmp or a member state), and supports the development of the european network of pediatric research at the european medicines agency (enpr-ema). ss • the committee for advanced therapies (cat) is a multidisciplinary committee established in accordance with regulation (ec) no / . it is responsible for providing scientific opinions on advanced-therapy medicinal products (atmps) and any scientific questions related to this field. for example, it prepares a draft opinion on each atmp application before the chmp adopts a final opinion on the granting, variation, suspension, or revocation of a marketing authorization for the medicine concerned. • the committee for medicinal products for veterinary use (cvmp) is responsible for preparing the agency's opinions on all questions concerning veterinary medicinal products. • the pharmacovigilance risk assessment committee (prac) is the last committee established by the ema to implement the new eu pharmacovigilance legislation. it is responsible for assessing and monitoring safety issues for human medicines. this includes the detection, assessment, minimization, and communication relating to the risk of adverse reactions, while taking the therapeutic effect of the medicine into account. it also has responsibility for the design and evaluation of post-authorization safety studies and pharmacovigilance audits. its recommendations are considered by the chmp when it adopts opinions for centrally authorized medicines and referral procedures, and by the cmdh when it provides a recommendation on the use of a medicine in member states. these ema scientific committees are comprised of members of all eu and eea-efta states (iceland, liechtenstein, and norway); some committees include patients' and doctors' representatives. they are supported by a number of working parties and related groups that have expertise in a particular scientific field. the committees consult with them on scientific issues relating to their particular field of expertise and delegate to them certain tasks associated with the scientific evaluation of an maa or drafting and revision of scientific guidance documents. in particular, the chmp is supported by an important number of groups (i.e., the biologics working party, the scientific advice working party, or the numerous scientific advisory groups specialized by therapeutic area); some are standing parties and some temporary groups. all these groups are made up of members selected from the european expert list maintained by the ema. indeed it is worth noting that the ema evaluation system works through a network of european experts made available to the agency by the national dras of all eu member states and of the three eea-efta states (iceland, liechtenstein, and norway). this system brings together the scientific resources and expertise of all these countries in a network of over , european experts who serve as members of the agency's scientific committees, working parties, or scientific assessment teams. the ema is today considered as the model of fruitful cooperation between national dras, working together within a community body to serve community purposes. also, to ensure that the european system is accessible to everyone, in the ema launched a dedicated office to provide special assistance to small-and medium-sized enterprises ( -for the collection, preparation, storage, distribution, and appropriate use of blood components in blood transfusions -for the transplantation of organs, tissues, and cells the role of the edqm is essential in europe in facilitating mutual recognition of quality control tests carried out on medicines and ensuring that patients receive the same quality of pharmaceutical products throughout europe. there is a substantial amount of interaction between the ema and the edqm. for example, the edqm representatives participate as observers of the ema's quality working party (qwp) and biologics working party (bwp) meetings, the gmp inspection services group meetings, as well as hmpc meetings at the ema. it is important to note that the european member state plays a significant role in the european pharmaceutical system. the ema works closely with the eu member states as well as the eea-efta countries (norway, iceland, and liechtenstein). member state representatives are members of the agency's management board while the agency's scientific committees and its network of , scientific experts are nominated by the member states. without their support and expertise, the ema would be unable to deliver on its responsibilities and mandate as laid down in european legislation. it is also important to realize that many medicines available in europe are not authorized by the ec on the recommendation of the ema. many products are still approved and supervised by the national dras via the mutual recognition procedure, the decentralized procedure, or national procedure. to coordinate their efforts, the member states established the heads of medicines agencies (hma) group, which is a network of the heads of the national dras. this hma is comprised of more than national agencies, some also having responsibility for veterinary products, medical devices, and cosmetics, and also pricing and reimbursement of products. the ema is also a member of the hma. the first meeting of the hma took place in amsterdam (the netherlands) at schiphol airport, on february , . the hma is focused on eu coordination and harmonization, decision making, and consensus on strategic issues of the european medicines regulatory network. its aim is to foster an effective and efficient european medicines regulatory system. more specifically, it works towards the following key objectives [ ]: ▸ addressing key strategic issues for the european medicines regulatory network, such as the exchange of information and sharing of best practices ▸ collectively being responsible for all areas of medicines regulation, including the mutual recognition and decentralized procedures ▸ focusing on the development, coordination, and consistency of the network ▸ supporting the network by providing high-quality professional and scientific resources ▸ providing a focus for making the most effective use of scarce resources across the network, such as developing and overseeing arrangements for work sharing to fulfill these objectives, the hma has been working on both general issues (i.e., strategy for telematics, and regulatory and scientific training) and technical and scientific topics (i.e., harmonization of clinical trials, coordination of products testing, and european risk management strategy) is support of the european medicines regulatory network. the hma's website contains the mri product index database, which includes all medicines approved in the member states according to the mutual recognition procedure. one interesting program that has been developed is the benchmarking of european medicines agencies (bema). the bema program assesses the systems and processes in individual agencies against a set of agreed-upon indicators. this is a good opportunity to exchange best practices and ensure harmonization of practices (i.e., assessment, inspection, etc.) between regulators within the network. coordination among the national competent authorities is not a simple task due to the heterogeneity of these national organizations. indeed, these authorities differ in size, historic origins, roles, resources, expertise, and funding. acknowledging these differences and also the legal, scientific, social, political, and financial challenges facing the network, the hma adopted a strategic paper that provides a plan of action for - [ ] . this second plan (the first one covered - ), highlights a number of key themes and areas of focus (i.e., pharmacovigilance, clinical trials, and communication) and also the need for international cooperation. the hma is supported by the heads of medicines agencies management group, the permanent secretariat, and working groups covering specific areas of responsibility. iceland, and liechtenstein) appointed for a renewal period of three years. observers from the european commission and accession countries also participate in the meetings. it also has many interactions with the ema to facilitate harmonization in several areas (i.e., pediatric regulation, variation regulation, and pharmacovigilance). it holds monthly meetings at the ema (which also provides the secretariat of the cmdh). in practice, approximately half of the time of the cmdh meeting is dedicated to discussions on procedural and regulatory issues, development of guidance documents, and oversight of the activities of the various cmdh subgroups and working groups, while the other half is devoted to trying to reach agreement for applications referred to the cmdh in the case of disagreement between member states. the gradual harmonization of pharmaceutical regulation in the eu has been dictated by the development and expansion of the community. it represents a good example of successful harmonization and also demonstrates the influence of the political and economical decisions on the harmonization process and its outcomes. ▸ the birth of the european union: the historical roots of the eu lie in world war ii. following this bloody, horrific war, several leaders in europe wanted to ensure that war could never happen again. their goal was to develop a peaceful europe and to stop the frequent wars via the promotion of cooperative projects. this initiative has been critical but not easily accomplished due to the post-war geopolitical situation and the beginning of the -year-long cold war that split europe into east and west. on september , , winston churchill called for a "kind of united states of europe" in a speech given at the zurich university. many attempts at cooperation were made in the following years (e.g., the customs convention between belgium, luxembourg, and the netherlands, and the organization for european economic cooperation). in , west european nations created the council of europe. uu it was a first step towards cooperation between them, but some countries wanted to go ever further. on may , , france's foreign minister robert schuman presented a plan for deeper cooperation and for the creation of an organized europe, which would prove indispensable to the maintenance of long-term peaceful relations. this proposal (known as the "schuman declaration") is considered to be the beginning of the creation of what is now the eu. may has since been designated as "europe day" to celebrate this event. the idea of this plan (inspired by jean monnet, top advisor of the french government) was to promote european peace by ( ) eliminating the age-old opposition of france and germany, and ( ) creating a framework and organization open to the participation of the other countries in europe. it proposed that the franco-german production of coal and steel be placed under a common high authority and that this new productive unit be open to all european countries willing to participate. the double objectives of this proposal were ( ) to set up common foundations for economic development as a first step in the federation of europe, and ( ) to make war materially impossible [ ] . based on the schuman plan, six countries (germany, france, italy, the netherlands, belgium, and luxembourg) signed the treaty of paris on april , to establish the european coal and steel community (ecsc) in order to run their coal and steel industries under a common management. it is important to note that the independence and the powers of the high authority have been critical, and differentiated the eu from other traditional intergovernmental organizations. indeed, the establishment of the ecsc was the first step towards a supranational europe. for the first time the six member states of this organization relinquished part of their sovereignty, albeit in a limited domain, in favor of the european community. building on the success of their first treaty, the six countries decided to expand cooperation to other economic sectors. on march , , under belgian minister for foreign affairs, paul-henry spaak, they signed the treaty of rome, establishing the european economic community (eec) (or "common market") allowing persons, goods, services, and capital to move freely across borders. the same day, they also signed a second treaty to create the european atomic energy community (euratom). despite the construction of the berlin wall in august , which increased the division between the east and the west, the cooperation between european countries continued to increase in different areas (e.g., food and agriculture, aerial navigation, the environment, etc.). on july , , the six countries created the world's largest trading group by removing customs duties on goods imported from each of the six countries to the others, allowing free cross-border trade for the first time. they also applied the same duties on their imports from outside countries. this eu internal market was reinforced in with the adoption of the "single european act" (which entered into force on july , ) to remove the final obstacles. in , the single market and its four freedoms (movement of goods, services, people, and money) had finally been fully established. additional agreements, such as the schengen agreement in , have since been signed to further facilitate movement within europe. today, this single market represents the core of the eu. in , following the collapse of communism across central and eastern europe and the dissolution of the pacte de varsovie, a decade began that would be critical for the future of europe. on december , , eu leaders agreed to start the process of membership negotiations with countries of central and eastern europe (bulgaria, the czech republic, estonia, hungary, latvia, lithuania, poland, romania, slovakia, and slovenia). the mediterranean islands of cyprus and malta were also included. in december , treaty changes agreed to in nice (france) and finally signed on february , were entered into force on february , and opened the way for enlargement of the eu by reforming its institutions and voting rules. this enlargement to the eastern european countries became effective on may , and january , . six years later, on july , , the accession of croatia brought the number of member states to countries. a single currency (euro [€]) was introduced on january , in countries (joined by greece in ) for commercial and financial transactions only. notes and coins were introduced in january . this introduction of the single currency followed a long stepwise process that started in the s with the creation of the "exchange rate mechanism" to maintain monetary stability. the next important step of integration (i.e., development of a political union with fully functioning institutions) took time and faced many challenges. the debate on the "constitutionalization" of europe started in when the european parliament adopted altiero spinelli's report proposing, in a "draft treaty on european union," a fundamental reform of the european community. in the s, two important treaties transformed the community: ▸ the treaty on european union (signed in maastricht [the netherlands] on february , , entered into force on november , ) represented a new stage in european integration as it opened the way to political integration. it was a major eu milestone, introducing the concept of european citizenship and setting clear rules for the future single currency and for foreign and security policy. under the treaty, the name "european union" officially replaced "european community." ▸ the treaty of amsterdam (signed on october , , entered into force on may , ), built on the achievements of the treaty from maastricht, laid down plans to reform eu institutions, gave europe a stronger voice in the world, and concentrated more resources on employment and the rights of eu citizens. building on this transformation of the community, the adoption of a european constitution and major institutional reform became an important topic of discussion for two reasons: ▸ succeeding treaties have spurred progress in the building and reforming of europe and its institutions. this long process marked by ever-closer integration progressively transformed europe from an economic community to a political union. ▸ the combination of the various treaties and protocols signed over years has made the european structure and legislation more and more complex. although the eu will certainly continue to grow, it is difficult to predict the next steps of integration due to the current geopolitical situation and the instability caused by the financial crisis. the evolution of pharmaceutical regulation harmonization and cooperation in europe represents an excellent example and model that needs to be analyzed in detail as it shows the different important steps necessary for harmonization success. a large body of legislation has been developed, with progressive harmonization requirements since the s. the first european directive related to pharmaceutical products (directive / /eec [ ]) was signed on january , . this text provides the european definitions of a "medicinal product" and a "substance" and set up some fundamental principles for the creation of the european pharmaceutical system such as: ▸ no medicine may be placed on the market of a member state unless a marketing authorization has been issued by the competent authorities following the review of an application submitted by the person responsible for placing that product on the market. ▸ quality, safety, and efficacy are the basis for the evaluation of an application by the competent authorities. ▸ the information included in the application should be updated on a regular basis. following this first directive, many texts followed over the years to further detail the european principles and requirements led by directive / /eec, to organize and structure the european system, and to add new requirements related to specific types of products or emerging problems. major texts and important steps in the development of the european pharmaceutical system are discussed below. however, it is important to note that many other legislative texts, guidelines, and other recommendations (including harmonized quality, and nonclinical and clinical requirements) have been prepared and released over the years to support the major legislatives texts listed in this section. directive / /eec was complemented by two additional directives (directives / /eec and / /eec) in may to provide further details on the analytical, nonclinical, and clinical standards and protocols to be applied during the development of medicines, and how the results of such studies should be presented in the maa. directive / /eec also established the idea of expert reports (that would later influence the structure of the ctd), the cpmp (that would later be part of the ema), and the first multi-state licensing procedure, which would then evolve progressively to become the current mutual recognition procedure (mrp). further clarification of requirements was provided by directive / /eec (which also modified the multi-state licensing procedure to facilitate its use), and directive / / eec (which established the notion of combination products and created a route for abridged applications in case of generics and literature-based applications). in , directive / /eec established the concertation procedure, which provided a simple community-wide licensing opinion (via a mandatory referral to the cpmp) for all new biotechnology products and optionally for high technology medicinal products [ ] . it was an important new step in building the european pharmaceutical system as this new procedure (the forerunner of the current centralized procedure) required further cooperation between national dras compared to the multi-state licensing procedure previously established. however, both procedures were still based on voluntary cooperation between the relevant national authorities, and each member state remained solely responsible for granting the marketing authorization. in , legislators extended the scope of the previous directives to specific types of products: vaccines, toxins or serums, and allergens (directive / /eec); radiopharmaceuticals (directive / /eec); and products derived from human blood or human plasma (directive / /eec). additionally, on april , , directive / /eec laid down the first common measures related to genetically modified organisms (gmos); several additional texts have since then been released on this topic over the years. finally, extension of the scope of the harmonization of homeopathic products was only made in via the adoption of directive / /eec. in , directive / /eec, which laid down the principles and guidelines of gmp, was adopted. in , four new directives covering the distribution of medicines were adopted to further establish the eu internal market and facilitate the free movement of products. they especially harmonized wholesale distribution (directive / /eec), the classification of products as subject to medical prescription or not (directive / /eec), the labeling of products (directive / /eec), and advertising principles (directive / /eec). despite all these texts adopted since , the resulting progress of completing the single market in pharmaceuticals was not satisfactory. it was therefore decided to fundamentally improve the authorization procedures. a new european pharmaceutical system was then created in (but only implemented in january ). this new system, still in place today, is based on two major texts that established, for the first time, "european decisions" binding to the member states: ▸ following the adoption of these european procedures, it was necessary to harmonize the system to vary the terms of marketing authorization. this was done via the adoption of two regulations in : regulation / (for the mrp) and regulation / (for the centralized procedure). additionally, acknowledging the increased complexity of the european pharmaceutical legislation, it was agreed to assemble all previous directives in one single text. this codifying directive, directive / /ec adopted on november , , was necessary because all the directives adopted since had been frequently and substantially amended. therefore, this directive regroups all legal requirements agreed-upon since (except requirements and legal provisions provided by regulation / ). this new directive has already been amended several times since its adoption, some of these amendments being the result of a major general review of the legislation and system discussed below. in , as directed by regulation / (article ), the commission conducted a major review of the operation of the new system implemented in . the goal of this audit, contracted out to independent auditors, was to review the extent to which the results achieved over the first five years have met the objectives (namely to enhance the creation of a single market in medicinal products, while ensuring the protection of public health and the development of the pharmaceutical industry). the audit report [ ] , known as the "cameron mckenna andersen report," includes the results of the extensive consultation carried out involving individual companies, all dras responsible for the authorization of medicines and the emea, patient and professional associations, trade associations, and relevant ministries. this audit highlighted the overall satisfaction with this new system, as both procedures had been perceived as contributors in both a qualitative and quantitative way to create a harmonized european community pharmaceutical market. ninety-two percent ( %) of companies and % of dras in the eu were satisfied or very satisfied with the centralized procedure. there was also general recognition of the very considerable contribution made by the emea and the eu telematics strategy to the successful operation of the system. however, this report also identified several issues and listed several possible improvements to the system. these criticisms were primarily directed towards the mpr for which it was agreed that the lack of real supervisory, management support, and liaison between member states had altered the application of the central principle of mutual recognition. concerned member states were continuing to assess applications. regarding the centralized procedure, it was felt that it should be opened up to a broader range of products and that the "decision-making process" of the commission (post-cpmp opinion) should be reduced and improved. finally, it was also interesting to note that the european procedures had not yet produced any real dividends in terms of cost efficiencies through economy of scale. there was also a need to reduce the administrative burden where there were no public health implications (e.g., in relation to minor variations to existing approvals). this evaluation of the regulatory processes was not only very timely with the emerging technical challenges (e.g., gene therapies, etc.), but also with the political challenges in preparation for eu expansion [ ] . indeed, there was little doubt that the upcoming major enlargement of the eu (in , and involving additional countries) would accentuate the weaknesses of the system if both the structural and process issues were not resolved by then. based on this review of the eu pharmaceutical legislation and various public hearings, the ec concluded that on the whole the system had proven appropriate and suitable for its purpose and therefore it was recommended that it keep its main principles and structures. however, the ec also proposed several adaptations of the system and legislation in order to better achieve four major objectives [ ]: ▸ assure a high level of public health protection, notably by increased supervision of the market through the strengthening of inspection procedures and of pharmacovigilance. ▸ complete the single market for pharmaceutical products, taking into account the stakes of globalization, and establish a regulatory and legislative framework that favors the competitiveness of european industry. ▸ respond to the challenges of the future enlargement of the eu. ▸ rationalize and simplify the system and improve its overall coherence and visibility and the transparency of its procedures. these proposals, such as opening up the centralized procedure to a broader range of products, establishment of a fast track procedure and conditional authorization, improvement of the transparency of the system, strengthening pharmacovigilance and supervision requirements, abolition of the renewal, control of the effective use of marketing authorization with the "sunset clause," improvement of the decision-making process after cpmp opinion, re-organization and increase of the role of the emea and its committees, major modifications to the mrp and creation of the decentralized procedure, and harmonization of data protection periods [ , ], have been further debated with the parliament and the council over subsequent years. most of them have finally been implemented via the adoption of new or revised legislation and/or guidelines. one of the major legislative impacts has been the adoption of regulation ( finally, in addition to these critical texts that created the european system and general requirements, it is worth mentioning the following additional legislative texts adopted over the past years on important specific subjects (see part i- . the current european pharmaceutical system has progressively developed over the years via the adoption of agreed-upon policies. since many texts have been adopted with the aim of achieving a single market for pharmaceutical products. as noted above, several european institutions and technical bodies, together with the eu member states, are involved in the harmonization of european pharmaceutical regulation. the european harmonization process lies in the adoption of eu laws [ ] that can be categorized as follows: ▸ the "primary" legislation: the treaties are binding agreements between eu member countries. they state eu objectives, rules for eu institutions, how decisions are made, and the relationship between the eu and its member states. they also form the basis or ground rules for all eu actions. this means that every action taken by the eu is founded on treaties that have been approved voluntarily and democratically by all eu member countries. for example, if a policy area is not cited in a treaty, a law cannot be proposed in that area. ▸ the "secondary" legislation: this is derived from the principles and objectives set out in the treaties. it includes the following texts: • regulations are the most direct form of eu law. as soon as they are passed, they have binding legal force throughout every member state and must be applied in its entirety across the eu. national governments do not have to take action themselves to implement eu regulations (i.e., regulations do not require any transposition by the national authorities). • directives are legislative acts that set out a goal that all eu countries must achieve. national authorities have to adapt their laws to meet these goals, but are free to decide how to do so. vv directives are used to bring different national laws in line with each other, and are particularly common in matters affecting the operation of the single market (e.g., product safety standards). they may concern one or more member states, or all of them. • decisions are individual acts relating to specific cases and are addressed to specific parties. they are binding only on those to whom they are addressed (e.g., an eu country or an individual company), and are directly applicable (no need for implementation into national law). decisions can come from the eu council (sometimes jointly with the european parliament) or the ec. vv each directive specifies the date by which the national laws must be adapted (giving national authorities room to maneuver within the deadlines necessary to take account of differing national situations). • recommendations are not binding, but allow the institutions to make their views known and to suggest a line of action (without imposing any legal obligation on those to whom it is addressed). • opinions are not binding. they are an instrument that allows the institutions to make a statement in a nonbinding fashion; in other words, without imposing any legal obligation on those to whom it is addressed. they can be issued by the main eu institutions (commission, council, parliament), the committee of the regions, and the european economic and social committee. the european parliament and the council of the eu share legislative power, which means they are empowered to adopt european laws (directives and regulations). in principle, it is the commission that proposes new "legislative texts," ww but it is the parliament and council that adopt them. the commission and the member states then implement them, and the commission ensures that the laws are correctly applied. the vast majority of european laws are adopted jointly by the european parliament and the council using a procedure known as "co-decision." xx this means that the directly elected european parliament has to approve eu legislation together with the council (the governments of the eu countries). in addition to this "ordinary legislative procedure," there are also other special legislative procedures (which apply only in specific cases) where the parliament has only a consultative role. the requirements and procedures for the marketing authorization of medicinal products, as well as the rules for variations to the terms of marketing authorizations and for the constant supervision of products after they have been authorized, are primarily laid down in directive / /ec and regulation (ec) no / (and their subsequent amendments). these texts additionally lay down harmonized provisions in related areas such as the manufacturing, wholesaling, or advertising of medicinal products for human use. in addition, various laws have been adopted to address the particularities of certain types of medicinal products and promote research in specific areas. in addition to the legal texts, many additional community or international documents and recommendations have been developed and support the harmonization and cooperation in the eu. the "introduction and general principles" of annex of directive / /ec, as ww the european commission is the only institution empowered to initiate legislation. before proposing a new text, it assesses the potential economic, social, and environmental consequences that they may have by preparing "impact assessments" (which set out the advantages and disadvantages of possible policy options) and by consulting interested parties. the commission will propose action at the eu level only if it considers that a problem cannot be solved more efficiently by national, regional, or local action. this principle of dealing with things at the lowest possible level is called the "subsidiarity principle," and has been reaffirmed in the lisbon treaty. xx the co-decision procedure was introduced by the maastricht treaty on european union ( ) , and strengthened and made more effective by the amsterdam treaty ( ) . with the lisbon treaty that took effect on december , , this procedure has been renamed "ordinary legislative procedure" and has become the main legislative procedure of the eu's decision-making system. amended, acknowledged these scientific and technical recommendations (i.e., "the rules governing medicinal products in the european community," ich guidelines, and monographs of the european pharmacopoeia). all community rules in the area of medicinal products for human (and veterinary) use are compiled in "the rules governing medicinal products in the european union" (eudralex), published by the ec. volume of this publication contains the body of the eu pharmaceutical legislation (i.e., regulations, directives, decisions, etc.). the subsequent volumes include guidelines yy developed to support this basic legislation: zz ▸ volume (also known as "notice to applicants"), first published in , contains all regulatory guidelines related to procedural and regulatory requirements (i.e., the presentation and content of the dossiers), and also the application forms. it was prepared and is regularly updated by the european commission in consultation with competent authorities of the member states and the ema. this notice has no legal power. in case of doubt, therefore, reference should be made to the appropriate community directives and regulations. also, in july , the information contained in chapter of volume a (concerning general information on procedures for marketing authorization) was transferred to ema and cmdh websites. ▸ volume consists of all the scientific guidelines for medicinal products for human use prepared by the committee for medicinal products for human use (chmp) in consultation with the competent authorities of the eu member states. the guidelines are intended to provide a basis for practical harmonization in the manner in which the eu member states and the ema interpret and apply the detailed requirements for the demonstration of quality, safety, and efficacy contained in the community directives. an updated list of scientific guidelines is accessible on the ema website. ▸ volume contains guidance for the interpretation of the principles of gmps for medicinal products for human and veterinary use. ▸ volume contained pharmacovigilance guidelines for medicinal products for both human use (volume a) and veterinary use (volume b). volume a was replaced by the ema "guidelines on good pharmacovigilance practice (gvp)" in [ ] . ▸ volume contains guidance documents applying to clinical trials. finally, in addition to the published rules listed above, a lot of other documents that do not have the status of a law or guideline (i.e., questions and answers [q&a], recommendations, public statements, position papers, reflection papers, etc.) are released by the ema to provide additional guidance. moreover, templates (e.g., assessment templates and guidance), internal standard operating procedures (sops), work instructions (wins), and policy covering both general and specific topics (e.g., pharmacovigilance, inspection, etc.) have been developed by the ema to improve consistency in activities and evaluations and to help ease the exchange of information. many technical requirements have been harmonized and published in europe to ensure that medicinal products throughout europe are of equal quality, safe, and efficacious. these are the three basic criteria that are always evaluated and taken into consideration when establishing the risk and benefit ratio. these criteria are evaluated through the quality, nonclinical, and clinical information included in all applications. of course, the level of quality/nonclinical/clinical documentation varies depending upon the type of products and the level of development, but they are always the basis of approval for the registration of a clinical trial or a new product. legal provisions related to these technical requirements are included in annex of directive / /ec and other relevant regulations or directives. in addition, scientific and technical guidelines are also prepared by the ema's committees (i.e. chmp, comp, pdco, etc.) and its working parties (in consultation with the competent authorities of the eu member states). guidelines developed by other technical bodies (e.g., the european pharmacopoeia) or international bodies are also used in europe. for example, europe is a founder and member of ich, and therefore all ich guidelines are also applicable in europe. ▸ quality: many european requirements are in place regarding the quality of the products (active substance, excipients, and finished products). detailed scientific guidelines have been developed to adequately cover pharmaceutical development, manufacture, packaging, control (i.e., specifications, analytical procedures and validation, and impurities), stability evaluation, and post-approval changes. moreover, guidelines for certain types of products (i.e., biologics, radiopharmaceuticals, medicinal gases, or herbal medicinal products) have been specifically released to take into account their specific challenges. these technical and scientific guidelines, together with the q&a document, provide a common interpretation of the european legislation and ensure harmonization of quality requirements. also, in addition to these guidelines, it is worth mentioning two other publications that have been critical in the harmonization of the quality aspect of medicinal products available on the european market: • good manufacturing practice (gmp) is one of the most important harmonized requirements that have been issued. as per directive / /ec and directive / /ec, all products (including investigational medicinal products) have to comply with the principles and guidelines of gmp. these gmp principles are laid down in directive / /ec. in addition, the ec has published detailed gmp guidelines in line with those principles in eudralex (volume ). this volume covers both the basic requirements for medicinal products (part i) and for active substances used as starting materials (part ii). particular considerations and conditions for specific products (biological products, radiopharmaceuticals, medicinal gases, products derived from human blood or plasma, herbal medicinal products, excipients, etc.) are also in place or under discussion. under this eu system, manufacturers and importers of medicines located in the eea are subject to a manufacturing authorization and come under the supervision of the competent authorities of the member states (the supervisory authorities), who are responsible for issuing the authorizations for those activities taking place in their territories. • the european pharmacopoeia (ep), established on july , by eight countries, aaa is a collection of standardized specifications, so-called monographs, which define the quality reference standard for medicines. today, the convention has been ratified by more than european countries and the eu. european directive / /ec refers to the mandatory character of ep monographs in the preparation of dossiers for maa in the eu. the ep is also applicable in all the signatory states of the convention for the elaboration of an ep, and is used as a reference by many other countries (there are more than observers). the ep is published by the edqm and covers active substances, excipients, substances or preparations for pharmaceutical use of chemical, animal, human or herbal origin, homoeopathic preparations and stocks, antibiotics, as well as dosage forms and containers. the texts of the european pharmacopoeia also apply to biologicals, blood and plasma derivatives, vaccines, and radiopharmaceutical preparations. ▸ nonclinical: all aspects of nonclinical testing and programs are covered under general guidelines (e.g., glp) bbb or discussions on nonclinical strategies to identify and mitigate risks for first-in-human clinical trials or guidelines specific to a type of testing (i.e., pharmacology, aaa belgium, france, germany, italy, luxembourg, the netherlands, switzerland, and the united kingdom. bbb the principles of good laboratory practice define a set of rules and criteria for a quality system concerned with the organizational process and the conditions under which nonclinical health and environmental safety studies are planned, performed, monitored, recorded, reported, and archived. pharmacokinetics, single and repeat dose toxicity, genotoxicity, carcinogenicity, reproductive and developmental toxicity, and local tolerance). most of these guidelines have in fact been developed under the auspices of ich. as for the quality requirements, specific nonclinical guidelines have also been developed for certain types of products. numerous clinical guidelines are available, which cover all phases of clinical development, from early on (i.e., clinical pharmacology and pharmacokinetics studies) to the design of phase studies (disease and patient characteristics, advice on selection of endpoint, duration, control groups, and choice of comparator, etc.). due to the specificities of each group of products, guidelines have been organized by therapeutic area, and some focus on certain types of products (herbal medicinal products or radiopharmaceuticals and diagnostic agents). additionally, general guidelines have also been released to provide advice on general considerations and topics during drug development that are not disease-specific (e.g., "guideline on missing data in confirmatory clinical trials," "extrapolation of results from clinical studies conducted outside europe to the eu population," "clinical trials in small populations," "data monitoring committee," "choice of a non-inferiority margin," and "excipients in the label and package leaflet of medicinal products for human use"). in addition to these numerous scientific guidelines, it is worth mentioning the development and implementation of gcp in europe for investigational medicinal products. this harmonization of gcp has been critical for the recognition of data between european countries, and therefore cooperation on clinical aspects of drug development. directive / /ec is the framework legislation that provides for additional directives, accompanying guidelines, and detailed guidance documents. these guidelines and guidance documents are published in eudralex (volume ). finally, it is important to note that there has been a lot of effort put forth in past years regarding harmonization of the european pharmacovigilance system. this system is coordinated by the ema, but also involves national competent authorities ccc and the european commission. it includes a broad range of activities such as the review of risk management plans (rmps) and psurs, the development and maintenance of the eu reporting and data warehouse system for case reports (eudravigilance), signal-identification activities in the eu, and the coordination of eu rapid alert and incident management systems for timely and adequate responses to new safety data. the eu legal framework of pharmacovigilance was provided in regulation (ec) / and directive / /ec. additionally, relevant ich guidelines have been implemented, and volume of eudralex has been dedicated to this key public health function. it included a number of detailed guidelines, definitions, standards, and information regarding the precise execution of pharmacovigilance-related procedures. ccc in some member states, regional centers are in place under the coordination of the national competent authority. in december , following a public consultation, the ec decided to further harmonize the system (to ensure it is optimally effective, robust, and transparent) via the adoption of two additional texts [ , ] . the final new legislation [ ] was finally published on december , in the official journal of the european union. on june , , the commission implementing regulation (eu) / was adopted, complementing the pharmacovigilance legislation that started to apply in july . finally, some pharmacovigilance incidents in the union have shown the need for further improvements of the legislation. these issues have been addressed by directive / /eu and regulation no / /eu, which started to apply in . due to the number and importance of improvements that need to be implemented [ , ] , many observers consider this new pharmacovigilance legislation as the biggest change to the eu legal framework since the creation of the ema in . the implementation of this new pharmacovigilance legislation required a lot of effort from the ema [ ]. this was a major activity because several processes needed to be established or amended (e.g., the establishment of a new pharmacovigilance risk assessment committee [prac] replacing the chmp pharmacovigilance working party). also, an important change of the new legislation is the increased direct involvement of the ema in the pharmacovigilance of nationally authorized products, in addition to the centrally authorized products. for example, the ema has released the "guidelines on good pharmacovigilance practice (gvp)", which replace volume of eudralex [ ] . this new set of guidelines applies to all medicines authorized in the eu, whether centrally or nationally authorized. the ema is also working with other groups to continuously improve the safety monitoring of medicines. this includes its central coordinating role in protect, ddd its support of the european network of centres for pharmacoepidemiology and pharmacovigilance (encepp), eee its work with the us fda on ae signal detection activities, and its notifications to the who of any measures taken in the eu on medicines that may have a bearing on public health protection in third-world countries. finally, the heads of medicines agencies have also put in place a multi-annual program (called the european risk management strategy [erms] ) which aims to strengthen european pharmacovigilance systems by putting in place efficient measures allowing for the early detection, assessment, minimization, and communication of a medicine's risk throughout its lifecycle. these guidelines apply to more than one specific area and have been prepared through the collaboration of several working parties. they provide advice and guidance on specific ddd protect is a project of the innovative medicines initiative (imi), which is aimed at strengthening the monitoring of the benefits and risks of medicines in europe by developing innovative tools and methods that will enhance the early detection and assessment of adverse reactions. eee encepp is a network that supports independent, post-authorization studies on the safety and benefit/risk aspects of specific medicines. important topics (i.e., pediatrics, cell therapy and tissue engineering, vaccines, biosimilars, gene therapy, and pharmacogenomics). the eu harmonization activities related to certain of these topics are further discussed in the following sections. it is also important to note that cooperation in the areas of inspection (e.g., gmp, glp, gcp, or phv) is critical. although the responsibility for carrying out inspections rests with the national competent authorities of member states, the coordination of these inspections by the ema (and the agreement of common standards) has been an important step that allows for: • increased cooperation between member states • reduced duplication of work (due to the recognition of inspections performed by other member states) • ensuring the same level of quality of medicinal products, and the data generated during their development, wherever the location of the manufacturing site or studies a european system for the authorization of medicinal products has been created with the objective of ensuring that safe, effective, and high-quality medicines can quickly be made available to all citizens across the eu. today, the european system offers several routes for the authorization of medicinal products: ▸ the centralized procedure (laid down in regulation (ec) no / ) is compulsory for certain types of products: products derived from biotechnology processes, advanced therapy medicines, orphan medicines, or products intended for the treatment of certain specific diseases. for medicines that do not fall within these categories (the "mandatory scope"), companies can also submit an application if the medicinal product constitutes a significant therapeutic, scientific, or technical innovation, or if it is in any other respect in the interest of public health. applications for the centralized procedure are made directly to the ema and lead to european marketing authorization. this authorization, binding in all member states, is granted by the ec (based on the opinion of the relevant ema committee). it is valid for the entire community market, which means the medicines may be put on the market in all member states. this is the ultimate integration model in this domain because there is a single application, a single evaluation, and a single authorization allowing direct access to the single market of the community. ▸ the mutual recognition procedure (mrp) (laid down in directive / /ec), applicable to the majority of conventional medicinal products, is based on the principle of recognition of an already existing national marketing authorization by one or more member states. should any member state refuse to recognize the original national authorization on the grounds of potential serious risk to public health, the issue is referred to the cmdh to find a consensus. in that case, the cmdh uses its best efforts to reach an agreement on the action to be taken (within the -day time period foreseen in the legislation). when this fails, the matter is then referred to the ema/chmp for arbitration (see below for details). at the end of the mrp and decentralized procedure, national marketing authorizations are granted in the member states involved, whereas the centralized procedure results in a single marketing authorization (called a "community marketing authorization") that is valid across the eu, as well as in the eea-efta states (iceland, liechtenstein, and norway). purely national authorizations are still available, but are limited to medicinal products to be marketed in one member state only. in addition to the above registration procedures, another european procedure called "referral" has been established. this community referral procedure is used to resolve disagreements (e.g. between member states during an mrp or a decentralized procedure), address specific concerns relating to the safety or efficacy of a medicine or a class of medicines, or when there is a need to harmonize national decisions across the eu. in a referral procedure, the ema is requested to conduct, on behalf of the european community, a scientific assessment of a particular medicine or class of medicines. the problem is "referred" to the chmp so that the committee can make a recommendation for a harmonized position across the eu. referral procedures can be started by the ec, any member state, or by the pharmaceutical company. at the end of the referral, the committee makes a recommendation, and the european commission issues a decision to all member states reflecting the measures to take to implement the chmp recommendation. finally, it is important to note that, in addition to the harmonization of procedures for the authorization of medicines, the system also ensures harmonization and coordination of the pre-and post-authorization activities: ▸ pre-authorization activities: companies can request scientific advice (or protocol assistance in the case of medicines for "orphan" or rare diseases) from the ema at any stage of medicine development, whether the medicine is eligible for the centralized procedure or not. this european procedure helps the company to make sure that it performs the appropriate tests and studies so that no major objections regarding the design of the tests are likely to be raised during evaluation of the marketing authorization application. ▸ post-authorization regulatory activities (i.e., variations or extensions and transfers of marketing authorizations, renewals, psurs, and notifications) have also been harmonized and are coordinated via the centralized, mrp, or decentralized procedures. this ensures that the same quality, safety, and efficacy of products are maintained during the entire lifecycle management of the products throughout europe (e.g., availability of new formulations, extension of indications, etc.). after years of extensive discussions involving ethical aspects [ ] , the european commission adopted a proposal on september , [ ] . this proposal led to new legislation (regulation (ec) no / ) that entered into force in the eu on january , . today, this amended text (and its several associated guidelines and other published information) [ ] sets up a system of requirements, rewards, and incentives together with lateral measures to ensure that medicines are researched, developed, and authorized to meet the therapeutic needs of children (representing over % of the total european population [ ]). in practice, this new regulation established an expert pediatric committee (pdco) within the ema, which is responsible for providing opinions on the development of medicines for pediatric use. the key objectives of the regulation are: • to ensure high-quality research in the development of medicines for children aged to years of age • to ensure, over time, that the majority of medicines used by children are specifically authorized for such use • to ensure the availability of high-quality information about medicines used by children in , a communication from the ec (communication /c / ) provided guidelines on the format and content of applications for agreement or modification of a pediatric investigational plan. many additional procedural and scientific guidance documents have also been released by the ema to facilitate the implementation of this new regulation. the eu introduced a new orphan medicinal product legislation in in order to provide incentives for the development of medicinal products for rare disorders. harmonization of requirements for these types of products is critical to allow for multinational clinical studies and to limit the development challenges due to the small number of patients. prior to this european legislation, a number of member states had adopted specific measures to increase knowledge on rare diseases and improve their detection, diagnosis, prevention, and treatment. however, these initiatives were few and did not lead to any significant progress in research on rare diseases. procedure for the designation of orphan medicines with the technical committee for orphan medicinal products (comp), which is responsible for the scientific examination of applications. designated orphan medicines are assessed centrally on a european level by the chmp, rather than in each member state separately. this regulation also put in place incentives for the research, marketing, and development of such products (e.g., fee waivers, a -year market exclusivity period postauthorization, and scientific assistance for marketing authorizations). following its entry into force and its associated rules and guidelines, the number of orphan medicines authorized has increased significantly [ ] . this directive's aim is to protect public health while securing the free movement of herbal medicines within the community. while most individual herbal medicines will continue to be licensed nationally by member states, the process for licensing and information on herbal substances and preparations will be increasingly harmonized across the eu. for example, in order to further integrate these special medicines in the european regulatory framework, a committee for herbal medicinal products (hmpc) was established at the ema in september (replacing the cpmp working party on herbal medicinal products). the major tasks of this scientific committee are to establish community monographs for traditional herbal medicines, and to prepare and maintain a list of herbal substances that have been in medicinal use for a sufficient period of time, and so are not considered to be harmful under normal conditions of use [ ] . the procedures for clinical trials in europe used to vary from one country to another. there were different national approaches regarding the approval and notification systems, documentation requirements, and timelines [ ] . in october , in order to coordinate the implementation of the new harmonized requirements across the member states, the hma established the clinical trials facilitation group (ctfg). the ctfg (attended by representatives from the national dras, ec, and the ema) acts as a forum for discussion on the agreement of common principles and processes to be applied throughout europe. it also promotes harmonization of clinical trial assessment decisions and administrative processes across the national dras. this group established a voluntary harmonization procedure (vhp) for the assessment of multinational ctas [ ] . during this three-phase procedure, dras from all member states involved assess the application, though each member state remains ultimately responsible for the approval of the cta in its own country. however, there is a coordinated validation phase (phase ) and voluntary cooperation of the member state during the assessment phase (phase ) before the usual formal national process (phase ). phases and of the procedure are coordinated by a vhp coordinator. the "acceptability statement" obtained through this vhp procedure is then included in the subsequent national cta applications. from march to april , applications were evaluated through the pilot vhp procedure; of these applications received a positive opinion [ ] . the average procedural time was days (which is significantly less than the average time of standard national procedures). the overall feedback from sponsors was positive, except that: directive / /ec and its associated texts and guidelines are a very important step in the harmonization of procedure for the registration and conduct of clinical trials in europe. implementation of this clinical trials directive into national legislation of all eu member states was completed in . principles like clinical trial authorization by the national dras within defined maximum timelines led to significant harmonization of the clinical trial approval process. however, it has been agreed that this new system needs further harmonization in order to achieve the ultimate objective [ ] . indeed, the actual assessment of a request for authorization of a clinical trial is done independently by the member states concerned. the legislation does not provide for a mechanism whereby the member states are obliged to reach a common conclusion regarding a clinical trial involving different member states. this lack of obligation and detailed direction implied different interpretation from member states and therefore created implementation issues. as a consequence, sponsors have to respond to the various required changes and adapt their protocol in view of diverging assessments by the dras. this situation requires additional time and effort by the pharmaceutical industry (without added value for the patients). in , following a public consultation and a long and thorough impact assessment ( the proposal has been submitted to the european parliament and the council who engage in ordinary legislative procedure. this proposal, once adopted by the eu-legislator, is going to replace the clinical trials directive. it is expected to come into effect in and to provide major revisions to the current system (e.g., single assessment outcome, simplified reporting procedures, etc.). finally, it must be noted that other important topics related to the regulation of medicines are also coordinated at the community level (by the ec and the ema) in order to have harmonized regulatory actions and enforcements, and to complete the single pharmaceutical market. these harmonization initiatives are at different stages of development: • to support cooperation and harmonization activities, the eu needed systems and knowledge management support. the implementation of this telematics (the integrated use of telecommunications and informatics) strategy, coordinated by the ema, is critical to increase efficiency and transparency across the european medicines regulatory network. in addition to the standards for electronic submissions (esubmissions) that were developed and published, a central set of pan european systems and databases was created. these systems and databases exchange information with systems of external stakeholders and dras, while staying separate from them. they also help provide high-quality information on medicinal products to the general public and support the monitoring of the post-authorization risk and benefit balance of medicines in the eu. the following critical projects and tools have been developed under this program (some of them are still under development): ▸ eudract: the community's electronic database for clinical trials containing information submitted by sponsors. it informs dras of ongoing clinical trials in all member states and eea countries, enabling an overview of multi-state trials. the system also alerts dras in the case of early interruption or termination. ▸ eudragmp: community database on manufacturing and import authorizations and gmp certificates. the ema launched the first release in april . this system is used by eu gmp inspectors to share information (i.e., gmp authorization, noncompliance with gmp information resulting from inspection activity, planned inspection activity, and "rapid alerts" arising out of faulty manufacture). ▸ eudranet: private electronic network linking the members of the european medicines regulatory network and ema. it ensures that both electronic mail between members of the network and their access to the eu telematics systems is secure. ▸ eudralink: the european medicines regulatory network's secure file transfer system used for exchanging information for regulatory purposes. it operates independently of eudranet, so that it can be used by applicants and marketing authorization holders, as well as the regulatory organizations within the network to transfer files. ▸ eudrapharm: the community's database of authorized medicinal products. some functionalities of this database are still under development. ▸ eudravigilance: system monitoring the post-authorization safety of medicines through safety reports (i.e., suspected adverse reaction reports). it is designed to receive, process, store, and make available information. one of the objectives of this system is the early detection of possible safety signals to facilitate the regulatory decision-making process (based on a broader knowledge of the adverse reaction profile of medicines). the ema to receive, validate, store, and make available information for review marketing authorization applications. the system's key benefit is its ability to take advantage of the lifecycle management functionality built into the ectd by easily allowing the full extent of the current valid documentation as well as its submission history. ▸ eu telematics controlled terms (eutct): central repository and publication system for a controlled term list used in the european medicines regulatory network. the establishment of the eu has not been easy, but it has represented the desire to end conflicts in europe. since its creation, the eu has been successful in delivering peace between member states and has reunited a fractured continent via the promotion of cooperative projects (i.e., economic and social). this cooperative initiative went beyond the initial objectives of its founders. ever deeper integration has been pursued while embracing new members. the membership of the eu has grown from to nations, bringing the eu's population to half a billion people. it has created stable institutions, a single market, and a single currency. despite numerous challenges, ggg the eu has survived, and is today a major economic and commercial power. although improvements are still needed in certain areas, the eu represents a unique model of successful cooperation, harmonization, and integration between countries of different languages, cultures, history, and levels of development. in the pharmaceutical sector, much has been achieved towards the consolidation of the european system of evaluation and supervision of medicines. several challenges have already been overcome, but outstanding issues still need to be resolved to further support and improve public health in europe, free movement and access to medicines in the community, and the competitiveness of the union. taking into consideration its successes and challenges, this section provides a balanced evaluation of the current situation. it demonstrates that harmonization of pharmaceutical regulation in europe can be considered a real and quick success in general (considering the major changes it required), but acknowledges some specific areas where work is still needed. for all these reasons, the development of the eu and its european pharmaceutical "regulation/ system" is a great example that needs to be further evaluated and discussed. although this model of harmonization and integration may not be fully applicable to other cases, this experience can certainly help other regional or global harmonization initiatives. since the adoption of the first pharmaceutical directive in , many topics have been harmonized. the past years have seen a gradual convergence of pharmaceutical legislation in europe. today, a considerable package of harmonized legislation (in the form of the pharmaceutical "acquis communautaire") is in place to support two objectives: the protection of public health and the free movement of products. these provisions/texts applicable to medicinal products are included in eudralex. they include binding legislation (i.e., regulations and directives), but also numerous technical guidelines and recommendations to facilitate the implementation of these common principles. a well-structured european pharmaceutical system has also been established. in addition to the european institutions necessary to harmonize and create the european pharmaceutical legislation, technical european bodies have also been established. today, the evaluation and supervision of medicines in europe is shared between european and national bodies that form a complex but well-organized network of approximately , technical and regulatory experts. words like "networking," "work sharing," and "harmonization" became common and remain crucial for the future. the establishment of the ema as a key coordinator of this system was an important decision for the integration and harmonization of practices and standards to support and promote the single european pharmaceutical market. the primary aim of this centralized system was to create conditions in which a single scientific evaluation of the highest possible standard would lead to rapid access to an integrated market of innovative and good cost-effective treatments. this objective, in large measure, has been achieved. the ema, which is comprised of experts provided by national dras, has today established itself as a leading world agency for the evaluation of medicines. its contribution to the effectiveness and efficiency of the eu system, and therefore to the protection of public health and to the achievement of an operational internal market, is well recognized by all stakeholders. the effectiveness of the system has been maintained despite its growing complexity. indeed, the increase in the number of centralized applications hhh and other procedures, eu enlargement, and new regulations have led to an increased workload and an enlarged scope of responsibility for the ema over the past years. these changes have led to the creation of new committees (comp, pdco, cat, hmpc, prac) that require the implementation of additional procedures and new tools. these structural changes and increased responsibilities should be monitored closely in the future to avoid risks of inconsistencies, overlapping, bureaucracy, and rigidity. also, it is critical to continue to monitor financial compensation of national dras and to regularly assess the involvement of each member state in the eu pharmaceutical system to ensure availability of appropriate resources and expertise [ , ] . within this legal framework and european pharmaceutical system, community authorization procedures (centralized, mrp, or decentralized) have been in place since the mid- s. the centrally coordinated tasks include assessments led by rapporteurs and co-rapporteurs, inspections, and pharmacovigilance through the medicine's lifecycle. although the national dras have prime responsibility for the efficient operation of mrps and decentralized procedures, national marketing authorizations, and clinical trial authorizations for human medicines, the ema has an important role in supporting these noncentralized functions. for example, the ema maintains the eudravigilance database and the eudract database, and supports a range of scientific committees and the coordination group for mrps and decentralized procedures [ ] . the criteria for the approval of medicines and other technical topics have been extensively harmonized within the eu. many technical and regulatory guidelines have been released in all areas (quality, nonclinical, and clinical). there has been a specific focus in recent years to improve the european pharmacovigilance system, to simplify the variation system, to harmonize the requirements for clinical trials, and to implement an advanced therapies regulation. the establishment of the european pharmacopoeia has also been very important to ensure standardization of specifications and quality of medicines in the eu. all these measures and actions described above have led to improved marketing authorization procedures, the harmonization of data protection in the eu, better access to medicines for children, orphan drug development, clinical trials, and a new regulatory framework for advanced therapies. lifecycle management of products has also been improved (i.e., the revised legislation on variations to reduce the administrative burden by streamlining the circumstances obliging industry to file applications). the next review of the european system will be noteworthy because it will evaluate if new measures (developed following the last review in ) improved the system and produced real dividends in terms of cost efficiencies through economy of scale (via the reduction of the administrative burden where this did not have public health implications). it is also worth mentioning that this european system is solid enough to stand the challenges of new therapeutics. the current structure, forum, and processes allow "proactive" harmonization. indeed, most of the harmonization initiatives are created to discuss existing disharmonies on specific topics. at the beginning, the european harmonization effort, related to pharmaceutical regulation, was focused on disharmonies between countries. today, even if disharmonies do still exist on some specific subjects, many topics have been successfully harmonized. the processes and structures that have formed over the years now allow the system to cover new subjects for which no national regulations and requirements have been developed yet. developing this new regulation at the eu level automatically creates harmonized requirements (this can be called "proactive harmonization"). . this group, which included ema staff and members of the chmp and its working parties, generated recommendations on how the ema should tackle these new emerging topics not covered by the existing national, regional, or global regulations and standards. ▸ ema innovation task force (itf): in order to provide support for medicine innovation in the eu, the ema established an internal horizontal cross-sectorial group to focus on emerging therapies and technologies. the itf brings together competences from the areas of quality, safety, efficacy, pharmacovigilance, scientific advice, orphan drugs, and good practices compliance, as well as legal and regulatory affairs. one of the objectives of the itf is to address the impact of emerging therapies and technologies on current scientific and regulatory requirements. its scope also encompasses areas for which there are no established scientific, legal, and regulatory experience. one of their tasks is to identify areas for legal, regulatory, and technical guidance preparation and proposals for consideration by the ema committees and working parties, and to contribute to relevant ec initiatives and legislation [ ] . the eu today is recognized as a major player in the international harmonization of pharmaceutical regulations. it has developed privileged relationships and initiated cooperation projects with other countries outside the european community (major developed countries and emerging markets). for example, the ema cooperates with many of the world's largest regulatory bodies outside the eu iii in areas such as inspections, safety of medicines, and exchange of information on issues of mutual concern. the establishment of the international and european cooperation sector, formed in february and responsible for the development, coordination, and implementation of the agency's international strategy and activities (including confidentiality arrangements with countries outside the eu), demonstrates the ema commitment to international cooperation [ ] . also, collaboration has been initiated with china, india, and russia on pharmaceuticals, and it is partnering with international organizations (i.e., ich, who, and pic/s). this work should continue and also be extended. it is indeed important to support the development of globally harmonized standards and requirements in order to ensure fair competition with other parts of the world for the development of medicines and to avoid delay in the availability of essential medicines for european patients. ensuring against falsified medicines, resolution of pandemic issues, product development in emerging markets, and reliability of clinical data produced outside europe are good examples where international cooperation is necessary to ensure adequate protection of public health in europe. in spite of all the above-mentioned major progress and regular improvement of legislation by the european commission, there is still room to improve the eu pharmaceutical system. on the regulatory side, issues dealing with the implementation and interpretation of community legislation by member states continue to create obstacles to the free movement of medicines. stakeholders continue to raise concerns regarding market fragmentation linked to disparities in national pricing and reimbursement schemes (despite the adoption of directive / /eec in the early days of the european pharmaceutical system), unnecessary regulatory burdens caused by divergences in the implementation of community legislation (e.g., clinical trials requirements), and a lack of commercial interest in national markets that are economically less attractive. european patients still suffer from inequalities in the availability and affordability of medicines. this situation could worsen and create significant inequalities between patients in accessing medicines if it is not resolved. additionally, europe has been losing ground when it comes to innovation and competitiveness in the pharmaceutical market. in its communication of december , [ ] , the ec recognized that further harmonization is necessary to resolve shortcomings in the eu pharmaceutical market in furthering increased globalization of this sector. to improve this issue, the ec confirmed its objective to continue to progress towards a single and sustainable pharmaceuticals market [ ] . to further support and improve the public health in europe and free movement of medicines within the community, and to maintain its competitiveness, the eu needs further harmonization in several areas, such as: novel medicines by patients, mainly due to increased pressure to cut healthcare budgets. in certain countries, medicines are not made available due to administrative requirements and poor economic rewards. a lack of transparency and harmonization with regard to pricing, reimbursement, and relative effectiveness remains a challenge [ ] . in contrast to the benefit-risk assessment carried out by regulators, national hta bodies compare the "relative effectiveness" of medicines and take their financial cost into account. this post-marketing national hta evaluation can lead to national differences due to different country needs. the addition of different requests (i.e., different type of studies) from regulators and hta bodies can also delay availability of new products. to resolve this major issue, the european network for health technology assessment (eunethta) was established to support effective collaboration between national htas. also, the ec gave the political mandate to the ema to begin interacting with hta bodies when it published the conclusions of the pharmaceutical forum in october . kkk since then, the ema has begun to collaborate with national hta bodies and with eunethta [ ] . this interaction focuses on centralized approved products and aims to facilitate communication between ema and hta bodies early in a medicine's development and throughout the medicine's lifecycle. as mentioned above, the harmonization of price and reimbursement evaluation is critical in supporting a european pharmaceutical market. however, it will be a very difficult and long process to implement due to political and budgetary aspects and differences in pharmaceutical markets and healthcare budgets existing between member states. the european clinical trials directive (directive / /ec) has been an important and necessary step in the harmonization of european pharmaceutical regulation. the principles defined in the declaration of helsinki (in ) and the ich gcp e guideline (in ) allowed some harmonization of clinical practices and protection of clinical patients. but, before this directive came into force, the rules for performing clinical trials (i.e., regulatory procedures and requirements) varied significantly in the european community as they were based on differing regulatory approaches in the member states. this new legislation promoted harmonization of clinical trial practices allowing important improvements related to the protection of patients (i.e., safety and ethical concerns) and reliability of data, and facilitated the exchange of information between dras. however, despite this progress, important negative effects of this new legislation have been reported (e.g., the increase in bureaucracy and administrative costs). the number of clinical trials carried out in the eu has fallen by % in recent years, while administrative kkk the pharmaceutical forum was set up in by the european commission as a three-year process in order to find relevant solutions to public health considerations regarding pharmaceuticals, while ensuring the competitiveness of the industry and the sustainability of national health care systems. more specifically, this forum analyzed three key themes: information to patients on pharmaceuticals, pricing and reimbursement policy, and relative effectiveness. costs and delays have doubled [ ] . it is still labor intensive and costly to duplicate largely identical administrative procedures for multinational clinical trials. additionally, sponsors spend a great deal of time retrieving the relevant national information and requirements and preparing customized applications without added value for the patient and the regulators (the core scientific information is the same, but the format and administrative information and forms differ). it is indeed a problem for a large pharmaceutical company, as it usually requires additional dedicated departments with the necessary resources to track differences in national requirements and follow the many parallel procedures. but it is even more problematic for smes or academic sponsors for whom these costs can reach prohibitive levels. this multiplication of parallel procedures also has an important impact on the dras. indeed, available resources are used in multiple assessments of the same core information in different member states, which clearly delays the start of clinical studies. it is important to note that this duplication of assessments does not necessarily increase the quality of the assessment, as the necessary specific expertise might not always be readily available in all the member states concerned. this is a nonefficient use of national resources without added value for the patients or science. this implementation problem is partly due to the legal framework that has been chosen for harmonization in this area. as with all directives, the clinical trials directive had to be transposed in national laws. unfortunately, in this case, the objectives of the directive were transposed into divergent national legislations, somewhat missing the harmonization goal and making multinational trials difficult to perform. in its consultation paper [ ], the ec proposed options to improve the situation. one of the best options is to continue with the harmonization process. this would mean creating a real european system of authorization for clinical trials to avoid duplication of assessment. it would avoid the inconsistent assessment conclusions and requests, encourage appropriate use of resources and expertise (for both the sponsors and dras), and ensure common implementation of the principles laid down in the clinical trials directive. the vhp initiative seems to be a good first step. it allows for a better implementation of the eu clinical trials directive principles and further harmonizes the conduct of clinical trials in europe. however, this procedure cannot be considered as the ultimate solution because it does not resolve all issues [ ] . more specifically: • there are still parallel cta assessments by multiple dras. • there are still major differences between countries regarding the time it takes to issue approval. • this is a voluntary cooperation and there are differences in the level of interest and responsiveness between countries. • the current procedure does not remove specific national requirements or differences between national assessments (this is a cooperation effort, not a harmonization of requirements). • this process does not accelerate the first patient enrolled (fpe) in europe. to resolve these outstanding issues, the current vhp procedure should be revised to become a real mrp where the assessment will be conducted by only one reference member state. the content of the dossier should also be fully harmonized between countries. the establishment of a centralized procedure through a new regulation (which will deliver a pan-eu approval) would also be very helpful for certain types of products that require specific expertise not available in all eu countries (e.g., advanced therapies), for orphan drugs, and/or for pediatric medicines. this centralized process for cta would be a good bridge between the ema scientific advice process and the centralized registration procedure. the system for registration of clinical trials would then mimic the system already in place for the registration of medicinal products with a combination of three types of procedures: • centralized procedure for specific products such as biotechnology and advanced therapies • mutual recognition procedure for other multinational clinical trials • national procedure for a clinical trial involving only one member state this reorganization of the system and procedures, supported by the ec [ ] and most of the shareholders involved in clinical trials [ ] , would utilize the current structures and expertise in europe, would build on the experience acquired with the registration process, and would facilitate patient access to clinical trials and to new technology within the community. it would allow the necessary flexibility and different levels of review for interventional trials (e.g., a small national study with a well-known entity does not need the same type of evaluation, organization, and bureaucracy as a phase study with a new fusion protein or a large multinational phase study). measures should be put in place to ensure that such reorganization would allow this flexibility and avoid any further increase of delay and administrative costs and burdens. for example, "recognition" of other assessments should be the focus, and "nonrecognition" should be limited to major issues (that should be clearly defined). these "nonrecognitions" of assessment by another country should be rare to avoid regular arbitration or appeals that would further delay the start of the clinical studies. selection of reference member states (rms) should also be defined because many parameters are involved in such selection (i.e., expertise, resources, balanced workload between countries, etc.). finally, this new cooperative system should not result in the simple addition of national requirements, but a harmonized scientific assessment that would be implemented equally in all member states. this next step in the harmonization of a clinical trial in europe would certainly be beneficial for patients, sponsors of clinical trials (pharmaceutical companies, but also small entities or academic centers), and dras. some of the above proposals have already been recommended by the european commission [ ] . the recent adoption of a "proposal for a regulation of the european parliament and of the council on clinical trials on medicinal products for human use, and repealing directive / /ec" [ ] by the commission represents an important step in the improvement of the current system. however, this process will take time to implement, and national interests will need to be overcome. finally, the assessments of ethics committees also need to be reviewed and improved. the clinical trials directive is based on the concept of one ethics committee opinion per member state concerned. however, several member states maintain a decentralized system where the single ethics committee opinion is based on the opinion of several local committees. as a consequence, in the eu there are approximately , ethics committees involved in the assessment of clinical trials [ ] . also, better harmonization of responsibilities between dras and ethics committees must happen across europe [ ] . it is agreed that ethical issues fall within the responsibility of member states. however, current practices need to be reviewed in order to smoothly integrate an improved harmonized system and to protect european clinical trials subjects. these programs are important to make new therapies available to patients as soon as possible. they should be handled on a european basis in order to ensure that every european person, wherever their location, has the equal right to access these new medicines at the same time. today, this difference in access within europe is clearly contrary to the overall european objective to ensure that all patients within the community have the same access to the same quality products throughout europe. of course, the harmonization of these requirements and procedures should be carefully implemented to avoid the creation of delays compared to the current situation. ▸ pharmacovigilance: the eu pharmacovigilance system demonstrates that cooperation and harmonization of regulations and practices in europe is beneficial to patients. indeed, merging the eu national pharmacovigilance systems into one network increases the quantity of data/reports/ information, which facilitates the early detection of possible safety signals, and therefore the monitoring of product safety. unfortunately, the mediator issue in france has shown that the eu pharmacovigilance system needs to be improved to be fully functional. this topic has been one priority of the european network. the ongoing implementation of the new legislation by the ema and the member states will be critical. although the mutual recognition and decentralized procedures have improved over time, challenges still exist, and the principle of these procedures (i.e., recognition of another country's assessment) is not always respected. in both these procedures, member states can only refuse to recognize other countries' assessments if they feel that this recognition could have a "potential serious risk to public health." unfortunately, this reason for disagreement is vague enough to allow flexibility for member states. in , a guideline was released [ ] to further clarify how this risk should be defined. however, some national dras continue to have a broad interpretation of "potential serious risk to public health," and trigger ema arbitrations for grounds that do not fall under this specific category [ , ] . in addition to the specific issues discussed above, more general challenges can also impact the harmonization of european pharmaceutical regulation. although these general considerations are not specific to the pharmaceutical sector, they can influence the establishment and implementation of pharmaceutical regulation. therefore, they need to be understood and integrated when developing implementation plans and timelines: • . this major difference in workload between countries demonstrates a big gap in work sharing and certainly highlights differences in national dras' expertise and resources and pharmaceutical companies' interests for each national market. • one of the complexities and difficulties of the eu system is the division of activities undertaken at the national level (e.g., clinical trial responsibility, scientific advice handling, etc.) and at the eu level (e.g., equal scientific advice handling, assessment of pediatric investigational plans, etc.). this requires many communications and infrastructures between the eu and national players. • external economic or political factors could also influence the harmonization of european pharmaceutical regulation. for example, the modification of european borders via new enlargement of the eu (even if the eu leaders have agreed to mark a pause for now, discussion on the accession of countries such as turkey, iceland, and serbia are still ongoing). additionally, the possible creation of a "mediterranean union" desired by past french president sarkozy could also impact the scope and timelines of the next steps of harmonization and integration. finally, it will be important to see if and how the two new functions created by the treaty of lisbon (president of the eu council and high representative of the union for foreign affairs and security policy) will benefit the eu. the first important dossiers after the creation of these two functions (global financial crisis, global security, and support to greece) have indeed still been handled by the political leaders of major member states (i.e., france and germany). it is clear that the european system is integrally linked to its own history. this model cannot fully fit every harmonization initiative in the world because every situation and need is different. however, it is worth reviewing the lessons learned from this plus years old initiative. this first regional harmonization initiative (rhi) overcame a lot of challenges, and has since developed into a strong regional harmonized pharmaceutical regulation and system. this success demonstrates that an organized cooperation and harmonization can facilitate the development of high standards and practices. more specifically, the european initiative clearly demonstrates that a structured stepwise approach is necessary: ▸ first, it is necessary to set up major principles (directive / /eec). ▸ second, it is critical to provide specific detailed requirements and to further detail the agreed principles (directives / /eec, / /eec, etc.). ▸ third, a structured and organized system is needed to implement the principles and requirements. technical bodies need to be established to control medicines and manage the establishment of common procedures (especially centralized types). in europe, it was key that the national dras provide expertise and resources to european bodies not only to ensure appropriate availability of resources, but also to ensure full adhesion of the countries into the system and adequate communication between all players of the system (national and european). ▸ when all the basic principles and a system are in place, additional more specific requirements can be discussed so that the system can take into account particular needs (i.e., specific requirements for specific products, population, etc.) in order to have a more coherent system. ▸ finally, it very important to monitor the system and regularly review the extent to which this system and measures support the harmonization goals and meet the predefined objectives. evolution of the environmental impact (i.e., globalization, change of membership, change of political commitment, and need for new requirements due to emerging problems, etc.) also has to be taken into consideration, and the regulation and system needs to be carefully adjusted to ensure its longevity. another lesson learned from europe is the importance of cooperation. to be successful and ensure effective functioning of this system, cooperation between the different entities of the system (ema, hma, national dras, ec) has been, and remains, critical. even if the european pharmaceutical system is complex, it is well organized. the provision by the member states of high-quality scientific resources for the evaluation and supervision of medicines is a critical factor for the success of the eu system. indeed, scientific excellence (as a result of eu-wide pooling of expertise and data) has been a key strength. in this respect, it should be stressed once again that such excellent progress has been highly dependent on close collaboration between the ema and the national dras within the context of the eu regulatory network, and in particular on the valuable input of high-quality specialist expertise provided by the member states. this provision of national resources, coordinated by the ema, is one of the features of the eu regulatory network. this success also relies on political support for this european harmonization initiative in order to support the creation of the single market. without this political commitment (and therefore associated funds and resources), it would have certainly been much more difficult and taken more time to create this system. it is recognized that other harmonization initiatives in the world are certainly suffering from the lack of such political commitment, especially when such harmonization is not driven by the willingness to create a single market (i.e., integration model). finally, the eu has also clearly demonstrated that better organization at the regional level is extremely critical to ensuring the success of global harmonization and cooperation. even if all regions are not working towards integration like europe, this example of better coordination and representation should be followed and discussed in other regions of the world. indeed, this example demonstrates that a well-organized and coordinated regional structure is beneficial to all stakeholders [ ]: ▸ individual countries via better representation and better access to international activities/agreements/decisions through regional structure (this is especially true for small countries with less expertise and resources). individual countries also benefit from the infrastructure (i.e., databases or training programs) and good practices developed at the regional level. ▸ regions because they allow better representation of interests (europe has more power than a combination of small countries' voices, and has an impressive network of experts). ▸ international cooperation and harmonization initiatives because they facilitate communication by reducing the number of contacts and seats at the international level (but provide a structure for dissemination of information). for example, having all eu countries represented at ich would not be possible. this regional coordination is very important for the future of global initiatives (such as ich or who projects), but it is even more important in the management of a worldwide health crisis (e.g., pandemic influenza). this european coordination system should be implemented in other regions of the world because the coordination of rapid and efficient communication of information and actions during such a crisis helps the overall coordination of the situation. for example, in the recent case of pandemic influenza, it was critical to have central coordination (not only global, but regional). the ema (using its "crisis management plan") allowed europe to respond rapidly and efficiently to the challenges of an outbreak of pandemic influenza by: ▸ the fast-track review of vaccines (using its best experts) ▸ monitoring the safety of centrally authorized pandemic-influenza vaccines and antiviral medicines ▸ liaising and coordinating activities with critical partners, including the ec, eu member states, other european agencies (such as the european centre for disease prevention and control), and international partners (such as who and regulatory bodies of non-eu countries) to ensure timely exchange of information and coordination of activities relating to the pandemic ▸ coordinating the communication of relevant information to the public, healthcare professionals, and the media all of these activities would be less efficient if performed by each individual country. political and economic development in the pan-american region has resulted in interest in regional economic integration. several subregional integration groups have emerged in this area since the s. harmonization of pharmaceutical regulations and technical standards is a component of this economic integration, but the degree of progress in this area varies a lot from one subregion to another (and even from one country to another). in light of these various economic integration initiatives, the need became evident for an entity in which the different countries of the region could share experiences and expertise. the pan-american network for drug regulatory harmonization (pandrh) was created in november . this is a regional initiative established to promote drug regulatory harmonization throughout the pan-american region within the framework of national and subregional health policies. this continental forum is not a supranational entity, and its decisions represent recommendations to be assimilated into the subregional integration initiatives. the mission of this network is "to promote the harmonization of pharmaceutical regulation covering aspects of quality, safety, efficacy and rational use of pharmaceutical products, the strengthening of national regulatory authorities (nra) capacity within the region of the americas based on the right of the population to access quality medicines, recognizing advances in science and technology and within the context of national and sub-regional realities" [ ] . the objective of this initiative is to facilitate regional harmonization of medicinal drug requirements and guidelines for specific regulatory issues. this objective is achieved by adopting recommendations for implementation at national and regional levels, and also by supporting the development of training on specific important topics. however, this initiative also has broader objectives such as: ▸ promoting and maintaining a constructive dialogue among dras, the pharmaceutical industry, and other sectors ▸ strengthening the dras of the region ▸ encouraging convergence of drug regulatory systems in the pan-american region ▸ facilitating technical cooperation among countries in collaboration with subregional integration groups. since , pandrh has been a member of the ich global cooperation group (gcg). this membership broadens pandrh's role because this regional harmonization initiative is now also involved in global harmonization. pandrh provides a way to disseminate recommendations on drug regulatory harmonization of global initiatives. it also ensures that regional specificities and challenges will be considered when new global recommendations are discussed. ▸ dras of all pan american health organization (paho) member states ▸ regional pharmaceutical industry associations: latin american association of pharmaceutical industry (alifar) and latin american federation of the pharmaceutical industry (fifarma). ▸ academia ▸ consumer groups and professional associations it also includes representatives from the five subregional trade integration groups within the americas (plate ) that are themselves multinational cooperation initiatives but are working on a broader integration with emphasis on political and/or financial interest: ▸ the andean community is a community established in (by the cartagena agreement) that currently regroups four countries (bolivia, colombia, ecuador, and peru). chile and venezuela have also been part of this initiative in the past and some others countries are observers. these countries decided voluntarily to join together for the purpose of achieving more rapid, better-balanced, and more autonomous development through andean, south american, and latin american integration. they also created a free trade area (including the four current members plus venezuela). this integration initiative is broad and regroups several areas, one of them being health. the integration of health is governed by the andean health body, which coordinates the actions aimed at improving the healthcare of member countries. it gives priority to cooperative mechanisms that promote the development of subregional supranational systems and methodologies. these actions are also coordinated with the other subregional, regional, and international organizations. discussions include many topics such as the development of a pharmaceutical policy model, the evaluation of medicinal products, and a surveillance network. ▸ sica (the central american integration system) is the institutional framework of subregional integration in central america. this is the latest step of a long integration process in the region. it was created in december (by the signing of the tegucigalpa protocol) by the states of belize, costa rica, el salvador, guatemala, honduras, nicaragua, and panama. this initiative also involves the dominican republic as an associated state and some regional and extra-regional observers (mexico, chile, brazil, china, spain, and germany). the headquarters of the general secretariat is located in el salvador. the first objective of this integration process in central america was to transform the area into a region of peace, liberty, democracy, and development, based firmly on the respect, tutelage, and promotion of human rights (following a history of political crisis, conflict, and dictatorial rule in the region). health topics are covered by the executive secretariat of the council of ministers of health in central america (se-comisca). several projects are under discussion in this subregion, such as the basis for quality assurance of drugs and a pharmacovigilance system. ▸ mercosur (the "common market of the south") was created in (by the signature of the treaty of asuncion) and encompasses five latin american countries (argentina, brazil, paraguay [which is currently suspended], uruguay, and venezuela). the purpose of this agreement was to set up a common market and eliminate trade barriers among the signatory parties. mercosur has been involved in several health projects (such as implementation of gmps with training and joint inspections and development of programs on vaccine regulation and control) to promote cooperation between its members and harmonization of specific pharmaceutical regulations in this subregion. to date, there is no mutual recognition system. ▸ nafta (north american free trade agreement) was implemented in january to remove most barriers to trade and investment among the us, canada, and mexico. the objective of this agreement was to establish procedures to facilitate trade and investment on the north american continent. this trade liberalization had some positive impact and created one of the largest trade blocs in the world, but some downsides have also been reported by economists (who have shown that nafta has not been able to produce an economic convergence). nafta has had a minor impact on the harmonization of pharmaceutical regulations in the region and has not been able to resolve the problem of parallel import of pharmaceutical products between canada and the us. one of the major components of this initiative is the pan-american conferences on drug regulatory harmonization held every two to three years. these conferences are the highest instance of the pandrh network. they serve to define priority areas for harmonization and to endorse standards, guidelines, and other recommendations, including norms and procedures and steering committee membership. they also provide a forum for discussing issues of common interest in drug regulation. participants include all interested parties such as the dras of all paho member states, representatives of the regional pharmaceutical industry associations, academia, consumer groups, professional associations, and representatives from the five subregional trade integration groups within the americas. the st pandrh conference took place in november (in washington, dc, us). pan-drh was then officially created during the nd pandrh conference in november (also in washington, dc). following these first two conferences, subsequent conferences took place to review ongoing activities of the working groups. pandrh mimics the ich structure. it is organized around three major bodies: ▸ the steering committee (sc), which ensures operational management of this initiative between conferences, is composed of: • seven members from five national dras (one from each of the subregional economic groups) and two industry representatives (fifarma and alifar) • seven alternate members from five different national dras (one from each of the subregional economic groups) and two industry representatives (fifarma and lifar) • regulators from other countries (not represented on the sc), representatives from nongovernmental organizations (ngos) recognized by paho/who, and other stakeholders invited by the sc who may also participate in sc meetings as observers members of the committee serve for a period of four years, with staggered rotation to maintain continuity. the sc meets at least twice every year. its primary role is ( ) to establish the agenda for the biennial pan-american conferences, and ( ) to follow up on conference recommendations by establishing and monitoring the progress of working groups. the responsibility of this group is to promote progress between conferences through the coordination, promotion, facilitation, and monitoring of the harmonization activities. ▸ the technical working groups are specifically formed to work on topics and areas identified for harmonization. the members are experts in their specific subject matter. a working group may include the following categories of members: • main members that represent the national dra of a country in each of the five subregional blocs, the regional industry associations alifar and fifarma, and those designated by the secretariat • alternate members designated to attend the meetings instead of the principal members • observers from any country generally nominated by a participating national dra (the observers do not retain voting rights) • expert resources (as needed) to support a specific activity of the group (expert resources do not have voting rights) the national dras of countries not represented in the working group can designate focal points to follow the activity of the group. each working group has a coordinator (and an alternate) who chairs and coordinates the meetings, leads the development of documents, and reports periodically to the sc on the progress of the group. in general, the first task of a new working group is to conduct a survey to identify the differences in regulatory requirements among countries in order to prepare a work plan. then, the group reviews international and regional and/or national recommendations and guidelines and prepares a harmonized proposal. when the harmonized standard is developed, the working group is in charge of designing training and helping in implementation of this standard by assisting countries in the dissemination and education concerning this new rule. technical working groups meet in conjunction with sc meetings or separately (determined by a work plan and resources). ▸ a secretariat, provided by paho, supports the initiative technically and administratively. it monitors the pandrh website, serves as a focal point for the coordination and dissemination of information, coordinates activities arising from recommendations of the conferences and sc, and acts as liaison and a representative of the network in global and interregional harmonization organizations (icdras, ich, etc.) as in other regions of the world, there is a need to promote harmonization of pharmaceutical regulations to facilitate the availability of safe, effective, and good-quality products and thereby protect public health. paho initiated communication among the different members of the pharmaceutical sector in the americas in order to facilitate communication among the different subregional blocs (and also the countries not already covered by these blocs) and organize regional harmonization. the first pan-american conference took place in november (in washington, dc, us). this conference was considered the first step towards the establishment of pandrh. during this first conference, the scope and the term "harmonization" were defined (as the search for common ground within the framework of recognized standards, taking into account the existence of different political, health, and legislative realities among the countries of the region). the structure and financial support of pandrh were also discussed at this first conference. however, pandrh was officially created during the nd conference (november in washington, dc) following a consultation in caracas, venezuela in january , and also several ad hoc discussions and meetings (meeting of americas' regulators in washington, dc in november , regional working group on bioequivalence in caracas in january , and regional working group on gcp in buenos aires in may ). during this second conference, the mission statement and objectives of the sc were agreed upon. this initiative was then officially recognized by the nd directing council of the paho in september . resolution cd .r , which was approved during this council, provided strong support from ministers of health of the member states in the region to pandrh and to the process of drug regulatory harmonization. during pandrh conference v (in buenos aires in november ), the regulations governing pandrh (mission, structures, and procedures) that were originally created during the nd conference were slightly modified to incorporate lessons learned during its first few years of establishment [ ]. harmonization proposals are developed by the technical working groups. these groups primarily use who documents as the basis for developing regional guidelines. other international guidelines including ich and selected regional (e.g., eu, american subregional) or national technical documents are also used as the basis for harmonization proposals and as reference materials. after a working group has agreed on a draft harmonized document, it is posted on the website for external comment. comments are reviewed by the working group to prepare the final version of the document that will be presented for adoption by the conferences through the sc. conclusions and recommendations of the conferences are to be adopted by consensus (if consensus cannot be reached, the different points of view have to be recorded). during its seventh meeting (in june in washington, dc, us), the sc established a system of phases and stages for its harmonization process. this system, which mimics the ich process, is composed of five phases, with each having substages: final technical documents are intended for use at the national level (through the subregional integration groups), but this implementation is at the discretion of each country. members of the sc are responsible for monitoring implementation in their subregion. pandrh is also discussing strategies to follow up the implementation of its recommendations at the national and subregional levels. in addition to the biennial pan american conferences on drug regulatory harmonization that allow for communication and exchange, pandrh is also committed to training all interested parties (including regulators and industry). such training covers major topics such as gmp inspection, gcp, glp, bioequivalence, etc. the initial priorities that the pandrh defined during the first conference were gmp (to facilitate the implementation of gmp in the region and ultimately to develop mechanisms for mutual recognition of inspection), bioequivalence, and gcp. additional topics were then added, each of these considered critical in the development of the network and in the protection of public health in all concerned countries. currently, there are areas of priority that have been selected by pandrh (for which working groups have been established): several recommendations developed so far are based on who recommendations. for example, who report was the basis for the discussion on gmps, and the who and ich guidelines were used to build consensus on gcps. most of the selected topics are technical and have been chosen in order to ensure the quality, safety, and efficacy of the products approved, and that these products are adequately promoted and maintained. the work on drug classification is also key to ensuring a common language and facilitating subsequent harmonization discussions. combat against drug counterfeiting has also been selected, as this is a major issue in this region directly affecting public health in all countries and requiring a multidisciplinary, multi-sectorial, and crossborder perspective. finally, the activity on drug registration is a broader project, and is very important for ensuring implementation of pandrh recommendations and for reaching full harmonization of pharmaceutical regulations. this is critical in ultimately developing a collaborative regional or subregional registration process and system and sharing of expertise and resources between countries. this group drafted a proposed list of harmonized requirements for drug registration in the americas [ ] . the current list of selected topics above will certainly be amended in the future if new emerging topics (creating potential health public issues in several countries of the region) need to be discussed and resolved at a regional level. for example, the working group on biotechnological products has been established following a roundtable session of the th pandrh conference. this roundtable session was organized to discuss biotechnological products (and also the specific issue of biosimilars). biosimilars present a clear risk for the patient (if they are not well controlled), but also a major opportunity for increased access to cheaper essential medicines (if they are well regulated). these biotechnological/biologic products have unique technical challenges that require technical and specific expertise. pandrh will have to work on this topic collaboratively with who, which has already released recommendations on this topic. pandrh's scope of harmonization and cooperation includes technical guidelines, regulatory processes, and the strengthening of national dras through harmonization of processes and standards to improve and assure drug quality. by adopting its recommendations and standards, countries in this region can clearly improve the quality of their regulatory system and provide access to quality, safe, and effective drugs. moreover, pandrh plays an important role in the global harmonization of pharmaceutical regulations. it is an important link between global organizations/forums and the regions. through its involvement in the ich gcg, it increases: ▸ the integration of the regional challenges/priorities/vision in the development of international standards ▸ the implementation of such international standards in the region this regional initiative is one of the most difficult to operate because it includes very different regulatory systems and structures (from the most developed system such as the us fda to the most undeveloped countries in the world). this initiative also has to take into account the existence of very different political, health, and legislative realities among the countries that correspond to very different priorities, interests, and resources. this reality creates difficulties in the management of projects and the establishment of consensus [ ] . however, this disadvantage also provides opportunities and benefits as the most developed dras can help to mentor the less developed ones. recognizing preexisting asymmetries in the region, pandrh has become a forum to discuss common issues on drug regulation and share knowledge and expertise. not all the countries are involved in actually developing the proposals, but all of them participate in the decision of adopting them via the conferences. by promoting the collaboration of experts from different countries/subregions, and also from both the public sector (authorities and academia) and private sector (industry), pandrh has developed quality recommendations (frequently based on who or other international reports and recommendations). it must be noted that pandrh is clearly dependent on paho/who. without this support and investment, pandrh would certainly not be viable. indeed, this financial, technical, and administrative support from paho/who, which represents an important recognition (both in and outside the region), is critical for the following reasons: ▸ as for all such multinational initiatives, one of the challenges of pandrh is funding. pandrh's budget is primarily supported by paho, but additional funds also come from governments, the pharmaceutical industry, international organizations, and registration fees from training courses. ▸ resources from involved countries are limited. paho, by providing a secretariat, has structured this initiative and allows the practical development of the harmonization projects. ▸ who provides critical technical help for the preparation of pandrh recommendations. most pandrh guidelines and documents are indeed based on who reports. the th conference of pandrh, held in july (which included over participants from countries), focused its discussions on the theme "strengthening national health regulatory authorities." several working groups presented the conclusions of their work and their recommendations and actions. the topics also addressed during this conference included the role of pandrh as coordinator of international cooperation, paho's recognition of national regulatory reference authorities (anmat-argentina, anvisa-brazil, invima-colombia, and cecmed-cuba), implementation of the pandrh guidelines in the subregions, and innovative activities of the national dras in surveillance or in treatment compliance. this conference concluded with the approval of a strategic orientations document. the main recommendations were aimed at developing more effective cooperation among countries to guarantee, inter alia, the adoption and implementation of the different technical documents produced. the major challenges for the future (what pandrh will be assessed on) is the implementation of both its own and ich's recommendations. this will determine if this initiative delivers on its promises and if the countries that form this initiative are committed to this harmonization. because dras of all countries in the region participate in the conferences, it is expected that recommendations and guidelines will be adopted and implemented by the individual countries and incorporated in the discussion at subregional economic groups. however, it may not always be so straightforward/automatic, and the implementation of its recommendations may become one of the major challenges of this regional initiative because its members have no obligation to implement harmonized standards. the decision to develop a - pandrh strategic plan to guide future development of the network, and ensure flexibility, scientific rigor, and representation of all stakeholders in the network [ ], will certainly strengthen this initiative. the gulf cooperation council (gcc), also known as the cooperation council for the arab states of the gulf (ccasg) is a political and economic union. established in , this trade bloc comprises six arab states of the arab gulf. it represents one of the wealthiest country groupings in the world due to its extensive oil and gas reserves. its population is approximately million and its gross domestic product (gdp) is estimated at approximately us $ billion [ ] . the gcc has been active in political affairs outside its territory. due to the instability of the middle east region, the gcc has been heavily involved in diplomatic discussions to solve the different conflicts and problems of the region (i.e., iraq/iran war, iraqi invasion of kuwait, iraqi situation after the breakdown of the former regime, israeli/palestinian war, etc.). the objectives are to avoid the expansion of war and eliminate violence and terrorism in the region in order to support regional development and modernization. in order to achieve unity, the gcc promotes the coordination, integration, and interconnection between its member states in various fields. one of the first objectives of the gcc is to formulate similar regulations in different areas, including health. cooperation and coordination in health are under the responsibility of the council of the gcc health ministers (chm). under its oversight, the gulf central committee for drug registration (gcc-dr) was established to provide gulf states with safe and effective medicines at a reasonable cost. this committee works towards this objective by promoting cooperation and harmonization among member states. this initiative covers prescription, nonprescription, generics, and biologics. on the international side, the gcc represents the region at the ich global cooperation group (gcg). the current gcc members are six arab states of the arab gulf (plate ): bahrain, kuwait, oman, qatar, saudi arabia, and the united arab emirates (uae). iran and iraq are currently excluded although both nations have a coastline on the persian gulf. yemen is currently not part of the union. this country is, however, involved in some gcc initiatives (i.e., activities related to the health sector) in view of a future accession. for example, yemen is a member of the council of the gcc health ministers (chm). the supreme council is the highest authority of the gcc and is formed by the heads of the member states. presidency of the gcc supreme council rotates, and it convenes annually in a regular session, though additional extraordinary sessions may also be scheduled. this supreme council is supported by the ministerial council, composed of the ministers of foreign affairs of member states or other ministers acting on their behalf. the ministerial council proposes policies, lays out recommendations, and coordinates existing activities in all fields. resolutions adopted by other ministerial committees are referred to the ministerial council, which in turn refers relevant matters to the supreme council for approval. the chm is the highest regional level of authority in the area of health. it consists of health ministers from each of the gcc member states (plus yemen, though presently not a member). it meets for two to three days twice a year, and these meetings are open to all regulators from the gcc member states and yemen. who (via its regional office for the eastern mediterranean, emro) also attends as an observer. the chm is supported by an executive board to whom an executive office general director reports. the executive office is located in riyadh, saudi arabia. at the working level, a gcc-dr was established to oversee the different activities in the pharmaceutical sector. the steering committee of the gcc-dr is composed of two members from each of the member states (including yemen), and meets at least four times per year. the membership is limited to government agencies or dras. the executive office also appoints two of its affiliates as advisors (nonvoting members) to the steering committee. this committee is responsible for the registration of the pharmaceutical companies and their products as well as for the preparation of technical regulations and guidelines. to develop a new guideline, the gcc-dr steering committee uses the resources of the member states by assigning the drafting of the specific guideline to either a single member state or several member states. technical working groups can also be set up to help in developing the guideline. within the executive office, a permanent gcc-dr secretariat was also created to support the organization. the role of this secretariat is to facilitate the harmonization activities through administration, coordination, and communication. it is also responsible for receiving and reviewing registration files for completeness and for preparing steering committee meeting agendas. the gcc was created on may , , and its unified economic agreement was signed by its member states on november , in riyadh, saudi arabia. the primary objective was to achieve "coordination, integration and interconnection between member states in all fields in order to achieve unity between them" [ ] . this integration plan was developed in detail during the first years following the establishment of the gcc. on december , , the gcc supreme council adopted, during its nd session in muscat, oman, a revised economic agreement that accelerated this integration. this revised agreement enhanced and strengthened economic ties and increased harmonization among member states. in chapter ii, the agreement defined specific areas that needed to be harmonized in order to support the gcc common market, health being one of these areas. article also promotes joint projects and adoption of integrated policies between member states. having finally completed all requirements, the gcc common market was declared in december and came into force as of january . this launch of the common market removed barriers to cross-country investment and service trade. gcc cooperation in the health sector began in the mid- s when the gcc health ministers held informal meetings such as the one held in geneva (may , ) during the general assembly of who. such cooperation was then formalized with the establishment of the conference of the health ministers of the arab countries in the gulf, which held its first meeting in february . since , it has been called chm. as mentioned previously, under the chm, the gcc-dr was established in to provide the gulf states with safe and effective medicines. the scope of the gcc-dr's harmonization and cooperation efforts in the pharmaceutical sector covers technical guidelines and regulatory processes. this includes the registration of pharmaceutical companies and products as well as good manufacturing practice (gmp) inspection. under the oversight of the chm, the gcc-dr steering committee is responsible for the selection and prioritization of topics, the assignment of the development of guidelines and policies, and the subsequent review and approval of the resulting recommendations. when a new topic is selected for harmonization, the gcc-dr steering committee assigns the development of the guideline/policy to either a single member state or several member states, and a technical working group is then established. the membership of this working group is at the discretion of the assigned member state(s). it may include regulatory, industry, and academic experts. technical working groups meet regularly (independently of the steering committee meetings). an annual meeting is also held with both the steering committee and relevant invited experts to discuss policy and regulations. ich guidelines are often used as reference material when developing gcc-dr guidelines. other international guidelines (including who recommendations), available national technical documents, and guidelines from other regions (e.g., eu) are also used. once developed by a working group, the draft guideline is posted on the gcc and the saudi food & drug authority (sfda) websites (http://www.sgh.org.sa and http://www.sfda.gov.sa/ en/pages/default.aspx). they are also circulated to all member states for comment. at the end of the consultation period, the working group reviews all comments received, finalizes the document, and proposes its adoption by the gcc-dr steering committee. following its adoption, the general director of the executive office submits the guideline to the chm for final approval. gcc-dr steering committee members are responsible for monitoring the implementation of the adopted guidelines in their countries. each country reports whether it encounters any problems in implementing the guidelines during an annual meeting where the gcc-dr activities are evaluated. standard practices and operating procedures have been developed to govern all steps of the harmonization process (i.e., selection and prioritization of topics, solicitation of comments, approval/ implementation of guidelines and responsibilities of the different bodies, as well as funding). additional procedures also cover the process in place for the registration of products and companies. the gcc-dr is financed by member states (using established quotas of contributions) and by registration fees. the status of its activities is communicated through its website, and also through presentations at national and international meetings, workshops, and conferences. although the executive office organizes gmp training, there is currently no official structured training program within this initiative. each member state is responsible for providing training to their regulators. the gcc-dr has initiated work on several general topics related to the development and registration of all medicinal products (gmp and gmp inspection, bioequivalence studies, stability, good laboratory practice [glp] , and clinical trials). the group also decided to harmonize practices on post-marketing activities via the development of guidelines on post-marketing surveillance (covering the counterfeiting problem) and pharmacovigilance. finally, recommendations on specific types of products (biosimilars, sera and anti-venom, vaccines, and blood products) are also under discussion. the guidelines listed above are at different stages of development (under discussion, drafting in progress, approved, or implemented). they are all based on ich, who, us fda, and/or ema recommendations. in addition to these guidelines, the gcc-dr also established a common central procedure for the registration of both the pharmaceutical companies and the pharmaceutical products. the establishment of a common system of registration and control of medicines was discussed at the first meeting of the chm in . this subject was a recurrent topic of discussion until actual implementation of this procedure in . since its implementation, the registration of both medicines and pharmaceutical companies has slowly transitioned from the national to the gcc registration procedure as shown in the table . under this procedure, dossiers (including fees) are submitted to the gcc-dr secretariat. each country reviews the dossiers and forwards its recommendations to the gcc-dr steering committee. the committee's resolutions are adopted by the majority of the attendant members' votes (four countries is the minimum that must be represented). gmp inspection and analysis of samples by the accredited laboratories are also part of this central procedure. after the central approval, each country must adopt this central approval nationally. as mentioned above, the gcc-dr is responsible for gmp inspections, but also for the approval of quality control laboratories and for the review of technical and post-marketing surveillance reports. all these central activities increase the harmonization and integration of the pharmaceutical sector. since its creation and the signing of its initial unified economic agreement in , the gcc has cooperated in many different fields (i.e., political, military, security, legal, economic, environment, and health) and developed common policies in support of achieving full integration. this integration goal was reemphasized in when the gcc supreme council adopted a revised economic agreement. in january , the launch of the gcc common market marked an important step in the gcc's integration. in the health sector, cooperation began earlier. before the signature of the unified economic agreement in , the health ministers had decided to cooperate in the area of health. since the initial discussions by the health ministers, many objectives have been fulfilled. the development of common guidelines, cooperation in the domain of gmps, and the establishment of a central registration procedure for companies and products are certainly the major achievements from this group. the unified purchase of drugs (i.e., common tenders concept) is also one of the most important achievements of the chm. it has ensured the purchasing of high-quality registered products from registered companies (national or international) for a more affordable price as it increased the amount of products purchased. but it has also ensured the use of the same products by all member states, which is indeed an important step in the integration process and the creation of the common market. this cooperation allows the member countries to implement common drug policies and adopt an efficient drug quality surveillance reporting system to monitor the efficacy and safety of the registered drugs [ ] . recognizing all the above achievements, and despite clear increases in cooperation, the gcc has, however, not yet fully achieved its goal of unity in the pharmaceutical sector. indeed, this group has selected an integration model that will require stronger ties between countries. for example, the central registration procedure still involves national reviews and is longer than the national registration [ ] . moreover, approvals delivered via this central procedure still have to be adopted by each member country. this integration process is not as advanced as in europe, where there is a rapporteur that conducts the review of the application on behalf of the group and where the ec approves drugs on behalf of all european countries. harmonization of the regulation (via both regional integration and international cooperation) is critical for this region for the following two reasons: ▸ first, this region is highly dependent on medicines developed and manufactured in other countries and regions. even if pharmaceutical companies (both international and regional) are increasing their investment in the middle east region, this region is still primarily an import-oriented market. all gcc countries share the same characteristic of being high importers of pharmaceutical products. more than % and % of pharmaceuticals consumed in oman and saudi arabia, respectively, are imported [ ] . it is critical for the region to ensure that products from other countries have been developed and manufactured following acceptable standards and requirements. ▸ second, we have seen that most of the gcc-dr recommendations and guidelines are based on other international work (i.e., ich, who, etc.). the gcc is therefore dependent on the resources and expertise of these international organizations to develop its own state-of-the-art requirements and standards. the next step in the integration process of the gcc region will certainly be a better and bigger sharing of resources and expertise. the challenges of this next step will be the development of an organization and infrastructure to support such evolution. today, the regulatory expertise in the different countries is varied, with saudi arabia being the leader in the region. this country represents the biggest pharmaceutical market of the region, with approximately % of the pharmaceutical sales of the gcc [ ], and its regulatory system is recognized as the most developed of the region. in , the regulatory agency in saudi arabia, the sfda, employed people in its drug sector with approximately reviewers, compared to less than in most of the other gcc countries. the ongoing development of a common and central system needs to ensure that the less developed countries of the regions will benefit from this cooperation without impacting the more developed countries in this sector. another challenge for this group, like for all other harmonization initiatives, is the implementation of the agreed-upon standards. the gcc needs to work on measures, including the development of a structured training program, to facilitate the implementation and follow-up of recommendations. today, the southern african development community (sadc) is comprised of southern africa states, and its headquarters are located in gaborone, botswana. each of the sadc member states is at varied stages of socio-economic development, but are predominantly underdeveloped. its aggregated gross domestic product (gdp) is approximately us $ billion, with south africa representing a significant portion of this amount. its estimated total population is approximately million [ ] , with an average population growth rate of . % and an average fertility rate of . births per woman of childbearing age. approximately % of this population lacks sustainable access to affordable and quality essential medicines. the average life expectancy is . years (the lowest in the world) [ ] . the sadc objectives (listed in article of the sadc treaty) support regional integration and increased economic, social, and political cooperation in order to promote peace and security, economic growth, well being of the population, and protection of the environment and natural resources of the region. to achieve this major and broad objective, the sadc has launched projects and defined specific actions (e.g., harmonization of policies and creation of appropriate institutions and mechanisms). additionally, the sadc has had major milestones, such as the formation of the sadc free trade area (fta) in , and set future goals, including the establishment of the common market by and the creation of a single currency by . the first achievement related to the formation of the sadc fta took place on august , at sandton, south africa during the th summit of sadc heads of state and government. acknowledging that regional cooperation was critical to addressing the health problems of the region, the sadc decided to include health in its program of action. the need for harmonization of registration and control of medicines was further justified in when the disparities of legal systems and levels of development affected the implementation of a regional bulk purchasing initiative (involving five medicines used to treat tuberculosis) [ ] . the sadc health program was developed taking into account global and regional health declarations and targets. to enhance this regional health integration within a legally enforceable framework, a protocol on health matters was developed. sadc also has access to the international network because it is part of the ich global cooperation group (gcg). the the summit, comprising all the heads and/or governments of sadc member states, is the highest regional authority and therefore the supreme policymaking institution of sadc. it is responsible for the overall direction and control of the community. its structure and functions are enumerated in article of the sadc treaty. the summit usually meets in the member state holding the deputy chairpersonship of sadc at the time (additional meetings can also be held if necessary). the main objective of the organ on politics, defense and security, under the oversight of the summit, is to promote peace and security in the region. the structure, operations, and functions of the organ are regulated by the protocol on politics, defense, and security cooperation, which was approved and signed by the summit at its meeting in august in blantyre, malawi. since , the sadc leadership has been based on the troika system, which includes the chair, incoming chair, and the outgoing chair of sadc (other member states may be co-opted into the troika if necessary). the troika represents the summit between annual meetings and makes quick decisions on behalf of sadc that are ordinarily made during the summit meetings. this system allows the organization to execute tasks and implement decisions expeditiously. it also allows the provision of policy direction to sadc programs and operations between regular sadc meetings. this troika system is applied at the summit level, but is also applicable for the organ on politics, defense and security, the council, the integrated committee of ministers, and the standing committee of officials. to support the sadc activities, a central secretariat was formed. this body is defined as the principal executive institution of sadc responsible for the coordination of the harmonization of policies and strategies to accelerate regional integration. it is responsible for the management of sadc meetings, and financial and general administration. it is also involved in strategic planning, management of sadc programs, and the implementation of decisions of sadc policy organs and institutions. one of the characteristics of the sadc is its emphasis on a decentralized institutional arrangement ( figure ) . following previous negative experiences and failures in regional discussions, the founder states agreed that member states should be the principal players in the formulation and implementation of policy decisions. therefore in addition to the central sadc institutions, sadc national committees were established by the sadc treaty. these sadc institutions at the national level are present in each member state and include key stakeholders from government, the private sector, and civil society. their functions are ( ) to provide national feedback and input in regional strategy and planning, and ( ) to ensure the proper implementation of these agreed-upon regional strategies, protocols, and programs at the national level. this southern african union was created in by nine founding member states (angola, botswana, lesotho, malawi, mozambique, swaziland, united republic of tanzania, zambia, and zimbabwe) with the adoption of the lusaka declaration on april , in lusaka, zambia. at that time, this alliance was called the southern african development coordination conference, and its main objective was to coordinate development projects in order to lessen economic dependence on south africa, then under apartheid. the formation of this alliance was the culmination of a long process of consultations begun in the s when it became clear to the leaders of the founder countries that the improvement of living standards would require regional cooperation. this cooperation was directed initially towards the political liberation of the region. following the decolonization and the political independence of southern african countries, and acknowledging the poverty and economic problems of the region, the leaders of these countries saw the promotion of economic and social development through cooperation/integration as the next logical step. on august , (in windhoek, namibia) , a new declaration and treaty was signed during the summit of heads of state and government. article of the treaty gave a legal basis to the organization and promoted it from a coordinating conference into a development community. the sadc was then established to spearhead economic integration of southern africa. this strengthening of the integration process in southern africa was aligned with the overall african continental efforts to promote closer economic relations (as defined in the treaty signed in to establish the african economic community). in march , sadc country heads of state and governments met in windhoek, namibia. during this extraordinary summit, many important decisions were made that triggered an amendment to the sadc treaty. first, the summit decided to restructure sadc institutions and to establish sadc national committees in order to facilitate the implementation of a more coherent and better-coordinated strategy. the extraordinary summit also approved the preparation of the risdp by the secretariat. the purpose of this -year plan (which was adopted in august and launched in march ) was clearly to deepen regional integration by providing sadc member states with a consistent and comprehensive program of long-term economic and social policies. this plan reemphasizes the major objectives of the organization, reviews the socio-economic indicators and challenges of the region, and analyzes all the important domains for the integration process (including health). it also provides objectives and specific targets for priority intervention areas, and specifies plans and timeframes for implementation and monitoring of its important measures. for example, in the health domain, the plan proposes to coordinate, harmonize, and monitor the implementation of regional policies and to standardize the qualification and accreditation systems. the cooperation in the health domain started in with the development of the sadc health program. three key policy documents were important in the implementation of this sadc health program: as defined in article of the sadc treaty, protocols were established in each area of cooperation. these protocols spell out the objectives and scope of, and institutional mechanisms for, cooperation and integration. each protocol (which is approved by the summit and is registered with the secretariat of the united nations organization and the commission of the african union) is binding for the member states that are party to the protocol. more than protocols have been developed in all domains of integration. the protocol on health [ ] covers all aspects related to health (from the control of major communicable and noncommunicable diseases to the health laboratory service and institutional mechanisms). article states that member states should cooperate in the harmonization of procedures for pharmaceuticals, quality assurance, and registration, and also in the production, procurement, and distribution of affordable essential drugs. the implementation plan of this protocol (which further defines and prioritizes the actions to facilitate implementation of the protocol) fixes the integration of regional regulatory processes and the establishment of a mutual recognition as a - past, present, and future milestone [ ] . in line with the sadc health protocol, a pharmaceutical program was developed to address issues related to the access to quality medicines in all member states. this program was approved in june . this sadc pharmaceutical harmonization initiative and cooperative activities include the development of technical guidelines and policies relating to the registration and control of medicines across the sadc member states. the initiative aims to improve the quality, safety, and efficacy of medicines circulating within the region, and to establish and maintain a regional shared network system for dras. the ich and who guidelines, as well as other guidelines, form the basis as reference materials for the development of regional guidelines, with agreement on the adoption of international guidelines whenever possible. potential topics for harmonization are identified at the level of the subcommittee of ministers of health, often with the input of senior ministerial health officials and mra forum experts. the process of harmonization is initiated through the sadc secretariat, which prepares and submits for decision an agenda to the ministers of health. within this context, the sadc pharmaceutical business plan was released in june . this - plan identified priority areas, objectives, and major activities that needed to be implemented both at regional and national levels to improve access to quality and affordable essential medicines (including african traditional medicines). for example, strengthening regulatory capacity (and ensuring that fully functional dras are in place with an adequate enforcement infrastructure) and facilitation of the trade in pharmaceuticals within the regions were key strategies developed in the plan. the monitoring and ongoing evaluation of this plan (its implementation was estimated at us $ million) was also described (see figure , which explains the relationship between the different players of the plan). under the oversight of the ministers of health, a group of designated senior officials monitored the implementation of the plan via the establishment of technical subcommittees or task teams. this group of senior officials (from the health departments of each member state) was also supported by the secretariat. the sector of the secretariat responsible for supporting the operations of the pharmaceutical harmonization initiative takes place under the directorate of the shd&sp. national health ministries also play a significant role (by coordinating and leading the implementation of programs at the national level), and report on progress through their sadc national committees. finally, other stakeholders (e.g., professional associations, research institutions, dras, etc.) are also involved and requested to provide expertise and feedback on specific actions of the plan. in , the medicines regulatory forum was created as a technical subcommittee to promote the harmonization and enhancement of the pharmaceutical regulations in the region. this standalone committee is made of the heads of the national regulatory bodies. the sadc has released guidelines on several topics. these guidelines regulate the following general areas: ▸ the conduct of clinical trials: these guidelines provide a framework (information to be submitted, review process, etc.) and refer to the entire ich gcp (this is not a replacement or subimplementation of the ich gcp). ▸ registration of medicines: "guidelines for submitting applications for registration of a medicine" were released in . an application form is also available. ▸ good manufacturing practices. ▸ pharmacovigilance (only basic rules are provided). ▸ advertising. ▸ recalls. ▸ registration of nutritional supplements, vaccines, and traditional medicines. ▸ bioavailability and bioequivalence. ▸ stability studies. ▸ import/export (with an emphasize on gmp). most of the above guidelines are based on, or cross-reference, ich and who guidelines and recommendations. these international bodies provide much of the technical assistance to sadc initiatives. when they exist, national rules and requirements are also used (e.g., the gcp requirements from south africa). guidelines have also been developed to cover the following topics that are of specific interest for the region: ▸ pharmaceutical wholesale ▸ hiv vaccine clinical trials ▸ donations of pharmaceutical products it should be noted that the sadc efforts in the pharmaceutical area include african traditional medicines. these products are an important part of the healthcare environment of these countries. one of the cooperation projects is to establish a regional databank of traditional medicines and medicinal plants, and to develop regional policies and legal frameworks for the practice of these traditional medicines. finally, sadc is trying to establish a joint procurement system and to harmonize standard treatment guidelines/lists among countries. these two actions will facilitate the use of the same medicines within the region and therefore allow further harmonization of the pharmaceutical environment. since its inception in april , sadc has demonstrated that regional cooperation and integration is possible and useful for southern africa. one of the foremost achievements of sadc has been to put in place a regional program (the sadc programme of action) with numerous projects covering cooperation in various economic sectors. the formation of the sadc fta on august , was an important first step in this ongoing integration process. the overall and ultimate goal of sadc is integration by ; this is a very ambitious plan. presently, the level of cooperation varies for each area. in some areas, this cooperation only aims to coordinate national activities and policies. in others, the cooperation goes towards real integration. for example, on foreign policy, the main objective is coordination and cooperation, but in terms of trade and economic policy, a tighter coordination is in progress with a view to one day establishing a common market with common regulatory institutions. in the health and pharmaceutical domain, many harmonization projects have been established despite challenges. indeed, as recognized in the sadc pharmaceutical business plan, the region has many weaknesses, such as weak regulatory systems (leading to many unregistered products), lack of adequate capacity and trained personnel, outdated medicine and intellectual property laws, and noncompliance to gmp (leading to inadequate availability of medicines and poor and inconsistent quality of these medicines in some member states). even if there is a political will, it is very difficult for the authorities of this region to resolve this situation as they are confronted by two major problems: ▸ the management of major diseases (such as hiv/aids, tuberculosis, malaria, etc.) ▸ the lack of adequate resources and finances to support all health initiatives the combination of the two above problems, common to all developing countries, slows down the development of other health activities. all the efforts and resources in the domain of health are rightfully dedicated to the prevention and treatment of the major public health concerns. activities such as the development of adequate regulatory function and framework or the development and harmonization of pharmaceutical requirements are therefore negatively impacted. even if all sadc member states have national medicine policies, legislation, and regulation in place, some of these policies have been draft documents for many years (up to years). a number of the laws date back from the s (some even to the s). it is clear that such policies and legislation need revisions to include recent developments and meet current standards in public health and medicines. such revisions and updates would help the implementation of the sadc harmonized recommendations and guidelines. however, despite the numerous weaknesses and problems that the region faces, the sadc was able to promote cooperation between member states in order to improve access to quality medicines. there have been several major accomplishments in the development and harmonization of pharmaceutical requirements, such as the development of pharmaceutical guidelines for the registration and control of medicines, the establishment of the pharmaceutical business plan, and the establishment of the "medicines regulatory forum." moreover, the sadc has now analyzed (with its pharmaceutical business plan) the weaknesses, opportunities, and overall priorities in the pharmaceutical domain (i.e., regulation and control of medicines). the road map includes the assessment and strengthening of dras (work performed in collaboration with the who), combat against the spread of counterfeit medicines, the development of regional training programs, and the establishment of accredited quality control (qc) laboratories. to support this road map and other areas of harmonization, the structure of the sadc institution will certainly have to be modified (as done in the past). in order to be successful, sadc will also need to continue to work with external organizations. support and technical assistance from ich and who will continue to be critical. but, communication and cooperation with other groups and regions (e.g., the new partnership for africa's development [nepad]) will also be necessary to coordinate the efforts on the entire continent and share the available resources, financial support, and expertise. this is especially important because some sadc members are also part of other african subregional initiatives. finally, the next important phase for sadc is the implementation of the agreed-upon standards, recommendations, and plans (e.g., how will the proposed actions to "strengthen national dra capacity to implement harmonized sadc guidelines" be managed?). implementation is a challenge for all harmonization initiatives. this is especially true for this region due to all the weaknesses carried by these countries and the lack of resources and finances. however, the lack of appropriate regulations in some countries may paradoxically become an opportunity; the coordination of the development of the regulation (based on the who and ich recommendations) can be viewed as an a priori harmonization. moreover, it is interesting to note that the sadc structure presents a specificity not found in other harmonization initiatives. in addition to the standard centralized bodies (i.e., summit, council of ministers, committee of senior officials, central secretariat, etc.), the sadc has established national committees. these national sadc contact points could become critical for this implementation phase. this unusual model may also be useful for other worldwide initiatives. the association of southeast asian nations (asean), established in , has very broad objectives. the aims and purposes of the association, stated in its declaration, include: ▸ the acceleration of economic growth, social progress, and cultural development in the region through joint endeavors in the spirit of equality and partnership in order to strengthen the foundation for a prosperous and peaceful community of southeast asian nations ▸ to promote regional peace and stability through abiding respect for justice and the rule of law in the relationship among countries in the region ▸ to promote active collaboration and mutual assistance on matters of common interest in the economic, social, cultural, technical, scientific, and administrative fields ▸ to provide assistance to each other in the form of training and research facilities in the educational, professional, technical, and administrative spheres ▸ to maintain close and beneficial cooperation with existing international and regional organizations with similar aims and purposes, and explore all avenues for even closer cooperation among them the asean region has a population of approximately million, a total area of . million square kilometers, a combined gross domestic product (gdp) of us $ , billion, and a total trade of about us $ , billion [ ] . its estimated annual pharmaceutical imports and exports is us $ . billion [ ] . among the three pillars of the asean community (political-security, economic, and socio-cultural) agreed upon by the asean leaders in the declaration of asean concord ii (signed on october , in bali, indonesia), the establishment of a single market by is an important goal. its objective is to allow the creation of a stable and prosperous asean economic region in which there is a free flow of goods, services, and investments in order to reduce poverty and socio-economic disparities. at the th asean summit in january , the leaders affirmed their strong commitment to accelerate the establishment of an asean economic community (aec) by and signed the cebu declaration on the acceleration of the establishment of an asean community by . in , in moving towards this ultimate goal, asean launched the asean free trade area (afta) and defined priorities (e.g., healthcare) where regional integration should be accelerated. one of the basic criteria to support afta, and ultimately a single market, is the harmonization of standards and regulations. therefore, recognizing the importance of the harmonization of standards to facilitate and liberalize trade and investment in the region, asean has established the asean consultative committee on standards and quality (accsq) to harmonize national standards with international standards and implement mutual recognition arrangements on conformity assessment to achieve its end goal of "one standard, one test, accepted everywhere." the accsq monitors the harmonization of standards and regulations in many different areas (i.e., pharmaceutical products, but also cosmetics, medical devices, food, electrical and electronic equipment, automotive products, wood-based products, etc.). harmonization in the pharmaceutical area is coordinated by the pharmaceutical product working group (ppwg). the objective of this group is to harmonize the technical procedures and requirements applicable to the asean pharmaceutical industry in the region, taking into account other regional and international developments on pharmaceuticals. since , asean has been a member of the ich global cooperation group (gcg). this membership helps asean to become an important component in the global harmonization process, as it constitutes a way to disseminate the ich recommendations on drug regulatory harmonization. it also ensures that asean specificities and challenges will be considered when new global recommendations are discussed. the the highest decision-making body of asean is the meeting of the asean heads of state and government (the asean summit) that is convened annually. additional ministerial meetings are also held regularly. committees of senior officials, technical working groups, and task forces have been created to support the asean summit and ministerial meetings and conduct the agreed asean activities. the accsq was established to coordinate the harmonization of national standards with international standards. this committee reports to the asean senior economic official meeting (seom) that is under the supervision of the asean economic ministers (aem). the ppwg, under the supervision of the accsq, was created to coordinate the harmonization activities related to the pharmaceutical area. the scope of activities of the ppwg includes the following: ▸ exchange information on the existing pharmaceutical requirements and regulations implemented by each asean member country. ▸ review and prepare comparative studies of the requirements and regulations. ▸ review the harmonized procedures and regulatory systems currently implemented in others regions in order to develop harmonized standards, regulations, and procedures for the region. for each specific topic selected for harmonization, the ppwg sets up ad hoc committees and assigns one of the member states as the project leader. membership of the ad hoc committee is on a voluntary basis. the core members of the ppwg are the chair and co-chair, representatives from the dras from each asean member state, a representative from the asean secretariat, as well as representatives from pharmaceutical industry associations. delegates from additional member states can also participate in ppwg meetings as observers. in addition, accsq members and invited experts may attend the annual ppwg meeting. the ad hoc committee meets prior to the ppwg meetings. additionally, the ppwg operates through self-sponsorship (i.e., each member state is responsible for its own funding for traveling or hosting meetings). who has also contributed to the process in the past. ppwg activities are supported by the asean secretariat, which was established on feburary , to coordinate the asean branches and to implement asean projects and activities. in , the mandate of the asean secretary-general was enlarged to initiate, advise, coordinate, and implement the agreed-upon asean activities. finally, it should be noted that another working group, the asean working group on pharmaceuticals development (awgpd) (under the supervision of the asean health ministers meetings), also participates in the regional harmonization of pharmaceutical regulations through its activities on traditional medicines, good manufacturing practices (gmps), good clinical practices (gcps), counterfeiting drugs, and pharmacovigilance [ ] . asean was officially established with the signature of its declaration (the bangkok declaration) on august , in bangkok, thailand by the five original member countries (i.e., indonesia, malaysia, philippines, singapore, and thailand). brunei darussalam joined on january , , vietnam on july , , laos and myanmar on july , , and cambodia on april , . the accsq was formed in to facilitate and complement the afta. efforts towards specific harmonization of pharmaceutical regulations have been initiated by the accsq since . the pharmaceutical product working group was then established in september in kuala lumpur, malaysia following a decision by the accsq during its th meeting (march in manila) . during its inaugural meeting during september - , , the ppwg formulated its terms of reference and set up a work plan (i.e., goals, strategy, activities, expected output, and status). subsequent meetings focused on the status review of ongoing harmonization activities, and discussion and adoptions of final recommendations. the asean also decided to develop relationships with other countries. they developed "bilateral agreements" with a number of countries (canada, india, the us, the russian federation, pakistan, etc.), other regions (europe, gcc, sadc, andean group, mercosur), and international organizations (united nations, unesco). but one of the most important developments was the creation of the "asean plus three" cooperation to promote the east asia region. this cooperation began in december with the convening of an informal summit among asean leaders and their counterparts from east asia, namely china, japan, and the republic of korea. it was then formalized in with the issuance of a joint statement on east asia cooperation at the rd asean plus three summit in manila, philippines. the asean plus three leaders expressed confidence in further strengthening and deepening east asia cooperation at various levels and areas, particularly in economic, social, political, and other fields. public health and harmonization of standards are topics under discussion among others. several bilateral economic arrangements have already been signed, and may be the basis for the possible establishment of an east asia free trade area in the future [ ] . in november , two important documents were ratified: ▸ first, the asean charter which spells out the principles to which all member states adhere to was signed. this legal framework, which entered into force on december , , serves as a firm foundation in formulating the asean community by providing legal status and an institutional framework for asean. it also codifies asean norms, rules, and values, sets clear targets for asean, and presents accountability and compliance. ▸ second, the asean leaders also signed the declaration on the asean economic community (aec) blueprint that provides the elimination of forms of nontariff measures and market access limitations in order to transform asean into a single market. the draft guidelines developed by the ad hoc committees are reviewed, discussed, and then adopted, by consensus, during the ppwg meeting. these standards are then endorsed by the accsq. the ppwg harmonization process includes the following steps: ▸ exchange and review of information on existing pharmaceutical requirements and regulations of the member states. ▸ compare the requirements and regulations to identify key areas for harmonization. ▸ create an ad hoc committee (and assignment of a lead country) to prepare the draft "harmonized product," which most of the time is based on guidelines or recommendations already available (in one of the asean countries, internationally, or in another regions). ▸ circulate the draft to all member states for comments. ▸ consolidate comments into the revised draft. ▸ discuss and adopt (by consensus agreement) the draft by the ppwg. ▸ endorsement of the document and recommendation by the accsq. ▸ dissemination of the adopted documents (via the asean website or seminars/ meetings). ▸ compulsory implementation of the recommendation by the member states. in order to organize, coordinate, and monitor the implementation of the agreed-upon recommendations and guidelines, the ppwg set up a specific task force and working group to focus on a mutual recognition arrangement (mra) and implementation. they developed a standard operating procedure (sop) and plan of action. they also assessed the status of the implementation of requirements (i.e., adoption into the national systems) in order to develop appropriate training (to government and industry) to increase understanding of the asean guidelines and fill the gaps among the member states. the first project of the asean ppwg was to compare the existing product registration requirements for pharmaceuticals of asean member countries in order to help define key areas for harmonization. this report was finalized in . following this assessment, the group developed the asean common technical requirements (actrs) for pharmaceutical product registration in the asean region. these requirements are sometimes based on the existing national requirements, who guidelines and recommendations from other regions (e.g. the asean guidelines for "the conduct of bioavailability and bioequivalence studies" were created from the ema/cpmp note for guidance). but most of the asean actrs have been developed via the adoption or modification of the ich guidelines. they cover all the quality, nonclinical, and clinical aspects already developed by ich. labeling requirements, administrative data (i.e., certificate of pharmaceutical product (cpp), letter of authorization, application forms, etc.), and the glossary have also been discussed. the final actrs were endorsed by the accsq at its st meeting (in march ) . guidelines to actr (e.g., process validation and stability) have also been developed. the group also developed an asean common technical dossier (actd) for pharmaceutical product registration. like the ich ctd, this initiative reduces the time and resources needed to compile applications for registration in different countries. regulatory reviews and communication with the applicant is also facilitated by a standard document of common elements. this actd is based on the ich ctd, but is organized into four parts only (the overview and summaries are included at the beginning of the relevant parts i, ii, and iii instead of being grouped under a separate section as in module of the ich ctd): ▸ part i: activities have also been conducted in the area of gmps. on april , , the asean economic ministers (at the th asean summit and related summits in pattaya, thailand) signed the asean mra for gmp inspection of manufacturers of medicines. this arrangement establishes the mutual recognition of gmp certifications and/or inspection reports (issued by inspection bodies) that will be used as the basis for regulatory actions such as granting of licenses and supporting post-marketing assessment of conformity of these products. the ppwg also worked on a bioavailability/bioequivalence study reporting format and a post-market alert system. the objective of the asean post-marketing alert system is to share information relating to defective or unsafe medicines, and also cosmetics, health supplements, and traditional medicines. this pilot project was launched in april and then adopted by the ppwg in february . the two major accomplishments of the ppwg are the actd and the atcrs. the actd is the common format for marketing authorization application dossiers, while the atcrs are the set of written materials intended to guide applicants to prepare application dossiers in a manner that is consistent with the expectations of all asean dras. a series of guidelines for the implementation of the atcrs is being finalized. most of the asean recommendations strictly follow the ich guidelines and recommendations. indeed, asean is a good example of the influence of the ich outside the ich regions and of the integration and implementation of ich standards outside ich frontiers. beyond these harmonized technical aspects of the pharmaceutical product registration that need to continue, the ultimate goal of the asean ppwg is clearly to implement a system where countries fully cooperate in enhancing mutual regulatory capacities and resources. with the ongoing challenges posed by the globalized economy, and in particular the huge economic growth of china and india, which may have specific impacts on the region, this association of countries is clearly committed to full integration (with the goal to establish an asean economic community by ) and moving towards the european community model. the ultimate steps in the pharmaceutical harmonization process will certainly be the development of asean pharmaceutical directives, the development of a pan-asean registration process (with a centralized procedure), and the establishment of an asean regulatory agency. but the full implementation of this supranational system will take time. it will only be possible when the asean has developed common legislation and structure (i.e., commission, parliament, etc.), as in europe. the harmonization of pharmaceutical regulations can, however, continue before such an organization is in place. the next logical step is the creation of an mra procedure. indeed, this type of procedure is not binding for the countries (and therefore does not require common legislation) and requests only a "facilitator body" and not a supranational evaluation agency. this procedure would be similar to the old "multi-state" procedure that europe established in as a first step towards the creation of the system that we know today. asean is also committed to increased relations with external partners. the creation of the asean plus three cooperation may indeed promote the harmonization of pharmaceutical regulations in the much broader asia region. outside the region, asean and its ppwg clearly want to increase relationships and cooperation with other regional organizations, and also international bodies (i.e., un, who, ich). this development, which is outside its current framework, could indeed strengthen this initiative by increasing its exposure on an international basis, therefore allowing this organization to play a pivotal role in the international community. the asia-pacific economic cooperation (apec) is a forum, established in , to facilitate economic growth, cooperation, trade, and investment in the asia-pacific region. this region accounts for approximately % of the world's population, approximately % of world gross domestic product (gdp), and about % of world trade [ ] . since its creation, this intergovernmental grouping has worked to reduce tariffs and other trade barriers across the asia-pacific region in order to liberalize trade and investment and facilitate business within the region. apec also works to create an environment for the safe and efficient movement of goods, services, and people across borders in the region through policy alignment and economic and technical cooperation. to support its "three pillars" (i.e., trade and investment liberalization, business facilitation, and economic and technical cooperation), apec has been active in a broad range of more than topics (from fisheries, agriculture, and tourism to terrorism, finance, and intellectual property). this broad range of topics, under which hundreds of specific projects have been developed, reflects the complex factors and issues related to economic development, growth, and the pursuit of open trade and investment for a region. several of these topics can influence the health and pharmaceutical sector (such as intellectual property or science and technology), but two specifically focus on this area: ▸ the health topic, managed by the "health working group," focuses mainly on the prevention and management of infectious diseases (naturally occurring or due to bioterrorism) in the region. this working group is not involved in the discussion related to pharmaceutical regulation. ▸ the life sciences topic, managed by the life sciences innovation forum (lsif), addresses key challenges in the health and pharmaceutical sector in order to create the right policy environment for life sciences innovation. the harmonization of standards and the regional and international cooperation are two of the tools used to achieve the objectives. as a member of the ich global cooperation group (gcg) since , apec lsif promotes the implementation of the ich guidelines through its workshops. it also keeps ich informed on the status of the different ongoing initiatives in the region. apec has member economies from the broad asia-pacific region, which spans four continents (plate the members of apec recognize that strong economies and harmonization initiatives are not built by governments alone, but by partnerships between government and its key stakeholders, including industry, academia, research institutions, and interest groups within the community. therefore, apec actively involves these key stakeholders in the work of the forum. at the working level, representatives from the private sector are invited to join many apec working and expert groups. this process provides an important opportunity for industry to provide direct input into apec's ongoing work. apec has official observers, the association of southeast asian nations (asean) secretariat being one of them. these observers participate in apec meetings and have full access to documents and information. apec operates as a cooperative, multilateral economic, and trade forum. apec policy direction is provided by apec leaders from the member economies. the life sciences innovation forum (lsif), under the committee on trade and investment, is a tripartite forum involving representatives from government and academia, and also from industry. it brings together scientific, health, trade, economic, and financial considerations to create the right policy environment for life sciences innovation. all the apec activities are supported by the apec secretariat, which is based in singapore and operates as the core support mechanism for the apec process. it provides coordination, technical, and advisory support, as well as information management, communication, and public outreach services. the idea of apec as a cooperative to enhance economic growth and prosperity, and to strengthen the asia-pacific community, was first publicly mentioned by the former prime minister of australia (bob hawke) during a speech in seoul, south korea in january . later that year, asia-pacific economies met in canberra, australia to establish apec. in november , apec's vision was reiterated by apec economic leaders during their meeting in bogor, indonesia. during this meeting, the economic leaders adopted what are referred to as the "bogor goals." these goals of "free and open trade and investment in asia-pacific no later than " were based on a recognition of the growing interdependence of the economically diverse region, which comprises developed, newly industrializing, and developing economies. due to the heterogeneity of the region, it was agreed that the pace of implementation would take into account differing levels of economic development among apec economies. in , a framework for meeting the bogor goals (referred to as "the osaka action agenda") was adopted. this action plan focused on three key areas: ▸ trade and investment liberalization ▸ business facilitation ▸ economic and technical cooperation following this first action plan, several other plans have been adopted over the years to support the implementation of the bogor goals. specific topics (such as climate change and severe acute respiratory syndrome [sars]) were also discussed. recognizing the global financial crisis as one of the most serious economic challenges ever faced, the leaders highlighted the importance of reducing the gap between developed and developing members. this meeting included discussions related to regional economic challenges (implementing a structural reform and food supply and price), the social dimension of globalization, the enhancement of human security in the region, and the problem of climate change. the lsif and the health working group held their first joint meeting in march in washington, dc, us to explore possible areas of cooperation. this meeting followed the recommendations of the apec senior official endorsing a new terms of reference for the steering committee on economic and technical cooperation. it was then agreed that the role and operations of the health working group would be reviewed with a view to merge, disband, or reorient this body. the lsif leads the activities related to the regulatory convergence in the pharmaceutical area within the asia-pacific region. both apec and the lsif have recognized the benefits of convergence related to the pharmaceutical standards within the region. to achieve this goal, these two groups rely on other regional and global harmonization initiatives. indeed, the lsif is working towards the adoption and implementation of existing harmonized international guidance and regulatory best practices. it also provides the ability to access funds to advance projects. unlike asean, the objective is not to proactively develop specific regional harmonized guidance. this practice is in line with the overall apec goals to facilitate cooperation and trade in the region, and to operate on the basis of nonbinding commitments and open dialogue. as already mentioned, apec has no treaty obligations required of its participants, and there is no plan for integration (unlike asean, which follows an integration model like europe). recognizing this specific context, the objective of lsif is "regulatory convergence" with gradual alignment over time between member economies. the distinction with "regulatory harmonization" is that "regulatory convergence" does not typically involve or require active harmonization of regulations that would be unrealistic within the apec environment. the objectives and priorities of the lsif, listed in its strategic plan approved by the apec ministers in , are very broad. this plan includes recommendations on four different sectors: research, development, manufacturing and marketing, and health services. the goal was to develop recommendations that would contribute to a more efficient, effective, and coordinated policy approach to support innovation and health in the region. these recommendations have applications in many different areas (legal, finance, scientific, regulatory, infrastructures, etc.). one of the recommendations from this strategic plan follows: "harmonization of standards for life sciences products and services and mechanisms for collaboration and exchange of information among economies were recognized as critical elements" [ ] . the principle was to review policies, standards, and regulatory mechanisms against international best practices in order to move towards regional convergence. the objective was also to achieve close collaboration and to facilitate the use of international standards and global best practices through collaboration with outside bodies such as the ich gcg. the lsif has been very active in sponsoring a series of workshops on anti-counterfeiting, ich quality guidance, clinical trials, and good clinical practice (gcp) inspection. however, it has been recognized that the lsif has not been used to its full potential to promote regulatory convergence and cooperation compared to some other rhis [ ] . what was missing was the engagement of regulators and the appropriate industry people in this equation, together with the lack of a more focused strategic framework and multiyear plan for medical products. in / , acknowledging the lack of strategic and effective approaches, the lsif decided to react and strengthen its organization: ▸ in june , the lsif took an important step towards harmonization by establishing, in seoul, south korea, the apec harmonization center (ahc). this followed a proposal from south korea in august (at the apec lsif vi in lima, peru) that was endorsed by the apec leaders in november in a joint ministerial statement. as an lsif organization, this center has its own structure (including a director, a secretariat, and an advisory board of lsif experts), and also its own website (www.apec-ahc.org). this organization includes representatives from government, industry, and academia. its mandate is to provide a platform to address and solve priority concerns of apec members on regulatory convergence. following the establishment of the ahc, several workshops took place. in general, they focused on the regional regulatory convergence, but also discussed specific problems such as multiregional clinical trials and the biosimilar concept. the purpose of these workshops is to allow government, regulators, academics, and the pharmaceutical companies to discuss and exchange information and views on the harmonization of standards. funding and support from the ahc has allowed for the delivery of more than a dozen workshops since june . ▸ in addition to the ahc, apec also decided to establish a regulatory harmonization steering committee (rhsc) within the lsif structure to strategically coordinate regulatory convergence in the region. the rhsc brought together senior officials from regulatory authorities and representatives from industry coalitions. this committee provides leadership and direction on regulatory priorities. during its inaugural meeting in seoul, south korea in june , the rhsc discussed and finalized its terms of reference and started to identify priority projects. since then, the rhsc has initiated several projects and developed a strategic framework on regulatory convergence of medical products by to coordinate activities [ ] . since the creation of the apec ahc and rhsc, considerable progress has been made with the design, development, and implementation of a more strategic, coordinated, and sustainable approach. this includes the strategic framework and the creation of priority work areas (pwas), each of which is associated with a roadmap that defines an overall strategy to achieve the ultimate goal of greater regulatory convergence by in the area of medical products. each project or activity undertaken must now support the roadmap and in turn move apec closer to the goal. this is a better-structured organization that moves away from individual, uncoordinated activities and workshops to a more directed, coordinated approach with parties and individuals that are in a position to effect change and commit resources. the workshops, organized and funded by the ahc and led by the rhsc membership, are now tied to a directed roadmap and strategic framework representing the collective efforts and commitment of many economies. these workshops served as a diagnostic of issues, challenges, and opportunities associated with a particular area of focus, with recommendations coming back to the rhsc for consideration. all workshops are championed by the regulators of various apec economies (for example, the us for medical product quality and supply chain integrity, korea for biotechnological products and pharmacovigilance, singapore for cellular-and tissue-based therapies, chinese taipei for good review practices and combination products, and thailand for gcp inspections). finally, this organization is partnering with other regional and international players in an effort to promote synergy and more effective use of resources. a good example here is the supply chain roadmap. this is a global issue and requires a global, coordinated approach. the rhsc roadmap is being implemented through the direction of an oversight committee that includes the who, ema, edqm, and the dra of nigeria. in doing so, apec takes account of and complements like initiatives, and can serve as a catalyst to global action. up to now, the apec did not proactively develop guidance or harmonized standards and requirements. the objective is to promote convergence via the dissemination of international harmonized information and recommendations (i.e., ich guidelines). to achieve these goals, the group has developed and funded several projects. in , the lsif released an "enablers of investment checklist," a voluntary guidance tool for member economies to assess and improve their innovative life sciences sector investment opportunities. one of the six principles covered by the checklist is "efficient and internationally harmonized regulatory systems." under this principle, the lsif promoted the development and implementation of focused efforts on harmonization towards international standards through recognized international organizations (i.e., ich). moreover, to support this objective, the lsif also proposed development of the following: ▸ a regulatory framework (transparent, predictable, and science-based) that allows for the quick introduction of new innovative products ▸ an efficient clinical trial regulatory system focused on safety, efficacy, and ethical standards ▸ an adequate number and level of training programs for regulatory personnel ▸ the publication of proposed regulations for stakeholder comments (which should be taken into account) ▸ laws providing for stakeholder consultation throughout the regulatory drafting and review process ▸ participation in international joint clinical trials performance metrics have also been defined to assess the implementation of the recommendations. finally, some of the other principles on this checklist also support cooperation and convergence as they assess the resources, exchange programs, intellectual property rights, and interagency coordination of life science policy and regulation. in addition to the "enablers of investment checklist," lsif has also developed projects focusing on specific topics of interest, such as: ▸ clinical trials: the area of clinical trials was selected as one of the lsif priorities in its strategic plan. assessment and improvement of the clinical trial system and regulation in each country has also been recommended in the lsif "enablers of investment checklist." the goal was to put in place an effective regulation infrastructure (by harmonizing regulatory practices and policies according to international best practices and standards). this activity includes work on regulatory process and framework (incorporating interagency review of new policies, guidances, and regulations), implementation and promotion of good clinical practices (gcps)/good manufacturing practices (gmps), protection and enforcement of intellectual property, establishment of clinical trials registries, and implementation of ich recommendations. to implement this goal and strengthen the dras' capacity to harmonize practices, a first workshop on "review of drug development in clinical trials" was held in march . several additional workshops concerning clinical trials and gcp (including clinical research inspection) have since been set up on this subject. the first workshop organized by the ahc in focused on the opportunities and challenges of multiregional clinical trials. each of the workshops serves to refine recommendations and showcase the china-japan-korea tripartite research initiative that is exploring possible ethnic differences between the three countries. as a result of workshops, two roadmaps have been developed: one for gcp inspection (under the leadership of thailand), and one for multiregional clinical trials (under the leadership of japan) [ ] . the focus will address gaps and needs not addressed by any other institution or regulatory authority to date. ▸ counterfeit medicines: another area of interest for lsif has been the increase in counterfeit medicines in the region. a series of seminars and workshops have been organized since january to examine ways to combat this problem. the lsif has also developed an anti-counterfeit medical product action plan. the objective of this plan is to share best practices in the detection and prevention of counterfeits to both dras and industry professionals, and organize systems to reduce the threat and occurrence of counterfeit medicines. finally, it is important to note that apec also promotes capacity building for its members. this objective is met through the organization of workshops, training courses, and seminars that enable people, businesses, and government departments to improve their skills and knowledge [ ] . the primary focus of apec is clearly the economy, and its objectives center on the facilitation of trade and business between member economies (with no integration plan). the asia-pacific region has consistently been one of the most economically dynamic regions in the world. since the establishment of apec in , the total amount of trade has grown significantly [ ] . apec's work under its three main pillars of activity has helped drive this economic growth. in , apec conducted an assessment to determine what progress has been made against the bogor goals of free and open trade and investment. the results were positive, showing that member economies have taken concerted action and progressed in a wide array of economic, trade, investment, and social areas. average tariffs in the region have been reduced from about % in to approximately . % in . nontariff barriers have also been reduced thanks to apec's work on trade facilitation. this progress by apec towards the bogor goals contributed to a more than five-fold increase in members' total trade (goods and services) between and (from us $ . trillion to us $ . trillion). finally, these activities contributed to real benefits for people across the entire asia-pacific region. over the span of years, from to , poverty was reduced by % (poverty levels are measured by calculating the population living on less than us $ a day) [ ] . apec represents a large region and approximately % of the world's population. this is obviously an advantage in facing the challenge of globalization. however, this size and magnitude can also be a disadvantage in terms of management. indeed, this region is very heterogeneous with countries at the two extremes of the development spectrum (i.e., very developed and very undeveloped countries). due to this disadvantage and the heterogeneity of this large region, it is difficult to adopt a treaty and to impose obligations on these members. for this reason, apec operates on the basis of nonbinding commitments where each country has the choice to implement the decisions. the implementation of economic measures (i.e., reduction of taxes and trade barriers to increase trade between members) is possible since it can quickly benefit all members. however, the lack of a treaty or obligations on members can sometimes be more challenging for more drastic long-term reforms (i.e., the harmonization of standards), as member economies have different priorities. the diversity of the apec region means that member economies will gradually move closer together in requirements and approaches, but not everyone will implement the measures at the same time. capacity and local realities must be taken into account. though technical cooperation is part of apec's objectives (i.e., apec is very involved on specific topics such as climate change), it is the second priority behind economic development. the health topic, managed by the health working group and the life science innovation forum, has clearly been funded because this topic has an impact on the economy. as stated on the apec website, "life sciences innovation is critical to growth and socio-economic development as healthy people produce healthy economies. efficient and effective delivery of patient focused products and services can improve a population's longevity, wellness, productivity and economic potential" [ ] . however, even if the above challenges are important, very positive outcomes have to be noted in terms of regulatory convergence in the pharmaceutical area. indeed, this organization supports convergence via the funding of projects and workshops. lsif was able to focus its effort on projects that impact all member economies (developed or developing), such as the coordination of multicountry clinical trials, the implementation of gcps, the quality of medicines, the counterfeit medicines problem, and the emergence of biosimilars. lsif also creates a forum that allows exchange of information between very different countries and between all the players (regulators, industry, and academia). this communication and dissemination of harmonized standards is very important, and is as essential as the development of the standards itself. in / , acknowledging a lack of strategic coordination, apec and lsif decided to better organize the activities. first, they established the ahc to facilitate the exchange of information and the creation of a network. second, they created the rhsc to strategically coordinate regional convergence. since this revision of lsif's structure and the creation of these two supporting bodies, significant progress has been made and apec has since declared that further harmonization to "achieve convergence on regulatory approval procedures" is targeted for [ ] . to support this goal, many important projects have been initiated on critical topics, such as product quality and supply chain integrity [ ] , good review practices [ ], gcp inspection [ ], pharmacovigilance [ ] , biotechnology products [ ], etc. all these changes and projects today represent great promise for this region, and the tools to be developed could also support global cooperation and convergence. the challenge is now to implement the plan and to continue to coordinate the projects in order to achieve the desired objectives. the recent establishment of the rhsc regulatory network (including dras not currently part of the rhsc) will certainly support the implementation of agreed-upon measures. many different types of bilateral cooperation have been established over the years. lll it would be difficult to list and discuss them all as several dozen exist. however, all these types of bilateral cooperation and agreements can be grouped into three categories based on their scope and objectives: ▸ cooperation between two developed countries: the objective of such cooperation is to exchange good practices and harmonize standards to avoid duplication of efforts (e.g., for orphan drugs). for example, the eu and the us developed a privileged relationship and the exchange of officials and staff between us fda and eu authorities allow for a closer collaboration, exchange, and therefore better understanding of each other. ▸ cooperation between one developed country and one developing country: this type of cooperation focuses on training, mentoring, and support from the developed country to the developing country. the objective is indeed to build expertise and capacity in the developing country based on the experience of the developed country. for example, the us fda has established several agreements with developing countries (e.g., brazil, mexico, south africa, taiwan, etc.) ▸ cooperation between two developing countries: by pooling experience and resources, two countries can better tackle issues of common interest. this type of cooperation allows for better allocation of sparse resources, and also increases interest for pharmaceutical companies (two small markets with different requirements would be less attractive to industry). for example, brazil has cooperation projects with cuba, dominican republic, mozambique, and several other countries [ , ] . one of the most advanced bilateral collaborations is between australia and new zealand. it represents a good example of a bilateral cooperation and harmonization model working towards a full integration of systems. indeed, after several years of convergence and harmonization, australia and new zealand agreed to establish a joint australia new zealand lll bilateral cooperation can involve two countries, but it can also mean the collaboration of a regional entity with another party. for example, the european union has been collaborating with australia, canada, the us, and japan, but also with the gcc group. therapeutic products agency (anztpa). this new agency will ultimately replace australia's therapeutic goods administration (tga) and the new zealand medicines and medical devices safety authority (medsafe). during the first meeting of the anztpa implementation ministerial council (melbourne, january , ), ministers from both countries agreed on key elements to establish the joint trans-tasman agency, and also how the joint regulatory scheme will be organized over a five-year period [ - ]. since then, the framework of the anztpa is under discussion . this cooperation/harmonization initiative was begun with the objective of sharing expertise and resources in order to provide health benefits for consumers by creating a world-class scheme. it is also expected that this single approval process for both countries will increase efficiency, improve the standards of medicines produced in the two countries, reduce regulatory costs for industry, and facilitate further economic integration [ ] . this initiative is a great example of successful bilateral harmonization and cooperation, and emphasizes the importance of a staged approach for this type of project. it also shows that such ultimate integration of systems is challenging. indeed, the agreement for a joint regulatory scheme was first reached in , but this project was not able to proceed because new zealand was unable to pass enabling legislation. negotiations between the countries were also suspended in july [ ] . the increased collaboration between europe and the us in the pharmaceutical domain is another interesting example of bilateral cooperation. though this cooperation does not follow an integration model, it is a well-developed bilateral initiative. it is a stepwise and structured program that is interesting as it provides a clear example of what such bilateral collaboration can achieve in a nonintegration process, and also outlines its limitations. it also provides examples of the measures and organization necessary to support such bilateral work. the european union (eu) and the united states of america (us), in addition to their collaboration within the scope of multilateral frameworks such as the international conference on harmonization of technical requirements for registration of pharmaceuticals for human use (ich), have also established strong regulatory and scientific bilateral cooperation in the pharmaceutical sector. this bilateral cooperation promotes public health, safer trade of products, and harmonization of regulations. over the years, the scope of this transatlantic collaboration has increased, and today represents a good example of what bilateral cooperation can achieve. this collaboration mainly involves the european commission (ec), the european medicines agency (ema) and the united states food and drug administration (us fda). however, it is important to note that the us fda also maintains an active relationship with national dras throughout europe. confidentiality arrangements with the us have been signed at the european level (ec and ema) and also at the national level with austria, belgium, denmark, france, germany, ireland, italy, the netherlands, sweden, and the united kingdom. this is particularly important for collaboration in the area of inspections. it also allows the us fda to exchange information on products not approved via the centralized procedure (this exchange is done through the relevant reference member states [rmss]). the leaders of the eu and the us agreed on a framework for advancing transatlantic economic integration and established the transatlantic economic council (tec) to oversee the efforts outlined in the framework, with the goal of accelerating progress and guiding work between eu-us summits. moreover, confidentiality agreements have been established to create a framework allowing the exchange of confidential information between the eu and the us fda as part of their regulatory and scientific processes. they include information on advanced drafts of legislation and regulatory guidance documents, as well as nonpublic information related to ensuring the quality, safety, and efficacy of medicinal products for human (and veterinary) use. an implementation plan has also been agreed upon between all parties to allow for a successful exchange of information and documents between the eu and the us fda in accordance with the terms of the confidentiality agreements. the objective of this implementation plan was to describe the processes by which each party will undertake information and document exchange as envisioned by the confidentiality agreements. also, to facilitate this transatlantic pharmaceutical cooperation, the us fda and the ema have established "liaison officials." these liaison officials remain employed by their home organizations, but their physical location in the partner agency is designed to facilitate collaboration. their role is to facilitate regulatory and scientific cooperation between the us fda and the ema, and to coordinate information exchange. they also increase awareness of interaction opportunities with the ema and the us fda, and potential new areas of common interest [ , ]. in , the scope of this bilateral cooperation intensified with the establishment of confidentiality arrangements between the parties. these agreements signed on september , were then extended on september , . in september , these confidentiality agreements were extended again, and are now in effect for an indefinite period without the need for further renewal. these two official statements of authority and confidentiality commitment [ , ] restate the agreement to pursue in-depth collaboration and exchange of confidential nonpublic information between the us fda and the ema. it is interesting to note that these statements reiterate that the shared information includes confidential commercial or trade secret information (the us fda is required by current legislation to ask pharmaceutical companies before sharing trade secret information with counterpart dras). at the eu-us summit on april , , further momentum was given to regulatory collaboration with the signature of the framework for advancing transatlantic economic integration between the european union and the united states of america by ec president josé manuel barroso, german chancellor angela merkel, and us president george w. bush. this document called for more effective, systematic, and transparent regulatory cooperation, and the removal of unnecessary differences between regulations. it also specifically requested the promotion of "administrative simplification in the application of regulation of medicinal products." the objective of this bilateral process is more towards cooperation than harmonization per se. exchange of information between the parties allows for a better understanding of each other's systems and requirements, and therefore builds confidence and recognition facilitating convergence. this eu-us cooperation also tries to avoid future disharmony by upstream regulatory cooperation on new medicines legislation [ ] . the exchange of information and practices are well structured and occur on a regular basis, but the exchange can also be done on an ad hoc basis if necessary. ▸ regular exchange: the ema and us fda exchange a list of specific information on applications (both pre-authorization of new molecules and post-authorization of marketing products), including decisions made for such applications on a quarterly basis. they also exchange other information such as a list of good clinical practice (gcp) inspections or pharmacovigilance topics (either product-or nonproduct-related issues). ▸ ad hoc exchange: in addition to the exchange of new drafts of final legislation or guidelines (prior to publication), the eu and us fda also exchange information relating to scientific advice, difficulties in relation to the evaluation of applications, and urgent drug safety issues and other issues impacting public health. these types of information are exchanged prior to their release into the public domain. meetings or workshops on regulatory issues of mutual concern are also organized on an ad hoc basis. finally, the ema and the us fda publish an annual report summarizing their interactions under the confidentiality arrangements. these arrangements also provide for annual meetings between the us fda, the ema, and the ec to monitor the operation of activities within the scope of the agreed-upon implementation plans. however, it should be noted that the sharing of product-related information is limited to medicinal products evaluated or authorized in accordance with the eu centralized procedure, as well as medicinal products authorized at the national level by the eu member states, which are subject to arbitration or referral in accordance with european community procedures [ ]. initiatives related to general topics are reported below. in addition to these initiatives, cooperation has also been established in certain specific scientific areas or for a specific type of product (i.e., oncology, pharmacogenomics, nanotechnology, advanced therapy medicinal products [atmp], blood products, and vaccines). under the auspices of the transatlantic economic council, on november , the ec hosted the "transatlantic administrative simplification workshop" in brussels, belgium, which was co-chaired by the ec and the us fda and organized in collaboration with the ema and the heads of the eu national medicines agencies (hma). the key objective was to identify opportunities for administrative simplification through transatlantic cooperation in the removal of unnecessary burdens of administrative practices and guidelines. this would allow more human and fiscal resources to be focused on greater innovation and efficiency in the development of quality products. it was agreed that this project should not require change to legislation, and of course, the simplifications should maintain or increase current levels of public health protection. as a follow up to the transatlantic administrative simplification workshop, a "medicines regulation transatlantic administrative simplification action plan" was published in june , outlining administrative simplification projects to be taken forward. this document promoted further cooperation and pilot collaboration programs in major areas such as inspections, biomarkers, counterfeit medicines, risk management (content and format), scientific advices, biosimilars, pediatrics, and advanced therapies. during the annual ec/ema-us fda bilateral meeting in september , it was agreed that the majority of projects in the original plan had been successfully completed and that most of the pilot projects had been extended and became "standard" cooperation [ ] . ongoing developments and new initiatives in transatlantic administrative simplification are now included in the annual reports on interactions between the us fda and the ema. several projects have been initiated to increase collaboration on gmp and gcp inspections. ad hoc exchanges on specific products, quality defects, product shortages, and on draft guidelines also took place. ▸ gmp inspections: several pilot projects were first initiated in the context of the transatlantic administrative simplification workshop deliverables. an initial project (established in cooperation with the european directorate for the quality of medicine and the australian therapeutic goods agency) was conducted between december and december and related to gmp inspections of active pharmaceutical ingredients (api) manufacturers [ ] . the project's objective was to determine whether greater international collaboration and information sharing could help to better distribute inspection capacity, thus allowing more sites to be monitored and reducing unnecessary duplication. the second project, related to finished products, allowed eu-us fda joint inspections and was aimed at developing ways of working together on joint inspections of routinely scheduled sites in the territory of the us or eu, to reduce duplicate inspections and the resulting burden on both the pharmaceutical industry and the dras. this pilot phase, conducted under confidentiality agreements, allowed the development of new tools for work sharing and the exchange of information in order to share inspection reports and to organize joint inspections. increased transparency and visibility of inspections performed by participating authorities allowed a successful collaboration between authorities on manufacturing sites of common interest. it also increased the number of inspections performed that were of value to more than one authority. this pilot phase confirmed that such collaboration in the area of gmp inspections led to a reduction in duplicate inspections, more efficient use of combined inspectional resources, and wider global inspectional coverage. following the successful conclusion of the pilot, it was agreed to maintain the cooperation established [ ] . in december , the us fda and the ema decided to further enhance their gmp inspection cooperation by moving from confidence building to reliance upon [ ] . this initiative, launched in january , allows the ema and the us fda to share inspections of manufacturing sites in each other's territories. this important step follows the positive experience acquired through the pilot joint inspection programs and other information sharing projects that have occurred over several years. this strategy allows some inspections on each other's territories to be deferred or waived completely based on a number of considerations and on a risk-based approach [ ] . this strategy is applicable to gmp inspections related to manufacturing sites located in the us and the european economic area (eea), mainly focusing on routine post-authorization and surveillance inspections as a first step [ ] . the result of this arrangement could free up inspection resources that would then become available for inspections to other regions. ongoing ema-us fda joint inspection pilot projects will continue according to the agreedupon procedures [ , ] as it remains important to maintain mutual confidence and build further mutual understanding of gmp inspection approaches. some successful pilot programs will also be expanded to new partners such as the ongoing collaboration on gmp inspections of active substance manufacturers [ ] . due to the increased globalization of pharmaceutical product clinical development, and based on previous positive experiences in the gmp field, the ema and us fda agreed to launch a pilot ema-us fda gcp initiative. the objective of this gcp initiative, conducted between september and march , was to reinforce and systematize periodic information exchanges on gcp-related activities between the us fda and ema. these included the exchange of gcp inspection plans to improve inspection coverage, the exchange of information on applications to help identify candidates for collaborative inspections, and the exchange of inspection outcomes and reports (both negative and positive) and their potential impact. conduct of collaborative gcp inspections and the sharing of information on interpretation of gcp (such as draft guidelines or policies) were also part of this project. the pilot initiative has been very productive. a considerable amount of information has been exchanged on many products [ ] , and this communication (which included teleconferences and four face-to-face meetings) has facilitated improvements in the inspection coverage and decision-making processes of the agencies. the collaborative inspections conducted under the initiative have contributed greatly to each agency's understanding of the other's inspection procedures. they have also led to the identification of potential improvements to these procedures. both agencies have learned several general lessons during the process [ ]. in addition, exchanges of views on interpretation of gcp documents have also been organized. during the pilot initiative, the ema and the us fda have shared different pieces of gcp-related guidance documents, position papers, and policies in order to harmonize the agencies' understanding of gcp and to standardize the requirements for industry wherever convergence would be beneficial for the clinical research process. at the end of the program, both parties considered this pilot initiative very successful and agreed to continue this collaboration, incorporating lessons learned with the broader aim of moving from "confidence building" to the mutual acceptance of inspectional findings. the agencies will also expand the scope of the initiative to sites outside the us and eu [ ]. although not defined as a cluster, interactions in the area of safety continue to play an important part in the ongoing collaboration between the us fda and the ema. ▸ videoconferences take place on a bimonthly basis and include product-related issues and issues related to risk management. usually five to six products are discussed at these teleconferences. ▸ regular informal teleconferences in order to exchange information on emerging safety and strategic issues. ▸ ema shares the early notification system on a monthly basis and the us fda sends advance notice of publication of its quarterly update reports on potential safety signals. ▸ joint projects have also been established, such as the collaborative project on the progressive multifocal leukoencephalopathy research agenda to stimulate research into this important safety issue that affects some biological agents. the objective of this program is to allow interaction between the ema and the us fda assessors and sponsors during product development. this dialogue between the two agencies' assessors and sponsors on scientific issues [ ] aims to optimize product development and avoid both unnecessary testing replication and unnecessary diverse testing methodologies. such a procedure can be valuable for products developed for indications for which development guidelines do not exist, or if guidelines do exist, the ema's and the us fda's recommendations differ significantly. experts from the ema and the us fda exchange views and discuss draft responses to questions from the applicants on their clinical development programs or on new biomarkers. general principles for this voluntary parallel scientific advice were published in by the ema and the us fda [ ] . it is important to understand that this is a parallel procedure, and unfortunately, not joint advice. the goals of the ema and us fda are primarily to share information and perspectives, rather than specific harmonization of study or regulatory requirements (although they recognize that harmonization is a beneficial outcome). after this procedure, the two agencies conduct their individual regulatory decision-making process regarding drug development issues and marketing applications. each agency provides independent advice to the sponsor regarding questions posed according to their own usual procedures and timelines. the advice of each agency may therefore still differ after the joint discussion. however, in many cases, these discussions between regulators achieved a high degree of alignment and helped industry move closer to a global development plan [ ] . in , following a rather slow acceptance in previous years (due to hesitation from industries to use this procedure that does not commit the two agencies to issue common advice), the ema and the us fda discussed seven new parallel scientific advice procedures. who experts were involved in two of these procedures, due to the therapeutic area covered by the request. in addition to the formal parallel scientific advice exchanges between the us fda and the ema, ad hoc informal scientific advice teleconferences between the agencies took place for five products in [ ]. "clusters" or specific areas of mutual interest have been identified, and a more structured working relationship has been established. these clusters (i.e., oncology, pediatrics, orphan medicines, pharmacogenomics, blood products, biosimilars, and vaccines) facilitate the exchange of information through teleconferences relating to applications for marketing authorization and extensions of indications, including risk management plans [ ] . the latest cluster established, with a focus on biosimilars [ ], significantly increased cooperation between the agencies. the recent announcement from the ema stating that the agency will now accept data from reference product batches sourced outside the eu for biosimilar product applications [ ] will certainly boost the eu-us cooperation in this domain and the global development of biosimilar products. this decision follows the us fda proposal to also accept comparative data referencing a product that is not approved in the us [ ]. the eu-us fda collaboration on orphan drug development has been important. discussions between the ema and the us fda usually include sharing of information on applications submitted in order to approach and discuss criteria for designation. a common application form has been designed and agreed to so that sponsors can apply for orphan designation (of the same medicinal product for the same use) in both jurisdictions using this common form, facilitating the exchange of information. since , discussions have also included analysis of different opinions. on february , , the us fda and the ema announced that they had agreed to accept the submission of a single annual report mmm from sponsors of orphan products designated for both the us and the eu [ ] . each regulatory body continues to conduct their own review of the annual report to assure the information meets their own requirements. the use of one single report benefits both the sponsor and the two regulatory agencies. the sponsors benefit from the elimination of duplication of efforts to develop two separate reports, and the regulators can better identify and share information throughout the development process of an orphan product. collaboration in pediatrics is governed by the principles agreed to in [ ] . this framework includes information exchange (product-specific and general issues) and invitation of the other party to relevant pediatrics meetings. the two main objectives are ( ) to avoid exposing mmm these reports provide information on the status of the development of orphan medical products, including a review and status of ongoing clinical studies, a description of the investigation plan for the coming year, any anticipated or current problems in the process, difficulties in testing, and any potential changes that may impact the product's designation as an orphan product. children to unnecessary trials, and ( ) to facilitate the development of global pediatric development plans that are based on scientific grounds and that are compatible for both agencies. in practice, the cluster on pediatrics organizes monthly teleconferences between the ema's pediatric team and the us fda during which pediatric investigational plans (pips) are discussed in detail and information between the two agencies is exchanged. in addition, more general questions have also been addressed, such as extrapolation, choice of endpoints, and patient/parent reported outcomes. from september until september , products and four general topics were discussed [ ] . since the end of , us fda representatives have been able to participate in certain ema pdco discussions and vice versa. the ema has also provided the us fda access to its internal database that includes scientific details on all pips. several guidelines have been developed at the ich level (ich q , q , q ) in order to facilitate the implementation of "quality by design." taking into account the global perspective of pharmaceutical manufacturing, the ema and us fda agreed that it would be beneficial if at this early stage of implementation assessors from the us and eu could exchange their views on the implementation of ich concepts and relevant regulatory requirements using actual applications. a three-year pilot program, operating under the us-eu confidentiality arrangements, started in april . this program allowed parallel evaluation of "quality by design" aspects of applications submitted to the ema and the us fda at the same time [ - ]. on august , , the ema and us fda published the lessons learned and q&a resulting from the first parallel assessment. both agencies found the pilot program extremely useful to share knowledge, facilitate a consistent implementation of the ich guidelines, and harmonize regulatory decisions to the greatest extent possible [ - ]. the bilateral collaboration between the eu and the us has been extremely productive, and today it is recognized as a very successful initiative. its scope has increased over the years, from the basic exchange of information and harmonization of format to close collaboration and discussion of divergent positions. the liaison placement in each organization has also been an important decision to facilitate such cooperation. this increase in interaction, in a relatively short period of time, has been driven in part by reaction to crises and in part by proactive measures to enhance ema-us fda communication and collaboration [ ] . the establishment of the transatlantic administrative simplification project in has also been beneficial as it initiated several pilot projects that further demonstrated the need for, and benefits of, such collaboration. in general, activities in all the clusters have increased over time, and there has been an overall increase in the number of ad hoc requests for teleconferences on specific products and topics. following a significant increase between and , the total number of monthly us fda and ema interactions (i.e., teleconferences, document exchanges, etc.) now averages about per month, excluding document exchanges relating to cluster and pilot activities. significant achievements have also been made in several critical areas for public health such as orphan medicinal products (with the agreement on a single annual report), drug development (with the establishment of the parallel scientific advice procedure and collaboration on pediatric development), gcp and gmp inspections (with several successful pilot projects that increased collaboration), and safety of products (with close collaboration and regular exchange of safety information, risk management, and safety alerts). exchange of draft regulation (before release in the public domain) has also facilitated harmonization of practices and exchange of opinions. finally, tools for more effective tracking have also been developed. all these achievements confirm that collaborations between countries have a positive impact on public health. it is particularly evident in certain areas such as orphan drug development (for diseases affecting a small population) or the exchange of information relating to urgent drug safety issues (to better assess and understand risks). it is also important to note that this successful collaboration allows not only for the convergence of practices, but more importantly, this exchange of information and communication builds confidence in each other's systems, practices, and evaluations, allowing for a sharing of activities in certain areas. this is already the case in the area of inspection. in december , ema spokesperson monika benstetter stated that "each agency is now relying on its partner for drug manufacturing facility inspection data." [ ] the success of this transatlantic cooperation is partly due to the fact that it has been well structured and organized over the years. the establishment of clusters and then the creation of the liaison officials' positions nnn strengthen regulatory cooperation between the agencies. these decisions have been extremely beneficial from the perspective of education and timely communication. a large number of staff visits and exchanges also took place, and there is now more routine involvement in the scientific work of both agencies. the us fda representatives take part as observers in committee for medicinal products for human use (chmp) discussions, and the ema representatives are provided with access to webcasts of us fda advisory committees. however, other parameters such as those listed below have also been critical for this success, and clearly demonstrate their importance of this type of cooperation and harmonization initiative: ▸ first, it is clear that the political commitment to increased cooperation has been important. indeed, closer collaboration was evident after the signing of the "framework for advancing transatlantic economic integration between the european union and the united states of america" in by ec president josé manuel barroso, german chancellor angela merkel, and us president george w. bush. ▸ second, the establishment of confidentiality agreements, which since are effective for an indefinite period, allow both parties to exchange inspection reports or other nonpublic product-related information. this was critical in the establishment of collaboration as this communication on specific practical cases allowed the parties to nnn since , the fda has seconded a permanent representative to the ema's office in london. since early , the ema has seconded a representative to the fda's offices. discuss the similarities and differences of opinion when assessing product applications and documentation. although necessary, sharing only public information (i.e., new regulations and guidelines) does not provide this opportunity. ▸ third, this bilateral collaboration benefited from the fact that both parties had the same level of maturity and development of their systems and regulations, and similar public health needs and challenges (even if they were not always identical). ▸ lastly, the step-by-step approach established has been helpful because it provided clear priorities (with the clusters), allowed progressive exchange of information (from ad hoc requests to regular teleconference and nonpublic product information exchange), and time for each party to evaluate the partner agency's system and practices (with several specific pilot projects and visits/exchange of staff). although it took some time and a lot of effort, these different steps were beneficial as they facilitated transparency and confidence building. this clear understanding of similarities and differences of practices is a prerequisite to foster a culture of convergence of each agency's assessments and evaluations. to conclude, this bilateral collaboration is now very developed and has moved from confidence building and exchange of information, to recognition of each other's information and data for decision making. its success so far supports the continuation of this collaboration and even its extension, as confidence in each other's system continues to increase. although it is recognized that each party will remain ultimately responsible for public health in their territories, closer cooperation and convergence are obviously possible in many domains. finally, it would be beneficial to continue to expand successful projects to additional partners (as has been the case for gmp inspections of active substance manufacturers [ ] ) in order to foster greater international collaboration and information sharing. in addition to the bilateral, regional, and global regulatory initiatives described in previous sections, other technical and scientific harmonization projects have also been initiated. although these projects do not enter in the scope of this research (as they do not specifically relate to regulatory harmonization), it is important to mention them, as the standards they develop are often used by the regulatory harmonization initiatives. the following organizations and projects ooo have indeed supported the harmonization of standards in the pharmaceutical domain: ▸ the pharmacopoeial discussion group (pdg) involves (since ) the european pharmacopoeia (ep), the japanese pharmacopoeia (jp), and the us pharmacopeia (usp) to harmonize pharmacopoeial standards (i.e., excipient monographs and selected general chapters). it works in collaboration with ich, and who became an observer in may . ooo this list of organizations/projects below is provided as an example and does not represent an exhaustive list. ▸ the international organization for standardization (iso) is the world's largest developer and publisher of international standards (with a network of the national standards institutes of countries and a central secretariat in geneva, switzerland). this is a nongovernmental organization that today has more than , international standards and other types of normative documents covering many technical areas. ▸ the pharmaceutical inspection co-operation scheme (pic/s) facilitates (since ) ppp cooperation and networking in the field of good manufacturing practice (gmp) in order to lead the international development, implementation, and maintenance of harmonized gmp standards and quality systems of inspectorates in the field of medicinal products. the pic/s activities include the development and promotion of harmonized gmp standards and guidance documents, the training of inspectors, and the assessment of inspectorates. this initiative currently includes more than worldwide pharmaceutical inspection authorities. ▸ the council for international organizations of medical sciences (cioms) is an international, nongovernmental, nonprofit organization that was established jointly by who and the united nations educational, scientific and cultural organization (unesco) in . it includes over international, national, and associate member organizations representing many of the biomedical disciplines, national academies of sciences, and medical research councils. one of the objectives of cioms is to facilitate and promote international activities in the field of biomedical sciences, and its activities include programs on drug development and international nomenclature of diseases. ▸ the world medical association (wma) is an international organization founded in to represent physicians. today, it includes national medical associations, and its goal is to achieve consensus on the highest international standards of medical ethics and professional competence. the declaration of helsinki (developed in ) is the wma's best-known policy statement. finally, other groups of experts have also worked and released recommendations on specific topics related to the harmonization of pharmaceutical regulations (e.g., the phrma's [pharmaceutical research and manufacturers of america] simultaneous global development project [ ] or the nonprofit transcelerate biopharma project [ ] ). all these projects contribute to the global convergence and harmonization of pharmaceutical regulations. many harmonization initiatives have been established over the past several decades because regulators understand that cooperation can help in resolving the new challenges brought on by globalization. understanding the importance and advantages of cooperation and ppp the pharmaceutical inspection convention (pic) had been operating since . harmonization in supporting their mandate to promote and protect public health, many countries and regions have strongly enhanced their collaboration with other countries bilaterally and multilaterally at the regional and global levels. the globalization of the pharmaceutical market has highlighted several problems that have been associated with data generated from foreign countries and with imported products. for example, in , deaths associated with heparin imported from china into the us was due to contamination of its pharmaceutical ingredients at a chinese plant, and in panama, the diethylene glycol found in cold and fever medicine killed many people [ ] [ ] [ ] . these problems have been a wake-up call, and they further increased the recognition of benefits to be derived from leveraging the activities and resources of foreign counterpart dras [ ] . for example, the us has strongly increased their international collaboration in the pharmaceutical domain. us legislators decided that such international cooperation and harmonization activities are an integral part of the us fda's mission. indeed, the food and drug administration modernization act of stated that one of the missions of the fda is to "participate through appropriate processes with representatives of other countries to reduce the burden of regulation, harmonize regulatory requirements, and achieve appropriate reciprocal arrangements" [ ] . since then, the us fda's international work has grown exponentially, especially over the past decade, to respond and adapt to the new global society [ ] . it has increased communication qqq and developed regulatory cooperation with other countries (bilaterally and multilaterally). the us fda's role in harmonization and multilateral relations is to coordinate and collaborate on activities with various international organizations (i.e., who, ich, pandrh, and apec) and individual countries on international standards and harmonization of regulatory requirements. in pursuit of appropriate international collaboration, the us fda utilizes a wide variety of international arrangements, including "confidentiality commitments" rrr and "memoranda of understanding and other cooperative arrangements." sss the ema is one of the us fda's closest regulatory partners. with china, uuu the us fda must increase its capacity for inspecting and analyzing chinese products before they are shipped to the us. in order to accomplish this, the us fda established an office in beijing, china in november and employed people (with additional employee hiring planned in the following years [ ] ). it has allowed for solid relationships with chinese regulators and exporters, and has trained more than , manufacturers and regulators on us safety standards in two years [ ] . finally, there has been increasing recognition within the us fda of the need to strengthen regulatory capacity and provide technical and scientific expertise to developing countries to ensure that products exported to the us meet us fda standards and adequate levels of patient protection. many cooperative initiatives have been established to meet this goal [ ] . other countries and regions, including the us, eu, australia, canada, singapore, and china. these bilateral collaborations are based on confidential agreements vvv and include information sharing. proactive exchange of staff has also been agreed upon with some dras ( ). japan's pmda has also developed privileged relationships with china and south korea following the pandemic influenza crisis [ , ] . since , this tripartite initiative has specifically cooperated on clinical research and promoted regional clinical trials [ , ] . in february , the advisory council approved the pmda international strategic plan as a framework for its international activities [ ] . this plan outlined the strategies for bilateral, regional, and global cooperation, and established an internal office in charge of international affairs. in line with this international strategic plan, further goals (to be attained by ) were published in november . finally, a roadmap for the pmda international vision was released in april . in this roadmap, the pmda defines more specific actions to support its international vision . the primary objective of this increase in international collaboration was to urgently resolve the "drug lag" www that has impacted the japanese pharmaceutical market in the past ( . years in ). many measures have been taken to improve the clinical testing environment (including the promotion of global clinical trials) and expedite drug approval decisions (via, among other measures, the increase of collaboration with the other worldwide dras). a global, simultaneous drug development approach has also been strongly recommended. many actions, including release of guidelines, have been taken to facilitate such global development [ ] . in addition to the us and japan, other major dras of developed countries (such as health canada and the australian tga) also recognized the important added value of global cooperation and therefore increased their involvement in international activities. the eu, based on its prior experience of harmonization and cooperation from the establishment of its own system, has also developed external bilateral and multilateral collaborations and is today an important international partner. although these diverse, coexisting, bilateral, regional, and global initiatives create complexity, it is important to note that they are complementary. global harmonization does not preclude having regional harmonization and regional harmonization does not preclude bilateral agreements. in fact these three levels of harmonization and cooperation bring about different added value: ▸ bilateral agreements allow for a bigger exchange of information, including productspecific data, through confidential agreements and the development of privileged relationships (and trust) between regulators as they allow for assessment of one another's vvv in the case of china, a cooperative agreement has been established. www drug lag is defined as the difference of availability of new medicines between the us and japan. systems and practices. xxx these assessments are indeed critical for confidence building and can ultimately support the signing of agreements, allowing for recognition of inspection or the exchange of nonpublic information (e.g., eu/us collaboration and confidentiality agreements). bilateral collaboration also helps strengthen relationships, which would be more difficult in the context of a multilateral initiative, and facilitates training and mentoring activities between developed and developing countries. ▸ regional harmonization allows for the harmonization of policies between countries that are usually closer in term of systems, cultures, and levels of development. it is indeed easier to harmonize closed systems and policies between countries of similar culture and environment (for example, it is more difficult to harmonize systems and policies between asia and north america because they have very different medical practices and cultures). this level is essential for global harmonization because it provides a structure. achieving global harmonization without a supporting regional organization structure is impossible. this regional level allows for inclusion of regional realities and difficulties in global discussions, and eases the diffusion and implementation of the global recommendations. ▸ global harmonization is the highest level of harmonization. compared to regional harmonization, the global harmonization initiative is not driven by economic objectives; the goal is not to create a free trade area or a single market, but to develop global consensus and standards in order to allow the world's population to have access to medicine and innovative therapies. to conclude, these bilateral, regional, and global cooperative activities have been beneficial as they supported the harmonization of requirements globally and therefore facilitated the availability of safe and efficacious medicines, critical in promoting global public health, on a worldwide basis. many topics and standards have already been partly or fully harmonized at a bilateral, regional, or global level. for example, most of the requirements regarding the conduct of nonclinical studies, and also the gmp and good clinical practice (gcp) principles, have been agreed on, allowing for joint inspection projects. a common format of application has been developed, and many technical aspects have been harmonized through the ich's work. collaboration has also been increasing in resolving major topics requiring global interaction, such as orphan drug evaluation yyy and development of medicines for the pediatric population. zzz confidence and trust have been built between developed countries through pilot projects, but xxx for example, bilateral collaboration allows two countries to assess their respective inspection systems or systems to control critical information (such as trade secrets). such assessments of each other systems could be possible in the case of multilateral collaboration, but would be more complex. yyy because only a small number of the population is affected by these life-threatening diseases or serious conditions, it is critical to have global requirements in order to facilitate global clinical studies. moreover, the pharmaceutical industry has been reluctant to invest in the research and development of medicinal products to treat these conditions. the development of global requirements allows quick access to the global market and therefore allows a better return on investment. zzz it is critical that countries cooperate in this area to avoid exposing children to unnecessary trials. also through the location of official liaisons in other dras to facilitate collaboration. this has been positive, and this new type of interaction is very promising as it increases relationships and allows for the better exchange of experiences and information. aaaa the establishment of liaisons in other countries also allows more proactive measures and risk analysis in the area of quality systems and inspections [ ] . exchanges of information between dras have also dramatically increased. this regular communication between regulators facilitates evaluation of risk (e.g., via exchange of safety alerts) and assessment of new medicines. finally, systems have been put in place to help developing countries (e.g., cpp scheme, prequalification of medicines, article of european regulation (ec) no / , etc.). however, without underestimating all these important positive outcomes, it is clear that differences still exist and that further efforts will be required to support this ongoing harmonization process. there are still differences between countries in terms of standards and strategies to assess compliance against standards. the conduct of global clinical studies continues to present many challenges (i.e., related to registration, conduct of the studies, and also the use of data), and there are still several clinical trial registries and databases in use. the safety of medicines has been one of the main focuses of dras in the past due to major problems and events, but there has not been a real effort toward worldwide harmonization regarding risk-mitigation strategies. additionally, new standards continue to be developed by different bodies (i.e., ich vs. regional organizations) in parallel that not only duplicate efforts, but also create disharmony (e.g., biosimilars requirements had first been developed by individual countries and also by who). there is also a significant difference in the level of implementation of harmonized standards (i.e., the ich recommendations/guidelines) between countries, and the ctd format has still not been implemented in all countries. it has also been reported that differences between developed and developing countries has in fact continued to increase in the past several years due to the increased complexity of technologies associated with the development of new therapies. even between two close partners like the us and eu, which have developed a privileged partnership and strong cooperation, there are still important differences in standards. for example, the us is still requiring two placebo-controlled studies to determine efficacy of a new medicine, while the eu is more interested in comparative studies using an active comparator. this difference is due to different legal requirements and scientific opinions regarding the value of such comparative data [ ] . this situation may change in the future with the growing interest in the us for "comparative effectiveness" promoted by the obama administration. finally, this complex worldwide harmonization context (with increased communication and exchange of experience, information, and good practices) requires good communication and coordination between all these ongoing initiatives. even if such communication was initiated by who and ich (with the gcg group), further improvement would still be needed. this enhanced coordination of international cooperation would indeed be beneficial, as it would provide the necessary transparency regarding the focus and responsibility of each initiative (i.e., development of standards, coordination of implementation of recommendations, etc.). aaaa exchange of information and best practices has been one of the most important outcomes of the eu/us bilateral collaboration. it would also facilitate the appropriate use of resources and expertise, and therefore avoid duplication of efforts or conflicting recommendations and actions. overlapping membership between the initiatives bbbb may not be fully efficient, and can create confusion and duplication of work. although the increased coordination of these diverse initiatives would be beneficial, it will certainly be challenging. it will need to be thoroughly structured and implemented, and it will also be critical that the coordinated body is a recognized and experienced entity, with appropriate mandate and power. under this directorate, the us fda's office of international programs serves as the agency's focal point for all international matters and is responsible for maximizing the impact of the us fda's global interactions. additional us fda reorganizations were also announced in to further respond to drug industry globalization [ ]. also, in addition to china, the us fda now has staff stationed permanently in india the total number of shipments of us fda-regulated products from china increased from approximately . million to . million. of the . million entry lines arriving in , % were drugs and devices, and % were human food products key: cord- -xt aa ur authors: schilling, stefan; fusco, francesco maria; de iaco, giuseppina; bannister, barbara; maltezou, helena c.; carson, gail; gottschalk, rene; brodt, hans-reinhard; brouqui, philippe; puro, vincenzo; ippolito, giuseppe title: isolation facilities for highly infectious diseases in europe – a cross-sectional analysis in countries date: - - journal: plos one doi: . /journal.pone. sha: doc_id: cord_uid: xt aa ur background: highly infectious diseases (hids) are (i) easily transmissible form person to person; (ii) cause a life-threatening illness with no or few treatment options; and (iii) pose a threat for both personnel and the public. hence, even suspected hid cases should be managed in specialised facilities minimizing infection risks but allowing state-of-the-art critical care. consensus statements on the operational management of isolation facilities have been published recently. the study presented was set up to compare the operational management, resources, and technical equipment among european isolation facilities. due to differences in geography, population density, and national response plans it was hypothesized that adherence to recommendations will vary. methods and findings: until mid of the european network for highly infectious diseases conducted a cross-sectional analysis of isolation facilities in europe, recruiting isolation facilities in countries. three checklists were disseminated, assessing items and specific questions. the median feedback rate for specific questions was . % (n = / ) (range: n = / ( . %) to n = / ( %). although all facilities enrolled were nominated specialised facilities' serving countries or regions, their design, equipment and personnel management varied. eighteen facilities fulfilled the definition of a high level isolation unit'. in contrast, facilities could not operate independently from their co-located hospital, and five could not ensure access to equipment essential for infection control. data presented are not representative for the eu in general, as only / ( . %) of all member states agreed to participate. another limitation of this study is the time elapsed between data collection and publication; e.g. in germany one additional facility opened in the meantime. conclusion: there are disparities both within and between european countries regarding the design and equipment of isolation facilities. with regard to the international health regulations, terminology, capacities and equipment should be standardised. the term highly infectious diseases (hid) defines mostly viral and bacterial infections that (i) are easily transmissible from person to person; (ii) cause a life-threatening clinical illness with no or few treatment options; and (iii) pose a threat for both health care workers and the public, thus requiring specific infection control measures and public health planning [ ] . included in this definition are viral haemorrhagic fevers (e.g. filo-, arena-or bunyavirus infections); some respiratory syndromes (e.g. severe acute respiratory syndrome (sars)-coronavirus, or pneumonic plague); as well as any (re ) emerging infectious agent transmissible from person-to-person [ ] . in europe, hids have caused several events within the last decade: sars affected eight european nations in ; lassa virus has repeatedly been imported to europe; and crimean congo haemorrhagic fever virus infections are increasing in several mediterranean regions, and have also been imported to central europe and the united kingdom (uk) [ ] [ ] [ ] [ ] [ ] . recently detected agents like the new human middle east respiratory syndrome coronavirus underline the continuous challenge faced [ ] . patients with suspected or proven hids should be cared for in a clinical environment that provides safe, secure, high-quality, and appropriate care with optimal infection containment, prevention and control procedures [ ] . consensus statements on the operational management and design of isolation facilities have been published in europe and the united states of america [ ; ] . in addition, the european commission has funded projects to enhance early recognition of cases by training front-line health care workers (hcws) and standardizing diagnostic methodology [ ] [ ] [ ] . despite such efforts, no pooled data on isolation facilities resources, such as infrastructure design, technical equipment, capacity and access to intensive care, do exist. the study presented was performed by the european network for highly infectious diseases, euronhid, and set up to compare the operational management, resources, and technical equipment among isolation facilities with recommendations published. due to differences in geography, population density, national response plans, and experience with hids in participating countries, it was hypothesized that the level of adherence to recommendations may vary. the objective of this article is to present data about infrastructure design and resources, technical equipment, available personnel, and access to intensive care. euronhid consists of infectious disease clinicians and public health officers with expertise in the management of hids identified via their national health authorities. administrative and scientific aspects are managed by an italian coordination team, and a steering committee, including partners from france, germany, greece, and the uk. both committees started their work in , and reported all organizational and scientific developments to the project members. names and countries of all project members are listed above.\ three checklists were developed and based on projectmember's experience, available literature, national preparedness plans, and guidelines of international authorities for the management of hids [ ] . each checklist contained questions addressing specific items of interest, and answers were structured as openended, closed, semi-open or free text options. overall, the checklists included items and specific questions. checklist one contained three major sections: infrastructure; technical equipment; and personnel management. a fourth section on emergency departments was not obligatory [ ] . all checklists can be downloaded after free registration from: www.eunid.eu. the checklists underwent a pilot application to identify structural gaps and sources of misinterpretation [ ] . the facilities of the steering committee members and one external facility applied the checklists and cross-checked all itmes for their applicability, but no external validation process was conducted. following minor changes, the checklists were disseminated to all eligible facilities until december and expected to be resubmitted within three months. data provided were cross-checked by personal on-site visits by a coordination team member in all but four centres enrolled. investigator (the national project representative) and observer (the coordination team member) bias may be ruled out, as all data recorded were cross-checked on-site and agreed with the facilities operational manager or director. until november , the coordination team collected data on eligible isolation facilities to be included into the survey. national health authorities of all european union (eu) member states were contacted to identify their will to participate. participating national health authorities were requested to identify clinical facilities responsible for the admission, assessment, and care of hid patients in their country, and to nominate a national project representative. euronhid was co-funded by the european commission/ director general for consumers and health (dg sanco) for an initial period of months and extended to months without additional funding. a technical and scientific report on all data collected during the project was delivered to the commission in december . for details please follow: http://ec.europa.eu/ research/health/infectious-diseases/emerging-epidemics/projects/ _en.html. until mid of , isolation facilities in european countries were recruited into this survey (table ). recruitment was closed by the end of with the exception of norway, who received questionnaires by end of and submitted data by spring of . data collection and on-site visits were finalised in spring . except for norway, all participating countries were eu member states. data were collected from all existing facilities in participating countries except for spain, where data were only collected in the catalonia region. overall, the median feedback rate for checklist was . % (n = / ), with a range from n = / ( . %) to n = / ( %) of centres providing valid answers to specific questions. all data provided in this article represent a minimum feedback from facilities (. %). the majority of facilities were constructed or underwent major re-construction in the year or later (n = / ; . %) and are located on the same campus or in direct connection with a general hospital (n = / ; . %). most facilities have designated rooms (n = / ; . %) or wards (n = / ; . %) located within other hospital structures, and seven facilities are located within a separate building (n = / ; . %). national guidelines for the construction of isolation facilities were available for facilities whereas were (re ) constructed in the absence of specific requirements ( % and %, respectively). three facilities accept paediatric ( . %), and ( . %) accept adult patients, only, whereas most facilities provide care for both (n = / ; . %). the number of isolation beds per facility ranges from one to , with a median of . beds for adult and . for paediatric patients. with respect to the overall population, the number of beds ranges from . to . per one million inhabitants (median: . ) ( table ). the majority of all facilities uses their beds on a daily basis for routine patients (n = / ; , %), while the remainder provide bed capacities reserved for hid cases, only (n = / , . %). except for the united kingdom, all countries (n = / ; . %) use barrier nursing techniques with high level personal protective equipment (hl-ppe) ( ) . facilities (n = / ; . %) have a response time of less than five hours after a case is notified, the remainder demand up to hours to be fully operational (n = / ; . %). only eight ( . %) of all facilities evaluated lacked any experience in the management of suspected or proven hid cases. among the remainder, most had experience with infections due to sars (n = / ; . %), and viral haemorrhagic fevers (n = / ; . %). essential technical requirements for isolation facilities includes: negative pressure in the isolation room(s); an anteroom; aerosoltight doors and windows; high efficacy particulate air (hepa) filtration of exhausted air; and a surface of walls, floors and ceiling withstanding disinfection procedures [ ] . all but five facilities enrolled (n = / ; . %) provide negative pressure in the isolation area, and the majority (n = / ; . %) is also equipped with all other essential equipment investigated. in facilities with two or less additional items, most often hepa filtration of exhausted air and anterooms were missing (n = and / facilities, respectively) ( figure ). additional technical requirements not considered essential, but benefitting infection control and hcw safety, includes: self-closing doors; audio and/or visual negative pressure indicators; an exclusive evacuation pathway; and an internal communication system (for hcw-hcw and hcw-patient communication) [ ] . at least additional technical requirements are found in the majority of facilities (n = / ; . %). most often, an internal communication system (n = / ; . %) and negative pressure indicators (n = / ; . %) were present, while an exclusive evacuation pathway was missing in half of all facilities enrolled (n = / , . %). thirty-three facilities ( . %) provide intensive care, available for patients within the isolation facility. twelve facilities ( %) rely on a designated isolation room within standard intensive care units, three ( . %) have no access to intensive care capacities ( figure ). equipment for the monitoring of vital parameters and advanced life support is available in all facilities. in contrast, mechanical ventilators are available either permanently in twelve ( %) or on call in thirty-two facilities ( . %), but not accessible at all in four ( . %). forty-two facilities ( . %) have either permanent or on-call access to blood gas analyzers, whereas six have not ( . %). hence, out of forty-five facilities reporting intensive care provision, one ( . %) lacks a mechanical ventilator, and three ( . %) have no ability to monitor their ventilation therapy. almost all facilities ( . %; n = / ) report permanent access to specifically trained infectious diseases doctors. / ( . %) facilities report providing intensive care, but only of those ( . %) have access to specifically trained intensive care specialists. in addition, out of facilities indicating to provide care for pediatric hid patients, only ( . %) report either permanent or on-call access to pediatricians. compared with available doctors, even fewer facilities report access to specifically trained infectious diseases nurses (n = / ; . %), but intensive care nurses are available in a comparable number of facilities (n = / ; . %) (figure ). all but two facilities indicate -hour access to specifically trained medical and non-medical staff (n = / ; . %), although . % (n = / ) only have specific protocols to contact staff responsible for the operation of the facility. a shift plan to limit the number of hcws exposed to patients as well as the duration of working under hl-ppe exists in . % of facilities (n = / ). permanent access to technicians was lacking in eleven facilities (n = / ; . %). all participating facilities were evaluated for their adherence to recommendations for the operational management of isolation facilities published [ ] . as hypothesized, the level of adherence to recommendations varied, both within and between participating countries. hence, three categories of isolation facilities representing different levels of adherence were defined: (i) high level isolation units (hliu), defined as operating independently from other hospital resources and specifically equipped; (ii) isolation rooms, defined as specifically equipped but only partially independent from other hospital resources; and (iii) referral centers, neither specifically equipped nor functionally independent. according to this classification, all facilities enrolled were categorized as shown in figure . few facilities completely fulfilled recommendations for hlius (n = / ; . %). six of those are located in germany, five in france, two in italy, and one each in the uk, finland, greece, ireland, and norway. such hlius are constructed and equipped for the admission, assessment and longterm critical care of hid patients. most facilities assessed did partially meet the recommendations, but lacked infrastructural components, or preconditions for personnel management (n = / ; %). such were labeled 'isolation rooms' considered effective to assess hid patients and provide short-term medical care until a patient can be transferred to a hliu. of these, five each are located in france, greece, and spain; two in germany; and one each in denmark, finland, ireland, luxembourg, malta, poland and slovenia. the remaining facilities (n = / ; . %) are considered to function as 'referral centers' where suspected hid cases can be assessed within daily routine, but even short-term critical care cannot be provided without an increased risk of nosocomial infections. two of such facilities are located in bulgaria and france, each, and one in austria. within the last decades, autochthonous outbreaks or imported cases of hids have affected europe with significant impact, and new pathogens are emerging. most european countries have established national response plans including specialised clinical care facilities for the management of such scenarios [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . this article presents the first standardised analysis on the operational management, infrastructure, and technical equipment of isolation facilities in european countries. data provided may support national authorities to assess their level of preparedness but have to be adapted to single member states' needs and resources. with regard to the implementation of the international health regulations [ ] , terminology, capacities and equipment accessible in isolation facility should be documented and standardised on a european level. although all eu member states were invited to join the project by dgsanco, only agreed to participate, thus data presented are not representative for the eu in general. a major limitation of this study is the time elapsed between data collection and publication: e.g. in germany one another facility has opened in the meantime. hence, a periodical re-assessment of facilities and countries should be achieved, aiming for a permanently updated record of national preparedness. in addition, the checklists applied did not undergo an external validation process (e.g. a delphi cycle) due to limitations of funds and project duration, and present published guidelines, only. hence, within the data collection process, misinterpretations occurred and items needed to be excluded from the data analysis presented here. half of all countries assessed have at least one hliu according to the recommendations [ ] . compared to the definition, facilities fulfilled the criteria applied, but the majority of all facilities enrolled could not operate independently from other facilities. the high rate of incompliance with the recommendations depicted here can be explained by the absence of legally binding documents by the time most facilities were constructed. after sars and the us anthrax attacks in , the political will was to enhance public preparedness, but no standard was agreed on. only few countries (e.g. the uk) had specific guidelines at hand, but the vast majority of facilities were constructed using laboratory-or operational theatre guidelines adapted to local needs and infrastructure prerequisites. an earlier inventory of isolation rooms in europe by our group faced this problem, when eu countries were asked to report on negative pressure rooms for hid patients [ ] : the overall number of rooms exceeded most expectations, but both technical and infrastructural conditions were not well defined. hence, an overreporting of any isolation room' with available led to the impression of a well prepared health system, although other aspects of equipment and personnel were not addressed. given the definitions for different levels of isolation facilities proposed now, a reassessment of the eu's capacities could lead to more accurate data. furthermore, the allocation of funds for hospital's biological event preparedness has decreased since the sars pandemic, and is not harmonised within the eu. hence, as our data show, the level of adherence to the recommendations mirrors the economical situation in the eu, with the highest level of adherence in the economical most stable countries. the situation in the united states of america is comparable: a consensus on the operational management of isolation units consistent with the european approach exists, but only three facilities fulfill those criteria, although hospital preparedness for biological events is mandatory [ , ] . the contrary can be found in israel, where the capability to manage hid patients is mandatory for every public hospital, and dedicated hlius are absent [ ] . as hlius are designed for single imported cases or small clusters in the beginning of an outbreak, only, they do not compensate for surge capacity planning in major biological events [ ] . besides infrastructure and equipment, additional features which support effective functioning of such specialised centres were addressed in this survey and recently published [ ] [ ] [ ] [ ] . access to a transportation system for hid patients plays an important role in distribution of isolation facilities and is reflected by the overall amount isolation facilities per country. balancing the necessary access to a focus of expertise with available transport logistics leads to either a centralised or a de-centralised approach to the location of specific facilities, and ambulance dispatch recording can be of use in early warning systems for outbreak detection [ ] . most documented hid cases demanded supportive intensive care [ , , ] , and access to such is considered essential for hlius [ ] . in order to allocate funds most effectively, a balance between infection control excellence and operational feasibility should be sought. basic specifications for hliu provision, including access to intensive care equipment and trained personnel, should be developed in order to facilitate a common standard of best clinical practice. key issues are the composition of a permanently available multidisciplinary medical and nonmedical team, the ability and physical fitness to work with ppe, and reliable timelines and shifts to prevent accidents or mistakes due to exhaustion [ , ] . data depicted in this article do highlight available personnel as the most crucial pitfall in operating isolation facilities: compared to the complicane with technical equipment recommendations, the lack of specifically trained staff is surprising, especially for hcws trained in intensive and paediatric care. it should be mentioned that all facilities with experience in proven hids provide more sophisticated human resources management such as shift-and surge capacity planning as an outcome of their experience. the overall investment for an isolation facility, and hlius in particular, is high, in terms of costs for construction and maintenance, medical and infection control equipment, human resources and training activities. even in time of economical constraints, employers ''are obliged to ensure the health and safety of workers'', as defined by the european commission [ ] . without adequate protection, hcws may acquire and transmit infections, with potential dramatic impact on the health system, and the economic solidity of a country, as documented during the sars epidemic in canada. for these reasons, we believe that specialized isolation facilities, and hlius in particular, still represent a critical infrastructure to which each eu member state should have access, within the country or through predefined agreements with neighbouring countries where such facilities are available. whether such capacities should be reserved for hid cases, only, or also used on routine basis, is part of an ongoing discussion. financial constraints within hospitals and the host country may benefit from a routine-use concept, although such facilities demand more frequent maintenence due to damage on technical equipment. in addition, access to routine use beds for training of hcws is reduced, and patients need to be relocated to other rooms once the facility receives an hid case. in contrast, reserved bed capacities demand additional personnel and material for routine functional checks, thus considered cost-ineffective. the major benefit from reserved capacities is a full access for training and no delay in patients admission since no evacuation of others is needed. the who ihrs define the need of preparedness for infectious diseases outbreaks, but a european, if not international, consensus on the funding and minimum number of isolation capacities is not in sight. although hids are rare events, they cause dynamic and often rapidly evolving issues in need of comprehensive solutions and may challenge the capacity of healthcare systems. leadership and funding at both national and european level are required to harmonize preparedness plans, terminology and communication to weaken the impact of future infectious disease outbreaks with cross-border potential. in order to achieve a balance between saving lives and protecting hcws in hazardous environments, national and international collaboration should continue to share experiences, and provide standardized training and equipment. the 'european network for highly infectious diseases -euronhid' is co-funded by the european commission/dg sanco under the eu contract number . neither the commission nor dg sanco had an influence on the study design, data collection, or interpretation of results. other members of the euronhid have supported the authors heli siikamaki (finland), christian perronne (france), all members of the ''stä ndige arbeitsgemeinschaft der kompetenz-und behandlungszentren -stakob framework for the design and operation of high level isolation units: consensus of the european network of infectious diseases infection control in the management of highly pathogenic infectious diseases: consensus of the european network of infectious disease severe acute respiratory syndrome: identification of the etiological agent a fatal case of lassa fever in london the first case of lassa fever imported from mali to the united kingdom first confirmed case of crimean-congo haemorrhagic fever in the uk crimean-congo haemorrhagic fever isolation of a novel coronavirus from a man with pneumonia in saudi arabia designing a biocontainment unit to care for patients with serious communicable diseases: a consensus statement a curriculum for training healthcare workers in the management of highly infectious diseases the european network of biosafety-level- laboratories: enhancing european preparedness for new health threats community research and development service european training and research centre for imported and highly contagious diseases euronhid checklists for the assessment of highlevel isolation units and referral centres for highly infectious diseases: results from the pilot phase of a european survey infection control management of patients with suspected highly infectious diseases in emergency departments: data from a survey in facilities in european countries acute liver failure, multiorgan failure, cerebral oedema, and activation of proangiogenic and antiangiogenic factors in a case of marburg haemorrhagic fever novel coronavirus associated with severe respiratory disease: case definition and public health measures implementing the international health regulations ( ) in europe isolation rooms for highly infectious diseases: an inventory of capabilities in european countries department of health and human services, health resources and services administration world health organization, regional office for europe ( ) assessment of heath-system crisis preparedness: israel. world health organisation personal protective equipment management and policies: european network for highly infectious diseases data from isolation facilities in european countries infection control practices in facilities for highly infectious diseases across transportation capacity for patients with highly infectious diseases in europe: a survey in nations diagnostic issues and capabilities in isolation facilities in european countries: data from the euronhid surveys surveillance of ambulance dispatch data as a tool for early warning on behalf of the ehec-hus consortium ( ) validation of treatment strategies for enterohaemorrhagic escherichia coli o :h induced haemolytic uraemic syndrome: case-control study on the introduction of measures to encourage improvements in the safety and health of workers at work key: cord- - pp l j authors: li, yan; liang, chao; ma, feng; wang, jiqian title: the role of the idemv in predicting european stock market volatility during the covid- pandemic date: - - journal: financ res lett doi: . /j.frl. . sha: doc_id: cord_uid: pp l j the main purpose of this paper is to investigate whether the infectious disease emv tracker (idemv) proposed by baker et al. ( ) has additional predictive ability for european stock market volatility during the covid- pandemic. the three european stock markets we consider are france, uk and germany. our investigation is based on the har and its augmented models. we find that the idemv has stronger predictive power for the france and uk stock markets volatilities during the global pandemic, and the vix has also superior predictive ability for the three european stock markets during this period. the world health organization (who) officially announced the outbreak of coronavirus as a global pandemic on march , . its outstanding characteristics are manifested as unknown etiology, no targeted drugs, and lack of sufficient experience reserves for treatment methods. all countries are responding to this epidemic in groping. it has caused considerable losses to the global economy. therefore, how to deal with the impact of the epidemic on the global economy is a hot topic that has been concerned recently (see, e.g., ashraf, ; corbet et al., ; goodell, ; sharif et al., ; wagner, ; zhang et al., a) . the modeling and forecasting of stock market volatility has always been a hotspot and difficulty in academic research (see, e.g., wei et al., ; wen et al., ; hong and lee, ; ma et al., ; liang et al., a; liang et al., b; zhang et al., b) . at the same time, it is very important for risk management and option pricing in practical applications. baker et al. ( ) design the infectious disease emv tracker to study the us stock market volatility during the global pandemic. in addition, an influential study of buncic and gisler ( ) shows that us stock market information has a superior predictive ability for the volatility of international stock markets. our motivation comes from this. the main purpose of this paper is to explore whether the idemv has additional predictive ability for european stock market realized volatility (rv) during the global pandemic. the three european stock indices we consider are the cac (fchi), the ftse (ftse), the dax (gdaxi). we use the har model as the baseline model, which is consistent with buncic and for more detailed introduction, please refer to baker et al. ( ). gisler ( . in addition to the har extension models used by buncic and gisler ( ) , we also consider two competitive models (i.e., har-usrv-idemv and har-all) to examine the predictive ability of idemv for the three european stock markets. the out-of-sample results suggest that the idemv contains useful information in predicting the rvs of the fchi and ftse indices during the global pandemic, while ineffective for german stock market. furthermore, as supposed to the whole out-of-sample periods, the vix has stronger predictive ability for the three european stock indices during the covid- , implying that the information in the us stock market still has a leading position during this period. finally, we check the model's predicted performance each month during the global pandemic, and find that the results are robust. this research is closely related to buncic and gisler ( ) and baker et al. ( ) . our contribution and the biggest difference are as follows. first, we examine the impact of u.s. stock market information on european stock market volatility during the global pandemic . second, we use the idemv to predict the rvs of the european stock markets and observe that the idemv has superior predictive power for the fchi and ftst indices during the global pandemic. third, the vix is more predictive for the three european stock indices during the global pandemic. thus, our study complements the research on european stock market volatility during the global pandemic, and is of some help to market participants and policy makers. the remainder of the paper is organized as follows. section provides econometric models and data. we present the out-of-sample assessment results in section . finally, section concludes. methodology and data the focus of this study is european stock markets realized volatility predictions. the related theory of the realized variance (volatility) can be found in the original studies of andersen and bollerslev ( ) and andersen et al. ( ) . the definition of rv is the summation of the intraday squared returns, which is given by where p t,k denotes the kth intraday price, r t,k denotes the kth intraday return, and f is the total number of fixed frequencies during the trading day. we employ the standard har-rv model of corsi ( ) as our benchmark model, the most important features of the har-rv model are that it effectively captures the characteristics of volatility and it is easy to implement and can be estimated with ols. the har-rv model can be written as where rv t , rv w , t , and rv m , t , represent daily, weekly, and monthly rv, respectively. following buncic and gisler ( ) , we add the us stock market information to the baseline model, that is, har-usrv-vix model, which is given by where , , , and , indicate daily, weekly, and monthly us stock rv, respectively. and , and , represent weekly and monthly vix. to investigate the role of the idemv, we replace the har components of vix in the har-usrv-vix model with the har components of idemv. thus, we obtain har-usrv-idemv model, which is expressed as where , and , represent weekly and monthly idemv. in addition, we consider both vix and idemv, namely, har-all model, which is written as, in this study, we employ daily realized variance to measure european stock markets volatilities. we use the -minute sampling frequency to compute the rv, because the -minute sampling frequency is widely used in volatility-related research and is superior to other sampling frequencies lin, , ; mei et al., we turn all international realized variances into annualised realized volatilities. in this section, we only report the out-of-sample predicted performance, because investors and policy makers are most concerned about the out-of-sample results. we use the rolling window method to generate the out-of-sample predictions, and the rolling window length is . first, to quantitatively evaluate the forecasting accuracy, we employ two robust loss functions of qlike and mse, which are widely used in various volatility prediction studies. the definition of the two loss criteria are where t rv is actual rv on trading day t, t rv represents the rv forecasts generated by prediction models, m and q denote the length of in-sample estimation period and out-of-sample evaluation period, respectively. second, we utilize the model confidence set (mcs) test proposed by hansen et al. ( ) to assess the out-of-sample results and determine whether the prediction models used have statistically significant differences in out-of-sample forecasting performance. a mcs is a subset of models that contains the best model with a given level of confidence. the significance level of mcs we choose is %. evidently, the larger the mcs p-value, the better the prediction ability of the corresponding model. in addition to the mcs test, we also employ out-of-sample r ( ) to assess prediction quality, which is defined as the in-sample estimation results are available upon request. for more detailed introduction about mcs technology, please refer to hansen et al. ( ) . are, respectively, the actual rv, forecast rv, and benchmark rv on day m+k, and m and q represent the lengths of the initial in-sample period and out-of-sample period, respectively. obviously, if the value of is greater than , the forecast from the model of interest is better than the benchmark model. insert table about here table shows the predicted performance for the fchi in january, february and march . obviously, the har-usrv-idemv model can produce significantly positive values in january, february and march , which are . %, . %, and . %. in addition, we find that the har-all model has the best predictive ability during covid- pandemic. especially in march , the har-all model can significantly beat other competing models. table presents the predicted performance for the ftse in january, february and march . it is evident that during the covid- the idemv contains useful information and the har-all model has the best predictive ability. from the results of table , we observe that the har-usrv-idemv model can pass the mcs test and produce a significantly positive value of . % only in january , however, the har-usrv-vix model is always the best prediction model. thus, our results are robust. insert table , table and table obviously, if the value of cumsfe is less than , implying that the benchmark model performs poor predictive ability. figure , figure , and figure show the cumsfe for the fchi, ftse, and gdaxi, respectively. we find the cumsfe values are negative for the fchi and ftse indices. however, for the gdaxi, we observe that the cumsfe value is negative only between har-usrv-vix and har-rv. therefore, our results are robust to the cumsfe. insert figure , figure and figure we declare that there is not conflict of interest. notes: the significance level of mcs we choose is %. *, **, and *** indicate significant at the %, %, and % levels, respectively. the following table is also consistent. answering the skeptics: yes, standard volatility models do provide accurate forecasts modeling and forecasting realized volatility stock markets' reaction to covid- : cases or fatalities? the unprecedented stock market impact of covid- global equity market volatility spillovers: a broader role for the united states the contagion effects of the covid- pandemic: evidence from gold and cryptocurrencies a simple approximate long-memory model of realized volatility forecasting the good and bad uncertainties of crude oil prices using a har framework the incremental information content of investor fear gauge for volatility forecasting in the crude oil futures market covid- and finance: agendas for future research the model confidence set a general approach to testing volatility models in time series which sentiment index is more informative to forecast stock market volatility? evidence from china is implied volatility more informative for forecasting realized volatility: an international perspective harnessing jump component for crude oil volatility forecasting in the presence of extreme shocks does us economic policy uncertainty matter for european stock markets volatility covid- pandemic, oil prices, stock market, geopolitical risk and policy uncertainty nexus in the us economy: fresh evidence from the wavelet-based approach what the stock market tells us about the post-covid- world forecasting crude oil market volatility: further evidence using garch-class models forecasting the volatility of crude oil futures using har-type models with structural breaks financial markets under the global pandemic of covid- forecasting global equity market volatilities key: cord- - p frlk authors: donell, simon t.; thaler, martin; budhiparama, nicolaas c.; buttaro, martin a.; chen, antonia f.; diaz-ledezma, claudio; gomberg, bruce; hirschmann, michael t.; karachalios, theofilos; karpukhin, alexey; sandiford, nemandra amir; shao, hongyi; tandogan, reha; violante, bruno; zagra, luigi; kort, nanne p. title: preparation for the next covid- wave: the european hip society and european knee associates recommendations date: - - journal: knee surg sports traumatol arthrosc doi: . /s - - -z sha: doc_id: cord_uid: p frlk purpose: to plan for the continuance of elective hip and knee arthroplasty during a resurgence or new wave of covid- infections. method: a systematic review was conducted using the terms “covid- ” or “sars-cov- ” and “second wave”. no relevant citations were found to inform on recommendations the plan. therefore, an expert panel of the european hip society and the european knee associates was formed to provide the recommendations. results: overall, the recommendations consider three phases; review of the first wave, preparation for the next wave, and during the next wave. international and national policies will drive most of the management. the recommendations focus on the preparation phase and, in particular, the actions that the individual surgeon needs to undertake to continue with, and practice, elective arthroplasty during the next wave, as well as planning their personal and their family’s lives. the recommendations expect rigorous data collection during the next wave, so that a cycle of continuous improvement is created to take account of any future waves. conclusions: the recommendations for planning to continue elective hip and knee arthroplasty during a new phase of the sars-cov- pandemic provide a framework to reduce the risk of a complete shutdown of elective surgery. this involves engaging with hospital managers and other specialities in the planning process. individuals have responsibilities to themselves, their colleagues, and their families, beyond the actual delivery of elective arthroplasty. these recommendations follow those from the european hip society and european knee associates recommendations for resuming elective hip and knee arthroplasty in the setting of the covid- pandemic [ ] . the aim is to help orthopaedic surgeons across europe (and a wider global audience) with a special interest in elective hip and knee arthroplasty prepare for the resurgence or new outbreak of covid- in their locality. this involves planning resources and mitigation procedures that minimise the negative effects of the necessary restrictions imposed on their patients, their families and themselves. the use of the term "wave" was first used in the - respiratory virus epidemic in russia, which had two phases [ ] . the term is inaccurate, since waves are preceded by troughs and are rhythmic in nature. the theory of second waves is based on the - "spanish flu" pandemic, which may have started in a us army camp in kansas, a british camp in etaples in france, or possibly german concentration camps [ ] . however, the data from this outbreak are not robust; it is not even certain that the cause was an influenza virus, nor the role of bacterial superinfections. influenza epidemics tend to be seasonal in nature. it is not known what pattern the covid- will take, and whether there will be a second wave [ ] . having said that, a "wave" follows the pattern of an increase and then a decrease in numbers affected. in a pandemic, numbers of deaths lag behind the number of infected people. the wave is completed when the mortality rate returns to the background rate. the disease activity is reported using the rt value [ ] . r t is defined as the average number of secondary cases generated by one primary case with symptom onset on day t. if r t > , the epidemic is expanding at time t, whereas r t < indicates that the epidemic size is shrinking at time t. complex mathematical local models have been developed to estimate the effectiveness of testing and tracing strategies to avoid a potential second wave of the covid- epidemic [ ] . close monitoring of the rt value is essential in predicting the risk of a resurgence or second wave of a pandemic. this requires adequate, timely, and transparent reporting of the positive patients by the local health authorities. sustained elevation of rt value above implies an increase in cases and should be considered as a marker for a resurgence of virus. the management of the covid- pandemic in the first phase, including the treatment of the patients affected, has proved very demanding. in response, health systems in most countries stopped undertaking elective surgery (both in the state and private sectors) to free up resources for managing the sickest patients [ , , , , ] . the pandemic has dramatically affected arthroplasty practice all over the world [ ] . even though there were some differences due to the severity of the spread of the infection in various countries, all elective surgery stopped, at least in europe [ ] . minor differences have been seen in the duration of the lockdown, which has lasted for a minimum of months [ ] . the economic impact on the health systems has been tremendous, and still to be realised. decrease in orthopaedic elective operations has occurred globally. in usa at the beginning of , almost , primary arthroplasties were being performed per week. assuming % have been cancelled, then , have been postponed per week. this was seen in when the global economic recession hit the usa [ ] . orthopaedic implant companies, private healthcare centres, and healthcare professionals, have all lost an unknown amount of income. the efficiency of the system has suffered a major loss, even though there are no more official restrictions to clinical volumes [ , ] . due to things such as strict prevention protocols, social distancing, team disruption, reallocation of beds, and shortage of personal protective equipment, the number of cases performed per day diminished, directing impacting elective waiting times. as a consequence, we are seeing an increase of costs coupled with a reduction in efficiency, whose real effects on healthcare economic viability are still to be evaluated. in this scenario, surgeons must still offer the best treatment for their patient whilst navigating a possible resurgence or second wave [ ] . most european countries have moved to a post-first wave period. a resurgence or a second wave of covid- , regardless of whether there are stringent or relaxed policies for mitigation, will then have a further severe impact [ , ] . adult hip and knee elective reconstructive surgery has restarted so as to avoid the serious effects of delaying surgery on patients' symptoms, function, and quality of life [ , ] . scientific bodies in both europe and north america have developed the guidelines for a progressive restart, paying attention to the safety of patients and staff [ ] . however, the scientific basis that underpins the guidelines is poor, despite the influx of covid- articles, most of dubious quality. as many of the countries begin to think about reducing restrictions to allow social an economic recuperation, the world health organisation (who) revealed that highest daily incidence of confirmed cases occurred on june [ ] . the numbers of cases of sars cov- are also increasing daily in south america, north america, africa, and india [ ] . overall, the number of cases continues to increase [ ] . detection of new cases in beijing has led to a renewed lockdown there. the potential for a second wave of infection has been highlighted in both the medical and non medical literature [ ] . potential reasons for this include: • relaxing the social distancing and population behaviour measures before adequate suppression of local cases from failure to detect all cases in a specific population. • introduction of new cases via travel or interaction with infected individuals who have not been screened. • initial failure to report or test populations at risk during the initial wave, e.g., in nursing homes [ ] . • failure of the population to acquire herd immunity. in most countries, the infection rate is less than % [ , ] . several of these factors, either in isolation or combined, can result in a resurgence of cases. further waves are possible in the future. whilst the timing and location of a potential second wave is difficult to predict, valuable lessons have been learned from the first wave, which can inform on the management of a resurgence or a subsequent wave. lockdown has been effective in reducing transmission of the virus. it has been estimated to have prevented or reduced million infections in several countries [ ] . screening and contact tracing and early detection have significantly reduced transmission rates [ ] . isolation and quarantine measures have reduced community and national spread, especially amongst at-risk groups (elderly, comorbidities, lower socio-economic, and certain ethnic groups). the use of personal protective equipment (ppe), by those at risk of exposure, has reduced the incidence of covid- spread in healthcare and other key workers [ ] . in this respect, the definition of 'key worker' has to be clear as, for businesses, healthcare institutions, and society as a whole to function, a variety of individual skills from cleaners to surgeons are required. a vaccine is being discussed, but this has not yet been developed. the important difference between applying these measures to a second outbreak will be to ameliorate the effects of the isolation of people, with its impact on their physical and mental health (in particular, populations at risk, and those in full-time education) and the reduction in economic activity. it is possible that 'normal life' will be adapted to have several of these intervention measures, including the daily routine. the covid- pandemic appeared suddenly and found the global medical community relatively unprepared despite the past experience of the spread of other viral infections. it is inevitable that a resurgence or new wave of codid- infections will have a further severe impact on elective arthroplasty services when restrictions are imposed on a locality. this paper reports the recommendations for individual surgeons on how to prepare for a new lockdown to minimise this impact on their elective hip and knee arthroplasty practice. the european hip society (ehs) and the european knee association (eka) formed a panel of experts to review the literature and to come to a consensus on the recommendations. a systematic review of the available literature was performed up to the rd july using the key words "covid- " or "sars-cov- " and "second wave". the following databases were accessed: pubmed, scopus, and google scholar. all available articles were accessed, and there was no language exclusion. sd and tk conducted the search. nc acted as the arbiter if there was any disagreement. a total of citations were found of which were letters or comments' warning of specific problems with a second wave and advising on getting prepared, three were about avoiding a second wave, and the rest had no relevance to preparation for a new wave, and none gave any details on recommendations. within these , the two orthopaedicrelated citations were principally concerned with the current management [ , ] . included was an open letter in the british medical journal to all the uk political parties' leaders, from a group of senior uk medical leaders. they concluded that rapid attention was needed in a number of policy areas [ ] : • governance including parliamentary scrutiny, and involvement of regional and local structures and leaders. • procurement of goods and services. • coordination of existing structures to optimise effective public health and communicable disease control infrastructure, resilience of the health service, and shielding of vulnerable individuals and communities. • the disproportionate burden on black, asian, and ethnic minority individuals and communities. • international collaboration. it was clear from the literature review that any recommendations would be based on expert opinion without any robust independent evidence to support them. the current pandemic is unique, since there are no postpandemic health care delivery system recovery studies, nor any theoretical studies of post-pandemic health care delivery system recovery operations [ ] . however, the healthcare systems are more prepared for a second wave, with improved logistics, experienced personnel, and treatment algorithms. however, as is clear from the open letter by adebowale et al. [ ] international, national, and local policies will drive the management of a resurgence or new wave for which an individual orthopaedic surgeon has no direct responsibility. looking specifically at arthroplasty, most countries who have resumed elective cases have created separate pathways for medically necessary covid-negative operations. this has been in the form of designated covid hospitals, or physically separating patients, wards, operating theatres, and personnel, coupled with increased vigilance in disinfection measures and identifying virus carriers. the allocation of resources and hospital beds to elective procedures will be titrated against the possible need for these resources if a resurgence of the pandemic occurs. this allocation will follow the guidance of local health authorities, international health organisations, and scientific societies. present and future economic limitations should not diminish the advocacy for high-quality healthcare, nor should it change the ideal indications for arthroplasty. the challenge is to provide the same standard of quality with fewer resources. a second wave will put a major stress on the system. efforts at mitigation need to be considered and put in place now, so as to maintain a high a rate of elective arthroplasty as possible, despite the strain of covid- on the system. the possibility of a new slowdown of elective surgery needs to be discussed with the patients, in particular in the most severe ones where delay will lead to a worse outcome. the goal is to avoid another complete lockdown of arthroplasty surgery, even if healthcare professionals must be ready for the worst from a both clinical and economic points-of-view. the disease will not be controlled fully until an effective vaccine has been found, or herd immunity in the population has occurred. this does not seem to be possible soon. post-disaster scenarios for healthcare management also point out a different second wave; the surge in demand for healthcare in chronic patients who have delayed or have been unable access treatment during the disaster [ ] . arthroplasty surgeons need to be engaged in the following: • prioritisation, limited amount of selected arthroplasty operations have been performed during the pandemic, depending on the type of procedure, status of the patient, and regional rules of patient prioritisation. arthroplasty for periprosthetic and femoral neck fractures were still performed, in % and % of suitable patients, respectively. however, % of aseptic arthroplasty revisions and % of elective primary total joint arthroplasty were postponed [ ] . indefinitely delaying all elective arthroplasty procedures is unacceptable. although prioritising patients, who are low risk for covid- disease transmission and postoperative complications, can be done, ethical aspects also have to be considered. older, more vulnerable patients will suffer most. the beneficial impact of total joint arthroplasty regarding mobility, social life, work capability, prevention of cardiovascular diseases, general health, patient satisfaction, decreasing pain, and increasing joint function, is clearly recognised [ ] . postponing total joint arthroplasty leads to an increase in the use of medication and more unsatisfactory overall outcome. the prolonged time of pain and social isolation, because of immobilisation, risks their mental health. if patients have limited access to total joint arthroplasty, or have to wait for an extended period on waiting lists, the direct and indirect costs of a nation's society will increase [ ] . • patient information, the individualised guidance that patients receive is the key to when deciding to undergo elective joint replacement [ ] . direct, succinct, and transparent information is recommended for communication through the coronavirus crisis [ ] . it is our duty as surgeons to be committed, use shared-decision making tools [ ] , and back up our recommendations with up-to-date scientific information supported by scientific societies and local governments. patients receive information from various sources, especially internet sites, which are not necessarily trustworthy [ ] . even though the peak of covid cases is declining, a recent study by chang et al. [ ] found that only % of the patients interviewed who were awaiting elective arthroplasty, agreed to undergo surgery. patients waiting for knee arthroplasty were more reticent than those for hip arthroplasty. it is remarkable that none of the patients who were waiting for a revision arthroplasty agreed to undergo an operation during this period. official information for patients, using an appropriate level for their clear understanding, should be available on institutional and orthopaedic societies' webpages [ ] . a patient's refusal to undergo elective surgery during this time should not be synonymous with declining a joint replacement. the surgeon should consider reoffering the surgical option after a reasonable time, ideally in a scenario with less risk of coronavirus transmission. • legal aspects, orthopaedic surgeons should be aware of the presence of several new legal issues during the covid- pandemic. there is a risk for claims against healthcare facilities, and, therefore, orthopaedic surgeon by association. the claims could be for: . failing to prepare and respond appropriately to a covid-positive patient. . fail to diagnose a covid- patient in a timely fashion and proper fashion. . negligence in taking appropriate precautions to prevent or limit exposure and spread of the virus. . negligence in treating a covid-positive patient in a timely and proper fashion. therefore, it is incumbent on the surgeon to be familiar with regional standards of care, in addition to national, state, local, and hospital protocols, for the medical and surgical treatment during the covid- pandemic. although there is often implied immunity given to treating physicians during the pandemic, these are usually reserved for "front line" staff. orthopaedic surgeons may not be viewed as such. it is important to understand the patient's legal rights during this time. appropriate information must be accompanied by a specifically designed consent form, which should be produced at the centre where the surgery is to be performed. patients must receive the message that elective procedures will be carried out to the highest technical standards whilst maintaining preventative measurements. the eventual modifications in peri-operative protocols (early hospital discharge, telemedicine, tele-rehabilitation, etc.) are intended to provide the best possible outcome. notwithstanding, patients should comprehend and accept that the occurrence of adverse events and complications can be minimised but not abolished. patients should understand that, in the event of an official declaration of a resurgence, or further wave, of covid cases in a given area, the scheduling of elective cases may be altered and even cancelled. in countries with a waiting list for joint replacement, patients should be informed that of the expected prioritisation decisions and how this will impact on their own waiting time. recommendations to prioritise, for instance younger individuals with a few comorbidities, could generate a delay in the surgical access of those with greater peri-operative risk. therefore, the possibility of a less successful functional result in those patients is a possibility [ ] . best practice is to document informed consent carefully. although there may be implied relief from documentation during the pandemic, the benefits of detailed documentation outweigh any intended relief. documentation is the single best way to protect the surgeon and the patient. the surgeon-patient discussion preceding arthroplasty should be documented; especially the changes from previous practice because of the covid- pandemic. several national societies have publicised a need for a specific, informed consent. at the very least, the following should be documented: . covid- has been declared a pandemic by the world health organisation. . the surgeon and facility closely monitor the local situation and put in place reasonable measures aimed to reduce the spread of the virus. . the patient understands that arthroplasty, except for impending or catastrophic failure, periprosthetic fracture, and infection treatment, is an elective procedure. . the patient acknowledges that, although they may have a negative symptom profile or coronavirus negative test, they still may have a covid- infection. should they develop covid- infection they have an increased risk of significant complications and death. . the patient acknowledges the option to delay surgery further was provided, and that the delay may lead to a worse overall outcome [ , ] . • the role of orthopaedic surgeons, during the next wave of the covid- pandemic, surgeons will have personal responsibilities. the actions necessary need to be planned for now (fig. ). there will be national guidance and local policies that need to be adhered to including around covid- screening for patients and staff, but surgeons themselves have obligations that must be considered and actioned during the phase between waves. there are professional responsibilities [ ] to oneself, to the multidisciplinary team, and to one's family. a period of reflection on what went well and what could have been done better during the first wave and the lockdown needs to be considered by oneself, the multidisciplinary team, and one's family. included in this is both the physical and mental wellbeing of all those involved, and any special requirements on individuals in the group from their personal risks factors, especially those with a black, asian, or minority ethnic background [ ] . it should include the type and availability of personal protective equipment, how to manage if sickness strikes, and such mundane things as how to get one's hair cut. on a personal level, the surgeon should consider what professional training they require, particularly if rostered into a new role from elective arthroplasty, e.g., providing cover for trauma. more training on managing ethical problems has been highlighted [ ] , although the legal ramifications may depend on the country of one's practice [ ] . one should also consider how to manage personal bereavement and its legal manifestations, including writing a will. the recommendations are reported in table . the recommendations provide a framework within which data collection and analysis are the key for the planning and refining of the response to a new phase of sars-cov- . further phases are expected, and new infective organisms with a global dissemination are expected. this means that the actual details of managing an epidemic at a local level will be changing over time as the resources needed to manage a particular outbreak will differ. this difference may be national or regional, and so, details cannot be assume to apply universally. although the details of a hospital's preparation for the next wave in maintaining elective primary arthroplasty are tied to national guidance, much can be done by an individual to mitigate on the problems that can arise at a personal and family level. it is here that action should be taken to relieve the burden and stress of a lockdown. the preparation for the next wave starts with reviewing the previous wave. between the waves preparation is needed at a personal, departmental, and hospital level and national levels. at a personal level, there are professional activities that need to be pursued, as well as to the multidisciplinary team and to ones family. during the next wave, accurate data collection with transparent dissemination is essential and needs to be planned for at this stage. conflict of interest one or more authors provide consultancy services to implant manufacturers. funding there is no funding source. ethical approval this article does not contain any studies with human participants or animals performed by any of the authors. open access this article is licensed under a creative commons attribution . international license, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons licence, and indicate if changes were made. the images or other third party material in this article are included in the article's creative commons licence, unless indicated otherwise in a credit line to the material. if material is not included in the article's creative commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. to view a copy of this licence, visit http://creat iveco mmons .org/licen ses/by/ . /. covid- : call for a rapid forward looking review of the uk's preparedness for a second wave-an open letter to the leaders of all uk political parties modeling the impact of social distancing, testing, contact tracing and household quarantine on second-wave scenarios of the covid- epidemic. medrxiv the role of the 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observational analysis key: cord- -dzma ye authors: rippin, holly l.; wickramasinghe, kremlin; halloran, afton; whiting, stephen; williams, julianne; hetz, kathrin; pinedo, adriana; breda, joão j. title: disrupted food systems in the who european region – a threat or opportunity for healthy and sustainable food and nutrition? date: - - journal: food secur doi: . /s - - -y sha: doc_id: cord_uid: dzma ye dietary health and sustainability are inextricably linked. food systems that are not sustainable often fail to provide the amount or types of food needed to ensure population health. the ongoing pandemic threatens to exacerbate malnutrition, and noncommunicable diseases (ncds). this paper discusses threats and opportunities for food environments and health status across the who european region in the current context . these opportunities and threats are focused around four key areas: ncds and health systems; dietary behaviour; food insecurity and vulnerable groups; and food supply mechanisms. food systems were already under great stress. now with the pandemic, the challenges to food systems in the who european region have been exacerbated, demanding from all levels of government swift adaptations to manage healthiness, availability, accessibility and affordability of food. cities and governments in the region should capitalize on this unique opportunity to ‘build back better’ and make bold and lasting changes to the food system and consequently to the health and wellbeing of people and sustainability of the planet. dietary health and sustainability are inextricably linked. food systems that are not sustainable fail to provide the amount or types of food needed to ensure population health. the current global situation is significantly challenging food systems at supranational, national and local levels. it threatens to exacerbate the current health crisis by increasing the problem of malnutrition, particularly in the form of noncommunicable diseases (ncds). within the who european region ncds (cardiovascular diseases, cancers, chronic respiratory diseases and diabetes) are the biggest causes of morbidity and mortality. overweight and obesitya major ncd risk factor affected % of adults in the who european region in (who ) . how are food environments and health status across the who european region transforming during the pandemic, and how will they continue to change after response measures have been lifted and in the months and years to come? millions across europe and the world are experiencing lockdown restrictions of varying severity, drastically changing the way we live, including our dietary and health-related behaviours. the future consequences of these changes are uncertain, but initial evidence shows that these measures contradict established public health advice, particularly regarding eating healthily and sustainably, and being active. the impacts of the covid- pandemic, together with the public health outcomes of widespread lockdown measures, create a perfect storm for ncd risk factors including obesity and poor diet, physical inactivity, alcohol consumption and tobacco use. in italy, % of deceased patients with covid- suffered co-morbidities, primarily ncds, and there is evidence for this elsewhere across europe and globally (istituto superiore di sanità ; instituto de salud carlos iii ; china cdc weekly ; cdc covid- response team ). evidence also suggests that body mass index (bmi) is positively associated with worse outcomes (peng et al. ) . covid- can present more severe symptoms in people with obesity-related conditions, due to the increased risk of obesity-related ncds such as diabetes (hussain et al. ) . in many who european region countries, more than % of adults are overweight or obese, putting them in a particularly vulnerable position (who a). further, individuals with obesity experience stigma and may suffer higher rates of mental health issues (emmer et al. ) , which may be exacerbated by self-isolation. the link between covid- and ncds means that despite the apparent immediate severity of this pandemic, its long-term health consequences have the potential to outweigh the short-term impact. health systems and society therefore need to fight both risk factors for and bias against overweight and obesity in order to improve health outcomes in a sustainable manner. if used correctly, the pandemic could represent an opportunity to shift towards a health system that better fulfills the role of health promoting settings. it could provide the impetus needed to move from a system built around retrospective reaction, to one that is proactive and based on prevention and promotion of wellbeing in a holistic fashionhighly relevant in a region with high ncd prevalence. in the immediate and short-term, health services should continue to provide essential nutrition services for vulnerable groups including pregnant women, newborns, the elderly and sick children. they should also provide appropriate support for mothers to breastfeed, including those with covid- , and communicate accurate information on maternal, infant and young child nutrition, particularly on complementary feeding. a modern and coherent food systems approach must be conducive to food and nutrition-related behaviors that contribute to a more efficient health system by reducing the burden of diet-related diseases. lockdown measures have shifted the dynamics of dietary behaviour. many national governments in the region forced the closure of non-essential services for an extended period, including food provision businesses such as restaurants and cafes. this could impact population dietary habits in various ways, both positive and negative, in the immediate and medium to long-term. people may engage in more home gardening and cooking out of necessity and pleasure, potentially leading to better quality diets (nielsen ) . this may also have a beneficial impact on mental health. conversely, lack of routine caused by restricted movement may encourage a 'holiday eating' pattern, potentially resulting in over-consumption (venema ) . however, whether these trends are playing out at all socioeconomic levels is also unclear. it is also uncertain as to whether these new patterns will be sustained as lockdown measures are relaxed and the out-of-home food sector reopens. alternatively, individuals may opt for food delivery rather than venturing outside to purchase food at supermarkets or fresh markets. even pre-pandemic, increasing numbers of urban-dwellers in the region were purchasing from online food delivery services due to time scarcity, a major inhibiting factor in the preparation of home-cooked meals. a pre-covid- non-representative survey of european countries found that every fifth meal was consumed outside of the home, of which % were from commercial outlets (iri ). many of these convenience options are high in fat, saturated fat, sugar and salt and are associated with ncd risk. however, lack of supermarket visits may reduce the bulk purchase of such unhealthy, ultra-processed foods and limit exposure to in-store promotions that might otherwise encourage over-consumption. while these adjustments may be temporary, evidence from previous intense social transformations shows that times of crisis can accelerate transformation in the food system, for better or for worse. there exists an opportunity to harness and encourage healthier cooking and eating, both at home and within the out of home sector as this reopens. communication and multisectoral collaboration will be required to facilitate this across all socioeconomic levels, ensuring that no population group is left behind. restricted movement and temporary business closures are affecting normal food-related practices. in a globalized food system, limited access to fresh foods due to panic buying, stockpiling and food shortages may compromise the ability to eat a balanced diet and increase consumption of highly processed foods, often high in fats, sugars and salt (world economic forum ; world health organization ). good nutrition is crucial for health, particularly in times when the immune system might need to fight disease. school closures negatively affect the food environment and possibilities for physical activity among children (world food programme ). among these effects are greater risk of food insecurity, as well as sedentary behaviours, both of which are linked to childhood overweight and obesity (rundle et al. ) . schools can play a vital part in mitigating the risks of both over and undernutrition, as there is evidence that food insecurity increases during school holidays, particularly for vulnerable groups (graham et al. ) . the health and economic implications of this crisis are intertwined. in the wake of the pandemic, many face job insecurity and reduced income, and the economic costs of social distancing tend to hit the poorest, most vulnerable and marginalized members of society. there have been reports of charities tackling such food insecurity, like food banks, experiencing higher demand across the who european region. over % of european food banks have seen increased requests for food aid and emergency food assistance (european food banks federation ). healthy and sustainable food systems must also serve these vulnerable populations. increased unemployment and financial insecurity may also lead to elevated alcohol and tobacco consumption, worsening ncd risk and outcomes (who b; henkel ). further, the elderly population and those with compromised immune systems across the who european region have been specifically instructed to strictly self-isolate for an extensive period. there are particularly strong associations of covid- incidence, hospitalization risk and poorer outcomes in the elderly and those with diabetes, obesity and hypertension. older people may face unique food and nutrition challenges, especially in contexts of isolation and/or poverty (starr et al. ) . elderly care homes are at high risk of residents contracting covid- , and there is evidence that deaths in care homes are increasing in some countries at alarming rates (office for national statistics ). additionally, older populations are often not adequately considered when food systems and nutrition issues are discussed. this double burden may put this group at particular risk of poorer outcome for covid- and ncds. the supply side of the food system is also impacted by covid- (torero ) . labour and logistics capacity issues mean that companies are struggling to meet consumer demand for fresh food (cbi ministry of foreign affairs ). restricted movement has prevented migrant seasonal workers from picking fruit and vegetables, resulting in food loss and waste, and potentially food safety risks. no food can be considered healthy if it is unsafe to eat, and it is important that neither health nor environmental sustainability are compromised due to spoiled food or increased food loss and waste. the covid- crisis may provide an opportunity to reassess our diets and food systems and to explore ways to add diversity. traditional and regional diets could play a significant role in responding to local and national challenges in ensuring healthy and sustainable diets, by promoting sustainable farming practices and smarter and shorter food supply chains to buffer against the multiple shocks created in times of crisis. a move towards such diets could also provide an opportunity for individuals to reflect and adapt their dietary choices related to food access and consumption. investment initiatives aimed at maintaining food systems as part of the covid- response and beyond should be an integral part of helping individuals and society recover better, both from a nutritional and an environmental perspective. the current pandemic poses significant challenges to food systems in the who european region, demanding from all levels of government, particularly cities, swift adaptations to manage the healthiness, availability, accessibility and affordability of food. there is no 'business as usual'we cannot afford to ignore the issues raised in this think piece. food security and nutrition are emerging as one of the biggest concerns as a collateral effect of the pandemic. immediate action is needed to address, monitor and understand this changing situation. governments in the region should capitalize on this unique opportunity to 'build back better' and make bold and lasting changes to the food system and consequently to people's health and wellbeing. work is urgently needed to gain a better understanding of and to help countries navigate this rapidly changing landscape. this includes a need to innovate surveillance systems to track the impact and to anticipate mitigation measures in terms of food systems for populations and at-risk subgroups in urban and rural contexts. we must leverage these opportunities to contribute to a more sustainable environment and to future proof against the next pandemic. author contributions all authors have contributed to the concept and design of the research and to the writing and/or revision of the manuscript and have approved the manuscript for submission. funding information this research was funded by the who regional office for europe. funding for the publication was received from the government of the russian federation within the context of the who european office for the prevention and control of noncommunicable diseases (ncd office). data availability all data and material are appropriately referenced. code availability not applicable. conflict of interest the authors declare that they have no competing interests for the content of this paper. jb is the head, who european office for the prevention and control of noncommunicable diseases. kw, sw and jw are technical officers, hlr and ah are consultants of the same office, kh and ap are interns of the same office. ah is also a postdoctoral researcher at the university of copenhagen. the authors alone are responsible for the views expressed in this article and they do not necessarily represent the views, decisions, or policies of the institutions with which they are affiliated. open access this article is licensed under a creative commons attribution . international license, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons licence, and indicate if changes were made. the images or other third party material in this article are included in the article's creative commons licence, unless indicated otherwise in a credit line to the material. if material is not included in the article's creative commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. to view a copy of this licence, visit http://creativecommons.org/licenses/by/ . /. dr. holly rippin is a consultant at the world health organization regional office for europe and is a registered nutritionist in the uk. she spent her early career in the food industry, before completing a phd and postdoctoral position in the university of leeds nutritional epidemiology research group. her research interests span european national dietary surveys, food portion sizes and trans fatty acids, sustainable and healthy diets and ncd prevention. rippinh@who.int high demand for processed fruit and vegetables due to covid- . cbi. the hague preliminary estimates of the prevalence of selected underlying health conditions among patients with coronavirus disease the association between weight stigma and mental health: a meta-analysis school holiday food provision in the uk: a qualitative investigation of needs, benefits, and potential for development unemployment and substance use: a review of the literature ( - ) covid- and diabetes: knowledge in progress situación de covid- en españa a de marzo de in meals eaten out of home as european consumers favour service over home cooking report sulle caratteristiche dei pazienti deceduti positivi a covid- in italia il presente report è basato sui dati aggiornati al marzo . covid- while still in lockdown, many europeans expect the impact of covid- to last another year number of deaths in care homes notified to the care quality commission clinical characteristics and outcomes of cardiovascular disease patients infected by -ncov covid- related school closings and risk of weight gain among children nutritional vulnerability in older adults: a continuum of concerns without food, there can be no exit from the pandemic europe's fresh food supply is being threatened by coronavirus european food and nutrition action plan - . copenhagen: who regional office for europe. world health organization overweight. european health information gateway stepwise approach to surveillance (steps) dr kremlin wickramasinghe mbbs, msc, phd is a technical officer on noncommunicable diseases (ncd) risk factors. his work focuses on integrating responses to addressing ncd risk factors in the region. before joining who, he was co-director of the who collaborating centre on population approaches to n c d p r e v e n t i o n a t t h e university of oxford, united kingdom. he was also course dir e c t o r o f t h e u n i v e r s i t y 's accredited short course on ncd prevention. he co-edited the text book "an introduction to populationlevel prevention of non-communicable diseases" published by the oxford university press. he has a special interest in multisectoral responses to health promotion, quantifying the outcome of health policies and implementation science. kremlin graduated as a medical doctor from the university of colombo. he holds an msc in global health science and a phd in public health from the university of oxford. wickramasinghek@who.int disrupted food systems in the who european regiona threat or opportunity for healthy and sustainable food and nutrition?dr. joão j. breda is the head of the ncd office in moscow, russian federation. this office leads who work on strengthening national capacity in all countries in the who european region to prevent and control ncds, promote an intersectoral approach and develop policies to tackle ncds. the office has been set up as the innovation powerhouse for ncds prevention and surveillance in europe and beyond.during the previous years before taking office in moscow joão breda was the programme manager: nutrition, physical activity and obesity at who regional office for europe based in copenhagen, responsible for providing support to member states of the who european region on the implementation of the european food and nutrition action plan - & the physical activity strategy for the who european region - , as well as evaluating their implementation process.joão breda is a phd in nutritional sciences from porto university where he defended a thesis focused on alcohol and young people. he has done his master degree in public health by the faculty of medical sciences of lisbon university and an mba from the european university in barcelona. rodriguesdasilvabred@who.int key: cord- -jdvvegp authors: bressan, silvia; buonsenso, danilo; farrugia, ruth; parri, niccolo’; oostenbrink, rianne; titomanlio, luigi; roland, damian; nijman, ruud g.; maconochie, ian; da dalt, liviana; mintegi, santiago title: preparedness and response to pediatric covid- in european emergency departments: a survey of the repem and peruki networks date: - - journal: ann emerg med doi: . /j.annemergmed. . . sha: doc_id: cord_uid: jdvvegp abstract study objective we aimed to describe the variability and identify gaps in preparedness and response to the covid- pandemic in european eds caring for children. methods a cross-sectional point prevalence survey, was developed and disseminated through the pediatric emergency medicine research networks for europe (repem) and the united kingdom and ireland (peruki). we aimed to include ten eds for countries with > million inhabitants and five eds for less populated countries, unless the number of eligible eds was below five. ed directors or their delegates completed the survey between march th and st to report practice at that time. we used descriptive statistics to analyse data. results overall centers from countries ( % response rate) completed the survey: % did not have an ed contingency plan for pandemics and % had never had simulations for such events. wide variation on ppe items was shown for recommended ppe use at pre-triage and for patient assessment, with % of centers experiencing shortage in one or more ppe items, most frequently ffp /n masks. only % of eds had negative pressure isolation rooms. covid- positive ed staff was reported in % of centers. conclusion we found variation and identified gaps in preparedness and response to the covid- epidemic across european referral eds for children. a lack in early availability of a documented contingency plan, provision of simulation training, appropriate use of ppe, and appropriate isolation facilities emerged as gaps that should be optimized to improve preparedness and inform responses to future pandemics. ever since the first human cases of the novel coronavirus were reported in wuhan, hubei province in china in december , the coronavirus disease (covid- ) pandemic has spread rapidly across the world. the epidemic in europe initially centerd around northern italy where there was a steep rise in the number of cases and case fatalities from february th onwards. while european countries were deciding upon or were enacting containment measures of varying degrees, the infection continued to spread across the continent with devastating impact on health systems, the economy and the society at large. within healthcare, it is crucial that the emergency department (ed), as the entry point to hospital care, is prepared to manage high risk covid- patients in an efficient and safe way, from triage to final disposition. the ed should respond to the epidemic surge in agreement with hospital contingency plans and guidelines from local and national health authorities, also learning from the experience of other countries. even though it has now become apparent that children are affected less frequently and with a much more benign disease spectrum than adults, , appropriate management in the ed of suspected and confirmed cases and their families is essential. , children may present with conditions not linked to covid- but some, when admitted for that condition, are found to have covid- positive swabs as an incidental finding. this may be a feature with the more widespread dissemination of covid- throughout the population. besides suspected or confirmed covid- patients, eds should also maintain the quality of care provided to children presenting with serious illnesses or accidents not related to the pandemic. pathways and protocols need to be in place to ensure that rapid appropriate care is provided to suspected covid- children, while avoiding delay in care of non-covid- patients. in addition, it is paramount to ensure adequate protection and minimize exposure of patients and staff to the infection. however, the dialogue between european pediatric emergency physicians who liaised through their european society and their research networks (research in european pediatric emergency medicine -repem and paediatric emergency research in the united kingdom and ireland -peruki) [ ] [ ] [ ] highlighted differences and challenges in ed preparedness and response between countries, as the covid- pandemic unfolded throughout europe. hence, we aimed to describe the preparedness and response to the covid- pandemic in european referral eds for children within the repem and peruki networks. we hypothesised that european referral eds for children would show variability and gaps in preparedness and response to the covid- pandemic, from which lessons could be learned for the current and future pandemics. we conducted a cross-sectional point prevalence survey. the survey was developed in english by the lead author and then underwent several rounds of review by the research team. the survey was distributed through the repem network, a research collaborative consisting of pediatric eds (peds) and eds of general hospitals with a separate pediatric section, serving as referral centers for children, and also the sites affiliated to the executive committee members of peruki. for each country, a country lead was identified on a volunteer basis, through the network, to disseminate the survey to centers meeting the above reported criteria. country leads were pediatric emergency physicians or paediatricians working in the ed. we also included israel as a european associated country, as israel has been part of the repem network since its foundation. to ensure balanced representativeness of participating countries and feasibility of the study, the research team, using a quota sampling method, agreed to include a pre-determined number of centers based on the population of participating countries. for countries with more than million inhabitants (namely italy, france, germany, spain and the united kingdom) participation of ten eds was sought. for countries with less than million inhabitants five eds were expected to participate, unless the number of eligible eds was less than five (i.e. estonia: four eds eligible; iceland: one ed eligible; latvia: one ed eligible; and malta two eds eligible). the denominator for our survey comprised centers. country leads were to decide on the strategy to approach eligible eds in their country. some country leads approached more than the pre-established number of centers to ensure a % response rate, accounting for the possibility that some contacted centers might not respond. by adopting this approach, some countries actually exceeded the expected number of participating centers per country. for calculation of the survey response rate the number of eds exceeding the pre-determined expected number of participating eds per country was not considered. ed directors or their delegates, staff members appointed by the ed director as most suitable persons to complete the survey, completed one survey for each participating center. the survey was open on march th and st . survey responses were collected in redcap, a validated online data collection system. respondents were asked to state their country of residency, but it was not mandatory to give the name of their hospital. each country lead recorded the name of the invited and participating hospitals. country leads communicated to the principal investigator the number of centers that completed the survey, without disclosing the hospitals' identities, ensuring the number of completed surveys per country matched the number of centers that actually completed the online survey. preparedness and response were defined, based on consensus of the research team, as organizational and operational actions taken by eds to face the pandemic, including contingency plans, training, screening of suspected cases, surge capacity, availability and use of personal protective equipment, ed infection control measure, care pathways and management of suspected cases, health professionals safety and sustainability of care. a first survey was completed by country leads to reflect the national situation of the covid- the participating ed survey completed by each participating center focused on organizational and operational aspects of preparedness and response as reported in the above mentioned definition. we followed strobe guidelines for reporting of observational studies. descriptive statistics were used to analyse the data, reporting % confidence intervals (ci) around each point estimate. data were analysed using stata (version , statacorp, college station, texas, usa). this survey accessed clinicians via a research collaborative to assess their departmental practice and therefore did not require formal ethics review, as determined by the ethics board of the university hospital of padova, italy. consent was implied by participation. a total of countries participated in the study. eighty-nine of the expected centers, based on the pre-established number of participating centers per country, completed the survey (response rate %). however, some countries exceeded the number of expected participating eds, leading to a total of eds participating to the survey ( table and approximately one third of centers ( %) did not have an ed contingency plan for pandemics and % had never organized simulations for such events. the majority of centers ( %) had not experienced mass casualty disasters or pandemics during the past five years. nearly all institutions had established a formal ed management plan for suspected/confirmed pediatric covid- cases, with daily updates in % of centers. surge capacity for pediatric suspected covid- cases was variable between centers at an ed, admission ward and intensive care level in terms of number of available rooms/beds ( table ). in one fifth of the institutions there was no intensive care availability for pediatric covid- patients. only admission ward surge capacity increased according to ed volume (supplementary figure ). adjustments implemented to best manage suspected/confirmed covid- patients included cancellation of planned activity (i.e. outpatient visits, surgery or hospital admissions) in % or more of participating centers; reorganization of beds in other pediatric wards in %, and telemedicine in nearly %. the distribution of responses showed variation in the timing of pre-triage set-up and training on personal protective equipment (ppe). variation was also observed in the recommended use of ppe to be worn during pre-triage and during patient assessment. if looking at recommended mask use during pre-triage, surgical masks were used in % of centers, ffp /n masks in % and ffp /n masks in %, while during patient examination, % of centers used ffp /n masks, % surgical masks and % ffp /n masks. recommended ppe use for patients was more consistent across centers with a surgical mask to be worn by patients in % of eds. recommended duration of filtering masks use was also variable. a shortage of both basic and aerosol generating protective ppe items was experienced by nearly two thirds of centers with ffp /n masks being the items most frequently missing ( table ) . contagion of healthcare workers was frequently reported at an institution level ( %), but less so at the ed level ( %). only % of sites endorsed a periodic active surveillance of ed staff. disposition of healthcare workers who had been in close contact with a confirmed covid- case varied between centers, with approximately one third allowing staff to work while asymptomatic and one third recommending quarantine at home. overall, ed physicians shift work had been re-arranged in nearly two thirds of centers with variable adjustments including both increase and reduction in staff, as well as different shift schemes to prevent cross-infection among staff ( table ) . and reorganized patient flow to accommodate suspected cases in separate dedicated areas. fewer than % of eds had isolation rooms with negative pressure. most eds performed swab testing for sars-cov- ( %). however, asymptomatic children with a history of close contact, who could be otherwise discharged, were not tested in the ed in the majority of centers ( %). at most sites suspected cases who were tested, but were fit for discharge, were sent home and swab results communicated to the family when they became available. in cases of positive test results for discharged patients, half of the centers could count on specific outpatient services to provide telephone follow-up. most eds experienced a substabtial reduction in pediatric presentations, by more than % in half of the centers ( table ) . centers further into the infection spread wave more frequently reported a larger reduction in the number of pediatric presentations (figure ). overall % of centers agreed ( %) or strongly agreed ( %) about the statement "my hospital was ready and prepared to handle covid- at the time the outbreak started in our country" and % agreed ( %) or strongly agreed ( %) when the statement was referred to ed pediatric care. the results of our study should be interpreted in the light of its limitations. although we included a large number of european countries, our survey does not provide a pan-european perspective. however, this is the first european dataset that provides a detailed snapshot of pediatric emergency care from within the pandemic, at a more granular level than any institutional channel has been able to provide so far. while the pandemic evolves in each country and accompanying adjustments are made, a repeat focused survey will capture the dynamic progress made from an organizational and operational perspective. we arbitrarily decided, as a research team, the number of centers to be included in each country to ensure a balanced representativeness and to obtain timely completion of the survey. the participating centers represent a subset of eds caring for children in europe and include referral centers for children, thus our findings may not be generalisable across different settings. although some countries exceeded the expected number of recruited centers, we were able to obtain a reasonable balance in terms of country representativeness. in addition, the objective of this survey was to explore common challenges and common learning points and not to compare responses between countries. lastly, while the authors themselves refined questions via review processes, the survey questions did not undergo a formal content validation procedure. given that most of the answers required fixed quantitative responses on practice in use/recommended at participating eds, a formal content validation would have likely had limited impact on the reliability of our findings. our survey provides a snapshot of preparedness and response of eds caring for children from european countries and a european associated country at one month after the covid- outbreak started in northern italy. overall, the findings of our study show high variation in time and in level of organisational responses to covid- of eds caring for children across europe and identified a few gaps that still need to be optimized to improve preparedness and inform responses to future pandemics. our data show that a written and documented contingency plan was still missing in approximately one-third of centers one month after the onset of the outbreak in europe. while the majority had not faced an epidemic or a mass casualty event in the past five years, nearly % had never run a simulation on how to manage such a crisis in the ed. a striking finding of our point prevalence survey was the wide variation in reported ppe use at pre-triage and for the assessment of suspected covid- cases, with % of centers experiencing shortage in one or more ppe items. in addition, a high percentage of centers reported infection in staff members, which may affect the sustainability of care provided. from a structural perspective, the low percentage of eds with negative pressure isolation rooms ( %) highlights opportunities for improvement, should renovation work be undertaken or new hospitals are built in the future. while participating countries were at different stages in the outbreak spread the different pace in the pandemic advancement represents an opportunity for healthcare systems to learn from each other by sharing experience and identifying areas for improvement. this may ensure a more rapid response in terms of implementation of infection prevention and control measures within healthcare in those countries that lag behind the spread wave. this is important at all levels of care within an integrated health care system, but it is paramount for frontline services such as eds. no ppe is required if preliminary screening does not involve direct patient contact. , nearly half of the centers reported a shortage of ppe, most often ffp /n masks. ppe use should be maximized to avoid shortage of supplies, which ultimately exposes staff and the broader community to an increased transmission risk. one third of respondents stated that respirators (n /ffp or ffp /n ) are disposed of after the assessment of each suspected case. this practice may contribute to shortage of supplies, as the same respirator could be used for more than one patient, as long as it is not damaged or soiled. approximately % of respondents were aware of infected healthcare workers at their institution, while one fourth reported infected staff in the ed. unfortunately, infection of healthcare workers has been reported as a major threat to the sustainability of healthcare in this pandemic. in fact, the disposition of healthcare professionals who had been in close contact with a confirmed covid- case varied between centers, possibly because of concerns regarding service provisions. manner. this is in contrast to barriers related to structural limitations and constraints affecting the organization of ed patient flow and isolation capacity, which may be difficult to overcome in a short time frame. infection control measures were more consistently reported in the survey, including re-arrangement of ed patient flow, changing of staff work shift to optimize resource utilization, reduction in the number of care givers allowed with the child and home quarantine for confirmed covid- pediatric cases fit for discharge. another interesting finding from our survey is the substantial reduction in pediatric ed presentations during the pandemic. centers from countries with a longer time since first case experienced higher reductions in the number of ed presentations. parents' fear of contagion in a healthcare environment, improved hygiene measures, reduced community transmission of communicable diseases, reduced opportunities to sustain injuries owing to the strict containment measures enforced by governments, and reduction in stress-related functional diseases may be the reasons underlying this phenomenon. reports from previous epidemics also showed an overall decrease in ped attendances. , the mers outbreak had resulted in a significantly higher proportion of high acuity ed pediatric presentations and an increase in delayed presentations. , despite its limitations, the provision of a timely report on preparedness and response in pediatric emergency care during the pandemic is useful to inform practice and policymakers to properly reorganize health systems while the crisis is still evolving. it provides an accurate objective historical dataset from which lessons can be learned for the future, including for adult eds. the collaboration of the repem and peruki european networks was instrumental to ensure wide representation of european countries and timely completion of this multinational point prevalence survey. the data provided highlight the importance of european multinational research collaborations to provide the best care to children in the frontline. in summary, we identified variability and gaps in preparedness and response to the covid- not reported ed=emergency department; ped= pediatric emergency department; uk= united kingdom § number of eligible eds was less than the pre-determined number of five expected participating centers for countries with a population of less than million inhabitants * referral ed for children # malta: general referral ed seeing children; portugal: secondary-care ped in a hospital for adults and children; spain: secondary-care ped in a hospital for adults and children; sweden: secondary-care ped for medical conditions. a secondary-care ped provides specialist care upon referral by primary care, pre-hospital emergency services or other smaller hospitals, but does not include highly specialized medical care, which may involve advanced and complex procedures and treatments performed by medical specialists in a tertiary-care state-of-the-art facility. ** belgium: centers did not know; germany: centers did not know; switzerland: center did not know; uk: center did not know he/she must be tested and in the meantime be in quarantine he/she must be tested and in the meantime can work with a surgical mask he/she must be placed in quarantine without being tested if he/she has no symptoms, can continue to work without being tested other don't know both parents/caregivers are allowed to be with the child only one parent/caregiver is allowed to be with the child there are no rules that establish the number of caregivers allowed in the ed % % % - % - % - % what is the patient flow for a suspected covid- in your ed/ped after pre-triage? patient is taken directly to an isolation room, with negative pressure patient is taken directly to an isolation room, but with no negative pressure patient is taken directly in a usual visit room patient waits in the usual waiting room patient waits in a dedicated waiting room for suspected covid- other - % - % . - % if you perform a nasal/pharyngeal swab for sars-cov- in the ed/ped to a clinically stable child (who would not otherwise require admission) i must keep the child in a dedicated isolation room until i receive the swab result i can discharge the child home and i communicate the family the result when available i have to admit the child to a regular pediatric ward until i receive the swab result children who do not need admission are not tested for sars-cov- in my ed other % % % % % - % - % . - % - % . - % if your only criterion for suspected covid- is close contact with a confirmed covid- case and the child is otherwise asymptomatic and well, do you test/will you test the child in the ed/ped for sars-cov- ? no, they are discharged home for isolation ( a novel coronavirus from patients with pneumonia in china similarity in case fatality rates of covid- /sars-cov- in italy and china european society for emergency medicine position paper on emergency medical systems response to covid- covid- : learning from experience clinical and epidemiological features of children with coronavirus disease (covid- ) in zhejiang, china: an observational cohort study epidemiological characteristics of pediatric patients with coronavirus disease in china coronavirus infections in children including covid- . the pediatric infectious disease journal diagnosis and treatment recommendations for pediatric respiratory infection caused by the novel coronavirus impact of middle east respiratory syndrome outbreak on the use of emergency medical resources in febrile patients supporting the health care workforce during the covid- global epidemic research priorities for european pediatric emergency medicine establishing the research priorities of pediatric emergency medicine clinicians in the uk and ireland pediatric emergency care in europe: a descriptive survey of tertiary medical centers a guide for the design and conduct of selfadministered surveys of clinicians the redcap consortium: building an international community of software platform partners the strengthening the reporting of observational studies in epidemiology (strobe) statement: guidelines for reporting observational studies pandemic planning and response in academic pediatric emergency departments during the h n influenza pandemic pandemic influenza and major disease outbreak preparedness in us emergency departments: a survey of medical directors and department chairs covid- infection in children severe acute respiratory syndrome coronavirus (sars-cov- ) infection in children and adolescents: a systematic review a novel coronavirus outbreak of global health concern sars-cov- infection in a pediatric department in milan: a logistic rather than a clinical emergency. the pediatric infectious disease journal covid- : protecting health-care workers protecting health care workers during the covid- coronavirus outbreak -lessons from taiwan's sars response infection prevention and control for covid- in healthcare settings -first update rational use of personal protective equipment for coronavirus disease (covid- ): interim guidance pandemic influenza planning: addressing the needs of children impact of the middle east respiratory syndrome outbreak on emergency care utilization and mortality in south korea the impact of middle east respiratory syndrome outbreak on trends in emergency department utilization patterns the authors would like to acknowledge all the respondents to the survey and who gave permission for their names to be included in the acknowledgment section of this paper: key: cord- - fnis ci authors: botella, leticia; tuomivirta, tero t.; hantula, jarkko; diez, julio j.; jankovsky, libor title: the european race of gremmeniella abietina hosts a single species of gammapartitivirus showing a global distribution and possible recombinant events in its history date: - - journal: fungal biol doi: . /j.funbio. . . sha: doc_id: cord_uid: fnis ci the population genetics of the family partitiviridae was studied within the european race of the conifer pathogen gremmeniella abietina. one hundred sixty-two isolates were collected from different countries, including canada, the czech republic, finland, italy, montenegro, serbia, spain, switzerland, turkey and the united states. a unique species of g. abietina rna virus–ms (garv-ms ) appears to occur indistinctly in g. abietina biotypes a and b, without a particular geographical distribution pattern. forty-six isolates were shown to host garv-ms according to direct specific rt-pcr screening, and the virus was more common in biotype a than b. phylogenetic analysis based on partial coat protein (cp) cdna sequences divided the garv-ms population into two closely related clades, while rna-dependent rna polymerase (rdrp) sequences revealed only one clade. the evolution of the virus appears to mainly occur through purifying selection but also through recombination. recombination events were detected within alignments of the three complete cp and rdrp sequences of garv-ms . this is the first time that recombination events have been directly identified in fungal partitiviruses and in g. abietina in particular. the results suggest that the population dynamics of garv-ms do not have a direct impact on the genetic structure of its host, g. abietina, though they might have had an innocuous ancestral relationship. the european race of gremmeniella abietina (lagerberg) morelet (anamorph brunchorstia pinea) is the causal agent of stem canker and shoot blight in many conifer species in europe and north america (donaubauer ) . it leads to epidemics depending on annual weather conditions (uotila ) . the most recent serious outbreak of g. abietina took place in sweden in (wulff & walheim ) , when more than ha of pine forest were severely affected. the up-todate taxonomy of this species comprises three biotypes. biotype b, or the small tree type (stt), is primarily found in northern european countries, but it appears to also be present in high mountainous areas of the lake district of turkey (botella et al. a,b; dogmus-lehtijarvi et al. ) . the alpine biotype has been described in the alpine region. these biotypes both grow in harsh conditions of snow and extreme temperatures and produce abundant pycnidia ( e celled) and apothecia in the field (hellgren & h€ ogberg ) . the third currently known biotype is the biotype a, or the large tree type (ltt). it is the most pathogenic biotype and shows the widest distribution, ranging from the italian apennines to northern sweden and north america, where it was introduced in the s and designated the european race (eu) (skilling et al. ) . conversely, it produces apothecia only rarely in forests (uotila ; kaitera & jalkanen ) , and its conidia are celled. population genetic studies performed through analyses of scar markers have consistently found highly similar alleles in the b and alpine biotypes (botella et al. ) , while biotype a displays fixed nucleotide differences from the other two (botella et al. ) . biotype a exhibits low overall genetic variability (hamelin et al. ) , with the exception of the spanish population, which is a clonal population in a warmer location, likely as a consequence of a founder effect (santamaria et al. ; botella et al. ) . artificial pairings between the a and b biotypes revealed low rates of successful germination and growth, and accordingly, no viable hybrids have been found in nature (uotila et al. ) , indicating that these biotypes should be separated into two different species (uotila et al. , uotila & hantula . tuomivirta et al. described the occurrence of two independent double-stranded (ds) rna banding patterns in a single g. abietina biotype a isolate, which were later shown to represent putative members of the families totiviriridae and partitiviridae (tuomivirta & hantula a; tuomivirta & hantula ) . only biotype b appears to harbour a taxonomically unassigned species with certain endornavirus-like characteristics , and both biotypes a and b host mitoviruses (tuomivirta & hantula b; botella et al. b) . mitovirus-infected isolates appear to present greater mycelial growth than mitovirus-free isolates at the optimal growing temperature for g. abietina of c (romeralo et al. ) . the viral communities appear to be separated into biotypes a and b (hantula & tuomivirta ) , except in spain, where two species of mitoviruses occur in the genetically unique population, which was most likely derived from biotype a. members of the family partitiviridae are doubled-stranded rna (dsrna) viruses that are hypothesised to have evolved from the genus totivirus (ghabrial ; sabanadzovic et al. ). the family partitiviridae includes dsrna viruses with bisegmented genomes, which encode separately encapsidated rdrp and cp segments as well as satellite rna molecules. partitiviruses infect fungal, plant, and protozoan hosts. the classification of partitiviruses has been recently revised, and they now comprise five families: alphapartitivirus, betapartitivirus, cryspoviruses, deltapartitivirus, and gammapartitivirus. the genera alphapartitivirus and betapartitivirus include viruses hosted by fungi or plants, while the genera gammapartitivirus and cryspovirus comprise only mycoviruses (nibert et al. ) . transmission of partitiviruses, similar to other mycoviruses, may occur vertically through sexual and asexual spores or horizontally via anastomosis between related fungal genotypes (ihrmark et al. ; tavantzis ) . however, a number of studies have demonstrated the capacity of partitiviruses to be transferred through somatically incompatible fungi under laboratory conditions (ihrmark et al. ; vainio et al. vainio et al. , vainio et al. , a and in nature (vainio et al. b) . fungal and plant partitiviruses appear to show many similarities in their virion structure and close evolutionary relationships. despite the lack of information about plant partitiviruses, a recent study based on rdrp amino acid sequences suggested that horizontal transfer of partitiviruses has occurred during the evolutionary history of fungi and plants (gabrial et al. ) . we previously analysed the european race of g. abietina (botella et al. ) and its community of mitoviruses (botella et al. a,b) . the present study builds on our prior objective of determining the evolutionary history of mycoviruses and their influence on the phylogeny of g. abietina, this time focussing on the family partitiviridae. more specifically, this study has the following three main objectives: ( ) to measure the prevalence of partitiviruses in the european race of g. abietina; ( ) to analyse their genetic diversity and population structure; and ( ) to assess evolutionary processes, such as recombination and selection, to better understand possible host-virus coevolution. a total of isolates belonging to the european race of gremmeniella abietina were screened for the presence of possible partitiviruses. overall, biotype a isolates were examined: four isolates from the czech republic, isolates from finland, six isolates from italy, five isolates from serbia, and one isolate from montenegro. for biotype b, there were isolates from finland, two isolates from turkey (biotype b/alpine), and one isolate from switzerland (alpine). from the spanish population, isolates were studied, and finally, from the european race in north america, there were isolates from canada and one isolate from the united states. the data collected for all of the strains are presented in supplementary table s , while only virus-containing isolates are listed in table . total rna extraction and cdna synthesis for population analysis all of the fungi were cultivated on malt extract agar medium (mea) supplemented with cellophane and incubated at c. after two weeks, total rna was extracted and purified using the commercial rneasy Ò plant mini kit (qiagen), with a few modifications. approximately mg of freshly grown mycelium was scraped and subjected to freeze using liquid nitrogen. the mycelium was first ground in ml rnase-free tubes with tissuelyser for min at hz. this step was repeated after adding the rneasy lysis buffer complemented with ml of b-mercaptoethanol. the rest of the process was developed following the instructions of the manufacturer. rna quality and quantity were tested through agarose gel electrophoresis and using a qubit Ò . fluorometer (invitrogen), respectively. a total of ng of rna was employed for the synthesis of first-strand cdna using the recombinant m-mulv reverse transcriptase (thermo scientific first strand cdna synthesis kit). the sample aliquot was denatured at c for min and immediately cooled in liquid nitrogen. the concentrations of pd(n) random hexamer primers, dntp, and  buffer were as recommended by the manufacturer. reverse transcription-polymerase chain reaction (rt-pcr) was successively carried out twice with phusion high-fidelity dna polymerase (thermo fisher scientific). in the case of both cp and rdrp rt-pcr, the final reaction volume was ml, with the following components: . u of the enzyme,  phusion hf buffer, mm dntps, . mm each specific primer (par_cp_ f/par_cp_ r and sprdrpgap f/sprdrpgap r) ( table ) and ml of cdna (< ng). the thermocycling conditions consisted of s at c for initial denaturation, followed by cycles of s at c, s at e c (cp) or c (rdrp), and s at c, with a final extension for min at c. the amplified regions were bp (cp) and bp (rdrp) (fig ) . the rt-pcr products were separated via electrophoresis in % agarose gels (serva) containing % tbe ( mm tris-boric table e specific rt pcr primers designed for the study. sequence - a genome region gagttcagcctcgaaaaagca rdrp a the following designations are used for degenerate sites: y (c or t), r (a or g), k (g or t),w(a or t). fig e representation of the genome organisation of garv-ms - and recombination events occurred between strains. *the actual breakpoint position is undetermined (it was most likely overprinted by a subsequent recombination event). minor parent, parent contributing the smaller fraction of sequence. major parent, parent contributing the larger fraction of sequence. unknown, only one parent, and a recombinant need be in the alignment for a recombination event to be detectable. the sequence listed as unknown was used to infer the existence of a missing parental sequence. acid at ph . , mm edta at ph . , mm tris base at ph . ) and ml of gelredä nucleic gel acid gel stain (biotium). the pcr products were visualised under uv light after h and min of running at v/ cm in % tbe buffer solution. gene rulerä bp dna ladder plus (thermo scientific) was used as a length marker. finally, the genejet pcr purification kit (thermo scientific) was employed to purify the pcr products, and the template dna concentrations were determined with a qubit Ò . fluorometer (invitrogen) following the instructions of the manufacturer. the spanish isolate of gremmeniella abietina h - was chosen for determination of the full-length sequence of a putative new strain of garv-ms (genbank accession numbers for the cp, rdrp, and the unknown protein iii: kj ekj ). the double-stranded rnas (dsrnas) of the two corresponding linear molecules ( . e . kb) were purified, integrating some modifications into the cellulose affinity chromatography protocol of morris & dodds ( ) and tuomivirta et al. ( ) . approximately g of freshly growing mycelium was collected and frozen with liquid nitrogen in ml rnase-free tubes. once all of the nitrogen was evaporated, the mycelium was homogenised with mm-diameter stainless steel beads (retsch, germany). the tube was vortexed on a standard vortex mixer at maximum velocity for min. then, . ml of lysis buffer ( . ml trisehcl ph . (carl roth gmbh þ co. kg, germany), . ml of % b-mercaptoethanol (serva electrophoresis, germany), . ml of edta (penta, petr svec, czech republic), ml of sds (ph , ; carl roth gmbh þ co. kg, germany), and ml of sterile distilled water (sdw) were added. the dsrna-binding cellulose added was sigmacell cellulose (c ) (sigmaealdrich; germany). the protocols for double-stranded (ds) cdna synthesis and cloning were developed as described in related works (tuomivirta & hantula a , b botella et al. b ). sequencing of the cloned cdna fragments and rt-pcr products for population analyses was conducted by macrogen europe. the universal primers m forward ( mer) and m reverse ( mer) were used. moreover, it was necessary to design a set of specific primers (table ) to fill some gaps within the sequences of cp and rdrp as well as a t rnaprimer for sequencing the ends. finally, to produce more reliable consensus sequences, the purified pcr products from two different reactions were sequenced in both directions. all of the sequences were submitted to genbank ( table ) . all of the obtained sequences were adjusted, compiled, and aligned using the geneious pro . . software package (biomatters ltd). ncbi protein blast (blastx) was employed to search for similar sequences and conserved domains. ncbi orf finder (http://www.ncbi.nlm.nih.gov/gorf/gorf.html) and geneious pro . . were used to search for orfs. the vienna rna web suite (gruber et al. ) was employed to predict secondary structures (http://rna.tbi.univie.ac.at). mega . . (tamura et al. ) was used to perform calculations to select the best substitution model for the construction of the phylogenetic tree based on the cp and rdrp amino acid sequences. the optimal substitution pattern was determined according to the bayesian information criterion of the program, which specified the jones-taylor-thornton (jtt) matrix-based method as the best model using a discrete gamma distribution (þg) with five rate categories and assuming that a certain fraction of sites are evolutionarily invariable (þi). the tree topologies obtained through neighbour-joining (nj) and maximum likelihood (ml) phylogenetic analyses were compared but were found to be highly similar. therefore, only nj trees are presented here (fig , supplementary s and s ). for these analyses, gaps and missing data were completely deleted from the alignment. there were a total of and positions in the final cp and rdrp datasets, respectively. dnasp v (librado & rozas ) was used to estimate genetic diversity parameters for the population. the average number of nucleotide differences per site between two sequences (pi), number of haplotypes (h), haplotype diversity (hd), number of segregating sites (s), number of total mutations (h) and the average number of differences per population were calculated. dnasp v was employed for the analyses of synonymous and non-synonymous nucleotide substitutions, according to the nei-gojobori model using the jukes & cantor ( ) correction (nei & gojobori ) . we calculated the rate of synonymous per synonymous or silent sites (d s ), the rate nonsynonymous per non-synonymous substitutions or nonsilent sites (d n ), and the ratio u (d n d s ). sites (or codons) with alignment gaps or missing data were not used. the occurrence of recombination was tested employing the default conditions of the suite of programs included in rdp . different detection methods are computed in this program: rdp, geneconv, bootscan, maxchi, chimaera, siscan, phylpro, lard, and seq. significant recombination events were considered to be those detected by at least three different methods, after bonferroni correction for multiple comparisons. rdp provides a detailed breakdown of recombination breakpoint locations and the identities of the recombinant and parental sequence. the presence of putative partitivirus molecules was previously demonstrated in the spanish population of gremmeniella abietina (botella et al. a ). among the positive isolates, h - was chosen for the determination of a possible new strain. following the nomenclature practices described by tuomivirta & hantula ( a and ) genome of garv-ms - was segmented into three putative particles (fig ) . the largest segment showed a length of bp and a relatively high gc content ( . %). it exhibited a unique open reading frame (orf) of bp, from position to . the and untranslated regions (utr) were bp and bp long, respectively. based on translation frame and the standard code, the rdrp presented an orf of amino acids (aa) and a molecular weight of . kda. the second longest segment presented a length of bp and a moderately higher gc content ( . %). there was only one orf of bp identified, from position to (fig ) , which encodes the coat protein (cp). the length of the utr was bp, while that of the utr was bp. the cp presented a length of aa and a molecular weight of . kda. the smallest segment was bp long and exhibited a gc content of . %. the orf, which encodes a hypothetical protein (unknown protein iii), was bp long, and the and utrs were and bp, respectively. based on frame and the standard translation code, the unknown protein iii consists of aa, with a molecular weight of . kda (fig ) . potential stem-loop and panhandle structures could be deduced from the rna sequences of the three proteins when the entire molecules and their individual and utrs were analysed in silico (data non shown). comparison of the cp, rdrp, and the unknown iii amino acid sequences of garv-ms - with the corresponding sequences of other partitiviruses in genbank (table ) revealed high similarity to garv-ms and , which were previously described in finnish isolates of g. abietina biotype a (tuomivirta & hantula b , , confirming that garv-ms - belongs to the same species as garv-ms - and garv-ms - (tuomivirta & hantula, b , . the genetic parameters of cp and rdrp in the garv-ms population indicated a relatively low degree of genetic variation. concerning the cp sequences, within a total of strains presenting a sequence length of bp (net sites), the total number of segregating sites (s ) was , including singleton and parsimony-variable sites. the number of mutations occurring in the population (h) was ; the number of haplotypes (h) was ; the haplotype diversity (hd ) was . ; the nucleotide diversity estimated based on the average number of differences per site between two sequences (pi) was . ; and the average number of differences was . . for rdrp, the nucleotide variability was even lower; the -sequence alignment displayed net sites (excluding sites with gaps/missing data). the total number of segregating sites (s ) was ; the total number of mutations (h) was ; the number of haplotypes (h) was ; the haplotype diversity (hd ) was . ; the nucleotide diversity estimated based on the average number of differences per site between two sequences (pi) was . ; and the average number of differences was . . taken together, these data confirm that garv-ms is highly conserved within the european race of gremmeniella abietina. phylogenetic relationships of garv-ms within the european race of gremmeniella abietina the phylogenetic nj trees based on the partial cp and rdrp sequences of strains of garv-ms are shown in fig and supplementary figures s and s . analysis of rdrp clustering did not reveal any major clusters supported with high bootstrap values. however, in the case of cp sequences, two major clusters were observed. the reliability of the phylogram was supported by the bootstrap test, as the differentiated clusters were confirmed in more than % of replicates. the first cluster was composed of eight strains belonging to gremmeniella abietina isolates from three taxonomically distinct groups: the eu race from newfoundland (canada), biotype a from finland, and biotype b from turkey. the second, more numerous cluster was composed of garv-ms strains hosted by biotype a and spanish g. abietina isolates from canada, italy, montenegro, serbia, and spain. the number of cp amino acid differences per site determined through calculations of the mean diversity between and within the two clusters in the phylogenetic tree were . and . , respectively (in both cases, se was . ). according to this result, the taxonomic complexity of g. abietina and garv-ms is not correlated. to obtain a better understanding of the evolutionary dynamics of garv-ms , the selection pressure on protein-coding regions was assessed based on the u ratio (d n d s ) for the partial cp and rdrp sequences (strains) of garv-ms and the three complete cp, rdrp, and unknown iii protein sequences. in the case of the partial cp strains, there were a total of . synonymous sites (ss) and . non-synonymous sites (nss) on average. the rdrp partial sequences hosted a total average of . ss and . nss. the average ss and nss within the complete rdrp sequences were . and . , respectively, and within the complete cp sequences, we observed . ss and . nss. in the case of the three complete unknown iii protein sequences from garv-ms - , , and , there were . ss and . nss. the substitution rate (u) was below one in all cases (table ) , suggesting the existence of active purifying selection on these sequences. no recombination events were detected within the garv-ms population when the partial cp and rdrp sequences were used for the analysis, nor were such events detected among the complete unknown iii protein sequences of garv-ms - , and . however, when the three complete rdrp sequences of garv-ms - , and were analysed, one recombination event was observed to have occurred between garv-ms - , as the 'major parent' (the sequence that contributed most to the recombinant), and garv-ms - , as the 'minor parent'. in this analysis, the minor parent was noted as "unknown", which means that only one parent and a recombinant are required in the alignment to detect a recombination event. garv-ms - may be the actual recombinant (fig a) . three detection methods resulted in significant p values: max-chi ( . À ), chimaera ( . À ) and, seq ( . À ). the analysis indicated possible misidentification of the initial breakpoint and recombinant. in the case of the complete cp sequences, two recombination events appeared. in both cases, the recombinant sequence was garv-ms - ( fig b) . the first recombination event showed significant p values when using three detection methods: geneconv ( . À ), maxchi ( . À ), and seq ( . À ). the 'major parent' was garv-ms - , which was detected as 'unknown'. the 'minor parent' was garv-ms - . the p values of the second recombination event were also significant for three detection methods rdp ( . ), bootscan ( . ) and seq ( . À ). in this case, the 'major' and 'minor' parents were garv-ms - and garv-ms - , respectively. occurrence of garv-ms in the european race of gremmeniella abietina among a total of gremmeniella abietina isolates analysed for the occurrence of garv-ms , (or . %) of the fungal isolates hosted the virus. when the occurrence of garv-ms was analysed within each population/biotype, the highest incidence was found in the spanish population ( %), followed by the eu race in na ( . %), biotype a ( . %) and, finally, biotype b (and the alpine biotype; only . %). the percentages varied slightly when the prevalence was analysed by geographical region considering the total studied isolates: region i (north of europe: finland), region ii (central europe: czech republic and switzerland), region iii (mediterranean europe: turkey, serbia, montenegro, italy, and spain) and region iv (north america: usa and canada). the highest prevalence occurred within region iii ( . %), followed by region iv ( . %) and region i ( . %). in region ii, no occurrence of partitiviruses was detected. most mycoviruses are 'cryptoviruses' (tavantzis ) ; that is, they do not produce phenotypic changes in their fungal hosts (ghabrial ) . however, some mycoviruses, such as cryphonectria hypovirus (chv- ), display parasite-like traits, such as hypovirulence (milgoom & cortesi ) , and have typically evolved with their hosts (bryner et al. ) . other mycoviruses, such as ustilago maydis virus h , are involved in symbiotic interactions, which strongly affect the evolutionary dynamics and community structure of both symbiotic partners (voth et al. ) . one of the basic relevant questions in evolutionary biology is the degree to which the diversification of parasites and/or symbionts is associated with the diversification of their hosts (g€ oker et al. ) . in this sense, the diversification of the european race of gremmeniella abietina into different biotypes, and possible species (uotila et al. , uotila & hantula , is mostly the outcome of geographical separation and ecological adaptation. however, the virus communities of these fungi could provide some insights about their history of dispersion. according to our results, only one species of gammapartitivirus (nibert et al. ) appears to occur in the european race of g. abietina (garv-ms ), and no host specialisation was observed. the nj phylogenetic tree based on partial cp sequences showed two very close clades of garv-ms strains, and the nucleotide variability of the entire population was quite low ( . ). the rdrp alignment exhibited even lower variability. highly similar strains appeared indistinctly in different countries and in both biotypes a and b (fig ) . the absence of codivergence between garv-ms and its host, g. abietina, might be due to an apparent lack of either negative (parasitic) or positive (symbiotic) effects. although a possible effect cannot be totally ruled out, tuomivirta et al. ( ) did not observe major differences between dsrna-containing (partitivirus-like patterns) and dsrna-free isolates. thus, garv-ms would not be acting as an evolutionary force driving the species diversification of g. abietina. nevertheless, combinations of events such as host switching, duplication, and parasite extinction may lead to topological incongruence and divergence (g€ oker et al. ) . virus transmission may have occurred from biotype a to b or vice versa, as hypothesised for gremmeniella mitovirus (gmv ) and (gmv ) found in the spanish population (specialised a population) and biotype b (botella et al. b) . transmission of partitiviruses between somatically incompatible fungi may also occur in nature (vainio et al. b) . however, in the case of garv-ms , it is difficult to assess where and when cell-to-cell contact might have taken place. the populations examined in our study are geographically separated, diminishing the possibility of recent transmission. purifying selection appears to be the main evolutionary force acting on garv-ms , as amino acid substitutions are silenced (kimura ) in the rdrp, cp and the unknown iii protein. this pattern is assumed to occur in other fungal viruses as well, such as ustilago maydis virus h (voth et al. ) , heterobasidion rna virus (hetrv ) (vainio et al. a) , and gmv and (botella et al. a,b) , whereas fungal viruses such cryphonectria hypovirus (chv ) are subjected to positive selection (feau et al. ) . additionally, recombination events were detected between the complete cp sequences of garv-ms - and , suggesting that one or more of the ancestors of garv-ms - was a recombinant. likewise, recombination was identified in the full-length rdrp sequences of garv-ms - and , with garv-ms - indicated to be a recombinant. because the donor sequences precisely substituted a homologous region of the receptor sequence without altering in its structure (nuss ) , we propose that homologous recombination occurred. rna recombination events are common among plant ssrna viruses (chare ; alabi et al. ; sztuba-solinska et al. ) and human and animal dsrna viruses such as rotaviruses (suzuki et al. ; phan et al. ), but there is less evidence of recombination in the dsrnas of plant (liu et al. ) or fungal viruses (carbone et al. ; feau et al. ; linder-basso et al. ; voth et al. ) , and this phenomenon is particularly scarce in fungal partitiviruses. nevertheless, some studies carried out on coronavirus have indicated that recombination might be more common than is often assumed but may go undetected because of the action of strong purifying selection, which will remove new, deleterious combinations of mutations (banner & lai ) . hence, in the case of garv-ms , we note that recombination could have taken place under specific conditions, permitting the virus to cross the species border. however, it is not possible to determine when this event occurred. limited information is available about possible factors enhancing viral recombination in fungi. nevertheless, the rna-silencing antiviral defence response has been shown to contribute to hypoviral rna recombination in cryphonectria parasitica (sun et al. ). the rna silencing pathway contributes to viral rna recombination by providing and fragments of viral rna (zhang & nuss ) . the prevalence of garv-ms was found to be moderate ( . %) and lower than that of mitoviruses ( %) (botella et al. b ). however, it was relatively high in comparison with other pathogens such as heterobasidion, where the frequency of partitivirus-infected isolates was shown to be ca. % (vainio et al. ) . the occurrence of garv-ms varied among the tested populations. biotype a and the spanish population in particular presented the highest rates, while biotype b and the alpine type displayed a lower occurrence of partitiviruses. the same phenomenon was observed in gmv and gmv (botella et al. b) , though these findings were most likely enhanced by the lower number of analysed isolates, especially from the alpine region; nevertheless, a further explanation may be based on their mode of reproduction. biotype a mainly reproduces via conidiospores, which is a known means of efficient transfer of mycoviruses (polashock et al. ; buck ; pearson et al. ), while biotype b and the alpine type show sexual reproduction more frequently. the percentage of partitivirus-infected sexual and asexual spores can fluctuate greatly within the same species, and it is apparently determined by the combination of fungal host and virus genotypes (tavantzis ) . furthermore, mycoviruses that flow through septal pores may also be constrained by fungal defence mechanisms (leslie & zeller ; cortesi & milgroom ; nuss ), such as the genetic self/nonself recognition system or rna silencing (nuss ) . evidence of such constraint was recently described after infection by rosellinia necatrix partitivirus (rnpv ) (chiba et al. ) , although no phenotypic alterations were observed. in conclusion, the partitivirus community of the european race of g. abietina appears to be composed of a single species (garv-ms ) with low genetic variability, which is seemingly stable within the different populations of the fungal host. this virus evolves not only through purifying selection but also to some extent via recombination. to the best of our knowledge, this is the first report of the occurrence of recombination in fungal partitiviruses. sequence diversity, population genetics and potential recombination events in grapevine rupestris stem pitting-associated virus in pacific north-west vineyards random nature of coronavirus rna recombination in the absence of selection pressure spanish population of gremmeniella abietina is genetically unique but related to type a in europe presence of viral dsrna molecules in the spanish population of gremmeniella abietina occurrence of two different species of mitoviruses in the european race of gremmeniella abietina var. abietina, both hosted by the 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mexico gremmeniella abietina: upptr€ adande i sverige. resultat fr an riksskogstaxeringen och skogsskadeinventeringen . swedish university of agricultural sciences (slu), department of forest resource management and geomatics, ume a e a host dicer is required for defective viral rna production and recombinant virus vector rna instability for a positive sense rna virus supplementary data related to this article can be found at http://dx.doi.org/ . /j.funbio. . . . r e f e r e n c e s key: cord- - gun ep authors: celi, giuseppe; guarascio, dario; simonazzi, annamaria title: a fragile and divided european union meets covid- : further disintegration or ‘hamiltonian moment’? date: - - journal: j doi: . /s - - - sha: doc_id: cord_uid: gun ep despite being symmetric in its very nature, the covid- shock is affecting european economies in a very asymmetric way, threatening to deepen the divide between core and peripheral countries even more. it is not covid- itself, however, but the contradictions within the eu’s growth model and institutional architecture that would be to blame for such an outcome. the dramatic impact of the economic crisis brought on by the pandemic and the threat that it poses to eurozone survival seem to have forced a reluctant germany into action: a minor step, but an important signal. this note analyses the crossroads currently facing europe—the risk of disintegration vis-a-vis the opportunity for a ‘hamiltonian moment’—discussing possible future scenarios in the light of past developments. like viruses, crises too can rapidly change their dna: the financial crisis of changed from international to regional, from financial to real, eventually turning into an existential threat to the whole european integration project. in the institutional context of the eurozone (ez), the financial crisis soon developed into a sovereign debt crisis, dragging the banks along with it. in the austerity environment that followed, the southern periphery (sp) never completely recovered the losses in output, employment, and fiscal sustainability. thus, the "symmetric" coronavirus shock hit countries that were in highly asymmetric conditions. in fact, not all the countries of the union have the resources needed to intervene in support of their economy, prompting concern that countries with the deepest pockets might be getting an unfair advantage in the eu's single market. far from triggering mutual protection, the covid- crisis seems to be paving the way for the same mistakes that followed the financial crisis. the centrifugal forces threatening disintegration of the european monetary union (emu) seem to have been defused, albeit only in part and only in extremis, at least for the time being. however, the survival of the union depends not only on responding to the severe financial problems caused by the epidemic, but also means addressing the long-term, structural problems that led to the increasing divergences among her members. as chancellor merkel herself acknowledged, "it is in nobody's interest for germany alone to be strong after the crisis". convergence is essential to put the union on a more solid basis so as to guarantee its long-term sustainability. what policies and what reforms should be implemented to pursue this objective? and are they economically and politically feasible? trying to answer these questions, we shall briefly review the institutional and structural causes of the increasing divergence between core and sp, shedding light on three momentous events: the creation of the monetary union, the financial crisis and the covid- shock. the first decade following the introduction of the emu saw continuity in the process of europeanisation embarked upon as from the formation of the common market, based on financial liberalization and market globalization. as argued in celi et al. ( celi et al. ( , , europeanisation meant eu-wide application of a policy of deregulation of goods, labour and capital markets that affected the timing, shape and direction of the european integration process, halting the process of convergence between the core and the sp of the eu. the more developed core (centred on germany) increased its productive and technological capacity; the sp, caught between product competition within the eu and cost competition from emerging economies in the international markets, saw a decline in its manufacturing capacity. with the fall of the soviet union and the entry of the former socialist countries of central and eastern europe in the eu, the eastern periphery (ep) became a key gear of germany's manufacturing matrix (stehrer and stollinger ) . a huge flow of direct investments, primarily in the automotive sector, transformed the economies merkel: germany must help other eu states get back on their feet, euractiv.com with reuters mag https ://www.eurac tiv.com/secti on/econo my-jobs/news/merke l-germa ny-must-help-other -eustate s-get-back-on-their -feet/. these diverging trends are likely to increase as a result of the slow, small and asymmetric response that europe is giving to the ongoing pandemic-driven economic crisis, as confirmed by the macroeconomic evidence provided in this forum by heimberger et al. of the visegrad pact (poland, hungry, slovakia, and check republic) into an essential source of intermediate goods (medium and medium-high quality) for the german industry. a well-qualified, extremely cheap workforce, generous subsidies and tax breaks, as well as geographical proximity and historical links, are among the determining factors of the increasingly tight links between the core and its ep. the impressive growth in manufacturing capacity in the east led to a restructuring in the hierarchical organization of the supply chains across europe: the weaker suppliers in the south were displaced by their cheaper competitors in the east, while the highly specialised suppliers of components in the industrial regions of the south maintained, and even increased, their close links with the german producers. the crowding-out of the less dynamic firms in the sp did not take the form of efficiencyenhancing market selection but rather a generalized reduction of production capacity, contributing to fuel a well-documented (see, among the others, guarascio and simonazzi ; dosi et al. ) process of 'poor tertiarisation' of the sp. on the other hand, the ep's industrial miracle was created by foreign, mostly german, direct investment, with the automotive sector taking the lion's share. so far, we have seen no comparable development of other productive sectors, nor has the automotive sector created spill-over effects in the rest of the economy (krzywdzinski ). on the contrary, the surge in the production of components for the automotive sector has partly displaced other productions, leading to an increasing 'mono-specialization' of these economies. despite a growing shortage of skilled labour, wages have remained modest. threats of production shifting further east, to romania, turkey, or to north africa, (pavlinek et al. ) are reflected in the adoption of a wage containment policy at home, driving young people with high educational qualifications to emigrate, and weakening the countries' skills base. with domestic demand subdued, the high growth rates recorded by these countries are entirely led by the growth in exports of local production by foreign multinationals (i.e., the so-called "integrated peripheral markets"). while their intensive specialisation in the automotive industry makes them totally dependent on the health of the german automotive industry, the foreign control of production decisions, innovation processes and markets makes it extremely difficult to undertake an independent, less unbalanced development path (celi et al. ) . to conclude, the two peripheries-the southern one, made up of the mediterranean economies, and the eastern one, with the prominent role of the visegrad countries-suffer from different fragilities, which descend from their common, albeit diverse, economic and financial dependence on the core. however, the core itself is dependent for its growth on the pattern of specialisation within the eu: the southern markets providing an outlet for its increasing surplus of manufactures, the eastern countries supplying cheap inputs for its industries. this combination of structural divergence and economic interdependence lies behind the fragility of the union as well as of the improbability of its disintegration given the high costs it would entail for core and peripheries alike. in the first period of the emu ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) , the core-sp structural divergence was partly hidden by massive financial flows to the periphery. the financial crisis, and the ensuing international liquidity crunch, prompted a "sudden stop" of capital flows and a collapse in demand and imports. at that point, the structural and institutional flaws of the emu became evident: the reaction to the crisis aggravated the divergence. with the blame for the crisis put squarely on borrowers, austerity policies were advocated (or imposed) to ensure debtor countries' public and private solvency. with austerity killing demand, growth and imports in the sp, germany, which had built most of its huge trade surplus between and by exporting to the periphery, had to find new outlets for its goods. special international conditionsnamely, china's huge growth, which gobbled up german capital goods and highquality durable consumer products (particularly cars), and the vigorous american recovery-supported germany's ability to redirect its trade flows, expand its market shares outside the emu, and make a speedy return to its pre-crisis production levels. the united kingdom, the united states, but above all china, became the most important markets for german exports. the rapid recovery of the german economy pulled the ep along with it: the visegrad countries recorded unparalleled growth in europe. with the abrupt change in the international scenario in , germany's (and the entire emu's) mercantilist strategy was up against the ropes. the brexit referendum, trump's election, and the u-turn in chinese economic policy inaugurated a phase of retreat in international trade. trade with the uk began to suffer due to the increasing uncertainty in future trade relations. when the united states took action to reduce the external deficit, china and germany, the countries with the largest trade surpluses vis-à-vis the united states, were caught in the crosshairs. trade tensions between the us and china put further pressure on international trade. the export-led growth model that had so far supported germany's leadership began to creak. the change in world trade took its toll on german (and eu) growth rates. from the second quarter of , the slowdown in german exports hit industrial production and the gdp, widening the growth gap with china and the usa and dragging the whole emu along with it ( fig. ) . as the escalation of trade disputes affected relations between the united states and germany (and by extension the eu), the negative effects on europe's (exportled) growth intensified. in the last quarter of , just a few months before the outbreak of covid- in the eu, germany's growth rate zeroed. income growth estimates for the rest of europe were consequently reduced. the pandemic arrived in europe from the south: italy was the first country to suffer the contagion. its abrupt, dramatic effects exposed the fragility of the periphery and the crippling effects of austerity policies. since , across the board cuts in social spending had hit the entire range, from health to education, from social assistance to social investment. figures , and show the evolution of the share of public expenditure on education and health (divided between general expenditure and hospitals) relative to gdp in the emu, germany and the sp between and . many hospitals had been closed, the number of beds reduced, medical and nursing staff cut back (for a detailed analysis of the impact that austerity policies had on the italian health care system, see prante et al. ). it is not surprising that the death toll was higher where intensive care facilities were scarcer. on the eve of the covid crisis, public health accounted for . percent of the social product in italy and spain, and almost % in germany, where per capita healthcare spending did not suffer cuts due to austerity (though it was not completely spared self-imposed restrictions). the covid- exposed another aspect of the 'divisive' union (celi et al. ) : different capacities to respond to the pandemic crisis. economic ideology shares with austerity the responsibility for the scant endowment of medical equipment and health staff. efficiency, understood as cost reduction, has been taken as the guiding principle. the obsession with competitiveness and reliance solely on the export-led growth model accounts for the almost exclusive emphasis on "tradable" sectors, to the detriment of "non-tradable" sectors (housing, health, education, welfare services in general), considered of lesser importance for international competition. this means that, in the era of austerity, these items have been the first to be sacrificed, in debtor and creditor countries alike. chazan ( ) reports that for years, politicians and health economists in germany have complained that the country has too many hospitals, with the bertelsmann foundation recommending halving the number of hospital, from to fewer than (chazan ). only such a radical consolidation-the bertelsmann study arguedwould "improve patient care and mitigate the shortage of doctors and nursing staff". the pandemic succeeded in transforming this "oversupply" into an asset. the same logic of pursuing the lowest cost guided the international location of production, which displaced domestic production and weakened production capacity in the sp. from a regional (european) point of view, this process resulted in a reorganisation of production and trade relations between core, ep and sp. on a global scale, core and peripheries entered into very long and complex gvcs that proved extremely vulnerable in the face of the interruptions prompted by the pandemic. personal protective equipment, respirators, medicines: the emergency has made it clear what it means to lose the capacity to produce domestically, both in quantity and quality, what is urgently needed, bringing the problem of self-sufficiency back to the attention of economists and policymakers. there is no such thing as a symmetric shock. in addition to the grim toll of victims and the incredible pressure on the health systems of all countries, the lockdown of activities to reduce contagion meant a tremendous plunge in production and incomes and enormous pressure on public finances all over the world. however, the lockdown is expected to affect economies differently. the central and eastern european countries have been less affected by covid than the western european countries: not trusting the resilience of their fragile health systems, they have had to rely on rigid social distancing (walker and smith ) . even within this group of countries there are differences: thanks to their more robust health systems, the czech republic and slovenia were less constrained by rigid social distancing and able to start economic recovery earlier. moreover, due to their strong productive links with austria-a country relatively less affected by the pandemic which came out of the lockdown earlier-and their favourable positioning in the development of digital economy (wiiw ) , their economic outlook is rather better. conversely, it will be tougher for the economies, like those of the sp, which are more dependent on services-tourism and hospitality in particular (fig. ) -and for cee countries and southern regions that rely to a greater extent on production of intermediate products for final producers, since the latter can better defend themselves from fall in demand by cutting down orders to their suppliers (the so-called "whip effect"). policies have also differed widely across countries and regions. while all the central banks of the developed world promptly intervened to provide almost unlimited . although the stability pact has been temporarily suspended, there are obvious differences in how much member states can spend, depending on their fiscal space. member states are making use of the new flexibility granted by the ec on state aid rules, strictly enforced beforehand to ensure fair competition within the internal market (rios ) . germany, which accounts for about a quarter of the eu's gdp, accounts for more than half ( %) of the emergency coronavirus state aid approved by the ec, prompting concerns that countries with the deepest pockets might be getting an unfair advantage by such a sudden (and temporary) abandonment of one of the common market's key pillar (france and italy each account for % of the total). an eu official, speaking on condition of anonymity, observed that "if you look at the scale of what germany in particular, but also some others, are doing-any notion of level playing field or single market integrity has gone out of the window." these concerns underpin the ailing south's demand for a joint eu financial plan. in the absence of a prompt and massive common effort, the sp will pay the highest price to the health crisis. indeed, the different firepower will entail a still greater asymmetry in the economic and power relations between the various member states. the ecb, alone among the eurozone institutions, is doing as much as it can to avoid breakdown of the emu. to address the covid- crisis, it launched a new asset purchasing programme: the eurosystem's balance sheet shot up from billion euros on february to billion by st may . despite this massive monetary injection ( billion in two months) the spread on italian bonds, which had fallen in mid-march following the ecb's announcements, again rose very rapidly, fluctuating in response to political developments. indeed, as tooze and schularick ( ) point out, if, in the crisis, the liquidity injected into the system by the ecb was enough to prevent deflagration of the banking system, the current crisis would require a coordinated fiscal policy of enormous proportions. despite some recent moves (inaugurated by a merkel-macron agreement), this still does not seem to be looming on the horizon. the newly released 'next generation' (ng) plan, based on the - budget, celebrated by some as a "hamiltonian moment", has yet to qualify as forerunner of an eu-wide up-to-the-challenge fiscal capacity. first of all, it is meant to be temporary and, moreover, it is too little, too late. the plan should mobilize billion euros, in the form of grants and in loans. quote reported by the website euractiv.com. the eurosystem balance sheet (the network of european central banks, guided by the ecb) rose from billion euros at the beginning of to billion euros by the end of ; that is, from barely % to almost % of the euro zone gdp ( billion euros). the ng plan money will be spent over the - period. with an even subdivision over the period, the package amounts to an annual . % of the eu's gdp, over four years. several parties, including most recently the president of the ecb, christine lagarde, are urging the ec to review the pact before its temporary suspension expires on december , . apart from the fact that these are gross figures-once the member states' contributions to the eu budget are subtracted, the net amount received by the neediest countries is much smaller-their disbursement will not start before , will be distributed over a -year period, with amounts that grow over time, and, as stated in the ec's "proposal for a regulation" the financial contribution will "be paid in instalments once the member state has satisfactorily implemented the relevant milestones and targets identified in relation to the implementation of the recovery and resilience plan" (ec , art. . .a). as darvas ( ) emphasizes, the incorporation of the ng plan into the eu's next multiannual budget would take advantage of a well-established framework, 'already subject to various checks and balances'. on the other hand, ng resources risk to be trapped in a 'slow-moving machine'. in order to be financed, ng-related projects need to be designed, approved and implemented as part of a process that can take several years. as a result, the timing of disbursements is just the opposite of what would be required to respond to the urgency imposed by the current situation and, even more so, by the expected collapse of incomes that the european economies are going to face. however, the commission expects that barely . % of the total new firepower for grants would be spent in - , when the recovery needs will be greatest (darvas ) . far from being a tool to counter the immediate effects of the crisis, the ng plan is more similar to the juncker plan, and shares all its weaknesses. it is highly unlikely that countries like italy, severely hit by the pandemic and in persistent financial distress, will be able to afford to refrain from asking for other funds (namely, esm, sure and others for a total amount of about billion euros) which could be paid out immediately, subject to the usual conditionality. the merkel-macron agreement has been hailed as the first step towards a more supportive union. behind the good intentions, there are the concrete interests of both france and germany for the survival of the emu: they look with growing concern at the rise of euroscepticism in the sp. the french economy has been hit hard by the pandemic, and was already in difficulty before. gdp forecasts for vary widely, but all agree in estimating a fall in the french gdp of much the same proportions as in the case of italy. on the other hand, germany was, together with the netherlands, the main beneficiary of the creation of the euro, and italy and france were the main losers (gasparotti and kullas ) . as chancellor merkel told the german lawmakers, "it is essential for germany, as an export nation, that its eu partners also do well". indeed, the history of the eu has taught that excessive german surpluses are deleterious for the south of the eurozone. greater government action, retreat from hyper-globalism, and lower growth rates predate the pandemic. the covid- crisis has given yet more voice to calls for protectionist and "beggar thy neighbours" types of policies. it has led countries to prioritize resilience and autonomy in production over cost savings and efficiency through global outsourcing. the same powerful german production platform, so disproportionately export-oriented and dependent on imports of intermediate goods, finds itself vulnerable to a type of shock (the covid- pandemic) that disrupts gvcs and threatens to change the existing economic order through permanent disruption of the patterns of demand and production. although transition from an industrial platform designed for export to one for the internal market (a sort of transition from a war to a peace economy) is a formidable challenge, this transformation would benefit germany itself, considering the winds of trade war and the growing uncertainty about the future developments of the global value chains. the european countries are at a crossroad between either letting the union dissolve or radically reforming it. today's darkened geopolitical environment requires europe to act as a whole. however, the emu will remain fragile as long as it chooses to continue to delegate control over its policies to market surveillance. a true "hamiltonian moment", which involves adopting a common fiscal policy in support of the common monetary policy is a matter of urgency. we still have a long way to go. divisions between member countries marked by opposition between debtors and "frugal" creditors, as well intra-country political struggles and conflicting interests, have-even in the face of this dramatic crisisled to the paralysis of the european institutions, with the one exception of the ecb. faced with what she sees as a serious threat to the eu's survival, the german chancellor (and the commission's president ursula von der leyen) have been driven to action. however, as we argued in sect. , little can be expected from the ng plan for immediate support. the ability of the sp to emerge from the crisis will increasingly depend on its ability to take advantage of the greater flexibility of eu rules for an efficient use of industrial policy, helping companies and the whole economy to respond to the challenge posed by social and technological innovation, the restructuring of production and the reorganization and shortening of gvcs. the pandemic will have significant repercussions on the international organization of production and gvcs (on this point, see also the contributions to this forum by strange and coveri et al.) . indeed, the countries initially most affected by covid (china, korea, italy) are among the most important suppliers of intermediate goods at the international level. studies on the propagation of economic shocks triggered by natural disasters (such as the earthquake that hit japan in ) along the value chains (boehm et al. ; inoue and todo ) found significant supplier substitution effects. anecdotal evidence signals numerous cases of supplier substitution in some countries as a result of the coronavirus (baldwin and tomiura ). the extent of these effects depends on the degree of complexity of the production chains, which affects the degree of input substitutability. propagation effects also depend on the presence of "hub" companies interconnected with a large number of supplier and customer firms (inoue and todo ) . future developments are uncertain, depending on the relative strength of two opposite effects. on the one hand, greater coordination afforded by digitalisation of production networks could favour substitution effects (especially in cases where value chains are less regionalised and the search for new suppliers is more difficult) (zhenwei quiang et al. ). on the other hand, processes of reshoring and shortening of value chains could occur, especially where production chains are less complex or automation is more advanced. the second possibility could represent an opportunity to reverse the processes of deindustrialization that have impoverished, above all, the productive fabric of the peripheral countries. a third perspective, probably utopian, could contemplate coordination of coalitions of producers across eu member states. in a situation of strong productive complementarities between countries, the fortunes of the producers (workers and firms) in one country are bound to those in the other. this would call for a coordinated industrial policy at the european level aiming at ensuring a balanced development of the economies of its members through their integration in the european production networks. in emergency situations where production activities are reduced or temporarily suspended (as in the case of coronavirus shock), bilateral agreements (mediated by governments) between producers in different countries should aim at stabilizing employment levels and pre-existing supply contracts between firms through "mutualisation" of the required financial effort. after all, having surprisingly spoken out in favor of the eurobonds, the ceo of volkswagen herbert diess could-at one remove-be also supportive of such a project! material. if material is not included in the article's creative commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. to view a copy of this licence, visit http://creat iveco mmons .org/licen ses/by/ . /. thinking ahead about the trade impact of covid- input linkages and the transmission of shocks: firm-level evidence from the tōhoku earthquake crisis in the european monetary union. a core-periphery perspective unravelling the roots of the emu crisis. structural divides, uneven recoveries and possible ways out un'unione divisiva. una prospettiva centro-periferia della crisi europea germany's oversupply of hospital beds aids pandemic fight, the financial times next generation eu: % of grants will have to wait until neodualism in the italian business firms: training, organizational capabilities, and productivity distributions proposal for a regulation of the european parliament and of the council establishing a recovery and resilience facility germany gains most from relaxed eu state aid rules re-prima -tranc he- -milia rdi- f -a - ea-b d-c d b b_previ ew.shtml ?reaso n=unaut henti cated &cat= &cid= &pids=po&credi ts= &origi n=https % a% f% fwww .corri ere. it% fint ernat ional % fpre mium% f _giugn o_ % frec overy -plan-tutti -ostac oli-fondi -europ eiralle ntato re-prima -tranc he- -milia rdi years of the euro: winners and losers. an empirical study, cep study a polarized country in a polarized europe: an industrial policy for italy's renaissance firm-level propagation of shocks through supply-chain networks globalisation, decarbonisation and technological change: challenges for the german and cee automotive supplier industry foreign direct investment and the development of the automotive industry in eastern and southern europe decades of tight fiscal policy have left the health care system in italy ill-prepared to fight the covid- outbreak eu countries use looser state aid rules to uphold troubled firms the central european manufacturin core: what is driving regional production sharing? (no. / - ) the shock of coronavirus could split europe-unless nations share the burden. the guardian why has eastern europe suffered less from coronavirus than the west? the vienna institute for international economic studies foreign direct investment and global value chains in the wake of covid- : lead firms of gcv acknowledgements open access funding provided by università di foggia within the crui-care agreement. key: cord- - sct t authors: kujundžić tiljak, mirjana; reiner, Željko; klarica, marijan title: is there a better future of healthy aging? date: - - journal: croat med j doi: . /cmj. . . sha: doc_id: cord_uid: sct t nan the future of aging is being problematized based on the multidimensional concept of healthy aging defined as "the process of developing and maintaining the functional ability," which enables well-being in older age. older people have more complex health needs and are susceptible to developing chronic diseases. frailty, a common clinical syndrome in older adults, marked by a greater risk for poor health outcomes, including incident disability and higher hospitalization and mortality rate, severely affects health care services and society as a whole ( ) . most health services treat acute conditions and manage health issues in a fragmented manner. such fragmentation, as well as suboptimal time-management, could be especially dangerous for organizing adequate care for the elderly. there is a need for a health system reform that would ensure that each and every older person has access to evidence-based medical interventions and timely organized shared care. this is the only way to prevent further health deterioration, disability, and complicated care dependency later in life. active and healthy aging is a common problem in all european countries but could also be an opportunity for europe to spearhead the quest for innovative solutions. aging is one of the crucial societal challenges in croatia, as reflected in the croatian presidency of the council of the eu priority of the ministry of health of the republic of croatia -lifelong health care with the emphasis on challenges of aging. the conference aims to promote research and innovations that improve health and well-being of the aging population, as well as to initiate discussion on transformation of health and care services into more digitalized, long-term, integrated, and personalized care models, while promoting innovative ecosystems in order to deliver a better quality of life among the elderly. the first objective of the conference is to address the issues of aging of biological systems and present the state of the art in the areas of regenerative medicine, neuroscience, clinical medicine, and other fields of medicine. the accent is on personalized and integrated medicine and innovative translational research leading to promising applications in regenerative medicine. the conference will also include discussions on advanced personalized diagnostic strategies enabling individualized therapy and accurate predic- tions of treatment outcomes. the implementation of modern sophisticated genomic methods in routine diagnostics for personalized medicine and the scale up of demanddriven innovations in health care systems includes organizational, economical, technical, and clinical aspects. if we want to extend healthy and independent living, we need to create robust and sustainable solutions potentially applicable in any eu state. the second objective of the conference is to assess the use of smart technologies in age-friendly ecosystems. the conference aims to initiate a discussion on the possibilities to scale up innovations and solutions for age-friendly environments, applications for independent living, and solutions for age-friendly buildings, cities, and environments, undertaken by different european cities, regions, or municipalities. smart ict solutions and advanced artificial intelligence (ai) can be used to provide personalized health care and social services, overcome immobility, cognitive, and vision problems, and improve general quality of life. digital technologies can encourage all groups of patients, and the elderly in particular, to assume a more active role in their health management. hopefully, the conference discussions will lead to discovering the ways how all european citizens can meaningfully use these new technologies and benefit from them. the third objective is to analyze the issues of aging and health care system sustainability at various levels -institutional, regional, state, and eu level. in the area of health systems sustainability, the conference focuses on the following themes and their relation to extended life and aging: health system financing, health system organization and structuring bottom up policy with successful examples, and demographical and ethical challenges. the role of digitalization in health care is vital in advancing solutions to challenges related to all three conference topics, particularly as health systems are often not keeping pace with the integration of the new technologies. the possible solutions to overcome disparities in the availability of technological developments and health and digital literacy of the elderly in different european countries and regions are particularly important. the bfha conference aims to create a platform for sharing the successful examples of using advanced technologies to increase the functionality of aging citizens and for learning from these examples. addressing the use of advanced technologies for improving the functionality and well-being of aging citizens to the benefit of a transformative and mission-oriented european research and innovation agenda goes beyond the traditional focus on scientific impacts of research. it emphasizes societal impacts, structuring impacts on policymaking and policies, as well as impacts on innovation and economy. unfortunately, at the beginning of who decade of healthy aging ( - ) the whole world is facing the coronavirus disease (covid- ) pandemic, which endangers mostly elderly and chronic patients. the pandemic outlines the importance of disease prevention using well known hygiene measures. however, it also reveals the unequal access to health services faced by older patients. in some countries, top ranking hospitals recommended that patients with covid- who are older than , between and years with one-organ failure, and between and years with two-organ failure should not be given priority if there is a lack of intensive care units beds. croatian bases its disease prevention and health promotion strategies on the ideas of dr andrija Štampar, one of the founders of the who ( ). he began his fight for better public health in the first half of the th century ( ) . at that time, sanitary and hygiene situation was poor, and the major health problem were infectious diseases, particularly malaria and tuberculosis ( ) . andrija Štampar became highly active in public health efforts in croatia and abroad. his definition of "health as a state of complete physical, mental and social well-being and not merely the absence of disease or infirmity" is still a part of the who constitution, which was adopted on april , . in his speech as acting president of the first world health assembly held in geneva in , Štampar said the following ( ): "disease is not brought about only by physical and biological factors. economic and social factors play an increasingly important part in sanitary matters, which must be tackled not only from the technical, but also from the sociological point of view.... health should be a factor in the creation of a better and happier life. since health for everyone is a fundamental human right, the community should be obliged to afford all its people' s health protection as complete as possible. medical science must adopt a positive rather than defensive attitude. great tasks await the world health organization in this field, and its future success will largely depend on its ability to put these ideas into practice. " in the light of his words, one can conclude that all people, regardless of their origin, should be granted the opportunity to live a long and healthy life. yet, the environments in which we live can promote health or degrade it. environment and spatial epidemiology play an important role in defining the health risks of a population. an-other important factor is the accessibility to quality health regardless of social status and other sources of deprivation. population aging makes these issues increasingly more visible. healthy aging is about creating the social climate and opportunities that enable people to thrive throughout their lives. the covid- pandemic has shown the importance of every person's involvement in creating a healthy environment, healthy relationships, building solidarity, and social awareness of health needs. let us use this newly obtained attitude to improve the care for the elderly and lead the world to a better future of aging. health, demographic change and wellbeing. brussels: european commission brussels: european commission barroso% % % % -% europe% % -% en% version brussels: european commission; c decade of healthy ageing - -a final proposal at the th who executive board meeting in february . geneva: who; the frailty syndrome: definition and natural history care for health cannot be limited to one country or one town only, it must extend to entire world: role of andrija Štampar in building the world health organization th anniversary of andrija Štampar school of public health -what can we learn from our past for the future? the th anniversary of andrija Štampar school of public health key: cord- -q dyc authors: mitter, hermine; techen, anja-k.; sinabell, franz; helming, katharina; schmid, erwin; bodirsky, benjamin l.; holman, ian; kok, kasper; lehtonen, heikki; leip, adrian; le mouël, chantal; mathijs, erik; mehdi, bano; mittenzwei, klaus; mora, olivier; Øistad, knut; Øygarden, lillian; priess, jörg a.; reidsma, pytrik; schaldach, rüdiger; schönhart, martin title: shared socio-economic pathways for european agriculture and food systems: the eur-agri-ssps date: - - journal: glob environ change doi: . /j.gloenvcha. . sha: doc_id: cord_uid: q dyc scenarios describe plausible and internally consistent views of the future. they can be used by scientists, policymakers and entrepreneurs to explore the challenges of global environmental change given an appropriate level of spatial and sectoral detail and systematic development. we followed a nine-step protocol to extend and enrich a set of global scenarios – the shared socio-economic pathways (ssps) – providing regional and sectoral detail for european agriculture and food systems using a one-to-one nesting participatory approach. the resulting five eur-agri-ssps are titled ( ) agriculture on sustainable paths, ( ) agriculture on established paths, ( ) agriculture on separated paths, ( ) agriculture on unequal paths, and ( ) agriculture on high-tech paths. they describe alternative plausible qualitative evolutions of multiple drivers of particular importance and high uncertainty for european agriculture and food systems. the added value of the protocol-based storyline development process lies in the conceptual and methodological transparency and rigor; the stakeholder driven selection of the storyline elements; and consistency checks within and between the storylines. compared to the global ssps, the five eur-agri-ssps provide rich thematic and regional details and are thus a solid basis for integrated assessments of agriculture and food systems and their response to future socio-economic and environmental changes. changes in climatic, environmental, socio-economic and technological conditions, whether gradual or abrupt, can be challenging for agricultural and societal systems to deal with, but at the same time may offer new opportunities for enhancing agricultural and food sustainability (e.g., bebber et al., ; knox et al., ; leclère et al., ; liu et al., ; reich et al., ) . future developments that bring https://doi.org/ . /j.gloenvcha. . received december ; received in revised form august ; accepted august about such changes are deeply uncertain, but society may be able to manage them better if some plausible developments have already been anticipated. here, we apply a scenario approach to develop socio-economic storylines describing alternative plausible future pathways for the european agriculture and food systems. scenarios can be qualitative storylines (i.e., narratives), quantified descriptions of alternative futures, or anything in between (ipcc, ; reed et al., ; swart et al., ) . they can be used to describe, explore and communicate how the future may unfold. starting from a defined initial situation for a specific region and thematic focus, scenarios look into the future by reflecting on an internally consistent set of assumptions about key drivers and their relationships (ipcc, ) . most often, a scenario matrix is used to structure future developments along two causally independent drivers, and to provide a basic frame for comparing and contrasting scenarios . scenario development has gained in importance over the last decades because of its wide applicability in research and practice. for instance, scenarios can inform integrated assessments of agriculture and food systems (feusthuber et al., ; harrison et al., ; hauck et al., ; holman et al., ; mitter et al., a; popp et al., ; schönhart et al., ) and can stimulate and guide research, public debate, education and communication (le mouël et al., ; Öborn et al., ; veland et al., ; vervoort et al., ) . they can inform strategic planning of private investments or designing of policies (butler et al., ; de lattre-gasquet, ; frame et al., ; harrison et al., ; mitter et al., b; rounsevell and metzger, ) and may thus help to steer towards a sustainable and resilient future. to successfully serve these purposes, a transparent and systematic scenario development process is key kunseler et al., ) . multi-perspective dialogue (borch and mérida, ) and stakeholder engagement strengthen scenarios to address the expertise and various needs of researchers, policy and decision makers in the public, private and educational sectors. finally, considering alternative futures makes planning more resilient to unexpected shocks and disruptions. in climate change research, scenarios are crucial for exchanging information across and within scientific communities , and for comparing research and modeling results across temporal and spatial scales. in recent years, a new global scenarios concept, shared socioeconomic pathways (ssps, o'neill et al., has been widely adopted by the climate change research community. these characterize five socio-economic pathways that encompass contrasting challenges related to climate change mitigation and adaptation and thus cover a large uncertainty space. the ssps provide qualitative descriptions as well as model-based quantifications of plausible global developments of socio-economic conditions including population growth, demographic change, urbanization, economic development and technological progress (see riahi et al., for an overview of the development of the ssps and the ssp database https://tntcat.iiasa.ac.at/sspdb for extended reporting of existing ssp scenarios). o'neill et al. ( ) and van ruijven et al. ( ) differentiate between 'basic' and 'extended ssps'. the former shall provide sufficient information to outline alternative future development pathways, while the latter build on the basic ssps to ensure consistency between scales or sectors and provide more details for sectoral or regional applications. diverse methodological approaches have been applied to develop extended ssps at the global and regional scales and with different thematic foci and time scales. at the global scale, the agrimonde-terra scenarios couple a qualitative approach based on morphological analysis (ritchey, ) and a quantitative modeling approach to explore land use and food security in (mora and de lattre-gasquet, ; mora et al., ) . similarly, lenzner et al. ( ) propose to combine stakeholder engagement and numerical models to derive scenarios on biological invasions for the st century. the diet-ssps are qualitative storylines that elaborate on food systems for diet, nutrition and health until . participatory approaches have been used to develop long-term global oceanic system pathways (maury et al., ) , and mathematical models have been applied to various sectors (e.g., energy and land use) to quantify the development of specific elements of the global ssps until popp et al., ; riahi et al., ; van vuuren et al., ) . at regional scales, extensions include qualitative storylines for europe until (eur-ssps; kok et al., ) ; representative agricultural pathway narratives (raps) for the u.s. dryland wheat-based systems until mu et al., ) and for kenya until (claessens et al., ) ; as well as quantified scenarios on agriculture and food security for west africa until (palazzo et al., ) . other examples include those for europe and central asia with a focus on biodiversity and ecosystem services , for europe's urban land use change (terama et al., ) , for new zealand to inform policy and decision making (frame et al., ) , for the barents region (nilsson et al., ) and for the u.s. southeast for adaptation planning (absar and preston, ) , for the mediterranean coastal zone with a focus on population projections (reimann et al., ) , for the baltic sea region and its environmental problems (zandersen et al., ) , and for a river delta in china to manage regional water use (yao et al., ) . we refer the reader to the iconics (international committee on new integrated climate change assessment scenarios) database for an extensive overview of publications related to the ssp framework and to the ssps (https://depts. washington.edu/iconics/publications/) as well as to the fore-sight food international collaborative initiative (https://www. foresight food.net/) and its platform for foresight activities related to agriculture and food systems. these regional extensions have added to the wealth of scenarios and to the methods used to develop them. yet, conceptual and methodological transparency and rigor lags behind and scenario development has often been criticized for insufficient scientific neutrality and replicability (beck and mahony, ; carlsen et al., ) including unsystematic stakeholder selection and engagement (reed et al., ) . we augment the existing set of ssps by systematically developing five storylines for european agriculture and food systems (eur-agri-ssps) following a detailed and stakeholder inclusive step-by-step protocol . the eur-agri-ssps describe alternative plausible future developments for the european agriculture and food systems advanced along the challenges to climate change mitigation and adaptation until . they aim to capture uncertainties in major socio-economic, environmental and technological drivers and include information on population and urbanization, economic development, policies and institutions, environment and natural resources. such information is useful for researchers performing integrated assessments of climate change challenges in agriculture and food systems at the european scale and under different socio-environmental developments. furthermore, the eur-agri-ssps may inform public policy and private business decision making. the article is structured as follows: in section , we describe how the collected data were analyzed by following the nine transparent and stakeholder inclusive working steps of the mitter et al. ( ) protocol; in section , we provide the main outcomes of the multi-year, participatory scenario development process; in section , we discuss application potentials of the eur-agri-ssps and major challenges encountered in the development process; and in section we conclude with lessons learned. we develop eur-agri-ssps in order to advance research on european agriculture and food systems in a transparent and systematic way. to extend the basic ssps, a consecutive, one-to-one nesting approach is chosen, i.e., the finalized basic ssps set the boundary conditions. in the following, we describe the methods applied to analyze the compiled data and resources. additional information and material for specific working steps is provided in the supplementary material (sm ii). the protocol for developing the eur-agri-ssps transparently defines nine working steps, specifies responsibilities of three multi-disciplinary scenario working groups (i.e., core, supporting, and stakeholder groups), and proposes methods as well as opportunities for stakeholder engagement . fig. summarizes how the protocol was applied to develop the eur-agri-ssps, whereby the arrows indicate the iterative character of the working steps. in our case, the core group comprises six scientists from three european research institutions, and the supporting group is composed of scientists from across europe. the stakeholder group is represented in the second step of the protocol and respective affiliations are listed in table sm ii. . using the protocol enables meeting pre-defined quality criteria such that the storylines are plausible, internally (i.e., horizontally) consistent and consistent across spatial scales (i.e., vertically consistent), salient or relevant for targeted users, legitimate in that they consider diverse interests, rich in detail, and creative or widely diverse. the goal and purpose, target groups, thematic focus, spatial and time scale of the storylines were agreed upon at the beginning of the process in order to guide the storyline development. the core and supporting groups defined these key characteristics of the eur-agri-ssps in moderated group discussions. the virtual and face-to-face discussions were supplemented by personal and digital exchanges regarding specific topics. engaging a broad variety of stakeholders in a storyline development process can increase creativity, salience, richness, and horizontal consistency . furthermore, it may reduce unintended bias from diverse personal backgrounds, interests and professional knowledge (ernst et al., ) . accordingly, the main criteria for stakeholder identification and selection defined by the core group were to cover a wide spectrum of knowledge types, roles in policy and decision making at the european and national scales, topical expertise, academic and non-academic backgrounds, geographic diversity, cultures, gender and age. diversity was achieved by including people from the public and private sectors, academia, advocacy groups and associations as well as inter-governmental and non-profit organizations working at european and national scales. the core and supporting group members performed a literature and online search for relevant stakeholders and reached out to their professional networks. in addition, the identified stakeholders suggested contacts to other relevant or potentially interested persons (snowball sampling approach). the storylines should address clearly defined elements that are particularly relevant and uncertain (alcamo and henrichs, ) for the future development of the european agriculture and food systems. the core and supporting groups applied different methods to identify, cluster and prioritize storyline elements. the core group conducted a literature review in order to specify the boundary conditions set by the fig. . overview of the research process based on the nine working steps defined in the protocol by . notes: for each working step (grey rectangle), the scenario working groups involved (green circles), the applied methods and the timing are given. the arrows indicate that the research process was iterative, i.e., some working steps were repeated until the final eur-agri-ssps were developed. (for interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.) basic ssps and the eur-ssps to determine the central elements of agriculture and food systems and to prepare subsequent stakeholder interactions. stakeholder engagement is key in this working step because it stimulates discussions and encourages non-linear and out-ofthe-box thinking. stakeholders were engaged in three workshops organized by the core group and via semi-structured interviews conducted by the members of the core and supporting groups. stakeholders were asked to describe drivers of future development in the agriculture and food systems, to rank these drivers by importance and uncertainty, and to estimate plausible directions of change over the next three decades, including 'extreme' developments and 'surprising' outcomes in order to derive a gradient of expressions for each driver (wright et al., ) . the core group was responsible for analyzing the qualitative data compiled during the workshops and semi-structured interviews, clustering the storyline elements, and summarizing potential directions of change. data analysis was supported by the qualitative data analysis tool atlas.ti. a prioritization for the storyline elements was guided by the objective to avoid potential conflicts with other spatial and (sub-) sectoral storyline extensions as well as with quantitative model outputs. guidance for differentiating between typical inputs and outputs of agriculture and food systems models was provided by the core and supporting group members' expertise and from the literature. the core group drafted the eur-agri-ssps by linking the identified directions of change from each storyline element under the framework of the ssps. while the stakeholders were explicitly asked for socioeconomic, environmental and technological drivers of european agriculture and food systems until and the ranges of change for the drivers, the core group's task was to combine the storyline elements and determine the directions of change for each element and eur-agri-ssp. similar to many previous scenario exercises (abildtrup et al., ; absar and preston, ; antle et al., ; busch, ; palazzo et al., ; reimann et al., ; valdivia et al., ; vervoort et al., ) , we differentiate increasing, decreasing and stable developments, compared to the initial conditions in the starting year. this implies that directions of change for individual storyline elements can be the same for two or more storylines even if the underlying development patterns differ. the core and supporting groups agreed on the directions of change in an iterative process. related storyline elements were identified in causal loop diagrams (mathijs et al., ) and the core group established development paths for each storyline, based on the ssps (narratives and quantifications), alternative scenarios for agriculture and food systems, and theoretical considerations. changes proposed by the supporting group members were considered if their argumentation was considered theoretically consistent and reasonable. consistency checks are important in order to increase robustness (priess and hauck, ; priess et al., ) because larger scale storylines set boundary conditions for smaller scale storylines (zurek and henrichs, ) and individual storyline elements influence each other (schweizer and o'neill, ) . we differentiate between vertical (with the global ssps) and horizontal consistency (internal or selfconsistency; weimer-jehle, ). while the core and supporting groups reviewed both vertical and horizontal consistency, stakeholders were asked to focus on horizontal consistency according to their specific expertise. systematic consistency checks required three iterative working steps. first, each eur-agri-ssp was cross-checked on the respective ssp narrative and related quantified storyline elements for vertical consistency. second, scientific theory (e.g., economic and behavioral theory), causal loop diagrams (mathijs et al., ) and intuitive logics (bradfield et al., ; wright et al., ) helped to understand causal processes, to identify relationships between storyline elements and to achieve horizontal consistency. experiences of agriculture and food systems modeling from the supporting group supported this step. third, the eur-agri-ssps were compared against each other to ensure contrasts while maintaining consistency. consistency checks were repeated several times and partly combined with peer and stakeholder reviews (section . . ). the core group updated the draft storylines if causality was not entirely clear or deviated from established scientific theories. contradictory comments were resolved in the core group via majority decisions. the core group developed visual and tabular presentation formats of the eur-agri-ssps to communicate the results to the supporting group and obtain feedback, as well as to increase their effectiveness for stakeholders for decision making purposes (padilla et al., ; tufte, ) . the tables provide a hierarchically structured overview of the storyline elements, i.e., each element is attributed to a topic. during a workshop, stakeholders were invited to reflect on the titles and pictures they had in mind for specific storylines in a stimulating environment. based on the collected ideas, three sets of titles were developed and the supporting group members agreed on one set by a majority vote. furthermore, the core group established the website https://eur-agri-ssps. boku.ac.at as a platform for sharing information, communication, education and interaction, and to serve as a freely accessible forum for the storylines to be openly discussed and commented. the core, supporting and stakeholder groups were involved in the review and revision processes which focused on the quality criteria as defined in the protocol, i.e., plausibility, consistency, salience, legitimacy, richness and creativity . overlaps between the broad review process and the more focused consistency checks arose (section . . ). four major review and revision rounds were necessary until no new comments came up and agreement on the final storylines could be achieved. the first storyline drafts were reviewed with written feedback by the core and supporting groups. according to the quality criteria, we specified six review tasks, ensuring that each task was covered by several group members. the revised version was discussed by the supporting and stakeholder groups in a two-day workshop. the workshop participants were purposefully assigned to review groups. each group evaluated one storyline by answering six questions referring to the quality criteria. in addition to the moderated group discussions, review sheets were distributed during the workshop and the participants were invited to write down their feedback. in the third revision round, the core and selected supporting group members provided written and oral feedback with a particular focus on clarity, comprehensibility and consistency. the feedback was incorporated into the storylines, and the draft eur-agri-ssps (including their summaries) were distributed amongst the core, supporting and stakeholder groups for a final revision. effective communication and different forms of dissemination for the storylines are important in order to reach potential users (van vuuren et al., ) and to make the storylines accessible to researchers, policy and decision makers, students and the interested public with varying (cross-) sectoral and (cross-) scale interests and perspectives. the core, supporting and stakeholder groups were involved in this step to increase the diversity of information, communication and dissemination channels. while the core group developed various dissemination formats, the supporting and stakeholder groups provided comments and feedback to increase the usefulness of the products. dissemination formats and channels were customized to the targeted users and to the purpose of the dissemination activity. evaluation is crucial to allow for methodological improvements and h. mitter, et al. global environmental change ( ) interpretation of scenario process and products. feedback on the storyline development process was collected after formal interactions with stakeholders in order to improve working relationships, increase productivity and enhance performance (see priess et al., ) . the evaluation methods were adjusted to the eur-agri-ssp setting, e.g., rating questions related to the content, targets, methods and interaction during workshops or stakeholder engagement in developing eur-agri-ssps in general. the development of the eur-agri-ssps was informed by the basic ssps and other scenario related exercises as well as by knowledge, expertise and expectations of a diverse group of researchers and stakeholders. the ssp literature was reviewed to specify boundary conditions for the eur-agri-ssps. major topics (e.g., demographics) and individual storyline elements (e.g., population size) were listed and storyline elements with similar meanings were summarized with one term. the major topics and storyline elements served as a starting point for analyzing the semi-structured stakeholder interviews, which may be interpreted as a deductive approach. future dynamics and development paths of selected storyline elements have also been based on the ssps, i.e., narratives and quantifications of plausible developments. in addition to the ssps, recently published scenarios related to agriculture and food systems and with a time horizon similar to the eur-agri-ssps were identified. the major purpose of scanning scenario exercises was to check for potential overlaps (e.g., storyline elements) and to build on existing knowledge of causalities and the exploration of trends. grey literature from relevant european and national organizations and institutions that provide future visions of agriculture and food systems and related sectors was primarily used to prepare subsequent stakeholder engagement activities. relevant sources and projects consulted in this research step are listed in table . stakeholders provided fresh inputs and creative ideas regarding potential future developments. stakeholder engagement was dynamic and depended on the aim of the respective working step and therefore it varied over time. qualitative data were a major input for specifying the storyline elements and development paths that are characteristic of the agriculture and food systems. in total, individuals from national and european organizations and institutions contributed to the eur-agri-ssps with their knowledge, expertise and expectations (table sm ii. ). data collection was organized through four workshops and semi-structured interviews. the aims of the activities are briefly described and the core workshop and interview material is available in sm ii. the eur-agri-ssps were developed using an iterative and participatory process. below we present major final outputs from the multiyear process. we refer the reader to sm ii for intermediate steps and results that highlight consensus and controversies. the major purpose of this scenario exercise was to develop five sector-specific socio-economic storylines for europe that capture the uncertainties related to the challenges to climate change mitigation and adaptation. the problem-focused storylines aim to describe alternative plausible pathways for the european agriculture and food systems until . potential users of the storylines are researchers working on climate change in agriculture and food systems as well as policy and decision makers from the public and private sectors. these framing conditions determine the key characteristics of the eur-agri-ssps as summarized in table . the eur-agri-ssps describe plausible changes in key socio-economic, environmental and technological drivers that affect climate change mitigation or adaptation options as well as sustainability levels in european agriculture and food systems. they bundle the diverse perspectives of different stakeholders into internally consistent development pathways, the effects of which (e.g., on land use change) can then be modeled quantitatively. they do not provide national details, nor development trajectories in related sectors, such as forestry. the eur-agri-ssps extend and enrich the basic ssps with a regional and sectoral component and mirror the structure of the ssps for reasons of vertical consistency (fig. ) . the storylines touch upon five major topics: population and urbanization; economy; policies and institutions; technology; environment and natural resources. summaries of the storylines are presented below. the comprehensive storylines are provided in sm i. in europe, social and environmental awareness increase steadily and significantly and are reflected by increasingly effective cooperation between public and private sectors and civil society. this is accompanied by tightened pro-environmental policies; abolished income support for farmers; rising public payments for the provision of regulation and cultural services; taxes on environmental damages; decreasing resource depletion; and technology developments towards low emissions, resource use efficiency and chemical pesticide-free agriculture. european domestic demand shifts towards plant-based diets h. mitter, et al. global environmental change ( ) and bio-based materials, whereas food waste and per capita demand for livestock-based products decrease gradually. markets are globally connected and trade agreements are strengthened. international trade decreases because short and transparent agricultural supply chains are preferred by consumers and external costs, e.g., for transportation, are internalized in agricultural commodity prices. challenges to climate change mitigation are low in the agriculture and food systems because of a decreasing demand for livestock-based products and technology development with a focus on reducing greenhouse gas emissions. challenges to climate change adaptation are also low because of increasing institutional effectiveness focusing on support for sustainable farming, public investments in infrastructure development, and cooperation along the agricultural supply chain. european development follows historical patterns resulting in slow but steady social, environmental and technological progress. cooperation between public and private sectors remains challenging with little progress in implementing further environmental standards and policy instruments. european agricultural policy is characterized by multiple support schemes to increase international competitiveness, productivity and efficiency, and improve environmental status. national policies complement the european policy framework. agricultural commodities are mostly traded within europe even though global market integration advances. demand for locally produced food, bio-based materials, regulation services and landscape amenities increases slowly, whereas per capita meat demand remains high. depletion of natural resources increases because of continuous growth of the agriculture and food economy, and pro-environmental regulations and resource-efficient technologies are only developed at a moderate pace. challenges to climate change mitigation are moderate because agriculture and food systems partly remain dependent on fossil energy sources, resource-efficient technology development is advancing gradually but is not expedited, and pro-environmental policy instruments remain inefficient. challenges to climate change adaptation in agriculture are moderate because of slow and insufficient development of european agricultural policy, and reduced investments in infrastructure in rural areas. mutual distrust and regional rivalry result in less efficient cooperation between national and european entities, more severe european and international trade restrictions, the emergence of national agricultural policies, and relaxed environmental standards. increased self-sufficiency concerns of individual countries influence demand patterns for food, feed and agro-fuels. public payments aim to maintain the national production potential, whereas demand for environmental services declines. access to international markets decreases, whereas neo-colonialism and land grabbing gain in importance. market concentration increases within countries and national governments keep agricultural production standards low. technology development and diffusion suffer from declining public and private investments and weak cooperation between governments and actors in the agricultural supply chains. challenges to climate change mitigation are high because of a lack of cooperation between the public and private sectors, decreasing environmental awareness, reduced public payments for environmental services, and slow technological progress that focuses on productivity instead of greenhouse gas emission reduction targets. challenges to climate change adaptation are high due to a combination of decreasing institutional effectiveness, reduced diffusion of new technologies between nations, tighter budget constraints, a dominance of national agricultural policies, and decreasing investments in infrastructure in urban and rural areas. increasing social disparities between and within rural and urban areas lead to social segregation. a business-oriented, wealthy upper class dominates european institutions, sets the policy agenda, controls agricultural supply chains, and stimulates technological uptake of energy efficient and renewable energy technologies, whereas the lower class majority are poorly represented in european institutions and agricultural supply chains. european agricultural policies increasingly support economic growth and technology development, from which the large, industrialized farms benefit the most and the interests of a large proportion of society are mostly ignored. agricultural markets are increasingly globally connected and demand for european agro-food products is increasingly diverse, with a stagnation in domestic demand table key characteristics of the eur-agri-ssps defined by the core and supporting groups. goal and purpose extend and enrich the ssps by providing a regional (europe) and a sectoral component (agriculture and food) in a systematic way provide a set of alternative future developments of the european agriculture and food systems provide a set of plausible storylines capturing future key uncertainties provide a solid basis for integrated assessments of agriculture and food systems to increase comparability of their results stimulate discussion between various actors with different interests, backgrounds and professional activities main target groups scientists from the climate change, agricultural, food and integrated assessment research communities working at european to national scales policy makers in european agriculture and food systems decision makers in the private sector (e.g., supply chain managers) thematic focus describe worlds in which socio-economic, environmental and technological drivers make it harder or easier to mitigate or adapt to climate change in agriculture and food systems or to tackle other sustainability issues spatial scale europe, with differentiations between nations or agri-environmental zones kept to a minimum time scale with optional extension to scenario type qualitative storylines, semi-quantitative specifications of trends, problem-focused quality criteria plausibility, consistency (vertical and horizontal), salience, legitimacy, richness, creativity h. mitter, et al. global environmental change ( ) for feed and non-food commodities. environmental standards decrease except for selected, scenic, hot spot regions. natural resources are increasingly overused. challenges to climate change mitigation are low in the agriculture and food systems because of effective european institutions, progress in technology development (towards low emissions and nitrogen efficiency), and rising public payments for technology penetration. challenges to climate change adaptation are high because of growing inequalities in access to institutions and production-related support, to globally connected markets, and to education. european residents share a growing faith in technology, materialintensive lifestyles and trade liberalization, which is reflected by improved international trade agreements, globally connected agricultural supply chains, accelerated technological progress and diffusion in the agriculture and food systems, and expedited structural change. individuals' affinity for technological innovation also affects increasing global demands for european agricultural products, particularly for biobased industrial raw materials as well as bio-based and innovative products. increased private investments in technological know-how and the education of employees in the agriculture and food systems boost economic growth, which is largely dependent on fossil energy sources. public payments to the agriculture and food systems are drastically reduced to conform with liberalized and integrated markets. environmental standards are considerably lowered, which results in overexploitation of natural resources in europe and abroad. challenges to climate change mitigation are high in the agriculture and food systems mostly because of decreasing environmental awareness, massively reduced payments for environmental services, and a growing reliance on fossil energy sources and other fossil-based inputs. challenges to climate change adaptation are low because of increasing investments in social and technical infrastructure, higher economic growth rates and professionalization in the agriculture and food systems. the storyline elements form an important basis for developing the comprehensive eur-agri-ssps. table gives an overview of the storyline elements and summarizes the directions of change for the five eur-agri-ssps. it is structured around the five major topics population and urbanization; economy; policies and institutions; technology; environment and natural resources. we developed five semi-quantitative scenarios for the european agriculture and food systems, called the eur-agri-ssps and extended the basic ssps in a structured and participatory process. however, we also experienced several challenges during the storyline development process and recognized limitations of the storylines. in this section, we first highlight application potentials of the eur-agri-ssps and then discuss experienced challenges clustered around five major topics, (i) extension of the basic ssps, (ii) participatory storyline development, (iii) interdisciplinary cooperation for storyline development, (iv) consecutive, one-to-one nesting approach, and (v) science-driven storyline development. where appropriate, we refer to the related quality criteria of plausibility, consistency, salience, legitimacy, richness and creativity as defined in . in sm ii, we provide additional material and intermediate results in order to increase process transparency. based on the feedback from the core, supporting and stakeholder groups, we have identified four major fields of application of the eur-agri-ssps, i.e., research, policy design and implementation, private decision making, and education. scientists have a strong interest to use the storylines as an input for integrated assessments of agriculture and food systems at the european scale (e.g., to inform modeling assumptions) and to expand the storylines for regional or sub-sectoral applications (e.g., kampermann, ) . as such, the storyline development process has already affected the research design of on-going projects (e.g., the bonares, dakis, salbes, and surefarm research projects). furthermore, the eur-agri-ssps can stimulate new research, and can serve as a reference for further scenario and strategic foresight studies as well as for the design of research programs, such as horizon europe, the next research and innovation framework program of the european commission, starting in . with respect to policy design and implementation, the stakeholder group highlighted the eur-agri-ssps as a potential planning tool for policies, especially in the context of rural and agriculturally dominated areas, nature-based solutions, land management, ecosystem services, bio-economy, international trade and the european single market as well as the sustainable development goals. furthermore, members from the stakeholder group referred to the usefulness of the storylines for explaining the role of the common agricultural policy, for comparing and informing national agricultural strategies, and for focusing ideas on historical and preferable future developments in agriculture and food systems. the eur-agri-ssps may inform the strategic orientation of private companies and may thus inform decision making. in education, the storyline development process is considered suitable for introducing students to foresight and scenario approaches. the storylines can be used to stimulate students' discussions on different, plausible futures. the basic, global ssps served as a starting point for developing the eur-agri-ssps. this basis was enriched by the eur-ssps, which are equivalent to the global ssps and thus provide a regional interpretation. we summarize similarities and differences between the basic, global ssps, the eur-ssps and the eur-agri-ssps in table . the applied ssp framework, which highlights challenges to climate change mitigation and adaptation, as well as ssp elements clearly influenced the specification of the eur-agri-ssps. however, differences emerged mostly because of the stakeholder engagement. for instance, the stakeholders emphasized the importance of future developments of the agriculture and food markets and of european agriculture, food and environmental policies. the economy, policies and institutions are thus given more weight in the eur-agri-ssps, compared to the basic ssps. stakeholders also articulated the risk of a scenario with 'medium' change, that is most often preferred by decision makers and represented by the ssp pathway, because potential for changes would be ignored. some even recommended to skip developing a eur-agri-ssp altogether. however, the core and supporting groups decided to deviate from the basic ssp pathway to develop selected agriculture-specific storyline elements that were defined as 'increasing' or 'decreasing' in the eur-agri-ssps, compared to the 'medium' changes of most storyline elements in the basic ssp . while vertical consistency was not jeopardized, this deviation increases the attractiveness of the eur-agri-ssp (agriculture on established paths) for stakeholders and at the same time enabled it to be modeled quantitatively in the scenario set. we have devoted considerable effort to stakeholder engagement during the storyline development process in order to acknowledge heterogeneous perspectives (i.e., legitimacy) and increase the comprehensiveness (i.e., richness), which are documented in sm ii. however, some challenges remain which could -to an extent -limit legitimacy, consistency and creativity of the storylines. first, the full range of actor perspectives in agriculture and food systems has most likely not been considered. even though we carefully h. mitter, et al. global environmental change ( ) 'neill et al., ) or the eur-ssps . arrows indicate directions of change, compared to the initial situation at a certain point in time, i.e., increasing (↗), stable (→), and decreasing (↘) developments. selected and invited a diverse group of stakeholders to engage in this exercise, we recognize that the perspectives of those who lacked the resources, or who were unwilling to participate in the multi-step process are missing. similarly, flick ( ) points out that 'good' cooperative stakeholders do not only possess the necessary knowledge and expertise, but are also capable of reflecting and articulating their perspectives, and can take the time to engage. we experienced a particular challenge in engaging stakeholders from organizations and institutions in east european countries (see table sm ii. ) . however, about one third of the involved organizations and institutions represent european interests and employ people originating from across europe. representatives of some companies may not be willing to discuss plans on future developments very openly with policy-makers, their competitors or clients. we have combined individual and group interactions in order to elicit stakeholders' long-term views on future developments, including sensitive topics. moreover, we have treated stakeholders' contributions as neutrally as possible in order to balance the dominant views and achieve a broad acceptance of the storylines by considering a plurality of futures. while open and unbiased interviews and workshop discussions were encouraged through the choice of neutral venues and the engagement of responsible scientists, professional facilitation of our meetings (with a particularly diverse and large audience) could have strengthened the coordination and alignment of actors' input in various future contexts. the requirement for nonjudgmental scientists who are able to switch between reflection, facilitation and intermediation has been highlighted in the process of participatory backcasting (sandström et al., ) and sustainability research (pohl et al., ) . second, interdisciplinary and participatory storyline development is often related to the 'intuitive logics model' (bradfield et al., ) which is widely applied (rounsevell and metzger, ) to analyze relationships between critical uncertainties, predetermined trends, and the behavior of actors (wright et al., ) , but is criticized for its limited transparency and reproducibility . we have followed suggestions by wright et al. ( ) for structuring and documenting the stakeholder interviews in order to augment the intuitive logics model and to ensure that major conclusions are robust and independent of the scientists and stakeholders involved. however, this process implicates normative judgements of scientists and stakeholders alike, meaning that another initiative to extend the ssps for european agriculture and food systems would likely result in different details within the storylines. however, major pathways and storyline elements would likely not deviate strongly due to the boundary conditions set by the ssps (vertical consistency) and the targeted horizontal consistency. third, and similar to frame et al. ( ) and kunseler et al. ( ) , we faced challenges to reconcile the abundance of stakeholder contributions on storyline elements and potential directions of change that did not always align. furthermore, stakeholders tended to address specific aspects of single storylines and found it more difficult to respond to the larger picture. rounsevell and metzger ( ) argue that differences in epistemologies or knowledge systems and incomplete mental models of the system in question may result in such conflicting stakeholder perceptions. even though we confronted the stakeholders with the ssp logic and the main ideas of each ssp, incomplete knowledge of the ssps further complicates stakeholders' inputs. we have based the storyline assumptions on scientific theory and have used causal loop diagrams (mathijs et al., and fig. sm ii. ) in order to handle stakeholders' conflicting perceptions on future development. furthermore, we have distributed the previous versions of the eur-agri-ssps, including summaries, to the supporting and stakeholder groups for feedback in order to achieve agreement on the final version (see sm ii). well-informed key stakeholders were able to identify caveats, logical flaws and weak points in the storylines which were clarified in a revision round in order to increase the usability of the final set of storylines. cooperation among scientists with complementary disciplinary backgrounds, in-depth methodological knowledge as well as inter-and trans-disciplinary expertise proved to be vital for systematic and scientifically credible storyline development, in particular for describing the agriculture and food systems and how they may develop under various future contexts. despite any formal agreement, our cooperation was effective throughout the storyline development process, which is mostly due to the common interest of the engaged scientists in the table relations between the basic, global ssps, the eur-ssps, and the eur-agri-ssps. kok et al., ; nakicenovic et al., ; gramberger et al., ; kok et al., ; pedde, , riahi et al., . application of the eur-agri-ssps for european modeling activities, regional or sub-sectoral extensions. strong leadership and continuous exchange of information are also of high relevance to keep an informal research consortium active. however, the iterative development of the storylines also required repeated review tasks in order to ensure their plausibility, richness, consistency and salience, which tied up resources. developing storylines that are vertically (i.e., with the global ssps) and horizontally (i.e., internally) consistent is key, but is also a considerable challenge. we have followed a consecutive, one-to-one nesting approach in order to ensure high vertical consistency and to limit the storylines to a manageable number (absar and preston, ; zurek and henrichs, ) . with consecutive development and one-toone nesting, the ssps serve as a starting point and represent boundary conditions for the eur-agri-ssps. however, applying the ssp logic and storyline elements to european agriculture and food systems also implies disadvantages for creativity and salience. previous exercises on participatory, nested storyline development confirm that stakeholders feel constrained by the framing conditions of storylines from predefined larger spatial scales (e.g., hagemann et al., ) . the consecutive, one-to-one nesting approach implicitly ensures that the european agriculture and food systems evolve in concert with global trajectories and that each global ssp manifests itself into a single eur-agri-ssp. this link affects the definition of storyline elements as well as the narrative logic. it constrains potential future developments and may thus discourage visionary thinking. however, the stakeholders were satisfied with the room to maneuver offered by the ssp framework and only mentioned limited room for creativity in the context of eur-agri-ssp (agriculture on established paths). in any case, unexpected shocks and disruptions to the agriculture and food systems, such as the exceptional increase in oil prices on the eve of the financial crisis or the outbreak of the covid- pandemic in / remain largely unreflected in the approach. such events are often referred to as 'wild cards' or 'surprises' in the foresight literature (van notten et al., ) and could occur under any of the eur-agri-ssps. however, the consistent integration of the systems' key drivers can substantially facilitate the analysis of possible long term effects of such external shocks and disruptions. potential users' interests may diverge from the ssp rationale, which may weaken the relevance (salience) of the eur-agri-ssps. with one-toone nesting, we implicitly assume that all ssps are plausible for the future of european agriculture and food systems, and that one eur-agri-ssp covers only one alternative global pathway. however, consecutive, one-to-one nesting has proven expedient for extending and enriching the ssps because the ssps are sufficiently disparate (absar and preston, ) . furthermore, the ssp rationale helps facilitators to manage strategic interventions or behaviors of stakeholders. the delineation of eur-agri-ssp (agriculture on unequal paths) was most challenging because the focus of the global ssp on inequality required to first define whether and how european agriculture and food systems were divided, and then to establish the differences from the other storylines. diversity of european agriculture and food systems and its consideration in the storylines was extensively discussed. for instance, a stakeholder suggested differentiating between western, southern and eastern europe, and some modelers stressed that regional heterogeneity can only partly be addressed in current model setups. finally, the core and supporting groups decided not to provide details on diversity of the agriculture and food systems within europe in order to leave room for eur-agri-ssp extensions and interpretation at national, sub-national and sub-sectoral scales. time mismatches between the ssps and the eur-agri-ssps are an additional potential challenge in a consecutive, one-to-one nesting approach (kok et al., ) . while the ssps describe futures for the st century, the eur-agri-ssps deal with the next three decades, i.e., until . the shorter period was chosen because of stakeholders' preferences for a time horizon consistent with their own actions and related effects and because it corresponds to previous storyline and scenario studies (e.g., gramberger et al., ; mylona et al., ; vervoort et al., ) . however, when using the ssp narratives, this problem does not materialise to its full extent. the ssps were presented as 'pathways in the st century' (which can be interpreted as protoscenarios, see table ) and o'neill et al. ( ) do not refer to a specific end point. not until the ssps are elaborated to full scenarios, do they have a specific time horizon. thus, the intentionally wide coverage of the ssps and their trends are applicable throughout the st century which allows for regional and sectoral extensions with a shorter time horizon (rohat et al., ) , as successfully demonstrated by, e.g., palazzo et al. ( ) . we do acknowledge that the eur-agri-ssps may have introduced developments that might be inconsistent with scenarios based on the ssps developed for . for improved process transparency and because of the particular interest in applying the eur-agri-ssps to research, we have chosen to follow a science-driven approach adhering to a protocol. accordingly, we chose 'scientists from the climate change, agricultural, food and integrated assessment research communities' as a main target group (section . ). 'policy makers in european agriculture and food systems' and 'decision makers in the private sector' were added as main target groups because they may use the eur-agri-ssps for designing policy or business strategies. therefore, stakeholder activities provided essential inputs for the storyline development process. however, defining three main target groups brings about challenges because of their partly different interests and needs, such as on the required level of detail of the storylines. the scientists expressed two concerns, on (i) the level of integration of storylines and quantitative model-based integrated assessments of agriculture and food systems, and on (ii) the translation of directions of change for quantitative model-based integrated assessments of agriculture and food systems. concern (i) refers to the extent to which the quantitative models can account for the storyline elements, and concern (ii) is about putting numbers on the directions of change, which can then serve as inputs for quantitative models. for both concerns, the suggested level of detail in the eur-agri-ssps is closely related to their application in integrated assessments of agriculture and food systems. thus, the storylines should provide sufficient detail for model parametrization and, at the same time, leave room for the modelers to implement in their specific study. in any case, it is advisable to document any remaining divergences. policy and decision makers emphasized their interests in which interventions particular developments will be stimulated or slowed down and therefore preferred a comprehensive description of storyline elements related to policies and the economy. this preference combined with modelers' needs for policy and economy specifications, resulted in an emphasis on policies and, in particular, on the economy. similarly, o'brien ( ) demonstrates that economic factors are often predominant in storylines and scenarios. science-driven, interdisciplinary cooperation and participatory engagement throughout our research process helped to diversify storyline elements, to agree on the final storylines, and also to balance the diverging demands. the eur-agri-ssps describe plausible but contrasting futures of the european agriculture and food systems, characterized by a number of socio-economic, environmental and technological drivers and their interactions, which have been identified as particularly important and uncertain for the european agriculture and food systems. these drivers are based on the ssps and eur-ssps but were strongly refined in a participatory process to satisfy researchers' and stakeholders' demands. both scenario methodological rigor and thematic depth facilitate a broad range of potential applications, including integrated assessments of agriculture and food systems, and policy and decision making. for their application in models, the eur-agri-ssps need to be translated into a set of quantitative assumptions that are required to run numerical models. further extensions of the eur-agri-ssps would be useful to address national and sub-sectoral futures. we suggest to follow the nine working steps in the protocol in order to maintain the high level of transparency and reproducibility and to adjust the methods and the level of stakeholder engagement to the respective needs. if resources are limited, we recommend to focus on defining storyline elements and consistency checks. from our experience, they are critical for the credibility and wide acceptance of the final storylines. however, protocol-based storyline development does not prevent researchers from taking reasoned decisions where stakeholders disagree or raise arguments that are contradictory to the underlying ssps. the authors would appreciate reports on any application of the eur-agri-ssps as well as comments on perceived contradictions to enrich the database at https://eur-agri-ssps.boku.ac.at for methodological learning and resource sharing. 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article. supplementary data to this article can be found online at https:// doi.org/ . /j.gloenvcha. . . key: cord- -dd z stt authors: giosa, penelope title: exploitative pricing in the time of coronavirus—the response of eu competition law and the prospect of price regulation date: - - journal: nan doi: . /jeclap/lpaa sha: doc_id: cord_uid: dd z stt nan • according to abg oil case, it can be argued that the firms selling goods in high demand due to the coronavirus outbreak have 'transitory market power' and so they hold a temporary position of market strength. • the issuance of 'commitment decisions' by the european commission is preferable to the imposition of fines to firms, because it could reset prices to a non-excessive level, rather than merely alleviating the harmful effects of excessive pricing. • price regulation is a much-disputed enterprise in contemporary regulatory practice, but in the absence of self-correcting excessive prices, the pragmatism calls for it. sellers who heavily increase prices on key coronavirus prevention supplies, if price gouging continues. however, price controls are 'one of the most intrusive forms of intervention in the market' and for this reason, they are heavily criticised, as they may undermine the institutional and ideological structure of the eu, which is built upon the idea of open and undistorted competition within the internal market. therefore, outside the merger context, antitrust enforcers in the eu proceed against excessive prices only when such activity violates the discrete prohibitions of articles and treaty on the functioning of the european union (tfeu). the european commission has stated it is bound to come across cases where 'dominant businesses are exploiting their customers, by charging excessive prices or imposing unfair terms' . in the same vein, national competition authorities (ncas) have sanctioned exploitative abuses of dominance, especially in network industries recently liberalised. however, what happens if the price gouging is genuinely unilateral and not collusive, due to an abnormal level of demand, in terms of both the number of consumers who desire the item and the sense of urgency that increases that desire? what happens if there is not only one dominant company in the market that sets excessive prices on goods and services in high demand? this article explores the provision of the eu competition law that the ncas and the european commission could possibly enforce against excessive pricing, in case the price gouging is not the outcome of a collusive agreement. it discusses the practical and conceptual difficulties that excessive pricing cases involve, especially under the current urgent circumstances that coronavirus has caused, and it assesses the remedies that the ncas and the european commission can impose in order to deal with price gouging. the article also recommends specific remedies that would be suitable to reset prices to a nonexcessive level in the time of coronavirus. in this context, it will also be discussed whether it is preferable for eu member states in terms of flexibility and promptness to rely on price regulation, which is an inherently political activity, or not. an illustrative example is france, which has imposed a ceiling on the retail price of hand sanitisers but also on the wholesale price to third-party merchants, after the coronavirus outbreak. hence, the article is divided into six sections. section i introduces the structure of the chapter. section ii discusses article (a) tfeu, the provision of the eu competition law that the ncas and the european commission could enforce in order to ensure that consumers are not paying inflated prices during the coronavirus crisis. section iii makes an analysis of the practical and conceptual difficulties that the use of this eu competition law provision involves, especially when it comes to restore prices during the current coronavirus outbreak. section iv examines the remedies that the ncas and the european commission can impose in order to deal with excessive pricing. this section also recommends the remedy of 'commitment decisions' that would directly address the excessive pricing in the time of coronavirus at the eu level, rather than alleviating its harmful effects. section v considers price regulation and to what extent it may be a permissible solution within the internal market during the coronavirus crisis. in section vi, the article ends with a summarising conclusion. outside the merger context, the ncas and the european commission show an interest in pursuing excessive pricing cases only when such activity violates the discrete prohibitions of articles and tfeu. the pharmaceutical sector is an illustrative example where competition rules 'did their bit to deal with excessive prices' . when more than one firms charge excessive prices without indulging in an agreement, the abuse of excessive pricing is a weapon in the armour of the european commission and ncas to fight the ongoing price gouging. article tfeu explicitly condemns unfair purchase or selling prices, which are set by an individually dominant firm. yet, the same article prohibits the abuse of dominant position 'by one or more undertakings' . specifically, article tfeu contemplates that: ' any abuse by one or more undertakings of a dominant position within the internal market or in a substantial part of it shall be prohibited as incompatible with the internal market in so far as it may affect trade between member states. such abuse may consist in: (a) directly or indirectly imposing unfair purchase or selling prices or other unfair trading conditions; (b) limiting production, markets, or technical development to the prejudice of consumers; (c) applying dissimilar conditions to equivalent transactions with other trading parties, thereby placing them at a competitive disadvantage; (d) making the conclusion of contracts subject to acceptance by the other parties of supplementary obligations which, by their nature or according to commercial usage, have no connection with the subject of such contracts.' for over a decade, the meaning of this reference, i.e. any abuse by one or more undertakings, had been quite controversial, as there was uncertainty whether it meant that the market power and behaviour of legally and economically independent firms or only of separate legal entities within the same corporate group could fall within article tfeu. until the italian flat glass , the narrow view of article tfeu was predominant, according to which the abuse by more undertakings only referred to an abuse attributed to undertakings within the same corporate group. the judgment in italian flat glass identified that article tfeu applies not only to single firm dominance but also to collective dominance. in paragraph of this judgment, the general court underlined the following: 'there is nothing, in principle, to prevent two or more independent economic entities from being, on a specific market, united by such economic links that, by virtue of that fact, together they hold a dominant position vis-àvis the other operators on the same market. this could be the case, for example, where two or more independent undertakings jointly have, through agreements or licences, a technological lead affording them the power to behave to an appreciable extent independently of their competitors, their customers, and ultimately of their consumers.' thanks to this important landmark reached in this judgment, the abusive conduct of two or more legally and economically independent firms, though not within article tfeu, can be controlled under article tfeu. hence, according to article tfeu, imposing unfair purchase or selling prices may constitute abuse of a dominant position, which undertakings in the market can hold either individually or collectively. this provision has been used to prohibit both excessive and predatory prices, meaning that in principle, if more than one undertakings charge excessively high or too low prices, they can be held to have abused their collective dominant position in the market. yet, contrary to predatory pricing, there have been only few investigations of excessive prices under article , most of which were motivated by single-market considerations, like impediment to parallel imports and exports in the eu. the conventional wisdom is that the focus should be on the behaviour of firms that seek to exclude competition and not that much on market outcomes, such as prices. thus, it may not be exaggerative to say that the 'abuse of excessive pricing has remained underdeveloped conceptually and in practice at the eu level' . this is primarily because the european commission's guidance on abuse of dominance does not give enough details on the topic. moreover, there have been formidable difficulties in analysing excessive pricing. these difficulties, as well as other practical hurdles that the european commission and the ncas will have to face under the current circumstances, if they want to rely on abuse of excessive pricing to fight price hikes, are discussed in the following section. all these difficulties are underlined by the fact that ncas are in a rush to respond to the ongoing coronavirus outbreak. this section will prove that there are considerable practical and conceptual difficulties that excessive pricing conduct which is directly exploitative of consumers, for example charging excessively high prices or certain behaviour that undermines the efforts to achieve an integrated internal market, is also liable to infringe article . the commission may decide to intervene in relation to such conduct, in particular where the protection of consumers and the proper functioning of the internal market cannot otherwise be adequately ensured. for the purpose of providing guidance on its enforcement priorities the commission at this stage limits itself to exclusionary conduct and in, particular, certain specific types of exclusionary conduct which, based on its experience, appear to be the most common' . cases involve, especially under the current urgent circumstances that coronavirus has caused. for this reason, it is necessary to investigate these difficulties, assess them and make recommendations, where it is possible, in order to mitigate them. before analysing them, it is also worth mentioning that if these difficulties are taken together, they may also create a substantial risk of type i (false convictions) and type ii (false acquittals) errors. a false conviction in excessive pricing cases may lead to loss of consumer welfare resulting from the lack of introduction of valuable products, due to the reduced incentives of falsely condemned firms to invest. a false acquittal may also lead to loss of consumer welfare, resulting this time from above-competitive prices, due to the wealth transfer from consumers to producers. the first step in an investigation to assess whether there is exploitative abuse of a collective dominant position by charging prices which are higher than they would be in a competitive market is whether the undertakings concerned are dominant or not. in order to assess dominance, it is essential to identify the boundaries of competition by defining the relevant product and geographic markets and then to see if each undertaking on the market enjoys a position of dominance. if a company has a market share of less than per cent, it is unlikely to be dominant. the market share threshold, i.e. per cent, which is required to establish the dominant position of a firm in the market, makes it quite difficult for ncas to intervene with excessive pricing in the time of coronavirus. this is because the sellers of goods in high demand due to the coronavirus outbreak are often small businesses and they do not have a position of dominance in the market. a counterargument to this seeming hurdle that ncas could face is the european commission's decision in abg oil case , in which the european commission relied for the first time on the concept of 'transitory market power' . in that case, the oil crisis that took place in - made certain oil firms to increase their prices for crude oil and to reduce their production. in the prevailing circumstances of that case, the european commission found that each of those companies was holding a dominant position relative to its customers and that their conduct constituted an abuse of their dominant position within the meaning of article of the ec treaty. specifically, the european commission justified its decision about the dominant position of the oil firms on the following grounds: 'for reasons completely outside the control of the normal suppliers, their customers can become completely dependent on them for the supply of scarce products. thus, while the situation continues, the suppliers are placed in a dominant position in respect of their normal customers' . though on appeal the above decision of the european commission was annulled for lack of abuse, the court of justice did not challenge the question of dominance. this means that the ncas could possibly follow the same line of reasoning as the european commission in the aforementioned decision and try to establish "transitory market power", in order to establish that the sellers of goods in high demand due to the coronavirus outbreak hold a dominant position in the relevant markets. the ecj in united brands held that a price is unfair when 'it has no reasonable relation to the economic value of the product' . economic value is a rather vague term to use as a point of reference for considering if a price is excessive or not. for this reason, the ecj proposed a two-fold cost-based test for the determination of excessive pricing. the first limb of the test regards comparison of actual prices, i.e. the selling price of the product/service, with the actual costs, i.e. the production costs. the second part of the test considers whether the price imposed is either unfair in itself or when compared to the prices of competing products or prices across markets regarding the first limb of the test, it is not easy to calculate the difference between the production cost and the price of the product/service in order to identify the profit earned by the dominant firms. this is because the calculation of the difference requires the identification of the producer's costs, which is a complex, demanding and time-consuming operation, especially in case of a collective dominant position, where the firms to be examined by ncas are more than just one. nevertheless, 'firms' diverse production and market operations incur various categories of costs' e.g. marginal cost, long-term average cost, total cost etc., which makes it difficult to decide which of the dominant firms' costs should be considered for the analysis of the excessive pricing. generally, the marginal cost of production is a determinant of price in competitive conditions. yet, marginal costs raise complications and it is hard for antitrust enforcers to calculate them in both dynamic and static markets. long-run welfare effects. additionally, even if we assume that, the identification of the dominant firms' production costs is eventually possible, this does not necessarily mean that dominant firms are cost-efficient. the dominant firms' costs may be higher than they would normally be, due to cost inefficiencies. this means that dominant firms' production costs will not be a credible benchmark for assessing whether there is excessive pricing or not. another major difficulty in assessing whether a price actually charged is excessive or not is the level of profit margin that should be acceptable, i.e. the maximum 'fair' price above which the price charged by dominant firms is excessive. the european commission and the ecj have not set a threshold above which profits become excessive, as what may seem excessive profit in static industries, e.g. a margin of per cent, may be legitimately justified in dynamic industries, where profit margins are usually much more significant. however, as the case may be, the assessment of the profit margin should not concern ncas too much, as many goods in high demand due to the coronavirus outbreak, like private label hand sanitisers, are being sold online for up to per cent more than the original price. a last point that should be raised is that the assessment of excessive prices may be even more complicated in case of collective dominance, as the dominant firms may have different cost levels but they can still charge similar prices. likewise, the profit margins of less efficient firms may be lower than the ones of more efficient firms, but this does not necessarily mean the absence of excessive pricing. either unfair in itself or when compared to the prices of competing products or prices across markets. in this way, the ecj made available to ncas several methods to analyse excessive pricing, which is good. yet, it has been argued that applying these tests in times of crisis is not easy, firstly because the current economic value of the products in high demand due to the coronavirus outbreak is high and so this justifies their high prices. secondly, because the geographical benchmark, i.e. the method that compares the prices of the dominant firms with those of other firms in a different geographic market, is inadequate, as everywhere in europe the prices of products in high demand due to the pandemic are currently high. though these arguments sound reasonable, it is submitted that they do not make impossible the application of article (a) tfeu to coronavirus profiteering. regarding the first argument, what should be clarified is that the definition adopted in united brands about an excessive price, i.e. 'a price that bears no reasonable relation to the economic value of the good or service, and is higher than such value' , cannot be taken literally. this is because even a monopolist in the market would never want to charge more than the value of the product, as in that case consumers would not want to buy it. 'the most the dominant firm can do is exploit its consumers' willingness to pay' . this means that an increase in the economic value of a product does not necessarily lead to a price exceeding the product's economic value to the consumers, because this would be economically detrimental to the relevant dominant firms in the market and would eventually result in low demand. this is also supported by the united brands decision itself, which clarifies that: 'it is advisable to ascertain whether the dominant undertaking has made use of the opportunities arising out of its dominant position in such a way to reap trading benefits which it would not have reaped if there had been normal and sufficiently effective competition' . additionally, the argument that the high economic value of products to consumers justifies their high prices may have merit in the ordinary market operation but not in cases of emergency. in these cases, the most urgent needs may go unmet because the needed products or services are sold at a price too high to be borne by vulnerable consumers. 'to take advantage of an individual, benefitting from her misfortune, and benefitting disproportionately relative to one's contribution' is deemed exploitation. the essence of exploitation is a failure to benefit the victim as much as fairness requires, and so even mutually advantageous transactions can be exploitative and wrong. regarding the second argument, it should be noted that the various methods of analysing excessive pricing are compatible with each other and the ncas can apply them in parallel, when possible, for a robust result. this means that if the geographical benchmark is not appropriate under the current circumstances, the ncas are not barred from using the historical benchmark, for example, which enables comparison over time in order to determine whether a price is unfair or not. the historical benchmark will be particularly useful for ncas in case they want to take measures against the excessive pricing caused by the coronavirus outbreak. moreover, for the purposes of article (a) tfeu, the comparison with competing products is not limited to the products/services strictly within the same market. it is possible for ncas to consider a wider range of products/services, as long as they are comparable and there is effective competition between the dominant undertakings at issue and their competitors. this approach gives ncas flexibility and easiness to analyse excessive pricing in times of crisis, such as the current ones. the above analysis as well as the united brands case highlight 'the major difficulties of proof associated with finding an abuse of excessive pricing, and probably explains the relative dearth of instances in which the commission has intervened in those cases' . yet, in case the ncas go over all the above hurdles and they are finally able to rely on article (a) tfeu in order to challenge excessive pricing during the coronavirus outbreak, it is hard to predict to what extent firms will stop charging excessive prices and how prices will evolve over time. the reason for this uncertainty is that the primary remedy that ncas have at their disposal when pursuing excessive pricing cases is the imposition of fines to the firms. this measure does not ensure that firms will comply with the decisions of ncas and will not raise their prices again in the future, as it amounts to 'suppressing the symptoms rather than curing the disease' . what is more, since fines are just a behavioural remedy, the monitoring of their effectiveness regarding the resetting of prices to a 'non-excessive' level will absorb many of the ncas' resources. apart from fines, ncas can also rely on other remedies, such as encouragement of consumers to switch to less expensive offers of new entrants, removal of entry barriers or settlement agreements with the firms setting excessive prices. relying on the first two remedies in times of coronavirus crisis is not easy, as they require new market players and plenty of time and resources on the part of ncas to take the relevant measures. coronavirus has a chilling effect on investments and thereby reduces new entries to the markets. structural measures, such as the removal or prohibition of entry barriers to the market, require prior investigation by the nca about the causes of excessive pricing. this kind of investigation needs time and resources, which means that the ncas cannot take urgent action and apply promptly the relevant remedies. besides, free entry does not eliminate by itself and immediately the power of incumbent firms in the market. in view of the aforementioned remedies that a nca has at its disposal in excessive pricing cases, it gets apparent that excessive price cases are among the most difficult and complex cases for ncas in terms of the design and implementation of suitable remedies. therefore, it is worth examining alternatives to the available remedies. at eu level, a good alternative to fines would be the issuance of 'commitment decisions' by the european commission, in order to motivate the relevant firms to set lower prices in the future and offer conditions that are more convenient for consumers. the european commission can issue 'commitment decisions' when it intends to adopt a decision requiring that an infringement be brought to an end, and the companies under investigation prefer to offer commitments in order to remove the european commission's competition concerns as expressed in a preliminary assessment. 'commitment decisions' may also be preferable if the case is not one where a fine would be appropriate or when efficiency reasons justify that, the commission limits itself to making the commitments binding, and does not issue a formal prohibition decision. the reasons why 'commitment decisions' could be a good remedy for excessive pricing under the current circumstances is that they can restore undistorted conditions of competition in the markets in a swift and effective manner, as the administrative process for commitment decisions is generally short. 'commitment decisions' can be either behavioural or structural and can be limited in time, meaning that the european commission is able to reassess them if a material change takes place in the meantime. they do not establish an infringement and do not require any admission by the parties, as they only require commitment to future behaviour. this can be an incentive for firms to pursue the commitment route, as they know that in this way they avoid any damage to their reputation as well as a formal finding of an infringement against them. in addition, the ecj does not require a strict proportionality test for assessing the remedies imposed by the european commission in a 'commitment decision' , as it would do in case of a prohibition decision. if a firm does not comply with the 'commitment decision' , the european commission is able to impose a fine up to per cent of the firm's annual turnover without having to prove any violation of the competition rules. from the viewpoint of the european commission, this provision saves money and time. moreover, the european commission is able to impose periodic penalty payments of up to per cent of the average daily turnover of a firm until it complies with the 'commitment decision' . given the above conceptual and practical difficulties that the ncas and the european commission may encounter in excessive pricing cases, competition law enforcement may be a rather risky and time-consuming form of intervention. therefore, some countries like france and cyprus proceeded to regulatory pricing frameworks in order to cap the wholesale and retail prices of some products in high demand due to the coronavirus pandemic. in the united states, many states have already laws against price gouging, which apply to actions taken during times of disaster or emergency and they regard certain classes of items, generally those that are necessary for survival or for coping with the problems caused by a disaster. hence, after explaining what price regulation means, this section is going to discuss the disadvantages and advantages of a regulatory pricing framework in order to see if, after all, it is preferable to competition law enforcement. price regulation refers to 'regulatory methods of determining and imposing controls on firms of industries' with the aim of restricting independent price setting. this may involve a fixed price or rate of return, a maximum price ceiling, or minimum price floor. the use of price controls and price caps has been generally criticised for being in contrast with competition policy for various reasons. within neoclassical economics, the free formation of prices, which relies on the interaction of supply and demand, is essential to the effective functioning of competitive markets and the efficient allocation of society's resources. this is because price is 'the most immediate parameter upon which undertakings compete' , it shows whether demand exceeds supply and it makes consumers estimate the value that a good or service holds for them. moreover, it is rather complex and difficult to determine a regulated price, as it must secure on the one hand the incentives of firms to keep on operating efficiently in the market, without distorting the optimal supply and demand levels, on the other hand the firms' profits, which should be neither excessive nor unviable. furthermore, from fairness perspective, it has been argued that price regulations do not take into account, as they should, the increased costs that the sellers might face because of the same crisis or disaster which put their customers in difficulty. according to this argument, the firms should not absorb the increased costs in order to benefit the consumers, particularly when the former have exercised good foresight and responsibility in obtaining a ready stock of goods that might be necessary in case of a crisis or a disaster. this argument holds truth today and in order to deal with this issue, the author suggests that price regulations should be flexible enough to enable price increases above the relevant price ceiling, in case the increased price is attributable to increased costs that the seller had to bear. apart from these general critiques, some academics have also argued that the application of price controls is contradictory to the broader eu context, particularly the pursuit of a competitive internal market. the internal market is built on open and undistorted competition, which presupposes the avoidance of unnecessary public restraints that may have an impact on efficiency or integration. hence, any state-imposed limitations on the free functioning of the price formation mechanism constitute an 'anathema to the underlying philosophy of the internal market' , as they are deemed to be 'a particularly strong limitation of rights to property and the freedom of economic initiative.' despite the aforementioned cons of price regulations, the free formation of prices, which 'constitutes the expression of the principle of free movement of goods in conditions of effective competition' , is not always the best choice, as sometimes it does not guarantee socially desirable outcomes. for instance, in the exceptional case of urgent needs, firms may set excessive prices for the necessary goods that are specifically important for the consumers' health and wellbeing. this may result in a transfer of wealth from consumers to suppliers as well as in a reduction in total welfare to society. the above exploitative nature of excessive pricing is like parallel accommodating conduct, where 'each rival's response to competitive moves made by others is individually rational, and not motivated by retaliation or deterrence nor intended to sustain an agreed-upon market outcome, but nevertheless emboldens price increases and weakens competitive incentives to reduce prices or offer customers better terms' . therefore, this behavioural exploitation can arguably justify regulatory intervention in the price formation process, as society should undertake the responsibility of protecting its members in time of crisis from this kind of conducts. in addition to this, where self-correction of excessive prices is not possible at all or within a reasonable timeframe, the likelihood that consumers will suffer high prices for a long time makes necessary the regulatory intervention. excessive pricing is self-correcting when it attracts new entrants to the market that deter in this way the dominant firm(s) from setting very high prices. in time of crises, like the coronavirus outbreak, the firm entry rate into the market is affected, as firms' incentives for investment and expansion are decreased in their attempt to survive the crisis and the resulting severe economic turmoil. for this reason, it seems that the current circumstances call for price regulatory intervention. moreover, it has been proved that in any case, excessive prices do not attract entry and so self-correction should not serve as an argument to justify non-intervention. if a potential entrant has gathered adequate information about the incumbent firms' advantages, and specifically their marginal costs, and the entrant considers that the incumbent firms are more efficient than it is, the entry to the market is unlikely, even if the incumbent firms charge an excessive price. this is because once such a market entry takes place, the incumbent firms will start a price war with the new entrant that could bring prices to levels that render entry unprofitable, taking into account the new entrant's cost-disadvantage. on the other hand, if a potential entrant considers that the incumbent firms are less efficient than it is, the market entry is likely, but not because of the excessive pricing. regardless of the preentry excessive pricing and the incumbent firms' expected post-entry price-cutting, the new entrant would know that its competitive advantage over the incumbent firms could enable it to make adequate post-entry profits. this article explored article (a) tfeu, the provision that the ncas and the european commission can use to prohibit undertakings from charging excessively high prices during the coronavirus outbreak for goods that are in high demand, such as hand sanitisers, disinfectants and disposable respiratory protection masks. the analysis showed that there are a number of difficulties of proof associated with finding an exploitative abuse of collective dominant position. some of these difficulties include the market share threshold, which shall be per cent and more, the calculation of the difference between the dominant firms' production costs and the price of the product/service, as well as the several methods used to analyse excessive pricing. despite all these hurdles, which are underlined by the fact that ncas are in a rush to respond to the ongoing coronavirus outbreak, it was explained that the application of article (a) tfeu to coronavirus profiteering is still possible. one of the suggested solutions to overcome these hurdles was the acknowledgement of a 'transitory market power' , as the european commission had done in abg oil case. in this way, it would be possible for ncas to establish that even small businesses, which do not have a position of dominance in the market, hold a temporary position of market strength in the relevant market. this is because these firms sell goods in high demand due to the coronavirus wrongful beneficence: exploitation and third world sweatshops - ; see also matt zwolinski 'the ethics of price gouging the law and economics of article tfeu see also scandlines sverige ab v. port of helsingborg(comp/ . ) of article -abuse of a dominant position motta and de streel (fn ), p. ; frederic jenny ' abuse of dominance by firms charging excessive or unfair prices: an assessment competition policy-theory and practice memo/ / -commitment decisions (article of council regulation / providing for a modernised framework for antitrust scrutiny of company behaviour) european commission 'cartel case european commission (fn ) opinion in case c- / the queen, on the application of vodafone ltd and others v secretary of state for business case c- / scotch whisky association and others v the lord advocate and the advocate general for scotland eu ) section , <www.justice.gov/atr/hori zontal-merger-guidelines- # \protect$
elax>$ accessed on imf") expressed the same opinion. see bbc news 'coronavirus: worst economic crisis since s depression, imf says outbreak and their customers are completely dependent on them for the supply of these products.then the article investigated the remedies that ncas have at their disposal when pursuing excessive pricing cases. the discussion focused on the imposition of fines to firms, which is a behavioural remedy that does not ensure the resetting of prices to a 'non-excessive level' . therefore, the article argued the issuance of 'commitment decisions' by the european commission, as a good alternative to fines. the 'commitment decisions' can address directly the excessive pricing in the time of coronavirus at the eu level, rather than merely alleviating its harmful effects.finally, the article dealt with the prospect of price regulation and examined to what extent it could be a good alternative to competition law enforcement. the discussion showed that price regulation is a much-disputed enterprise in contemporary regulatory practice, but the pragmatism calls for it, especially in the absence of selfcorrecting excessive prices.doi: . /jeclap/lpaa key: cord- -r ypur authors: simpson, shmona; chakrabarti, ajoy; robinson, david; chirgwin, keith; lumpkin, murray title: navigating facilitated regulatory pathways during a disease x pandemic date: - - journal: npj vaccines doi: . /s - - - sha: doc_id: cord_uid: r ypur in , the bill and melinda gates foundation convened over thirty subject matter experts in clinical development, manufacturing, and regulatory assessment to determine how the development and approval of medical countermeasures could be accelerated in the event of disease x. disease x is the result of a presently unknown pathogen with epidemic or pandemic potential. a key opportunity to accelerate the scientific assessment and regulatory approval of medical countermeasures exists within efficient navigation of facilitated regulatory pathways. it was identified that not all stakeholders will be able to skillfully navigate the facilitated pathways offered by the various regulatory agencies during a public health emergency. to democratize this knowledge, we have written an overview of the facilitated approaches which have been developed and refined by stringent regulatory authorities and the world health organization for the primary assessment of medical products. we discuss the conditions necessary for use of these approaches, scenarios in which certain pathways may be applicable, and the pros and cons of these approaches. we also address opportunities available to developers in, or developers who wish to access, low-income countries that may have nascent regulatory frameworks. the - ebola epidemic significantly impacted the global health community. between guinea's index case in december and the epidemic's end in june , there were , cases and , deaths across eight countries . despite years of prior research, no products were ready to deploy in time to save these lives. the question then arose: how many lives would have been saved if effective medical countermeasures had been made available sooner? the who created the research and development blueprint initiative , which asked multiple agencies how to shorten the time to development of medical countermeasures for the world's most deadly pathogens. the bill and melinda gates foundation attempted to answer this question for disease x-the result of a presently unknown pathogen with epidemic or pandemic potential. in december , the foundation convened a disease x working group-comprised of experts in emerging infectious diseases; bioterrorism agents; non-clinical studies; clinical trials; chemistry, manufacturing, and controls; and regulatory scientific assessment . the goal was to understand how the next epidemic could be controlled using medical countermeasures that arrive sooner than has been the experience to date. the control of disease x requires improvements in many areas, including disease surveillance, public health infrastructure, laboratory capacity, product manufacturing, and delivery -however the remit of this working group was to focus specifically on the expedited provision of medical countermeasures. one of the key challenges identified was a perceived difficulty in navigating facilitated regulatory pathways. in this perspective, we summarize some facilitated regulatory pathways available to innovators tackling disease x and suggest how these may shorten product development and regulatory assessment timelines. we also discuss the pros and cons of these approaches and suggest in which situations they may be most applicable. so far, we have identified facilitated regulatory pathways (listed in supplementary note ), in countries around the world. while many of these have critical nuances and are at various stages of refinement , , they cannot all be adequately described in this short perspective. a broader set of considerations are available elsewhere , . rather, some "stringent regulatory authorities" (as classified by who) have completed primary assessment of thousands of products using these facilitated approaches. these include products used in epidemic emergencies. many other national regulatory agencies (nras) have mirrored these processes, and/or rely on the scientific outcomes of these approaches to assist in their own authorization practices. this perspective provides a high-level overview of the facilitated approaches which have been developed and refined by stringent regulatory authorities and the world health organization for the primary assessment of medical products. it is possible that these represent the approaches most likely to be relied upon in the next epidemic or pandemic. we hope this paper will enable academic innovators and small and medium enterprises to navigate the flexibility that exists within regulatory approaches for products that address life-threatening diseases of unmet need. we encourage developers to make early contact with regulators in their focus countries to discuss which programs may be applicable. several potential regulatory scenarios may exist and co-exist during an epidemic: for example, (a) de-novo candidates requiring rapid development and regulatory assessment (b) de-novo products requiring assessment when the typical package of clinical efficacy data may not be available, (c) approval of de novo or repurposed products for "emergency" use only in specific populations (d) for compassionate use in specific (e.g., "named") individuals of an unauthorized medicine (e) conditional or accelerated authorization before the completion of efficacy studies or, (f) use of a licensed product outside of its approved use (e.g., for another indication, dosage regimen, or population). many regulatory agencies have instituted various programs to help navigate these scenarios (table ) . these pathways are generally reserved for products that address a serious or life-threatening condition where there is unmet clinical need, or where the current treatment options are unsatisfactory. eventual authorization through one of these pathways depends on an evaluation of the known clinical benefits and risks of the product-in the context of the known risks of the disease. for this, regulators employ a variety of complex modeling and analytic techniques to conduct an assessment of the benefits and risks of the product and any remaining uncertainties, and compare these to the risks of the disease and the available options at the time of application , . in an outbreak, these parameters can be highly variable; they may change depending on the pathogen, co-morbidities, evolution of the epidemic, populations and geographies affected. however, these agile facilitated regulatory pathways recognize the higher tolerance for unknown risk the community has in these situations: they allow for flexibility on the depth of certain routine data requirements given the specifics of the disease being considered. not all of these pathways facilitate a shorter assessment time: some allow for assessment to be conducted at an earlier phase of the typical product development lifecycle in instances where the benefits, at that point in time, outweigh the risks (fig ) . a critical element for successful use of these facilitated pathways is the engagement of regulators early and often: scientific advice and pre-submission meetings are invaluable. most authorities allow rolling submissions of data and rolling reviews under these processes. this early engagement allows for on-going alignment on development plans as further data become available. it also allows sponsors to focus on the critical data requirements, identify opportunities for additional product development acceleration, and ultimately save substantial time. depending on the product and the clinical situation, a product may be eligible for any or all these facilitated programs and can be candidates in more than one program simultaneously. without active regulator engagement, it is often difficult to navigate these approaches and understand the challenges inherent in each. regulation through reliance and regionalization are critical elements for broadening the utility of facilitated pathways. reliance allows for nras to rely on the work product of a trusted authority to inform their own regulatory decision. regionalization extends the utility, allowing neighboring regulatory authorities to workshare, and share work product, within their economic or cultural blocs. this may mean that regulatory assessment and authorization in one jurisdiction, can de facto facilitate accelerated authorization in another jurisdiction thereby avoiding duplication of effort . a list of regulatory harmonization initiatives is provided in the supplementary note . the following pathways are available under certain circumstances to expedite product development and marketing application assessment. the united states food and drug administration's (fda) priority review process provides feedback on a marketing application, i.e., an authorization or complete response, within, generally, months, instead of the standard months. this faster application review is for products that purport to demonstrate significant efficacy or safety over a currently available therapy related to the treatment, diagnosis, or prevention of a serious condition. this designation, which comes at the time of submission, does not affect the length of the clinical trial period. many other regulatory authorities have expedited review timeframes for similar situations, and one should always check to see if the medical countermeasure would qualify in the country in which the product is intended to be used. accelerated assessment, under similar situations, is also available at the european medicines agency (ema), reducing review time from days to days. japan's pharmaceutical and medical devices agency (pmda) also provides a priority review option, available to products that address ( ) serious diseases, ( ) conditions of unmet clinical need, or where superior safety and efficacy can be provided, and ( ) orphan designated products. this option reduces review time from to months , . fda's fast track is a program for products that have some initial evidence of efficacy or improved safety over an available therapy . this designation provides an opportunity for frequent meetings and communication with the fda. most importantly, this program allows a "rolling review" process, in which the marketing application is submitted in pieces as each segment is completed rather than having to assemble the entire application and submit it all at once. this allows fda to assess each segment as it is submitted, and thus fda only must review the last segment when it is completed rather than the entire application. fda's breakthrough designation is granted where a new candidate demonstrates substantial improvement over an available therapy on a clinically significant endpoint. candidates demonstrating an improved safety profile over an available therapy with similar efficacy are also considered. efficacy can be demonstrated using a pharmacodynamic biomarker, surrogate or intermediate clinical endpoint providing they strongly suggest a clinically meaningful effect. for example, pfizer's -valent pneumococcal conjugate vaccine candidate and janssen's prophylactic vaccine for the prevention of respiratory syncytial virus both achieved breakthrough designation following a phase and b studies, respectively , . a breakthrough therapy designation enables fast-track designation, intensive guidance on a drug development program, and organizational commitment. like fda's breakthrough program, investigational products to address unmet medical need are eligible for consideration under the european union's priority medicines (prime) scheme, providing early clinical data demonstrate potential benefit. in addition, applicants from the academic sector, small and medium enterprises can engage with the ema quite early based on compelling non-clinical data and tolerability data from initial clinical trials. not exclusive to these applicants, the ema also offers scientific advice and protocol assistance to ensure the most expeditious experience with applying for market authorization . prime also ties in to accelerated assessment if the data ultimately demonstrate the level of improvement needed for accelerated assessment designation. the following pathways are reserved for candidates when the benefit risk analysis indicates that access should be granted even if the entire clinical trial process has not been completed. generally, these are temporary authorization statuses and are not intended to replace or circumvent ultimately finalizing the clinical trials required to support full market authorization. as a condition of approval under these pathways, it is general a requisite that the clinical trials be continued until adequate clinical efficacy and safety either are or are not demonstrated. fda's accelerated approval allows for authorization where efficacy is demonstrated via an unvalidated surrogate endpoint or an intermediate clinical endpoint . these surrogate endpoints are likely predictors of clinical benefit and require that this be demonstrated by "adequate and well controlled" studies. ultimately, the requirement remains to confirm clinical benefit in post-authorization confirmatory trials that validate the approved endpoint. when these studies are completed, the fda will review the data and decide if the approval can be converted to a full authorization. the accelerated approval may be revoked if clinical benefit relative to the risks cannot be confirmed. to date, compounds have been approved under this pathway. these range from numerous antiretroviral compounds in the 's, to janssen's levaquin for aerosolized bacillus anthracis, to the recent approval of ismed's arikayce for treatment of mycobacterium complex in -all averaging an initial accelerated approval time of six months . priority medicines with accelerated assessment scientific advice and protocol assistance (prime) •a strongly substantiated mechanism of action, preclinical data, and human tolerance data •academic, small and medium enterprises may apply earlier for advice and protocol assistance serious or life-threatening condition for which therapy is inadequate. rapid assessment, based on early clinical data japan pharmaceuticals and medical devices agency (pdma) priority review •available for orphan designated products, and those that receive conditional approval for diseases of unmet clinical need rapid assessment, decision received within months conditional term-limited approval fda's expanded access (ea) is a program designed for patients with an immediately life-threatening disease to access a product that has clinical trial data (putatively showing an acceptable benefit-risk profile)-but does not yet have marketing authorization. because the authorization has not yet been granted, the product is considered investigational and therefore written informed consent of the patient must be obtained when used. generally, ea is used in situations where alternative therapies are not available. which ea program is chosen reflects the perceived need for the product in terms of number of patients. under such programs, the product can be used for: ( ) a single patient, ( ) immediate-size populations that occur after the fda has received a number of requests for single patient use, or ( ) under a "treatment investigational new drug," designation. these are generally used during the time period after the completion of pivotal trials, but before the authorization is granted. during this period, there may be large numbers of patients that might benefit from the product during the time the marketing application is being assembled and/or the product is under review . options for international expanded access exist in cfr . (b)(ii) which allows for the export from the united states of investigational products for national emergencies elsewhere, with nra approval in the receiving country. for example, following promising animal and early-phase clinical studies (prevail ii trial) the fda supported the use of zmapp through a standing expanded access protocol prior to completion of the submission, which allowed countries to retain access to vital therapeutic agents . manufacturers who provide access to product under one of the "expanded access" programs may only recover the direct costs of manufacturing their investigational product and may not recoup additional costs or make a profit. in these cases, access to product under an "expanded access" program allows for patients to receive the product when the potential benefit outweighs the known risk in the specific context of the patient(s). this sometimes provides further valuable data to help support a full marketing application. the ema also offers programs to bridge the clinical trial and full authorization gap. unlike a typical full marketing authorization, a conditional marketing authorization (cma) can be granted in instances of unmet medical need where the benefit-risk assessment is positive. this is based on early data suggesting that the sponsor will be able to provide more complete data within an agreed timeframe to validate the tentative positive benefit-risk profile of the product. these data generally include more comprehensive clinical efficacy and safety data . cma's are generally granted for a -month period, after which they are further reviewed in light on any new data available. if deemed of benefit to public health, the conditional authorization can be extended. a large number of antiretroviral compounds - and treatment for multi-drug resistant tuberculosis such as bedaquiline and delamanid continue to be marketed conditionally by the ema as trials are ongoing. ultimately, this pathway this may provide a route to full authorization for many of these candidates. these provisions, both in the us and eu, require postauthorization infrastructure so that further, more comprehensive data can be captured and so that any requisite safety monitoring can be performed. such infrastructure often does not exist in lowincome countries. this makes use of these pathways sometimes difficult, if not impossible, in countries without such needed infrastructure. a key feature of these mechanisms is that they often feed into each other and are not meant to be mutually exclusive. for example, in , degrees pharmaceutical's tafenoquine for malaria prophylaxis was awarded both fast track and priority review designations . similarly, a medicine in the prime scheme could also be granted conditional marketing authorization during clinical trials, while still benefitting from accelerated assessment and eventual full authorization . japan's pdma offers two conditional approval options; the first is for regenerative products which grants a term-limited approval based on exploratory phase i and ii safety and efficacy data . confirmatory studies are required post-market, and applications must be submitted for complete market authorization within years. this is described as conditional "term limited" approval. the second conditional approval is applied in instances of a serious disease, disease of unmet clinical need, or where it is too difficult or excessively lengthy to conduct efficacy studies. in this instance, early phase clinical studies must show some safety and efficacy, and post-market requirements will include surveillance or clinical studies . japan's sakigake (or pioneer) program is available for diseases of unmet clinical need where the sponsor agrees to file the marketing application first in japan (or simultaneously first in japan and another country). for a candidate to qualify for sakigake, there must be some early phase clinical data and strong non-clinical and mechanism of action data. this program offers pre-application consultation, expedited review of around six months, and superior support from pdma. the re-examination of clinical safety and efficacy data can be lengthened for an indeterminate period of time, in order to strengthen the likelihood of full market authorization . when conducting clinical trials presents insurmountable logistical, safety, or ethical challenges, the following regulatory approaches allow for efficacy to be demonstrated using an animal surrogate or clinically indicative endpoints alongside human safety data. these are reserved for when public health need is significant and human efficacy testing is not possible. in all instances, every attempt is made to procure follow-up analyses of human experience with the product post-authorization. the fda's animal rule provides a pathway to approve novel candidates in the absence of the demonstration of efficacy in clinical trials. where clinical trials are impossible or unethical to conduct, determinations of efficacy are primarily based on studies in well-characterized animal models . the investigator must provide: • poof that the animal model can provide plausible inference for human efficacy, • proof that the mechanism of action is the same in the model, as it would be in humans, • rationale for the dose provided to humans (which may come from human pharmacokinetic studies and animal efficacy studies). in these situations, safety is demonstrated using traditional toxicology studies and human experience data (usually from phase trials in healthy human volunteers). the chemistry, manufacturing, and controls information needed to support an animal rule application follow fda guidelines for a routine new drug or biological license application. this program allows a promising candidate to be authorized for human use where a major public health need is justified. this authorization is rare: to date only and drug and biologic approvals, respectively, have been granted under this provision . built into the "animal rule" is an iterative process where post-authorization human safety and efficacy data can be obtained and used as the product is employed to further refine the product label. aside from de-novo product authorization, the animal rule may also be used to gain authorization for a new indication. in addition to several medicines that have been approved under this provision, the anthrax vaccine biothrax was approved in for post-exposure prophylaxis making it the first vaccine to receive approval for a new indication based on the animal rule . in the european union, an analogous option exists within the provisions of the "marketing under exceptional circumstances" pathway. in exceptional circumstances, when an applicant cannot reasonably be expected to conduct clinical trials, they may submit as much clinical safety as efficacy data as possible, alongside a proposal for post-approval studies that would support the original safety and efficacy claims. inability to supply a complete efficacy package may occur due to the rarity of the disease, limitations in the present state of scientific knowledge, or when it would be unethical to conduct. once marketed under exceptional circumstances, the candidate will be supplied with a summary of product characteristics stating that information regarding the candidate is incomplete, and the label may be updated . in , the smallpox vaccine imvanex was marketed under exceptional circumstances after exploratory data demonstrated that protective antibodies could be triggered, with only mild side effects (including in patients with hiv or atopic dermatitis). to date, the vaccine's benefits and risks continue to be studied in vaccine recipients. ultimately true efficacy could only be demonstrated if there is an outbreak of the disease in the future . the following pathways are reserved for when a national or global public health emergency has been formally declared. fda's emergency use authorization (eua) is enabled once the united states secretary of health and human services declares a specific national public health emergency . it considers whether the "known and potential benefits of the product outweigh the known and potential risks" providing that no reasonable alternatives exist for treating the cause of the specified national public health emergency. data to support the application may include domestic and/or foreign clinical trial data, in vivo animal data, and in vitro data that provide plausible support for clinical efficacy. access to a product under an eua is limited to the duration of the national health emergency and the specific access caveats imposed. thereafter the product is again considered investigational. for example, during the h n influenza outbreak, euas were granted for the dispensing of tamiflu (oseltamivir phosphate) and relenza (zanamivir), and intravenous peramivir. these were discontinued in once h n was no longer deemed a threat to the united states . this pathway is applicable to medicines, biologics, vaccines, and medical devices, including in vitro diagnostics. so far, the eua has been utilized primarily for diagnostic tests for several infectious agents including influenza, anthrax, coronaviruses, ebolavirus, and zika viruses. for those developing products to be used in countries with nascent or under-resourced regulatory agencies, the who has provided the emergency use listing (eul) process to provide guidance on product quality and use during a public health emergency of international concern (pheic) or other specified public health emergency. only the director general of the who can authorize the use of the eul process. this program was developed and first used for ebola diagnostics during the - west african ebola outbreak and was previously referred to as the emergency use assessment and listing (eual) process. since then, it has been further refined and renamed, with the most recent guidance issued in early . this process was used extensively in both the ebola and zika outbreaks for in vitro diagnostic products, although the process is clearly intended for medicines and vaccines also. the first example of a vaccine advancing into the eul process is the new oral polio vaccine for type virus (nopv ). this genetically modified (designed to improve safety relative to the existing oral polio vaccine, mopv ), oral polio vaccine is being developed by bio farma (indonesia) and path. on-going transmission of type vaccine-derived polio virus (vdpv) in certain regions of the world has resulted in who maintaining its long-standing pheic for polio. successful completion of the eul process would facilitate the use of nopv in field campaigns to control vdpv events once an outbreak has been identified. use of the eul process would allow this novel vaccine to be used prior to who prequalification, enabling an improved vaccine to be utilized in this emergency in advance of obtaining all the data required for traditional product licensure pathway. during this use, data will be gathered to support the traditional who prequalification process and national product licensure pathways. while the gmp requirements are generally the same as required for the who prequalification program, the efficacy of a vaccine, for example, may be demonstrated by pre-clinical efficacy data in a suitable animal model, alongside clinical immunogenicity that is reasonably predictive of human clinical efficacy. a plan to monitor safety and efficacy in the field must be included, and an eul is granted initially for months. the manufacturer's history of successfully prequalifying products may contribute to the decision -especially if specific manufacturing site inspection is difficult to obtain in a timely manner. specific to vaccines, though the who has provided the eul, at least one qualified nra is responsible for providing "oversight of batch release and other post-eul product safety and manufacturing quality assurance requirements ." this is usually the agency in the manufacturing country. some countries where epidemics occur are not equipped to do this for certain vaccines. to mitigate this, national and regional regulatory agencies generally should engage with their global regulatory counterparts and who prequalification in a collaborative approach to product assessment and oversight under emergency circumstances, especially when novel outbreak etiologies and novel therapeutic and prophylactic modalities are being proffered. when clinical trials have been conducted routinely, there is little motivation for manufacturers to produce initial quantities of product in excess of what is required for the clinical trials program. however, in a public health emergency, manufacturers may be called upon to supply larger quantities of product much more quickly. this may deplete clinical trial supplies. if adequate forethought about production has not been undertaken, there may only be enough product for early clinical studies: this results in significant delay to commencement of later phase trials or early larger scale use of the product under one of these pathways, especially during an epidemic (fig. ) . mitigation may involve rapid and significant investment of resources in manufacturing, at risk, prior to clinical proof of concept in order to be able to meet demand if the early data support wider use of the product in emergency situations. when clinical trials have not been conducted a key element to utilizing the animal rule and analogous facilitated programs is the need for a well-validated animal model. a well-validated animal model may not always exist for "disease x" and establishing a well-validated animal model can be challenging. in addition, these pathways that utilize animal surrogates may not be applicable to most outbreak scenarios involving new pathogens. once an outbreak is underway it typically would be feasible and ethical to conduct clinical trials at which point authorization under the animal rule becomes less relevant. conversely, after an outbreak ends, the opportunity to evaluate efficacy in humans also ends, but at that point the level of urgency also decreases. for novel pathogens, these approaches remain best reserved for instances where probability of human efficacy is higher and apparent at an earlier stage. without this, the chief value of this approach may be prior to an outbreak or after the outbreak has ended with the aim of supporting use rapidly in a potential future outbreak. the advantage of many of these pathways is that demand for a product with reasonable presumption of efficacy and safety may be met earlier during a public health emergency, than in a traditional product development and marketing application assessment timeline. in these situations, specific use under emergency authorization may include use in first responders, use in those infected, use in ring programs, and/or use in mass distribution as the situation warrants and as the caveats of the specific emergency use authorization dictate. manufacturers must commit to collecting further clinical efficacy and safety data, often including clinical trials where feasible and ethical, in conjunction with this emergency usebased field use. depending on the scientific robustness with which such data are collected and analyzed, these data may provide primary and/or supplementary data for later licensure. a key downside of these approaches is the data may not help differentiate the potential clinical safety risks of the product from the underlying clinical complications of the disease. uncontrolled use may confound the long-term safety and efficacy assessments of a product due to the high morbidity and mortality rates during these emergencies. the studies may also be confounded by other comorbidities and other factors typical of the geography, concomitant use of other interventions, or availability (or lack thereof) of other healthcare and medical supplies. mitigation of these challenges involves early discussions with regulators. this ensures that there is a common understanding about the natural history of the disease (where possible) and thus a way to try to differentiate drug risk from disease risk and differentiate product efficacy from disease natural history. in addition, where possible, these candidates should be deployed in a controlled clinical study to ensure that the efficacy and safety of the product is appropriately evaluated . despite their challenges, these pathways can be used to potentially expedite medical countermeasure availability in a public health emergency for candidates with positive pre-clinical efficacy signals. while the abovementioned regulatory agencies have instituted various programs to help expedite the development and assessment of products for use during public health emergencies, many low, and lower-middle income countries have nascent and underresourced regulatory agencies. for products manufactured in, or used in, countries that cannot assure quality standards, who prequalification is a system developed to help procurers of prequalification eligible products determine if the products they are procuring meets international regulatory standards for product efficacy, safety and manufacturing quality. many low-income countries rely on prequalification listing and the assessment and inspections documents who provides them to inform their own national regulatory decision on that specific version of the product. this is done through the who-nra collaborative process. who cannot "authorize" a product; rather, it "lists" the versions of the products when the who assessment determines that the clinical and manufacturing data meet international standards. like routine product authorizations, routine who prequalification is generally not used in public health emergencies. several early engagement opportunities and facilitated accelerated pathways exist when one is focusing on regulators in lowincome countries. these national agencies are engaging more with their global regulatory counterparts and who prequalification program staff in a collaborative approach to assess and authorize products (both clinical trials applications and marketing applications) under emergency circumstances, especially when novel outbreak etiologies and novel therapeutic and prophylactic modalities are being proffered. who has instituted a number of pre-emergency activities, described in the latest version of the eul process . the preemergency activities involve establishment of platforms for collaborations between who, subject matter experts, nras with special expertize, and nras where the products will be used (where they differ). who establishes expert advisory committees to support each stage of the eul. in addition, these platforms are used for pre-submission meetings/activities, selection of products, and assessment of submitted data. these activities allow for accelerated decision making upon declaration of a pheic or other covered public health emergency. a key benefit of these activities is that the nras are involved at early phases of product development and participate in the assessment process. together, all nras and who can align on appropriate non-clinical model and clinical study design. this allows for phase b and trials to commence quickly and with the appropriate assistance levels. this can clarify clinical trial endpoints that would be supportive for eul by who and in the target countries. an additional opportunity for clinical trial discussions are regional health agreements in sub-saharan africa - , south america , and asia : these facilitate work-sharing and joint decision-making. for example, the africa vaccine regulatory forum (avaref) is a continental platform of regulators sponsored by who afro and who-geneva which coordinates the joint regulatory and ethics board assessments of multinational clinical trials in africa. innovator engagement with avaref, for example, allows the benefit of joint scientific advice and clinical trial assessment meetings. in addition, avaref has established an emergency clinical trial assessment process for multi-country clinical trials. originally designed only for vaccine trials, avaref, despite its anagram, is also available for use with multinational clinical trials application assessment for medicines in africa. outside of the who eul, two other pathways exist to specifically support lower income countries with facilitated assessment, particularly of novel products. they both bring the resources of an agency with specific expertize and who to conduct scientific assessments of clinical development programs and marketing authorization applications with opportunities for engagement in the discussions by nras from the countries where the product will most likely be used. the use of such pathways brings low income country regulators into the development and assessment processes as a partner so that use of the outputs of the process can be utilized more readily by nras where the product will ultimately be used. article of european commission's regulation no / is a specific framework, in collaboration with who, designed to support lower income countries regarding products to be marketed outside of the european union . the ema will assess products of major public health interest, collaborating with the who and the nras in the countries of use. regulators, experts and observers from lower income countries participate in the scientific review of the product -both during the development phase and during the marketing application phase. the ema publishes a scientific opinion regarding the marketing application. this allows under-resourced nras to make a decision that leverages the ema's assessment (including good clinical and manufacturing practice inspections) and their engagement with ema and the who during the development and marketing application assessments. recently, an option has been provided for total or partial fee waivers for the manufacturer . while not marketed in the eu, the malaria vaccine rts,s/as received a positive scientific opinion under article following trials in seven african countries . likewise, in a special program to support access to innovative products in low-income countries, swissmedic deploys the marketing authorization for global health products (maghp) program. it performs similarly to the ema's article regarding assessment of product development packages and marketing authorization applications in conjunction with who and the nras from the countries where the product will be used. the difference is that the product, if the assessment is positive, will receive a swiss marketing authorization even if it will not be used in switzerland. in the eu, as the product is not intended for use in the eu, the result of the process is a positive opinion, but not a european marketing authorization. both of these programs aspire to facilitate a reduction in timelines for development and authorization of products intended solely or primarily for use in lower income countries, thus making needed medicines available faster . in the near future, regulatory agencies in lower income countries, that have not yet done so, must also become equipped to undertake post-eul oversight and vigilance surveillance requirements. in addition, they must develop local frameworks to allow emergency use authorization of products during public health emergencies, and that allow the use of candidates that may lack human efficacy data but have both recognized animal efficacy data and initial human safety data. accelerating availability of effective, safe, quality products is essential in a public health emergency. depending on the context, the feasibility of clinical trials, the strength of animal or clinical surrogate data, and the initial safety profile of the product, one facilitated pathway may be pursued over another or several of these pathways may be pursued simultaneously or sequentially. in these situations, regulators are generally quite willing to discuss putative development plans and regulatory pathways with product developers. developers should take advantage of such opportunities: these are key to accelerated product development, marketing authorization assessment, and patient access under these facilitated pathways. generally, this is an iterative process, with decisions being made and modified as further data regarding the emergency and product become available. meeting demand via these pathways in the case of a large public health emergency will require robust pre-clinical studies and significant at-risk investment in scaling manufacturing ahead of clinical proof of concept. because of the rapidly changing nature of public health emergencies, and the requirement for a well-validated animal model, certain pathways may not be able to be utilized in a public health emergency. historically, most regulatory pathways used in public health emergencies rely on some human efficacy data. pathways that bring together the manufacturer, nras where the product is going to be used, nras with specific needed expertize, who, and regulatory and clinical experts will accelerate the availability of needed novel medical countermeasures. the benefits of such rapid development could have major impacts, both in terms of lives saved and reduction in disease spread and intensity. received: march ; accepted: september ; understanding ebola: the epidemic the world health organization accelerating the development of medical countermeasures for the next pandemic the next epidemic-lessons from ebola experiences with and challenges afforded by expedited regulatory pathways accelerating access to new medicines: current status of facilitated regulatory pathways used by emerging regulatory authorities fda facilitated regulatory pathways: visualizing their characteristics, development, and authorization 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investigational drugs for treatment use-questions and answers guidance for industry code of federal regulations title . code of federal regulations title drugs for human use expanded access program information on the zmapp expanded access program the european medicines agency. conditional marketing authorisation guideline on the scientific application and the practical arrangements necessary to implement commission regulation (ec) no / on the conditional marketing authorisation for medicinal products for human use falling within the scope of regulation (ec) no the european medicines agency the european medicines agency the european medicines agency. european public assessment report-intelence the european medicines agency the european medicines agency the united states food and drug administration the european medicines agency. support for early access japan pharmaceutical and medical device agency. correspondence to the drug conditional early approval system the united states food and drug administration product development under fda's animal rule: understanding fda's expectations and potential implications for traditional development programs the united states food and drug administration. cder drug and biologic animal rule approvals first vaccine approval under the fda animal rule guideline on procedures for the granting of a marketing authorisation under exceptional circumstances, pursuant to article ( ) of regulation (ec) no the european medicines agency. european public assessment report-imvanex-modified vaccinia ankara virus emergency use authorization of medical products and related authorities-guidance for industry and other stakeholders c. a. termination of the emergency use authorization (eua) of medical products and devices- - h n the world health organisation. emergency use listing procedure, version the world health organization. emergency use assessment and listing procedure (eual) for candidate vaccines for use in the context of a public health emergency considerations for use of investigational drugs in public health emergencies the west african health organization. harmonization of medicines registration in the ecowas region east african commission medicines regulation harmonization the new partnership for africa's development. african medicines regulatory harmonisation (amrh the pan american health organization. pan american network for drug regulatory harmonization the asia-pacific cooperation the world health organisation regional office for africa. the african vaccine regulatory forum medicines for use outside the european union defining the strategic vision for the ema 'article ' process the european medicines agency. first malaria vaccine receives positive scientific opinion from ema guidance document scientific advice maghp author contributions l contributed equally to the conceptualization of the manuscript. s.s. designed and authored the manuscript, table and figure. a.c. and m. m.l critically revised and approved the final version the authors declare no competing interests. supplementary information is available for this paper at https://doi.org/ . / s - - - .correspondence and requests for materials should be addressed to s.s. publisher's note springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.open access this article is licensed under a creative commons attribution . international license, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons license, and indicate if changes were made. the images or other third party material in this article are included in the article's creative commons license, unless indicated otherwise in a credit line to the material. if material is not included in the article's creative commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. to view a copy of this license, visit http://creativecommons. org/licenses/by/ . /. key: cord- - abupl authors: vokó, zoltán; pitter, jános györgy title: the effect of social distance measures on covid- epidemics in europe: an interrupted time series analysis date: - - journal: geroscience doi: . /s - - - sha: doc_id: cord_uid: abupl following the introduction of unprecedented “stay-at-home” national policies, the covid- pandemic recently started declining in europe. our research aims were to characterize the changepoint in the flow of the covid- epidemic in each european country and to evaluate the association of the level of social distancing with the observed decline in the national epidemics. interrupted time series analyses were conducted in european countries. social distance index was calculated based on google community mobility reports. changepoints were estimated by threshold regression, national findings were analyzed by poisson regression, and the effect of social distancing in mixed effects poisson regression model. our findings identified the most probable changepoints in european countries. before changepoint, incidence of new covid- cases grew by % per day on average. from the changepoint, this growth rate was reduced to . %, . % increase, and to . % and . % decrease by increasing social distancing quartiles. the beneficial effect of higher social distance quartiles (i.e., turning the increase into decline) was statistically significant for the fourth quartile. notably, many countries in lower quartiles also achieved a flat epidemic curve. in these countries, other plausible covid- containment measures could contribute to controlling the first wave of the disease. the association of social distance quartiles with viral spread could also be hindered by local bottlenecks in infection control. our results allow for moderate optimism related to the gradual lifting of social distance measures in the general population, and call for specific attention to the protection of focal micro-societies enriching high-risk elderly subjects, including nursing homes and chronic care facilities. after a million confirmed and , fatal european cases, the covid- pandemic started declining in europe in april (european centre for disease prevention and control ). this much awaited decline was headed by the introduction of unprecedented "stay-at-home" national policies in most countries, including border closure, public gathering bans, school and workplace closure, and temporary restrictions on free internal movements of the citizens (european centre for disease prevention and control ). these economically and socially disruptive control measures are not sustainable on the longer term (petersen et al. ) , and gradual restart of economy and social life is now on the political agenda throughout europe (european centre for disease prevention and control ; european commission ). a european roadmap to lifting the coronavirus containment measures has been framed, proposing a gradual, slow approach backed with adequate monitoring and healthcare capacity to ensure sufficient control of potential flareups (european commission ). planning the consecutive steps is supported by general provisions and considerations of the roadmap; however, ultimately, it remains a trial-and-error-based process due to the high uncertainty in possible consequences of any change in containment measures. importantly, all national epidemic containment measures were introduced within a narrow time period in most countries (european centre for disease prevention and control ; hale et al. a) . hence, the contribution of unique interventions to the overall impact on covid- spread is hard to estimate retrospectively (imai et al. ; imperial college covid- response team report ). as an overall measure of policy response intensity, the blavatnik school of government proposed a composite stringency index, integrating rigor and scope of multiple containment and closure policies (school and workplace closure, restrictions on gathering, international and internal movements, public transport, cancelation of public events, and information campaigns) into a single numeric parameter in the - range (hale et al. a) . the same team organizes global data collection on all included indicators, as well as on economic responses and adaptations of public health systems, providing a freely available but very precious tool to overview and visualize global policy efforts. however, this stringency index has important limitations when tested as an explanatory factor of sars-cov- infection spread. first, the adopted categories of policy rigor and scope may aggregate heterogeneous policies (as illustrated by an amendment on april -see the details at (hale et al. b) ). more importantly, the stringency index is based on sterile policy decisions, while the compliance of the population with the corresponding restrictions may vary across countries and over time. the google covid- community mobility reports provide an alternative option to derive a composite stringency measure of multiple containment and closure policies (google ) . these community mobility reports provide daily, country-level (and subregional) aggregated anonymized data on time spent at different categories of places such as retail and recreation, groceries and pharmacies, parks, transit stations, workplaces, and residential areas, compared with a baseline period before the epidemic. telemonitored mobility trends are dynamic in time and reflect real-world changes in social behavior, making them promising explanatory factors in sars-cov- infection spread control analyses. recently published covid- microsimulation models based on social network data in the uk and usa revealed that epidemic suppression would require a complex intervention package including social distancing of the entire population, home isolation of cases, and household quarantine of their family members, supplemented with school closure, in intermittent periods adjusted to epidemic intensity and unoccupied critical care capacity (imperial college covid- response team report ; kucharski et al. ) . however, adaptation of these microsimulation models to other countries would require rich and solid input data on local social networks. a semi-mechanistic bayesian hierarchical model of social distancing interventions across european countries was also reported, calculating daily infections from observed death rates (imperial college covid- response team report ). the authors inferred that the combined application of five intervention types (lockdown, public events ban, school closure, self-isolation, and social distancing) could prevent about , covid- deaths in the investigated countries until the end of march . nonetheless, individual contributions of the five intervention types to the overall effect showed high uncertainty, probably because many interventions occurred on the same day or within days of each other. important limitations of this study were the assumptions on identical effect of interventions across countries and over time, and the possible over-representation of countries with more advanced epidemics (imperial college covid- response team report ). additional reassurance whether the covid- transmission was truly slowing has been warranted. to estimate the effect of social distancing on the time trend data of the epidemic, interrupted time series analysis is an alternative approach (aminikhanghahi and diane ) . this analysis can be extended to a broader range of european countries without need for sophisticated local input data collection and assumptions on between-country similarities; hence, it can broaden our current understanding of the epidemic and its association with changes in population social distance patterns. our research aims were to identify the date when the covid- pandemic started declining in each european country and to evaluate the association of the level of community mobility restrictions (social distancing) with the observed extent of decline in the national epidemics. establishing an association of telemonitored population mobility patterns with a decline in covid- spread may support policymakers in assessing the benefits of previously implemented stay-at-home policies, and in planning the gradual lifting of current restrictions. daily incidence of new covid- cases by countries was obtained from the open-access database of the european centre for disease prevention and control (european centre for disease prevention and control data ). data from european union member states and the european free trade association countries were included to focus our analyses on countries with similar sociocultural characteristics and reliable estimates of changes in daily covid- incidence. data from cyprus, iceland, and liechtenstein were dropped, due to the lack or scarcity of related google community mobility reports. data from latvia have also been dropped, because it covered only days in the observation period (see below). accordingly, the analyses included data from countries: austria, belgium, bulgaria, croatia, czech republic, denmark, estonia, finland, france, germany, greece, hungary, ireland, italy, lithuania, luxembourg, malta, netherlands, norway, poland, portugal, romania, slovakia, slovenia, spain, sweden, switzerland, and the uk. within the study period of february to april, the first day of observation was defined in each country as the last day when the number of the new cases was at least following days with less than new cases. due to missing data for some calendar days, the start of observation period was postponed to march in finland and to march in luxembourg (table ) . the most likely changepoint date was determined in each country by linear threshold regression models of the logarithm of daily cases over time, replacing zero daily cases in these analyses by one, and looking for threshold in the - percentile range of the country time series, using the threshold application of the statistical package stata . (statacorp ). the threshold regression using the logarithm of cases in linear regression with the usual gaussian, homoscedastic and independent errors is a correct method to identify the threshold, but when the extent of change is estimated the count nature of the data needs to be taken into account as the non-gaussian errors might give incorrect standard errors of the regression coefficients. therefore the extent of change at the most probable changepoint in the reported daily incidence was estimated by country via poisson regression models using poisson application of stata . . independent variables in the models were time from start of observation and time from the estimated changepoint. as the observations by country were not independent, huber/white/sandwich variance estimator was used to estimate confidence intervals. results of changepoint identification and poisson regression are summarized in table and fig. . the google covid- community mobility reports provide daily, country-level (and sub-regional) aggregated anonymized data on time spent at six different categories of places, compared with a baseline period before the epidemic and controlled for the weekday effect (google ) . in the investigated countries, largest reported decline in staying in retail and recreation, grocery and pharmacy, parks, transit stations, and workplace areas were − %, − %, − %, − %, and − %, respectively, while highest reported change in staying in residential areas was + %. these dimensions of community mobility were integrated into a social distance index. first, data on staying in parks was omitted, since its implications on social distance were considered ambiguous: staying in parks may reflect either individual or social activity. as a next step, daily change from baseline in each mobility report dimension was normalized between baseline and international maximum (see above). finally, the normalized values were averaged, yielding a country-specific daily social distance index with a baseline of and a theoretical maximum of . country-specific social distance index data over time are shown in fig. . for multivariate regression analysis, the average social distance index was estimated for a -day incubation period ending at the changepoint for each country, separately, and countries were grouped by four quartiles of this parameter ( table ) . effect of social distancing on the spread of the epidemic daily new cases were modeled via mixed effects poisson regression with gamma random effect (sutradhar and jowaheer ) in the xtpoisson application of stata . , using countries as random effect. fixed effects in the model were time from the start of observation period, time from changepoint, and the interaction between the latter and the quartiles of the average social distance index in days ending at changepoint, reflecting an incubation period of - days before diagnosis of new cases (european centre for disease prevention and control ). all analyses were conducted in stata . (statacorp ), and double-checked in r (r core team r ) using packages chngpt (fong et al. ) and hglm (rönnegård et al. ; alam et al. ) . most likely changepoints and the estimated extent of change are summarized in table and depicted in fig. . the identified changepoints were associated with statistically significant alteration in daily covid- incidence in of the investigated countries, and all significant findings exhibited a decline in epidemic spread. findings of the multinational regression analysis are summarized in table . translating the model coefficients into incidence rate ratios shows that before changepoint, incidence of new covid- cases grew by % per day (irr . ) on average. from the changepoint, this growth rate reduced to . %, . % increase, and to . % and . % decrease by increasing sdi quartiles. the beneficial effect of higher social distance quartiles (i.e., turning the increase into decline) was statistically significant for the fourth quartile. our analysis identified the most probable changepoint in the flow of the covid- epidemic in european countries and found a clear dose-response association of the observed flattening of the epidemic curve with increasing social distance index derived from google community mobility reports. countries in the highest sdi quartile achieved a statistically significant decline of the epidemic, with less and less new cases every day, while countries with the least stringent sdi increase also greatly reduced the initially high growth rate of incident covid- cases. accordingly, it can be inferred that the unprecedented "stay-at-home" national policies meaningfully contributed to the suppression of the covid- pandemic in europe. countries which achieved on average only % of the maximum observed level of the decrease in social contacts showed already a large reduction in the spread of the epidemic. on the other hand, restrictions on internal movements of the citizens are obviously not the only contributors to this decline: contact tracing and isolation, widescale use of individual protective equipment, keeping safe interpersonal distance in public places, and proper hand hygiene are all plausible contributors to stopping the first wave of the pandemic in europe (european commission ; imperial college covid- response team report ). notably, the importance of local micro-epidemic chains in the overall covid- epidemic is better and better recognized. nursing homes are known to be predisposed to having high transmission rates for infectious diseases for many reasons including crowding, sharing bathroom facilities, social contacts, and low preparedness for infection control. unfortunately, covid- does not seem to be an exception in this respect (davidson and szanton ; trabucchi and de leo ) . according to a who report on april, up to half of those who have died from covid- in the european region were resident in long-term care facilities (world health organization statement ). to sdi social distance index, sd standard deviation prevent covid- transmission in nursing homes and other chronic care facilities enriching high-risk elderly patient groups, effective local infection control measures are clearly more relevant than general interventions targeting the country population as a whole, without specific focus on critical hot spots of the epidemic. such a discrepancy between global and local containment measures may also explain the relatively small difference in the slowing of the epidemic by different level of social distancing. therefore, in parallel with the gradual lifting of country-level covid- spread control measures, special attention must be paid to ensure adequate local infection control in nursing homes and chronic inpatient care facilities, in compliance with the european roadmap to lifting coronavirus containment measures (european commission ) and the corresponding recommendations of the centers for disease control and prevention (centers for disease control and prevention key strategies ). the unprecedented "stay-at-home" national policies meaningfully contributed to the suppression of the covid- pandemic in europe, which can be detected in macro level time trend analysis. however, the importance of several other interventions introduced in parallel must be noted as well, and our findings could be shaped also by the important distinction between country-level and institution-level preparedness. our findings allow for moderate optimism related to the gradual lifting of social distance measures in the general population, and call for specific attention to the protection of focal micro-societies enriching high-risk elderly subjects, including nursing homes and chronic care facilities. code availability (software application or custom code) threshold, poisson, and xtpoisson applications of the statistical software of stata . were used for the analysis as described in the methods. author contributions jgp initiated the project and prepared the data for the analysis. zv developed the analysis plan and performed the analysis in stata . . jgp checked the analysis by repeating it in r . . . the first version of the manuscript was drafted by jgp. both authors contributed to the interpretation of the results and to the writing of the successive versions of the manuscript. funding information open access funding provided by semmelweis university (se).data availabilitypublicly available data was used for the analysis (see in the acknowledgments). conflict of interest the authors declare that they have no conflict of interest. open access this article is licensed under a creative commons attribution . international license, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons licence, and indicate if changes were made. the images or other third party material in this article are included in the article's creative commons licence, unless indicated otherwise in a credit line to the material. if material is not included in the article's creative commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. to view a copy of this licence, visit http://creativecommons.org/licenses/by/ . /. fitting conditional and simultaneous autoregressive spatial models in hglm a survey of methods for time series change point detection centers for disease control and prevention key strategies to prepare for covid- in long-term care facilities (ltcfs). nursing homes and covid- : we can and should do better rapid risk assessment: coronavirus disease (covid- ) in the eu/eea and the uk-ninth update european centre for disease prevention and control data on the geographic distribution of covid- cases worldwide a european roadmap to lifting coronavirus containment measures chngpt: threshold regression model estimation and inference google covid- community mobility reports covid- government response tracker. blavatnik school of government oxford covid- government response tracker. what's changed? adoption and impact of nonpharmaceutical interventions for covid- estimating the number of infections and the impact of nonpharmaceutical interventions on covid- in european countries impact of non-pharmaceutical interventions (npis) to reduce covid- mortality and healthcare demand effectiveness of isolation, testing, contact tracing and physical distancing on reducing transmission of sars-cov- in different settings covid- -we urgently need to start developing an exit strategy a language and environment for statistical computing. vienna: r foundation for statistical computing hglm: a package for fitting hierarchical generalized linear models statacorp. stata statistical software: release . college station: statacorp llc on familial longitudinal poisson mixed models with gamma random effects nursing homes or besieged castles: covid- in northern italy world health organization statement -invest in the overlooked and unsung: build sustainable people-centred long-term care in the wake of covid- key: cord- - rxradwz authors: kohl, claudia; kurth, andreas title: european bats as carriers of viruses with zoonotic potential date: - - journal: viruses doi: . /v sha: doc_id: cord_uid: rxradwz bats are being increasingly recognized as reservoir hosts of highly pathogenic and zoonotic emerging viruses (marburg virus, nipah virus, hendra virus, rabies virus, and coronaviruses). while numerous studies have focused on the mentioned highly human-pathogenic bat viruses in tropical regions, little is known on similar human-pathogenic viruses that may be present in european bats. although novel viruses are being detected, their zoonotic potential remains unclear unless further studies are conducted. at present, it is assumed that the risk posed by bats to the general public is rather low. in this review, selected viruses detected and isolated in europe are discussed from our point of view in regard to their human-pathogenic potential. all european bat species and their roosts are legally protected and some european species are even endangered. nevertheless, the increasing public fear of bats and their viruses is an obstacle to their protection. educating the public regarding bat lyssaviruses might result in reduced threats to both the public and the bats. the european continent is inhabited by hibernation. many bat species migrate over vast distances while others are rather territorial. all bats in europe utilize echolocation to navigate. contrary to the worldwide efforts in protecting bats, they have been increasingly gaining attention as potential reservoir hosts of some of the most virulent viruses we know. various publications reviewed bats globally as carriers and potential reservoir hosts of human-pathogenic and zoonotic viruses [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] , while hardly anything is known about human-pathogenicity of european bat viruses apart from lyssaviruses. in this review, we discuss a selection of viruses as possible threats posed by european bats to the public from our point of view. a summary of viruses that have been detected in european bats is given in table at the end of the manuscript. a more comprehensive and up-to-date list of bat-associated viruses can be found online at the database of bat-associated viruses (dbatvir) [ ] . european bat lyssaviruses (family rhabdoviridae) are the most important zoonotic bat-borne viruses in europe and have been comprehensively reviewed by banyard et al. in this special issue on bat viruses (title: lyssavirus infections of bats: emergence and zoonotic threat) [ ] . therefore, we will provide a short overview. [ , ] nyctalus noctula rhinolophus ferrumequinum hungary pcr [ ] myotis myotis germany pcr [ ] astroviridae myotis myotis germany pcr [ ] mamastrovirus italy pcr [ , ] the postulates drafted by jacob henle and robert koch in the late th century constitute a framework regarding the principles of cause-and-effect in microbiology [ ] . back then, it was comparatively straightforward to limit cause-and-effect to four postulates, although viruses had not yet been discovered nor was molecular biology developed ( table ). all of the postulates are hard to fulfill for viruses, as they do not grow on nutrient media, but require living cells for replication. when looking for viruses on a molecular level, it is necessary to consider that only the first postulate can be accomplished. studies identifying a host-pathogen relationship solely at the molecular level do not take into consideration that detection does not equal etiology. even though polymerase chain reaction (pcr) screening and metagenomic studies are indispensable and valuable tools, virologists should stay close to the henle-koch postulates when assuming a possible virulence of viruses detected in bat hosts. a plethora of coronaviruses has been detected in bats, mostly belonging to the alphaand betacoronaviruses [ , ] . the genus alphacoronavirus hosts human-pathogenic strains (i.e., human cov e and nl ); however, in this review, we will focus on selected highly human-pathogenic betacoronaviruses and their european bat virus relatives [ ] . from november until july the world was confronted with the first pandemic of the new millennium, caused by a novel coronavirus (cov) inducing the severe acute respiratory syndrome in humans (sars) [ ] [ ] [ ] . the pandemic spread from its origin, a wet-market in the guangdong province in china, through countries on five continents resulted in more than infected humans of whom more than eventually died [ , ] . the search for the animal reservoir began, identifying masked palm civets and bats as possible sources. subsequently, a plethora of diverse coronaviruses of distinct groups have been detected in various bat species around the world via molecular-biological techniques. in , another human-pathogenic coronavirus, called middle east respiratory syndrome coronavirus (mers-cov), began spreading from the arabian peninsula, so far resulting in globally laboratory-confirmed cases of infection with mers-cov, including at least deaths [ ] . dromedaries and bats are suspected as reservoirs for mers-cov [ ] . recent findings support the plausibility of dromedaries as reservoir species [ ] . although numerous studies in european bats report the presence of sars-like-cov and mers-like-cov sequences [ , [ ] [ ] [ ] ] , no final conclusion can be drawn regarding their zoonotic potential. a related virus detected in bats cannot necessarily be considered as zoonotic. a few alterations in the sars-cov spike protein enabled its binding to the host receptor ace- , thus sars-cov became capable of infecting humans [ ] . so far, the sars-like cov detected in european bats lack these alterations and thus are not predicted to be capable of infecting humans. although virus strains might be similar or related on a nucleic acid level, the distinct function of proteins is crucial when determining the host range. therefore, mere similarity is not sufficient to examine the potential of viruses to infect humans or even predict their virulence. it took ten years from the emergence of sars-cov for the first bat cov to be isolated from rhinolophus bats in china, that displayed the human ace- receptor, which enabled the virus to infect human cells [ ] . these findings provide evidence for the reservoir theory. from the european perspective, nevertheless, no sars-like cov or mers-like cov has been isolated from any european bat, nor has any transmission of sars-like cov or mers-like cov to humans been reported. the case of mers-cov is slightly different, as a sequence of base pairs with % identity to mers-cov was detected in a bat (taphozous perforates-the species identification performed was not beyond doubt, as it was based on exclusion criteria (no cytochrome b sequence of taphozous perforates is available in genbank [ ] )) in saudi arabia [ ] . this finding initiated a controversy among leading cov experts, as the journal nature recently reported [ ] . they discussed that the complete genome sequence of mers-cov obtained from the bat should confirm that the virus was indeed identical and not coincidentally just a short conserved region of the virus genome. furthermore, a prevalence study might provide insights into the distribution of mers-cov in bat populations. although taphozous perforates are not abundant in europe, climate change and environmental factors may have an effect on the future distribution of this bat species (figure ) [ ] . the case of mers-cov emergence impressively demonstrates the necessity of virus discovery and prevalence studies. with the first sequence of mers-cov that became available, bats were suspected as reservoir hosts, not only because mers-cov is a sars-cov relative, but also because previous bat virus discovery studies had provided eligible sequences of bat cov to genbank, allowing for correlations with the novel mers-cov. recently, a quasi-species of mers covs was recovered from nasal swabs of dromedaries of the kingdom of saudi arabia [ ] . the mers cov consensus genome variants from dromedaries and humans are indistinguishable, supporting the plausibility of dromedaries in the role of transmission [ ] . in , the first reported outbreak of filovirus, named lloviu virus (llov), in a european bat population occurred in france, spain, and portugal [ ] . several colonies of schreiber's bats (miniopterus schreibersii) suddenly declined due to an unknown disease. llov was found in animals that showed signs of viral infection, but not in healthy bats co-roosting in the caves (myotis myotis). llov is distinctly related to filoviruses found in african bats and was classified in as type species of the novel genus cuevavirus [ ] . unfortunately, the lack of successful isolation of llov prohibits the experimental infection of schreiber's bats to clarify whether llov is the first filovirus capable of inducing disease in bats. this would challenge the hypothesis of bats as potential reservoir hosts for other filoviruses like ebola and marburg virus. schreiber's bats are distributed in distinct lineages throughout oceania, africa, southern europe, and south-east asia (figure ) [ ] . they are thought to transmit and maintain llov across different lineages throughout their habitats, although no studies are available to prove this hypothesis. consequently, the sole demonstration of a novel filovirus sequence does not provide evidence of a possible public health threat. following the henle-koch postulates, the virus should be isolated and further characterized to draw conclusions on the evolution of filoviruses in their respective bat host. as most filoviruses are described as highly pathogenic for humans, the occurrence of llov should be carefully monitored by prevalence studies in the highly abundant miniopterus schreibersii (figure ). in , three distinct paramyxoviruses were detected in german bats, two of which were related to the proposed genus jeilongvirus (myotis mystacinus, pipistrellus pipistrellus) and one was related to the genus rubulavirus (nyctalus noctula) [ ] . another study published in the same year described another different paramyxoviruses in bats from germany (myotis bechsteinii, m. daubentonii, m. myotis, and m. mystacinus) and bulgaria (myotis alcathoe and m. capaccinii), all of which belong to the genus morbillivirus [ ] . none of the novel bat paramyxoviruses are closely related to viruses of the highly pathogenic genus henipavirus or other human-pathogenic paramyxoviruses [ , ] . there is no evidence to suggest that any of these novel paramyxoviruses are capable of infecting humans. similar to the case of the llov filovirus, virus isolates and prevalence studies in both humans and bats could improve knowledge and clarify their zoonotic potential. few studies have documented the negative results from pcr testing of european bats for other human-pathogenic viruses. for instance following generic pcr screening for flavi-, hanta-and influenza-a viruses in european bats in [ ] , testing of another central european bats for influenza-a viruses [ ] and testing european bats for hepadnaviruses in did not lead to the detection of any viral nucleic acids [ ] . pcr screening of european bats for orthopoxviruses has not revealed any known or novel virus sequences [ ] . so far, the only virus isolates (beside lyssaviruses) obtained from european bats are one bunyavirus, one adenovirus and orthoreoviruses [ , , , ] . these represent the only isolates that would allow for further characterization and potential clarification of their zoonotic potential. nevertheless, recombinant viruses, constructed on sequence information, are also valuable tools to study prevalence and pathogenicity in vitro. toscana virus (tosv) was isolated from a bat's brain in , while simultaneously tosv was isolated from sandflies in the laboratory [ ] . as tosv has never been reported in bats afterwards and no hemagglutination-inhibiting antibodies has been initially found in the bat, there is a reasonable chance that this tosv isolation may have been a cross-contamination [ ] . bat adenovirus (bat adv- ) was isolated from a bat's intestine in [ ] , and the whole genome was obtained and circumstantially analyzed [ , ] . bat adv- displays a monophyletic relationship to the adenoviruses of canids (cadv). moreover, open trading frames (orf) in the bat adv- genome and the cadv are identical and not present in other members of the mastadenoviruses. the closely related canine adv contribute to the severe kennel cough syndrome in canids and show an unusually broad host range [ ] . this provides evidence suggesting an ancestral inter-species transmission of mastadenoviruses between bats and canids. like in the case of rabies virus, which is prevalent in both bats and terrestrial mammals (e.g., dogs, raccoons, skunks, and foxes) of the americas, a continuing exchange and transmission between bats and canids or other terrestrial animals might be possible [ ] . there is no evidence of a zoonotic potential of bat adv- . in , three novel orthoreoviruses were isolated from plecotus auritus and myotis mystacinus in germany [ ] . a subsequent pcr screening obtained identical viral sequences also in other bat species: pipistrellus pipistrellus, pipistrellus nathusii, pipistrellus kuhlii, and nyctalus noctula. at the same time, a group in italy detected further orthoreoviruses in myotis kuhlii, rhinolophus hyposideros, tadarida teniotis, and vespertilio murinus [ ] . summing up the data for the reovirus isolates from germany and italy, a close relationship was revealed to the genus mammalian orthoreovirus (mrv), in particular to an orthoreovirus obtained from a dog (strain t /d ) with hemorrhagic enteritis in italy [ , , ] . no ancestral relationship was assumed here, but rather an opportunistic -behavior‖ of the novel closely related mrvs, as they were detected in various different bat species. moreover, the newly isolated mrvs are phylogenetically related to viruses capable of inducing severe meningitis in humans [ ] . recently, a study published by steyer et al. described the detection of an mrv from a child hospitalized with acute gastroenteritis in slovenia [ ] . the causative agent was determined to be an mrv with the highest similarity of . %- . % in the respective segments to a bat mrv (t /bat/germany/ / ) [ ] . this might indicate a human-pathogenic potential of strain t /bat/germany/ / . as the case of sars-cov has shown that even small changes in the genome are important for determining the host range, this has to be determined for the bat mrvs in further studies. interestingly, no contact was reported between the infected child and bats, but contact to a domestic dog was assumed [ ] . the isolated viruses will allow for a seroprevalence study (cross-reactivity and cross-neutralization with other strains) in humans, which shall be initiated to examine the prevalence of specific antibodies to bat mrvs in germany and italy (where these viruses have been found) to clarify their zoonotic potential. this is especially interesting as asian bat orthoreoviruses of the genus pteropine orthoreovirus have already been linked to potentially zoonotic respiratory diseases in humans [ , ] . rhabdoviruses of the genus lyssavirus that have been detected in europe are considerably harmful and truly zoonotic agents, inevitably causing the death of unvaccinated humans if not treated in time before onset of the rabies disease [ ] . even though bat-transmitted lyssaviruses have a fatality rate of virtually % and are suspected to be transmissible by bat biting and scratching, the reported total number of human fatalities in europe is low (n = - since ) [ ] [ ] [ ] . all described hosts of european bat lyssaviruses (eblv- and eblv- ) are synanthropic, hence sharing their habitats with humans [ ] . eblv- has been predominantly detected in eptesicus serotinus and e. isabellinus in europe, both living in buildings, roofs, and attics usually in the southern regions of europe (e. serotinus until ° north, e. isabellinus in southern portugal-e. isabellinus is a north african population of e. serotinus that is controversially but not concludingly discussed as a novel species [ ]), and male bats are reported to co-roost with multiple bat species [ ] . eblv- was also detected in v. murinus, m. schreibersii, m. myotis, m. nattereri, r. ferrumequinum, and t. teniotis. whether these bat species constitute accidental hosts infected by spillover from co-roosting e. serotinus species, or whether they are additional reservoirs, has not yet been determined [ ] [ ] [ ] [ ] ] . two human cases described by johnson et al. were confirmed as infected with eblv- , which is prevalent in european m. daubentonii and m. dasycneme [ , ] . m. daubentonii is prevalent in north-eastern europe and is frequently found co-roosting with p. pipistrellus and m. nattereri, whereas m. dasycneme is found throughout europe and in the mediterranean, co-roosting with m. capaccinii. so far, none of the co-roosting bats have been reported to carry eblv- [ ] . however, spillover transmission to other animals (stone-marten, sheep, and cat) was described for eblv- [ ] [ ] [ ] . overall, lyssaviruses prevalent in european bats pose a risk to public health, and preventive measures have already been implemented by many european countries for decades (e.g., surveillance, vaccination plans, and post exposure prophylaxis) [ ] . especially the high-risk occupational groups (i.e., bat workers, bat carers in bat bat hospitals) are at increased risk. however, lyssavirus prevalence in european bats is very difficult to determine and results are very heterogenic [ ] . the lyssavirus prevalences are considerably low, but changes of behavior as a result of a lyssavirus infection may be more likely to bring bats into contact with humans. however, it is necessary to balance the risk with the total number of fatal human cases during the last years (five cases in million people living in greater europe) [ ] . accordingly, the risk is relatively low and would probably fall to zero if people were educated appropriately. direct contact (bites and scratches) with certain bat species might be risky and require post exposure prophylaxis. only few of the european bat species are known to be reservoirs of eblv- and eblv- , but all of the european species are endangered or close to extinction. relocation or culling of bat colonies, in spite of being an obvious solution from the viewpoint of the general public, increases the risk of lyssavirus exposure and transmission and should not be considered [ ] . only education can channel public fear to avoid further threats to the bats and the general public. alexander von humboldt discovered the latitudinal gradient in species diversity as early as [ ] : the richness of species is subject to a global diversity gradient, abating from the species-rich tropics toward the higher latitudes [ ] . bats influence this gradient significantly. more than bat species have been described worldwide. although they are abundant worldwide except for the polar regions, a steep diversity gradient is present from the tropics towards the poles [ ] [ ] [ ] [ ] . are fewer viruses prevalent in european bats because of the lower abundance of species in the more temperate europe? and is the zoonotic risk posed by bats decreased accordingly? only few studies on the biogeography of microorganisms are available. these studies indicate that the latitudinal diversity gradient has either no or a top-down effect on microbial diversity [ ] [ ] [ ] [ ] [ ] . two studies hypothesized that the local diversity and dispersal of viruses is very high, though overall, the viral diversity is limited on the global scale [ , ] . therefore, no assumptions can be made regarding the viral diversity in species abundant in temperate climates. as the total number of abundant species might not be essential, the change in biodiversity might play a role. the effect of decline in biodiversity on the emergence of diseases is subject of numerous publications [ ] [ ] [ ] [ ] [ ] [ ] [ ] . basically, there are arguments in favor of two controversial theories; reduced biodiversity could either increase (dilution effect) or decrease the risk of disease transmission. for almost half of the zoonotic diseases that have newly emerged by spillover since , a preceding change in land-use, agriculture, and wildlife hunting was reported [ ] . all of the above-mentioned effects contribute to changes in biodiversity and increased contact situations between human and animal hosts, also in europe. once spillover in novel hosts has occurred, a high density of the novel host population eventually facilitates the establishment in the novel niche. thus, human overpopulation and a decreased biodiversity might be mutual factors promoting the establishment of emerging infectious diseases. in conclusion, the baas becking hypothesis from might still be appropriate: -everything is everywhere, but the environment selects‖ [ ] . until now, lyssaviruses have been the only proven zoonotic viruses in european bats and may cause rabies in humans. however, only few bat species are known to transmit lyssaviruses in europe, and the number of human cases is rather low. nevertheless, education of the general public should be intensified to avoid easily preventable infections. although viruses with zoonotic potential have been detected in european bats, no clear assumption can be made without further studies. sero-prevalence studies should be conducted on the orthoreoviruses isolated from european bats, especially as a closely related virus was detected in a diseased child in slovenia [ ] . other bat viruses detected by using molecular techniques should be isolated (e.g., mers-like cov or bat bunyavirus) to allow for characterization and follow-up sero-prevalence studies. in general, bats are special reservoir hosts because of their biological features, long-time co-evolution and high diversity of viruses that can be found. furthermore, there is neither a clearly decreased risk in the emergence of zoonotic viruses in temperate climates compared to the tropics nor a decreased risk in regions of lower biodiversity. in conclusion, many drivers of emergence in the tropics also have validity in europe. however, european bats are endangered species, and some of them are threatened by extinction. although lyssaviruses are prevalent in european 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transmission of infectious diseases biodiversity loss and emerging infectious disease: an example from the rodent-borne hemorrhagic fevers biodiversity loss and the rise of zoonotic pathogens pangloss revisited: a critique of the dilution effect and the biodiversity-buffers-disease paradigm a meta-analysis suggesting that the relationship between biodiversity and risk of zoonotic pathogen transmission is idiosyncratic host range and emerging and reemerging pathogens population biology of multihost pathogens geobiologie of inleiding tot de milieukunde the authors are grateful to ursula erikli for copy-editing and the two anonymous reviewers for their valuable comments. both authors reviewed the literature and wrote the manuscript. the authors declare no conflict of interest. key: cord- -v pvlka authors: navajas-romero, virginia; díaz-carrión, rosalía; casas-rosal, josé carlos title: comparing working conditions and job satisfaction in hospitality workers across europe date: - - journal: int j hosp manag doi: . /j.ijhm. . sha: doc_id: cord_uid: v pvlka job satisfaction is important in the tourism sector since workers’ satisfaction is key to providing high-quality service, which is very important in determining organizational success. the working conditions that influence job satisfaction depend to a large extent on the institutional context, which shows similarities in some european countries. this research aims to compare working conditions and job satisfaction among european country blocks that have similar institutional characteristics. unlike previous studies, this research adopts a comprehensive approach by considering institutional and organizational factors in the analysis of employees’ perceptions of job satisfaction. the sample is made up of workers in european countries. the results demonstrate the existence of three different models of working conditions in europe leading to differing levels of job satisfaction in tourism. these models do not correspond to the clusters identified by the previous literature, which adopts an institutional perspective. and internationally (lee and chelladurai, ) in different sectors such as in banking and the public or hospitality sector (ariza-montes et al., ; kong et al., ) . most of these studies were conducted without taking into account the institutional context. economic conditions, unemployment rate, and national level of inequality of a national territory, among other institutional factors, generate similar working conditions among countries in terms of salaries, working hours, job security, and flexibility (posada-kubissa, ; tangian, ) . working conditions are particularly context-sensitive due to their strong linkage to the industrial relations system of a country, unemployment rate, etc. (van dierendonck et al., ) . despite the existence of a supranational government in the eu, the institutional context differs across countries, and therefore working conditions and employee satisfaction are also different across europe. previous studies have classified countries according to their institutional context and identify different models of human resource management in europe (e.g., brewster and tregaskis, ; ignjatović and svetlik, ; nikandrou et al., ) . it is interesting to complement these studies that present an institutional focus with a perspective centered on organizational practices and employees´perceptions. for employment practices to create value for companies and society, they must generate job satisfaction. due to the importance of job satisfaction at individual, organizational, and societal levels, including employeesṕ erception of their job satisfaction, the analysis becomes crucial. a deep understanding of the differences in job satisfaction across europe could set the basis for a deeper discussion and formulation of novel hypotheses regarding the influence of institutional factors on working conditions. this understanding could lead companies and policy-makers to propose policies for improving working conditions in order to enhance job satisfaction and social welfare. although some studies that compare job satisfaction across european countries can be found in the literature (e.g., eskildsen et al., ; millán et al., ; pichler and wallace, ) , comparisons are made across national territories without considering the homogeneity that may exist among european countries. according to the literature, these countries can be grouped by blocks according to the similarities in their approach to the welfare state-which impacts, among its main facets, working conditions. the welfare state model of each country is determined, among other aspects, by public policies, labor regulation, and organizational practices-fundamentally, human resources management practices. hence, the literature establishes blocks of countries based on their similarities in their institutional setting and their prevailing organizational human resources management models (e.g., albareda et al., ; brookes and barfoot, ; filella, ; ronen and shenkar, ; tangian, ) . the underlying premise is that there is some convergence toward homogeneity of these characteristics of countries within the same cluster and differences with respect to the rest of the blocks. studies that analyze whether this convergence leads to homogeneity in workers´perception of labor conditions and job satisfaction across europe are rare. this study tries to contribute to this end by exploring working condition models in europe from an organizational perspective and considering workers´perceptions. this might allow identification of possible deviations between the institutionally established regulations at the national or supranational level and the patterns of interaction of the workers and organizations in the labor market. this can help us understand which models lead to higher levels of job satisfaction and whether there is convergence in this aspect in the european context. the research seeks: (i) to analyze the different models of working conditions-what likely leads to differences in perceived job satisfaction-that exist in europe; and (ii) to explore whether these models differ among the clusters of countries based on institutional characteristics identified in the previous literature. from these objectives, the following research question is derived: does the clustering of european countries according to institutional characteristics correctly reflect the differences in labor conditions and subsequently job satisfaction across europe? this article is divided into six sections. first, a review of the relevant literature is presented in the second section. next, the methodology of the research and the results are explained in the third and fourth sections. finally, a discussion of the results and the conclusions, which includes the limitations and suggestions for future research, are detailed in the fifth and sixth sections. job satisfaction is an essential aspect for firms to gain a competitive advantage in all sectors, given the central role that employees play in business success (kramar, ) . however, despite the importance of job satisfaction, there is no general agreement regarding its definition. different authors have contributed to its clarification. among the mostcited definitions is the one given by spector ( ) , who emphasizes that job satisfaction refers to the way employees feel about their job and depends on different factors. mahdieh and sotoudehnama ( ) affirm that job satisfaction depends on factors such as personal, organizational, managerial, academic, professional, and economic variables. goetz et al. ( ) underline four factors as determinants of job satisfaction: professional development, interpersonal relations, economic expectations, and working conditions. there are principally two methodologies for assessing job satisfaction: the integral measurement of a single factor and the comprehensive multidimensional measurement. the difference between the two methods lies in the fact that while the former relies on a single item to measure job satisfaction, the latter employs several factors. most research on job satisfaction at the national level adopts a multidimensional measurement approach. for instance, the descriptive work index (jdi) developed by locke et al. ( ) includes different dimensions of the job such as promotion, payment, and relationships with managers and colleagues. spector ( ) created a job satisfaction survey (jss) that contains nine dimensions: salary, promotions, additional benefits, incentives, superiors, colleagues, operating environment, intrinsic work characteristics, and communication. parent-thirion et al. ( ) developed their job quality index (jqi) from seven variables (earnings, prospect, social environment, physical environment, work intensity, skills and discretion, and work time quality) that are related to the multidimensional nature of work. the jqi has been considered for the present investigation because it is comprehensive in coverage, transparent in method, and widely employed in the research on job satisfaction and the quality of work in the european context (e.g., erro-garcés and ferreira, ; punzo et al., ; soriano et al., ) . it is the basis for the development of the sixth european working conditions survey (ewcs) which, according to grimshaw et al. ( ) , yields solid and reliable information. in , the eu launched the european employment strategy with the aim of creating more (quantity) and better (quality) jobs (ariza-montes et al., ) . ewcs asks workers about the intrinsic characteristics of their jobs: salary, hours, participation, organization, and security, among others. the ewcs has been used in previous studies in which the impact of working conditions on satisfaction is analyzed, but using different perspectives such as new technologies (castellacci and viñas-bardolet, ) , gender issues (brinck et al., ; gómez-baya et al., ) , and workers' age (berde and rigó, ; okay-somerville et al., ) . the tourism sector is characterized by high levels of seasonality, which leads to labor practices that do not favor workers' commitment and permanence in the company in the long term (hofmann and stokburger-sauer, ) . the characteristics of the job positions in the tourism sector are related to higher levels of job dissatisfaction compared to other industries, which explains why more than half of the workers in the tourism sector are dissatisfied and consider moving to other sectors (stamolampros et al., ) . factors explaining the low levels of job satisfaction observed in the tourism sector are related to characteristics of job positions and to the lack of professionalization of the human resources management in this industry (jovanović et al., ; lillo-bañuls et al., ; zopiatis et al., ) . on the one hand, the characteristics that make this sector present low levels of job satisfaction compared to other sectors are related to low salaries (earnings), long working hours (work intensity), low job security, and the scarcity of promotional possibilities (prospects) (zopiatis et al., ) . the low work time quality of the jobs in the tourism sector is associated with the continuous relationship with customers, shift work, unsocial hours, and night work (lillo-bañuls et al., ) . this, together with the scarcity of occupational health and safety practices that favor an adequate physical environment, make employees working in this sector experience difficulties maintaining a work-life balance and a healthy lifestyle that would prevent stress and not lead to low levels of job satisfaction (hofmann and stokburger-sauer, ) . this stress is increased by the lack of perceived organizational support and autonomy that characterize jobs in the tourism industry (loi et al., ; tongchaiprasit and ariyabuddhiphongs, ) . the low levels of employee recognition, centralization in decision-making, and presentism that characterize this industry are associated with a lack of professionalization of human resources in the tourism industry (nickson, ) . the degree to which employees perceive social support from their superiors (the quality of the social environment at work) and are provided with autonomy to perform their job (skills and discretion) highly determine employees' level of satisfaction and work engagement since the social support of managers and supervisors influence workers´perception of justice at the workplace (jovanović et al., ) . to obtain a comprehensive view of job satisfaction and its antecedents in the tourism sector, different dimensions must be considered. this study combines different factors that determine the quality of work (earnings, prospect, social environment, physical environment, work intensity, skills and discretion, and work time quality) to provide a holistic view of working condition that allows the comparison of the quality of work and the level of job satisfaction across europe by relying on the employee's own perspective. the eu´s regulations favor workers' mobility within europe. labor mobility is the result of different levels of national unemployment rates, salary level, flexibility, etc. (fahri and werning, ) . taking into account that the quality of employment varies across european countries, factors that strongly explain workerśmobility and differences in job satisfaction depending on the country can be observed, as indicated in the literature (e.g., leineweber et al., ; salpigktidis et al., ; thite et al., ) . these differences can be explained by the distinct institutional settings of each territory (salvatori, ) . as derived from the premises of institutional theory, coercive pressures-especially national regulations-highly determine human resource management practices, so they might lead to differences in working conditions across countries (western, ) . the different labor legislations across european territories, despite european countries sharing a supranational government, influence working conditions and job satisfaction (brewster and hegewisch, ) . according to institutional theory, in addition to the coercive pressures exerted by legislation in a country, there are normative pressures, which are related to the appropriate and desirable norms of behavior for both organizations and individuals that predominate in a country (acemoglu and johnson, ; dimaggio and powell, ; scott, ) . these pressures also vary across territories and can be determinant in working conditions. countries that present similar institutional contexts-that show similar coercive and normative pressures-might present differences in terms of employee job satisfaction. this could be the case in countries such as denmark and norway, which present both institutional and cultural similarities but significantly differ in their working conditions (bech et al., ) . reviewing the literature, it can be observed that previous research has made efforts to identify blocks of european countries according to their institutional context (e.g., albareda et al., ; brookes and barfoot, ; filella, ; ronen and shenkar, ; tangian, ) . one of the most commonly used classifications identifies four clusters of countries in europe: anglo-saxon (ireland and the united kingdom), central european (austria, belgium, germany, the netherlands, and switzerland), latin (france, greece, italy, portugal and spain) and nordic (denmark, finland, norway and sweden) (filella, ; ronen and shenkar, ) . numerous aspects of institutional context determine working conditions. pichler and wallace ( ) emphasize the key role played by four institutional factors in working conditions: economic conditions, unemployment rate, the national level of inequality, and the degree of unionization. economic conditions of a territory highly impact the labor market in terms of job rewards in both extrinsic (average wage level, working hours, etc.) and intrinsic terms (meaningful, high-skilled jobs, etc.). the national unemployment rate and the national level of inequality also influence working conditions and job satisfaction. high levels of unemployment hinder job mobility regardless of a workers' level of satisfaction. employees, even those who are dissatisfied, will remain in their jobs because of the lack of opportunities in the labor market. the scarcity of job opportunities and the excess of job demand might lead employers to offer poorer conditions in terms of salary, working hours, etc. socio-economic inequality is also a determinant of job dissatisfaction if employees perceive that similar jobs lead to great differences in economic outcomes. the degree of unionization in a country seems to be highly determinant of the average wage level and other conditions of work that influence the welfare of employees. in highly unionized countries, employees are more likely to find better jobs in terms of salary, working hours, etc. accordingly, working conditions are generally better in countries that present a solid economic situation, a low unemployment rate, and a high level of unionization. this is the case for companies in the nordic cluster, which have good working conditions in comparison with the rest of european companies (eskildsen et al., ) . this can be explained by the high level of trade union intervention in those countries, where labor reforms encourage workers' representatives to negotiate working conditions with trade unions. as indicated in the literature, another institutional characteristic that determines working conditions is the country level of regulation (gialis et al., ; keune and jepsen, ) . the level of regulation is closely related to the level of flexibility in the labor market and to the degree of job security (posada-kubissa, ). labor flexibility is negatively associated with job satisfaction and employees´physical and psychological health since flexibility is associated with low levels of job security (carr and chung, ; probst et al., ) . flexibilization comes from deregulation; job security pursues the maintenance of social advantages through a compensatory system. both depend on the country and are not only affected by economic conditions, but by collective agreements, and by the agents involved: governments, employers, and trade unions (tangian, ) . in this line, sapir et al. ( ) identified four different social systems within europe according to the level of flexibility of each country. gil-alana et al. ( ) affirm that a robust social security system is associated with low levels of inequality. from the aforementioned two premises are derived: (i) that the institutional context strongly influences working conditions and that these become a determinant factor in job satisfaction (williams and hall, ) ; and (ii) that since institutional pressures are similar in each country block-anglo-saxon, central european, latin, and nordic-similar working conditions within each cluster (intra-group similarities) and differences across clusters are expected (inter-groups differences). this is because, among other aspects, government regulations determine an organization's freedom of action regarding employees' minimum wages, training and development investments, working hours, etc. (vaiman and brewster, ) . although the influence of the institutional context on working conditions is expected, companies' freedom of action within the framework of labor regulations is also expected to determine working conditions. in this way, workers in the tourism sector of countries with similar institutional settings could present discrepancies in their working conditions and, subsequently, in their job satisfaction. providing evidence about this would justify the need to group countries according to their working conditions model, a categorization that would more accurately show the reality of the labor market from an employee's perspective. in order to address the research objectives, the methodology used to develop the empirical analysis is presented below. to investigate differences in job satisfaction and in the quality of work among countries that show significant institutional differences, we have focused on the tourism sector due to the relevant role it plays in the european economy. the data used for the research were extracted from the sixth ewcs (the most recent available). this survey contains data on , working individuals years old or older residing in private homes in one of the european countries studied ( countries of the eu; albania; the former yugoslav republic countries of macedonia, montenegro, and serbia; and turkey). this survey was developed by the european foundation for the improvement of living and working conditions ( ) (dependent on the european commission) to obtain information on the quality of work and employment in europe. to perform the analysis, countries that present significant institutional and organizational differences were selected (filella, ; ronen and shenkar, ) . the sample includes the following countries and country clusters: the united kingdom and ireland (anglo-saxon); austria, belgium, germany, the netherlands and switzerland (central european); france, greece, italy, portugal, and spain (latin); and denmark, finland, norway and sweden (nordic). the sample used in this study is formed of employees of european countries that work in the tourism sector. table shows the number of observations for each country cluster. to select workers from the tourism sector, the statistical classification of economic activities in the european community, nace codes were used. according to eurostat, the following codes were included as part of the tourism sector: (passenger rail transport and interurban); (other passenger land transport); (sea and coastal passenger water transport); (inland passenger water transport); (passenger air transport); (hotels and similar accommodation); (holiday and other short-stay accommodation); (campgrounds recreational vehicle parks and trailer parks); (restaurants and mobile food service activities); (beverage serving activities); (rental and leasing of personal and household goods); (travel agency and tour operator activities); and (other reservation service and related activities). filtering by these criteria, employees ( . % of , ) made up the sample. we based our research on the sixth edition of the ewcs, which includes the dimensions of the european jqi developed by parent-thirion et al. ( ) . this index is formed of seven dimensions that determine working conditions: earnings, prospects, social environment, physical environment, work intensity, skills and discretion, and work time quality. all the constructs used in the analysis except salary (expressed in euros) and job satisfaction (expressed on a four-point likert scale) are numerical variables expressed on a scale of values between and . according to parent-thirion et al. ( ) , the constructs were defined as follows: earnings: the importance of earnings as a motivational factor has been widely studied in the literature (suzuki et al., ) . this construct is defined as the net hourly earnings of workers. prospects: this refers to the job characteristics that contribute to a person's material and psychological needs, encompassing the need for income and for employment continuity. de witte et al. ( ) point to these factors as determinants of job satisfaction. skill and discretion: this dimension refer to the skills required for the job and the level of job autonomy. both are pointed to in the literature as relevant factors influencing job satisfaction since they enhance job identification and commitment (fregin et al., ; mateos-romero and del mar salinas-jiménez, ) . social environment: this dimension measures the social support perceived by employees (good social relations with line managers and fellow workers) and the absence of abuse in the company, which becomes especially important for workers' welfare as it moderates the negative impact of stressors (wisse et al., ) . this construct includes two constructs: adverse social behavior and social support. physical environment: this dimension refers to environmental hazards and to factors related to posture-related risks, which become relevant factors in the health of employees, a fundamental aspect of job hygiene and satisfaction (devonish, ; koh et al., ) . work intensity: this dimension refers to the intensity of work demands. high work intensity is associated with a risk of suffering high levels of occupational stress, which in turn is associated with low levels of job satisfaction (iranmanesh et al., ; rushton et al., ) . work time quality: this dimension refers to the organization and length of working time. the number of working hours, shift work, night work, etc., are determinant for the achievement of a good work/life balance, subsequently playing a significant role in job satisfaction (eagan et al., ; roy, ) . job satisfaction: the level of satisfaction is a variable included in the sixth ewcs survey and is measured as a four-point likert scale. the question is: "in general, are you very satisfied, satisfied, not very satisfied or not at all satisfied with your working conditions?". all the items used for the construction of the variables are included in the sixth ewcs and are shown in the appendix, together with the results of the reliability tests obtained with the cronbach alpha coefficient for the tourism industry. the main objective of the empirical analysis is to determine whether the classification of countries based on the institutional context adequately reflects the different models of working conditions-and subsequently differing levels of job satisfaction-existing in europe in the tourism sector, and if not, to propose a more appropriate classification of countries. to do this, based on the classifications of filella ( ) and ronen and shenkar ( ) , a comparison of working conditions among countries of the same clusters (intra-group comparison) is made. the existence of a high heterogeneity among countries of the same block would indicate an inappropriate grouping of countries located within the same institutional block. this analysis will be completed with an inter-group comparison, in which a high homogeneity in the working conditions of countries of different blocks would indicate a reduced discriminatory capacity among the blocks. therefore, a high intra-group heterogeneity and a reduced inter-group heterogeneity would allow us to conclude that the classification made by previous studies does not correctly classify countries according to the labor conditions perceived by workers. next, through a two-step cluster analysis, a new classification is proposed that improves intra-group homogeneity and inter-group heterogeneity. the suitability of this new group of countries will be evaluated using the methods previously described. the normality of these variables was previously checked for the selection of the method of analysis. to address the research objectives, both inter-group and intra-group differences have been analyzed for both job satisfaction and working conditions. first, the analysis of intergroup differences-among country blocks-has been performed using the mann-whitney test. this technique allowed a comparison of the level of job satisfaction among country clusters (anglo-saxon, central european, latin and nordic). second, the existence of significant intragroup differences among countries within the same cluster in the level of job satisfaction have been studied using the mann-whitney and kruskal-wallis tests due to the ordinal nature of this variable. as the mann-whitney test can only be used to make comparisons between two groups, it has been employed to test the intra-group differences in the level of satisfaction within the anglo-saxon cluster (between ireland and the united kingdom). since the kruskal-wallis test allows comparing more than two groups, it was used to analyze the existence of intra-group differences for the central european, latin, and nordic clusters. working conditions have also been compared among country blocks (inter-groups) and among countries within the same block (intragroups). first, the analysis of inter-group differences in working conditions has been performed using the t-student test. this technique allowed the comparison of the working conditions among all the country blocks. second, as working conditions (earnings, prospects, social environment, physical environment, work intensity, skills and discretion, and work time quality) are numeric variables and normally distributed, t-student and analysis of variance (anova) have been used to analyze the intra-group differences. as the t-student test can only be used to compare two groups, it was employed to analyze the intragroup differences of working conditions within the anglo-saxon cluster. as anova allows comparisons among more than two groups, it was used to assess the existence of intra-group differences among the central european, latin, and nordic blocks. the effect sizes have been estimated with the statistic proposed by rosenthal ( ) for the mann-whitney contrasts ( . , . , and . are used to indicate small, medium, and large effect sizes); cohen's d statistic for t-student contrast ( . , . , and . are used to indicate small, medium, and large effect sizes), and η statistic for the anova test ( . , . , and . are used to indicate small, medium, and large effect sizes) proposed by cohen ( ) . a statistic is used for the kruskal-wallis test ( . , . , and . are used to indicate small, medium, and large effect sizes) (tomczak and tomczak, ) . the existence of significant intra-group differences and limited differences among blocks of countries that present different institutional settings justifies the need for a new classification of european countries. to create this new grouping, a two-step cluster analysis has been developed. to confirm the validity of the proposed clusters, the intra-group and inter-group differences in the level of job satisfaction and in working conditions have been analyzed using the same statistical techniques previously explained. the descriptive analysis of the data shows that the average age of employees of the sample is heterogeneous, standing at just over years, with a standard deviation of . years. the male gender is slightly predominant; they represent . %, compared to . % of women, which contrasts with the existing proportion in this sector at the european level, where these proportions are inverse. secondary education is the predominant level of education among workers in the sample ( . %), followed by university studies ( . %), and primary education ( . %). the most represented sub-sectors in the sample are "beverage serving activities," which represent the majority group ( . %), "passenger rail transport and interurban" and "other passenger land transport" ( . %), and accommodation ("hotels and similar accommodation," "holiday and other short-stay accommodation," and "campgrounds recreational vehicle parks and trailer parks") ( . %). following the international standard classification of occupations (isco- ) based on oecd ( ), . % of the workers in the sample are "white collar" employees, of which less than a quarter are highly qualified. within the "blue collar" employees-who represent . % of the total sample-only . % are considered highly qualified. presuming that the institutional environment is a factor that could significantly affect the degree of satisfaction of workers, in particular those who work in the tourism sector, we have explored the levels of job satisfaction across country blocks that present institutional differences. using the mann-whitney test, we analyzed the differences among working conditions in country blocks with different institutional contexts. the results show that there are mainly significant differences in the level of satisfaction in the latin countries with respect to the rest of the blocks, while the differences among the rest of the blocks are not significant. in addition, the effect size is very small, even in the case where the differences are significant (table ) . hence, there is a high homogeneity in job satisfaction across country blocks that present different institutional settings. when analyzing the intra-block differences, within the nordic cluster, denmark and finland do not present any unsatisfied employees. about % of employees present high and medium-high levels of satisfaction in austria and switzerland (within the central european cluster), the latter not presenting any unsatisfied employees (see table ). the analysis of intra-group differences shows that these differences are significant; therefore, a lack of homogeneity in job satisfaction among countries in the same block is observed, mainly in the central european and nordic blocks, in which the effect size is medium. inter-groups differences in job satisfaction. mann-whitney test. p-value (effect size). accordingly, differences in the degree of job satisfaction among countries within the same block are found, indicating high intra-group heterogeneity. the differences in working conditions among the blocks of countries identified in the literature-based on their institutional characteristics-were also studied. the results show that the latin cluster presents significant differences with respect to the rest of the blocks in all the analyzed variables (except the social environment variable), with some effects of medium size. the results show the absence of significant differences between the anglo-saxon cluster and the central european block in all the variables studied. the same is observed when comparing the former with the nordic group, except in the labor expectations and the physical environment variables, although with a small effect. the differences between the nordic and the central european blocks are reduced since, in addition to finding differences in the previous variables, significant differences are also observed in the skills needed to develop the work, although with a small effect (see table ). hence, there is a high homogeneity in the working conditions across country blocks that present different institutional settings. comparing the working conditions of the countries within each cluster, ireland and the united kingdom (anglo-saxon block) show a great homogeneity in all variables except salaries-workers in the tourism sector in the united kingdom receive higher salaries than in ireland. however, differences among countries of the same block are significant if we analyze the rest of the blocks, as can be extracted from the results of the intra-group anova test (see table ). among the nordic countries, significant differences are observed in the prospects, physical environment, work intensity, and skills and discretion variables. the differences found among the countries of central europe are also significant. a high disparity in wages across countries within this block can be observed, motivated by the high average salary in switzerland, followed by the significant differences in job prospects, in the social environment, and in the skills required for the jobs. hence, the results indicate the existence of a high degree of heterogeneity in the working conditions of countries within the same block. based on the previous results which show differences in working conditions among the countries of the same block and scarce differences among blocks established according to their institutional characteristics (with the exception of the latin cluster), we propose the creation of a classification of countries according to the similarity in their working conditions in the tourism sector, specifically from the seven jqi dimensions (earnings, prospects, social environment, physical environment, work intensity, skill and discretion, and work time quality). to create this new clustering, a two-step cluster analysis was performed (see table ). the results of the cluster analysis show that, on the one hand, there are countries such as greece and spain (group ) that show worse working conditions and, consequently, lower levels of job satisfaction in comparison with the rest of the countries. at the other extreme are denmark, finland, france, and sweden (group ), which present the most advantageous working conditions and the highest degree of job to confirm the validity of these results (average silhouette value is greater than . ), the working conditions of the groups created and the job satisfaction among blocks and within blocks are analyzed. regarding the latter, significant differences between clusters in terms of working conditions and job satisfaction are observed (see table ). comparing job satisfaction among blocks, significant differences are observed. likewise, analyzing the working conditions among blocks, differences among all of them exist, with the exception of clusters and , which show similarity in their work time quality; and between clusters and , which show similarity in physical environment and work intensity. there is a high heterogeneity in the variables related to working conditions in the three groups identified, endorsed by mediumhigh effect sizes in many of the comparisons that are also higher than the effects found in the original blocks identified in the literature. regarding the differences across countries within each cluster, it can be observed that there are no significant differences in job satisfaction among the countries that are part of the same block (see table ). regarding working conditions, a high degree of homogeneity is observed. countries in groups and show the greatest homogeneity in working conditions. although there are significant differences, especially in group , the effect sizes are small. this is observed in the anova test (except in earnings and, to a lesser extent, in prospects in group ), and in the rest of the tests performed, as shown in table . the workers in the three defined blocks are homogeneous in terms of characteristics such as age, gender, seniority in the company, and the percentage of self-employed people, as shown in table . studying the working conditions of each block, a great disparity between the salaries of groups and is observed. the group composed of greece and spain presents lower values in all variables except social environment and work intensity. groups and show similar results, but working conditions are slightly more favorable in group . this group presents better results with respect to the rest in skills and discretion, intensity, and prospects, while group shows more favorable conditions in the social environment and physical environment dimensions with respect to the rest. this research identifies a novel grouping of european countries according to the working conditions prevailing in the tourism sector. the differences among country clusters are manifested in different levels of employee satisfaction since the institutional context greatly influences working conditions, which in turn determines job satisfaction (salvatori, ; western, ) . despite the relevant role of the table proposed country blocks according to their working conditions in the tourism sector. group austria -belgium -germany -italy -ireland-netherlands -norway -portugal -switzerland -united kingdom group denmark -finland -france -sweden group greece -spain variables group group group work intensity physical environment work time quality prospects skills and discretion social environment earnings institutional context-where planning and policymaking occur-in shaping working conditions, this issue has received little attention in the literature on tourism. studies focused on institutional context and working conditions and job satisfaction in the tourism industry are rare. according to western ( ) , working conditions are highly influenced by national regulations-and especially by labor regulations-and therefore by the institutional context. the strength of unionization becomes an important factor influencing job satisfaction because employees' wellbeing is highly determined by salary and work intensity, among other working conditions, which are especially influenced by the levels of unionization (pichler and wallace, ) . since strong unionization in a country can lead to better working conditions, the relevance of the institutional context as an antecedent of working conditions and job satisfaction must be highlighted. classifying european countries according to their working conditions can set the basis for a deeper understanding of the factors that determine job satisfaction in the tourism industry in different territories. as has been concluded from the analysis, a classification of countries based on their institutional characteristics as proposed by the previous literature (e.g., albareda et al., ; brookes and barfoot, ; filella, ; ronen and shenkar, ; tangian, ) does not group countries correctly according to working conditions and job satisfaction perceived by workers. few differences in worker satisfaction among countries that have different institutional settings and large differences among countries of the same institutional context have been found. similarly, countries of different institutional environments have similar working conditions, while countries of the same context present large differences in working conditions. these results point to the need to propose a new classification or clustering of european countries according to their prevailing working conditions and job satisfaction levels. although the comparison of job satisfaction across european countries has been studied by academics, previous studies have analyzed individual countries without considering the existence of homogeneity among countries and the existence of differentiated blocks in terms of their institutional setting. this research proposes a novel classification of countries according to prevailing labor conditions in each territory-what marks differences in job satisfaction across country clusters. one of the key aspects that determines working conditions is labor flexibility, and this depends to a large extent on institutional context (posada-kubissa, ) . tangian ( ) affirms that policies that enhance flexible employment are incompatible with achieving employment security. carr and chung ( ) propose that in countries where the levels of labor flexibility are high, employment security policies should be implemented to increase employees' security. therefore, different levels of employment protection and labor flexibility determine different social systems. despite the eús supranational government, there are differences in social systems across countries (brewster and hegewisch, ) . sapir et al. ( ) identified four social models in europe, each emphasizing security versus flexibility to a different extent: flex-insecure, inflex-secure, inflex-insecure, and flexsecure. according to our analysis, group corresponds to two groups of inflexible countries according to sapir's classification: the continental cluster (inflexible and secure: austria, belgium, germany, italy, norway, the netherlands, and switzerland) and the countries included in the anglo-saxon block (inflexible and insecure: ireland, portugal, and the united kingdom). the former are countries characterized by high income inequality, low-wage jobs, high levels of employment protection, low job security, and by early retirement pensions (sapir et al., ) . according to the previous characteristics and inspired by sapir et al. ( ) , we propose to call group as inflexible group. according to probst et al. ( ) , this model was considered to be effective in reducing poverty but ineffective in job creation in the long term. on the contrary, the anglo-saxon model is characterized by low-wage jobs, low job security, and high levels of income inequality. this model was effective in creating employment opportunities but ineffective in reducing poverty. group resulting from our analysis corresponds to the scandinavian model (denmark, finland, france, and sweden) . this country cluster is characterized by a robust social security system. although job protection is low, employment security is high in comparison to the rest of the blocks. this model enhances job creation and a high standard of living. the countries grouped in this cluster present similar levels of employment protection and low levels of inequality (gil-alana et al., ) . therefore, following sapir et al. ( ) , we propose to call this country block as flex-secure. group resulting from our analysis, the so-called mediterranean working conditions and demographic characteristics in the proposed blocks. model (greece and spain), emphasizes employment protection and early retirement pensions (probst et al., ) . inspired by sapir et al. ( ) , this cluster could be called flex-insecure because both countries in this group show high levels of flexibility and insecurity. according to our results, greece and spain show homogeneity in their working conditions. these countries experienced a deep recession after , leading to an economically inferior position within europe. they are characterized by their weak institutions and the fiscal balance programs that have been implemented by their governments following the recession. both countries have been highly affected by prolonged austerity policies and present the highest levels of unemployment in comparison to other european countries ( . % in greece and . % in spain), according to eurostat ( ) . this can be an important factor that determines the differences found in this research in comparison with the blocks identified by the literature, which groups these two countries according to the institutional and organizational characteristics. while filella grouped italy, france, and spain within the mediterranean cluster in , the socioeconomic development of each country has been different in the past decades. while france and italy have improved their working conditions, spain has remained among the countries with low job security and high flexibility in its labor market, which is reflected in the lowest levels of job satisfaction, showing more similarities to greece in terms of working conditions and job satisfaction than to italy and france. two main motivations led us to focus our analysis on the tourism sector: its high weight in the economy of european countries (world tourism organization, ) and its characteristics that entail high levels of precariousness (jovanović et al., ) . the results of the empirical analysis show that classifying countries according to their institutional setting does not properly reflect the differences in working conditions and job satisfaction across europe. this study proposes a novel classification of european countries according to working conditions in order to understand the differences in job satisfaction in different european countries from an employee perspective. the results point to differences among countries that present similarities in their institutional context. this is observed in the higher levels of satisfaction that countries such as france, italy, and portugal present in comparison with spain and greece (all of them belonging to the same block according to previous studies). the great differences among countries that belong to the same block and the small differences in working conditions among the countries of different blocks (with the exception of the latin cluster) lead us to posit the need to propose a novel classification of countries according to their working conditions. our research results show the existence of different models of working conditions in europe that go beyond the national borders of each country. the existence of three differing working conditions models-and subsequent differences in the levels of job satisfaction-are determined not only by institutional factors, which are similar among some european countries, but by other factors that need to be further analyzed such as companies' freedom of action in labor policies and workers' perceptions. this follows from the results of our study, which show that the grouping of countries according to their institutional context does not correspond to the grouping of countries according to their working conditions. therefore, it can be inferred that working conditions are not only a reflection of the institutional characteristics of the territories, but that other factors must be explored to understand the differences in working conditions and job satisfaction across europe. although previous classifications of european countries according to their institutional context and the model of managing employees exist (e.g., brewster and tregaskis, ; filella, ; ignjatović and svetlik, ; nikandrou et al., ) that take into account different aspects such as regulatory framework, economic and legal characteristics, and the type of educational system prevailing in each country, our research highlights the need to complement these studies with the employee's perspective. human resources policies are instruments that seek to ensure the proper functioning of organizations, but this will not be achieved if these policies do not generate job satisfaction. hence, the relevance of complementing studies that adopt an organizational perspective with the employee's perception of their working conditions and level of job satisfaction. the research makes several contributions to the literature. first, studies on the relationship between the institutional framework and working conditions in the tourism sector are rare. previous research does not explore the differences between the framework in which the working conditions are developed (which is highly influenced by the institutional context where the company operates) and the labor conditions developed at the organizational level, both determining job satisfaction. previous works that classify countries according to their institutional characteristics have only considered the framework in which working conditions are developed, ignoring that organizational management highly determines working conditions. in this vein, this study complements existing literature by proposing a novel classification of european countries based on the working conditions developed at the company level and by considering workers´perceptions about these conditions and their job satisfaction. on a practical level, the research shows how european countries are grouped according to workers' perceptions of their working conditions in the tourism sector. the results show that, although the institutional context is decisive in working conditions, these conditions are not determined entirely by these factors since there are territories with similar institutional settings but with substantially different working conditions. grouping european countries according to their homogeneity in working conditions is particularly interesting for understanding international differences in job satisfaction since work satisfaction is a direct reflection of organizational policies and practices and the extent and character of institutionalized labor norms and regulations. these results have implications for organizations and policy-makers. for organizations, assuming the freedom of movement of workers in europe, companies can attract talented employees from different european countries if they improve their working conditions by assimilating them to the territories with higher levels of job satisfaction. for european policy-makers, interesting conclusions might be drawn from this research. to advance the eu convergence, it is necessary to homogenize the working conditions of the european countries, aiming to reach those conditions that achieve the highest degrees of job satisfaction. this will have benefits not only at the individual level, but also at the organizational and social levels. this need is especially emphasized in the uncertain context in which the tourism sector finds itself due to the covid- pandemic. it is difficult to predict the structural changes that the economic crisis expected after the pandemic will generate in the tourism sector, but it is expected that demand could contract in the near future due to the economic crisis predicted by international organizations such as the international monetary fund ( ). the expected contraction in demand could be seen as an opportunity to create a more sustainable tourism model that prioritizes quality over quantity, a more balanced tourism model that distributes its value more equitably and fairly among the different stakeholders. considering the fundamental role played by employees in the quality offered in the tourist service and their important contribution to business success in this sector, a model based on quality must be accompanied by better working conditions that result in greater employee wellbeing. despite the usefulness of this study, the results should be taken with caution due to the following methodological limitations. in the first place, job satisfaction is measured through self-perception, which can generate some bias in terms of the use of variables with an objective nature. second, the problem of comparing countries involves the bias that is introduced regarding different variables such as salary, which cannot be compared in absolute terms without considering the cost of living, and the expectations of employees in each country. future research could include perception variables about satisfaction with a salary instead of the salary in absolute terms to make the data comparable across countries. the classification of countries proposed by this study sets the basis for a deeper discussion on the factors-beyond the regulatory pressures that shape the institutional context-that influence working conditions. therefore, future research could explore factors such as the culture that might be similar in each of the clusters identified and that can be determinants of job satisfaction. finally, exploring job satisfaction in sectors different from tourism might lead to different groupings due to the specific characteristics of each sector. therefore, future research could replicate this study in other industries. unbundling institutions public policies on corporate social responsibility: the role of governments in europe the price of success: a study on chefs' subjective well-being, job satisfaction, and human values decent work as a necessary condition for sustainable well-being. a tale of pi (i) gs and farmers a 'civic turn' in scandinavian family migration policies? comparing denmark, norway and sweden job satisfaction at older ages: a comparative analysis of hungarian and german data institutional pressures and 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desde el modelo abolicionista (critical reflections on prostitution from the abolitionist model) psychological capital: buffering the v longitudinal curvilinear effects of job insecurity on performance job satisfaction in the "big four" of europe: reasoning between feeling and uncertainty through cub models clustering countries on attitudinal dimensions: a review and synthesis parametric measures of effect size banana time" job satisfaction and informal interaction burnout and resilience among nurses practicing in high-intensity settings burnout syndrome and job satisfaction in greek residents: exploring differences between trainees inside and outside the country labour contract regulations and workers' wellbeing: international longitudinal evidence an agenda for a growing europe. the sapir report the adolescence of institutional theory mediating role of job satisfaction, affective well-being, and health in the relationship between indoor environment and absenteeism: work patterns matter! job satisfaction: application, assessment, causes, and consequences job satisfaction and employee turnover determinants in high contact services: insights from employees' online reviews earnings, savings, and job satisfaction in a laborintensive export sector: evidence from the cut flower industry in ethiopia is work in europe decent? a study based on the th european survey of working conditions is europe ready for flexicurity? empirical evidence, critical remarks and a reform proposal internationalization and hrm strategies across subsidiaries in multinational corporations from emerging economies-a conceptual framework the need to report effect size estimates revisited. an overview of some recommended measures of effect size creativity and turnover intention among hotel chefs: the mediating effects of job satisfaction and job stress how far do cultural differences explain the differences between nations? implications for hrm linking hrm practices and institutional setting to collective turnover: an empirical exploration institutions and the labor market tourism and migration: new relationships between production and consumption catering to the needs of an aging workforce: the role of employee age in the relationship between corporate social responsibility and employee satisfaction european union tourism trends job involvement, commitment, satisfaction and turnover: evidence from hotel employees in cyprus key: cord- -sxl g p authors: mathews, fiona title: chapter zoonoses in wildlife: integrating ecology into management date: - - journal: adv parasitol doi: . /s - x( ) - sha: doc_id: cord_uid: sxl g p zoonoses in wildlife not only play an important ecological role, but pose significant threats to the health of humans, domestic animals and some endangered species. more than two‐thirds of emerging, or re‐emerging, infectious diseases are thought to originate in wildlife. despite this, co‐ordinated surveillance schemes are rare, and most efforts at disease control operate at the level of crisis management. this review examines the pathways linking zoonoses in wildlife with infection in other hosts, using examples from a range of key zoonoses, including european bat lyssaviruses and bovine tuberculosis. ecologically based control, including the management of conditions leading to spill‐overs into target host populations, is likely to be more effective and sustainable than simple reductions in wildlife populations alone. bovine tuberculosis. ecologically based control, including the management of conditions leading to spill-overs into target host populations, is likely to be more effective and sustainable than simple reductions in wildlife populations alone. parasites and pathogens in wildlife are a natural part of biodiversity. an abundance of theoretical studies indicate that they have key roles in ecological and processes, including the regulation of population size may and anderson, ) and the maintenance of genetic diversity (may and anderson, ; read et al., ) . occasional field experiments have also demonstrated impacts of sublethal infections on reproductive rates (munger and karasov, ) and susceptibility of wildlife to predation (hudson et al., ) . yet the vast majority of empirical studies consider wildlife pathogens and parasites only if they threaten the health of humans or their domestic animals, and often overlook their natural history. partly this reflects the priorities of funding agencies; and their concerns are not trivial. more than % of emerging (or re-emerging) pathogens of humans are thought to have wild animals as their natural reservoirs (taylor et al., ) . examples include sars-cov (severe acute respiratory syndrome coronavirus), avian influenza a (h n ) virus (bird flu), nipah virus, hantavirus and west nile virus. the economic imperative for controlling zoonoses that affect domestic stock can be very strong. for example, bovine tuberculosis (btb) in the united kingdom undoubtedly has a reservoir in wild badger populations, and the direct cost of the disease to agriculture is projected to reach £ billion by (department for environment, food and rural affairs (defra), ). it has been recognised relatively recently that disease can also pose a serious threat to the survival of endangered wildlife (lyles and dobson, ; may, ; smith, ) . this can either be through direct mortality, where losses are greater than the birth rate, or through effects on birth rate, longevity and survival, which suppress the population size to a level that renders it susceptible to extinction by stochastic effects (table . ). generalist pathogens with a wide host range are particularly problematic, since even virulent species can persist in alternative hosts while driving the rare host to extinction (begon and bowers, ) . although remaining low on the list of priorities compared with other threats such as habitat loss, efforts are therefore sometimes now made to control infectious diseases for conservation reasons. examples of recent successes include the control of canine distemper virus in black-footed ferrets (mustela nigripes) (williams et al., ) , and rabies in african wild dogs (lycaon pictus) (hofmayer et al., ) and ethiopian wolves (canis simensis) (haydon et al., ) . in europe alone, there are at least zoonotic parasites and pathogens in wildlife that are known to be important either to public health or the agricultural economy (artois et al., ) . for many other infectious agents, such as cryptosporidium parvum, the epidemiological role of wildlife is unknown. despite the many attempts to control zoonoses in wildlife, the success rate is poor. typically, measures are adopted as crisis management (usually in the form of culling) following an outbreak, with little understanding of the ecology of species or its relationship to the pathogen. crisis management also means that proper scientific designs with appropriate controls are often lacking; it is therefore difficult to evaluate the effectiveness of a given intervention. even where there is a long history of attempts to control a disease through the management of a wildlife reservoir, the results have not been encouraging. for example, efforts over the last years to control btb in cattle in the united kingdom by culling of badgers has failed to yield significant benefits, with analyses of the recent randomised controlled trial of badger culling concluding that culling could not contribute meaningfully to future control strategies for btb (donnelly et al., ) . similarly, the culling of foxes has been discounted as a means of rabies control in western europe (blancou et al., ) . however, success has been achieved through the use of widespread vaccination (administered via bait) (aubert, ) . rabies is currently the only example of a widespread strategy of vaccination being favoured over the control of the host species (artois et al., ) . this review considers the ecology of zoonoses in wildlife and the links between infection in wildlife and humans or livestock. it proposes that a shift to ecologically based control, explicitly considering the natural history of wildlife hosts and their pathogens, is crucial in minimising the risk presented to humans, domestic animals and endangered species from zoonoses. this approach will also yield benefits for the conservation and welfare status of wild animals. notwithstanding the complexities of specific relationships, the probability of a zoonosis being passed from wildlife into another host population, be it humans, domestic stock or an endangered wild species, is always influenced by several key parameters. these are the intensity of infection in the reservoir hosts; the size, or depending on the case, the density, of the infected host population; the degree and nature of the contact between infected individuals (or infectious particles in the environment such as infected faeces); and the susceptibility of the in-contact animal. (for indirectly transmitted parasites and pathogens, the role of vectors and/or intermediate hosts must also be considered.) whether the zoonosis persists after initial invasion is also determined by the new host's population size. a great variety of models has been developed to describe the transmission dynamics of macro-and microparasites, taking into account the nuances of particular host and parasite population structures (diekmann and heesterbeek, ; heesterbeek and roberts, ; scott, ; scott and smith, ). yet empirical research has lagged behind the theoretical advances. the legacy of researchers like elton and chitty (chitty, (chitty, , elton, ; elton et al., elton et al., , , who sought not only to describe pathogens but to understand their ecological role, has not been sustained. (there are a few notable exceptions, including the long-term studies of small mammals in the north of england (beldomenico et al., a,b; feore et al., ) ; rodent reservoirs of hantavirus in the united states (calisher et al., ; mills et al., ) and macroparasite infections in laboratory models (ehman and scott, ; scott, ; scott and anderson, ; scott and smith, ) .) this deficiency was noted in a key text in the field in (grenfell and dobson, ) and again in the follow-up publication in (hudson et al., ) . we lack even species lists of parasites and pathogens for most, if not all, wild animals. while pathogens that affect international trade are reported to the world organization for animal health (oie), and many of these affect wildlife (see artois et al., ) for the lengthy list of those likely to affect wildlife in europe), there is no agreed systematic programme of surveillance (kulken et al., ) . even where programmes exist, they lack integration with surveillance in humans and domestic animals at both local and international scales. disease surveillance in wildlife is usually driven by outbreaks in humans or domestic animals (childs, ) . virulent pathogens are, therefore, more likely to be detected than more benign ones (williamson et al., ) . such studies are also, by their nature, not designed to screen for a range of pathogens, so opportunities to investigate the epidemiology and ecology of coinfections are often lost. systematic surveillance of representative samples of the population is difficult and time consuming. yet prevalence estimates can be seriously skewed if the only data available are derived from passive surveillance of carcasses. not only are estimates likely to be too high if they are based on samples of wildlife found dead or sick by the public, but even road kills and game bags are likely to over-represent certain population classes (such as dispersing juvenile male mammals) and animals in compromised health. disease-responsive surveillance also offers little information on the frequency with which transfer events are likely to occur. for example, many of the 'spectacular' epidemics derived from bat viruses, such as hendra virus, nipah virus, sars-cov-like virus, have been observed only a small number of times. we do not know why this should be the case. is transfer of zoonoses from bats to terrestrial vertebrates generally rare due to a lack of appropriate contact? or is there regular inter-specific transmission of other viruses but these go undetected because they lack the extreme pathogenicity of hendra and nipah viruses to stimulate screening efforts? pro-active surveillance of wildlife and of apparently healthy human or livestock populations could help answer these questions. structuring and species specificity of a pathogen in wildlife hosts? screening for european bat lyssaviruses in europe is an exemplary case of research stimulated by public health concerns. the first recorded european bat rabies case was in hamburg in and several other cases were identified subsequently (king et al., ). yet surveillance of bats was not really pursued until a woman in denmark was bitten by a serotine bat (eptesicus serotinus) infected with european bat lyssavirus (eblv ). since then more than rabies-positive bats have been identified across europe; the vast majority being serotine bats infected with eblv- (harris et al., ) . in the united kingdom, screening efforts were intensified following the death of a man in scotland from eblv- in , after apparent contact with many bats in the united kingdom and europe (fooks et al., ) . in contrast with classical rabies (rabv) there is now good evidence that at least some bats (and possibly other animals) can produce neutralising antibodies and survive eblv infection for at least years (serra-cobo et al., ; van der poel et al., ) , and experimental models suggest that eblv- might be inherently less virulent than eblv- (vos et al., ) . eblv- also appears to have a much more restricted geographical range than eblv- , and small numbers of positive bats have been identified in the united kingdom, switzerland, the netherlands, denmark and germany (department for environment, food and rural affairs (defra), ; racey et al., ; vos, ; vos et al., ) . these cases have all been in the closely related daubenton's (myotis daubentoni) and pond bats (m. dasycneme). structuring of eblvs therefore is apparent from these data both across geographical areas and across species. the serotine bat occurs over most of europe, extending north to latitude (england south of the wash estuary, denmark and southern sweden); daubenton's bats are common across europe; and the pond bat is present in a wide band across central and eastern europe (between and latitude; absent from the united kingdom (schober and grimmberger, ) ). yet neither eblv- nor eblv- appears throughout their hosts' ranges. some suggest that in the case of eblv- , this may be because long-distance travel is uncommon in serotines, the primary host (vos et al., ) . yet the species is widely distributed, and it is unlikely that there are gaps between populations that could not be travelled with relative ease; dispersing movements of up to km have also been recorded (hutterer et al., ) . interestingly, eblv- also appears to have a patchy distribution, despite its host species, at least in continental europe, being migratory over long distances (vos, ) . whether the geographical distribution is, in reality, less patchy than it currently appears requires co-ordinated surveillance effort and a willingness by statutory authorities to publish test results even if they are negative. it is clear that active surveillance (systematic screening of bats in the wild) has been undertaken in a few countries only, and passive surveillance (submission of dead bats by members of the public for screening) has involved few, if any, animals in a number of european countries, including portugal, ireland, greece, the czech republic and slovakia (racey et al., ) . a range of european bats, most of which are common and widespread, has been identified as having active eblv infections in addition to the key hosts (table . ). it is striking then that the vast majority of reported cases come from just three species. undoubtedly, the numbers of bats of each species submitted by the public does not match their abundance in the wild, but is influenced by the closeness of their contact with humans (and their cats, which are a major cause of bat mortality). for example, few woodland specialists have been submitted, whereas bats that frequently roost in houses, particularly pipistrelles, long-eared bats and possibly serotines, are over-represented (harris et al., ) . even active surveillance does not attempt comprehensive surveys of all species in proportion to their abundance: instead, it focuses on the three species already identified as being important sources of eblvs, potentially failing to estimate properly the prevalence in others. despite these limitations, the data clearly suggest that species partitioning occurs. the common pipistrelle bat is known to be susceptible to experimental infection with eblv- (kuzmin and botvinkin, ) . yet none of the more than , pipistrelles (p. pipistrellus and also p. pygmaeus, which is cryptic with p. pipistrellus) surveyed in the netherlands, france and the united kingdom (harris et al., ; picard-meyer et al., ; van der poel et al., ) has proved positive for the virus. whether structuring across bat species driven by differing immunoresponsiveness to particular eblv types, by a lack of transmission opportunities or by other mechanisms, is unclear. multi-species summer, and particularly hibernation, roosts are known, though the amount of inter-specific direct contact appears to vary by season and species. for example, bats in houses and trees tend to use single-species roosts, even if more than one species is present at the site (park et al., ) . there may be more potential for inter-specific contact at key underground sites used by bats. in a survey of more than , bats of different species roosting in caves in turkey, it was noted that multi-species clusters frequently occurred in the post-hibernation season, but not during hibernation; and the horseshoe bats (rhinolophus spp.) only ever formed single-species clusters (furman and Ö zgü l, ) . many bat species also use swarming sites-enclosed areas often in and around caves-for display purposes. at these sites, hundreds or thousands of bats of mixed species congregate (glover and altringham, ; parsons et al., ) . the amount of contact, for example, via urine or aerosol droplets, between species at these events is unknown. more field research is needed to investigate the opportunities for disease transmission across bat species, and across geographical barriers. interestingly, in the united kingdom, only a single case of exposure to eblv- has been found (the test was able to detect exposure rather than live virus), whereas in other european countries with eblvs, the apparently more pathogenic eblv- is more common (defra, ; racey et al., ; vos, ) . to date, seven daubenton' bats (myotis daubentoni) in the united kingdom have been found to have eblv- infection (defra, ) , the latest case being diagnosed in may . it is notable that although a low prevalence (around %) of seropositivity was detected during active surveillance of daubenton's bats in scotland, live virus was not isolated from any of them (brookes et al., ) . similar results were found in an active-surveillance study in spain, which found that up to % of individuals in some colonies were seropositive for eblv- , but the prevalence of active infection was less than . % (serra-cobo et al., ) . it is currently difficult to interpret these results, but the vertical transmission of antibodies, as well as acquired immunity, is a possibility. while it is clear that eblv- and eblv- can cause deaths in unvaccinated humans, whether natural immune responses and cryptic recovery (i.e., without the virus having invaded the central nervous system and become symptomatic) are possible remains unknown. it is unfortunate that there was been no serological testing of bat workers in the united kingdom to establish the natural prevalence of neutralising antibodies to eblvs prior to . since that date, following the fatality in scotland from eblv- , it has been officially recommended that bat handlers be vaccinated against rabies. the take-up rate of vaccination has been very high. this understandable management of the public-health crisis means that it is now not possible to gather information that would have helped indicate the pathogenicity of eblv- to humans, and also whether exposure was more widespread than the single fatal case. it certainly appears that despite other species being susceptible in experiments, natural spillover into other non-bat hosts to produce clinical symptoms is rare, with the only known case for eblv- being a single stone marten . there are no reports of spill-over for eblv- . the apparently simple task of establishing the prevalence of a pathogen in wild animals can be fraught with difficulty. even assuming that a reasonably random cohort can be sampled, there is usually no opportunity to repeat 'live-tests' in cases of diagnostic uncertainty. establishing values for other key parameters is equally problematic. fundamental data on the sex-and age-distributions of infection are often not recorded. sometimes this is because the surveys (particularly for 'crisis management') were not designed with research in mind. sometimes it is practically difficult for the data to be acquired. bats, for example, can live more than years, yet it is impossible in the absence of long-term banding studies on the particular population being surveyed, to judge the age of animals with much greater accuracy than 'juvenile', 'young of the year', and 'adult'. weight is frequently used as a surrogate for age or maturity, particularly in studies of rodents, but there can be difficulties in distinguishing age from dominance effects, since both are correlated with body mass. the size of the population (or its density) is also often estimated with huge margins of error, as surveyors simply lack the time to undertake detailed ecological studies in addition to collecting clinical samples. distinguishing between different burdens of infection (particularly for macro-parasites) and stages of infection (particularly for micro-parasites) is frequently overlooked. this makes it difficult to use the data to parameterise epidemiological models. for example, animals infected with btb but in which the bacilli are encysted present no risk of transmission at that particular time point, yet these groups are often combined when data from post-mortem examinations are used. the fact that the disease may reactivate at some future time (measured by the overall prevalence) is not relevant to the calculation of the basic reproductive rate of the disease r . by conducting large-scale surveys of representative populations of wildlife on british farms, workers were able to build deterministic models to investigate the likelihood of the disease persisting in each host species. initially they assumed that no between-species transmission was present. using the prevalence of infectious individuals, together with field data on population structure and density derived from the same sites, they computed the basic reproductive number r for each of the species. the analyses showed that even when the maximum likely prevalence was assumed (based on the upper % confidence limit), the r (the basic reproductive rate of the disease) ranged from just . in wood mice to . in rats. (the lower confidence intervals for prevalence always gave r values that were < . ; mathews et al., b .) it is therefore unlikely that the disease would persist within single-host systems in the wild: the animals are unable to pass on the infection to their conspecifics at a rate high enough to maintain the disease. the findings are robust to underdiagnosis of infection: to affect the r materially, the prevalences would need to be have been underestimated very substantially. if, instead of single-host models, we assumed multiple-host systems, then higher prevalences should have been observed in the field than those recorded. alternatively, to achieve the prevalences seen in reality, the withinspecies transmission rate would have to be even lower than the very low value calculated. they have therefore been able to conclude that multi-species transmission of btb within farmland wildlife communities appears unlikely. perhaps the best example of long-term epidemiological studies in wildlife leading to epidemiological models of value to human health comes from studies of hantavirus infection in the united states. large-scale studies of several thousand rodents were conducted by four separate research teams, but were co-ordinated by common methodologies (calisher et al., ; easterbrook et al., b; glass et al., ; mills et al., ) . using longterm datasets, with repeated trapping at set grids, the teams were able to explore key components of the transmission pathway. seropositivity was higher in males and in heavier animals, suggesting horizontal transmission among adult males. decreasing prevalence with age among the youngest deer mice suggests that infected dams confer passive immunity to pups. in the main host of sin nombre virus, the deer mouse (peromyscus maniculatus), gender, age, wounding, season and local relative population densities were linked with the period prevalence of antibody (used as a marker of infection). nevertheless, antibody prevalence and some of the risk factors associated with antibody prevalence, such as relative population density, gender bias and prevalence of wounding, varied significantly among sites and even between nearby trapping arrays at a single site. this suggests that local micro-site-specific differences play an important role in determining relative risk of infection in rodents and, consequently, in humans. these data are now being used in spatially explicit models of the risk of human disease outbreaks (eisen et al., ) . as described for bat lyssaviruses, the contact rates between infectious and susceptible individuals (or a vector and a susceptible) is a critical step in the transmission pathway. yet compared with the effort that goes into improving, for example, the accuracy of a diagnostic test, very little attention is paid to measuring it in the field. this failure may offer some explanation for the difficulties faced in attempted disease control programmes. for example, disease is a primary threat to the survival of the critically endangered ethiopian wolf (canis simensis). since the early s, outbreaks of rabies and canine distemper virus (cdv) have had significant impacts on wolf population dynamics randall et al., randall et al., , . these diseases are maintained in local domestic dog populations, and a programme of dog vaccination was therefore introduced in , with the aim of reducing the population of susceptible dogs and hence the risk of transmission to wolves. attempts were made to achieve coverage of more than % of susceptible dogs during annual vaccination campaigns. this was not an easy task since dogs in ethiopia are used for guarding cattle and are not tame. rabies vaccines have high efficacy, and in theory, this level of coverage should prevent rabies outbreaks % of the time (coleman and dye, ; world health organization, ) . over , vaccinations have taken place, and at least initially, the number of rabies cases in dogs declined (randall et al., ) . nevertheless, a rabies outbreak occurred in wolves in , and could be linked with more than sympatric dogs with clinical symptoms consistent with rabies . mathews has, therefore, been analysing the reasons for the apparent failure of the vaccination strategy, focusing on the population dynamics of the domestic dogs, using data collected by the ethiopian wolf conservation programme. the key factor appears to be the growth of the dog population, which, as in other african countries, is keeping pace with, or even outstripping, human populations (cleaveland et al., ; rhodes et al., ) . eighty-six percent of all households owned dogs, rising to . % in rural areas. virtually nothing is known about the true contact rate between domestic dogs and ethopian wolves. it is clear that interactions do occur as wolf-dog hybrids are seen. we might speculate that diseased dogs, and aggressive dogs that are difficult to vaccinate, might be even more likely to interact with wolves than would healthy ones. some data are available on the demography of the dogs surrounding the bale mountains national park-one of the strongholds of the remaining ethiopian wolf population-as a result of a questionnaire survey administered by the ethiopian wolf conservation programme. the rate of increase in the dog population size appears to be around % per annum, and the turnover rate is also high. this creates a constant influx of new susceptibles into the dog population. it is difficult to keep pace with these, given the financial and logistical constraints on the numbers of visits veterinarians can make to each village. there also appears to be some geographical clustering of vaccination effort, and the implications of pockets of unvaccinated dogs on the probability of rabies transmission to wolves is currently being explored. the vast majority of efforts to control zoonoses in wildlife hosts, rather than in domestic animals, rely on culling strategies. in simple terms, the idea is to depress the population of the reservoir host to a level at which the disease can no longer be sustained, because the density of infected and susceptible hosts is too low. few of these culling programmes have systematically examined either the total population size or the level of population reduction likely to be required to achieve the desired endpoint. even where this has been done, it can be difficult practically, as with vaccination, to achieve the level of coverage desired. the strategy to control btb in badgers and cattle in the united kingdom has had the culling of badgers as its cornerstone for more than years. when it became evident that gradual badger culling was having little or no impact on the incidence of the disease in cattle, a formal review of the programme was introduced, culminating in a large-scale randomised trial of badger culling. this has demonstrated that there is no clear-cut reduction in btb in cattle. on the contrary, whereas the incidence of btb in farms at the centre of km badger culling zones fell by around %, the incidence in farms up to km away from the borders of these zones increased by around % (donnelly, ) . similar results were found in comparable areas where clusters of badgers were removed reactively, following nearby outbreaks of btb in cattle (donnelly et al., ) . at least part of the explanation for the failure to achieve effective btb control is likely to be the alteration in contact rates between infected and susceptible badgers, and also between infected badgers and cattle, as a result of the culling. there has been only one detailed study of m. bovis epizootiology in undisturbed badgers (culling having been suspended at the site, woodchester park in gloucestershire, england in ; delahay et al., ) . this study showed that btb does not spread rapidly at high incidence through badger populations, but rather is distributed patchily among a minority of individuals. social groups are relatively stable, and long-term dispersal movements are uncommon, though shorter movements do occur more regularly (rogers et al., ; vicente et al., ) . there is a correlation between rates of inter-group movement and the incidence of new infections (rogers et al., ; vicente et al., ) . while spatial clusters of infection exist, there is no strong synchrony between neighbouring groups, suggesting that there is only limited transmission between adjacent social groups (delahay et al., ) . both individuals and groups are more likely to be incident cases where the social group was diminishing in size, although there is no apparent relationship with group size itself, suggesting that it is the change in group size, and possibly the associated social dynamics, that influences disease risk (vicente et al., ) . badger culling operations have clear impacts on the behavioural ecology of the survivors. woodroffe et al. ( ) found that badger social group ranges increased among survivors within reactive and pro-active culling areas and along the perimeters of pro-active culling areas. their finding, at a large scale, accords closely with the observations of of individual and group behaviour in two zones of badger removal in england, as well as those of o' corry-crowe et al. ( ) in ireland. in all cases, the spatial organisation of social groups was considerably altered following the culls, with a large increase in the extent of overlaps between social groups (e.g., fig. . ) . the numbers of ranges with which each group overlapped also increased. there was a rather chaotic alteration in population densities (e.g., fig. . ) . in the examples shown, culling was conducted in , largely targeting areas of highest badger density (in effect, the largest social groups). one year later badger density was, unsurprisingly, lower in the culled areas, whereas there had been some increases elsewhere. in , although the population as a whole had not grown, the density remained low, or even fell further, in two removal areas, but increased elsewhere. by and , the distribution of badgers in the study area was radically different from that at the outset, with some previously high density, but culled, areas remaining depauperate . thus, while the population density recovered as a whole, the badgers built up in a different place. this sort of radical redistribution has not been reported in undisturbed populations. the changes have not only implications for absolute contact rates, but also the nature of contacts. for example, bite-wounds-an important route of btb transmission-were more common in the macdonald et al. ( ) study following social perturbation. (post-cull) figure . badger social territories before and after the selective removal of social groups following btb incidents in local farms. social structure also plays an important role in hantavirus transmission. in deer mice, both wounding and sin nombre virus antibody prevalence increased with mass. although it occurred in both sexes, the a c e b d badgers removed in bro increase was much more pronounced in males. wounding was more frequent in adult males than in adult females, and adults had more wounds than juveniles. the highest rate of infection was seen in individuals with the most wounds. similarly, in rats (rattus norvegicus) hantavirus infection (seoul virus) was associated with both wounding and elevated testosterone levels (easterbrook and klein, ; easterbrook et al., a) . it is therefore evident that changes to social structure-for example, by the removal of a dominant male-could have important implications for the epidemiology of a disease. rather than attempt to control disease by vaccination or culling, an alternative approach is to understand the factors leading to disease outbreaks in the first place and to manage these (dobson, ) . habitat changes that lead to alterations in population structure or migratory patterns, for example, are likely to affect the risk of zoonotic disease transmission (dobson and may, ) . the effect of habitat fragmentation on disease processes has rarely been investigated, but it has recently been shown that trypanosoma cruzi infection rates are higher in fragmented than continuous atlantic forest (vaz et al., ) ; and the risk of lyme disease in new york is also apparently increased by fragmentation (allan et al., ) . interestingly, the division of endangered ethiopian wolf population into small sub-populations, joined by habitat corridors, has been shown to allow rabies control to be achieved using a low-coverage vaccination strategy (haydon et al., ) . the strategy operates by eliminating the largest outbreaks of disease, and so enhances meta-population persistence, rather than by the conventional objective of reducing the reproductive number of the disease to less than one (haydon et al., ) . human activities that artificially increase, rather than decrease, animal densities also influence disease processes. these increases can be the result of losses of absolute habitat area, or from the provision of supplementary food or water. in the united states, the practice of supplementary feeding of house finches and white-tailed deer has lead to an increase in the incidence of mycoplasmal diseases and btb, respectively (hartup et al., ; schmitt et al., ) , presumably because of greater opportunities for disease transmission, and possibly also immunosuppressive effects of aggression at the feeding sites. btb has also been an intractable problem in the british cattle herd, with the incidence rising inexorably since . with a cull of badgers recently being ruled out, somewhat controversially, as offering no meaningful contribution to the long-term control of the disease (donnelly et al., ) , it is worth asking whether consideration of the ecology of the badgers and cattle might help generate workable solutions. over the past years, along with the increase of btb, there has also been an increase in badger densities, and it is likely that this contributes at least in part to the disease in cattle. so why have badger populations risen? might the answer lie in changes in land use? macdonald and newman ( ) speculate on a possible role for climate change, with milder winters and hence greater earthworm availability improving survival rates. changes could also have occurred in the susceptibility of badgers to btb and/or of cattle to btb. for example, the average milk production of a dairy cow rose from , l in to , l in (farm animal welfare council, ) , possibly to the detriment of the animal's immune status. similarly, stress resulting from cull-associated social perturbations, or from other changes to habitat, food availability or population density, may have influenced the innate immune response of badgers. little is known of the physiological responses of free-living wild mammals to poor environmental quality or other potential stressors. an argument has been made for polychlorinated biphenyls (pcbs) and other pollutants contributing to phocine distemper outbreaks (ross et al., (ross et al., , , but this is has been questioned (o'hea, ) . while the role of toxicants is not clear, recent work indicates that high population densities in wild field voles is associated with compromise in haematological and immunological indices. poor body condition appeared to affect the inflammatory response (as indicated by lower neutrophil and monocyte peaks) and lower immunological investment (as indicated by lower lymphocyte counts (beldomenico et al., a,b) . i have found, with co-worker jon blount, preliminary evidence of increased oxidative stress (measured by serum malondialdehyde concentration) among non-infected badgers from farms with recent btb in cattle, compared with those at sites free of btb ( fig. . ). there is also a considerable literature from farm (moberg and mench, ) , laboratory (galloway and handy, ) , and free-living aquatic animals (liney et al., ) showing that environment has a strong impact on stress responses, and that these can lead to pathological and pre-pathological alterations in immune function and overall health status. while it might be difficult to intervene directly to reduce the causes of stress in animals, ecologically based interventions that reduce both disease susceptibility and the opportunities for transmission may be possible. for example, in a study of british farms it has been shown that habitat management and cattle herd size were strongly associated with the risk of btb in dairy cattle (mathews et al., a) . reduced risk of btb was associated with the management of farmland in ways favourable to wildlife, including greater hedgerow availability, a lack of gaps in hedgerows, increasing hedgerow width and the presence of ungrazed wildlife strips adjacent to hedgerows. all of these measures are encouraged by recent european common agricultural policy reforms ( ) . broadly, habitat could influence cattle contact rates or be associated with agricultural management practices in ways relevant to btb transmission (such as reduced herd size). favourable habitat may lower the susceptibility of badgers to btb or reduce the number of inter-group excursions; alternatively, cattle on hedgerow-rich farms may be at reduced risk of ingesting contaminated soil. taking for simplicity just one of the parameters contributing to the effects-the total length of hedgerow-an increase of km/ ha was associated with a decrease in the odds of btb by about . % ( % confidence interval . % increase to . % decrease) in univariate analysis. in absolute terms, this equates to the annual risk of btb changing from the current rate of . % ( , confirmed incidents in , herds in ) to . % ( , incidents) for herds in the west of england if a policy of moderate hedgerow density increase were adopted. this would mean a reduction of infected herds per year. by comparison, systematic badger culling appears able to reduce the odds of btb by a maximum of about % and may even increase the prevalence in neighbouring areas (donnelly et al., ) . change in land use has also been linked to the emergence of two henipaviruses, nipah virus and hendra virus in the s, and land use management may therefore offer part of the solution. both viruses appear to be asymptomatic in their natural hosts, fruit bats (genus pteropus). they are amplified in domestic animals, pigs and horses, respectively, where they cause mortality, and can then be passed on to humans (chua et al., ; halpin et al., ) . the closeness of rna sequence match between pteropus sp., livestock and human isolates of each virus suggests that a sudden change in virulence is a less likely explanation of their rapid emergence into domestic animals and humans than is the ecological change that have affected the habitat of their natural hosts. many flying fox species are in decline, with roosting and feeding sites being deforested, and converted to agricultural or urban use. a number of hypotheses have been proposed to explain exactly how nipah virus emerged (see breed et al., ) , all of which involve the establishment of piggeries in previously forested regions still used by fruit bats (chua et al., ) . increased contact rates are also the likely explanation for the emergence of hendra virus in australia, with many pteropus populations having relocated into urban areas (hall and richards, ) . with no vaccine available, and pteropus in need of conservation, ecologically based strategies to limit contact rates between bats and livestock offer the best prospects of controlling the disease (field and mackenzie, ) . managing the risks from zoonoses to the health of humans and domestic animals is complex. it is also fundamentally important: virtually all emerging infectious diseases have originated in wildlife. superficially, the simplest method of control is via a reduction in reservoir host-disease prevalence, this being achieved by culls of host populations. however, effective reductions in population densities can be difficult to achieve in practice and may be undesirable where the target is of conservation concern. for example, most bat species are threatened, and yet they appear to be particularly important sources of emerging viruses (calisher et al., ; dobson, ) ; and despite not being endangered, badgers in the united kingdom are legally protected. an alternative, and possibly complementary, strategy is to manage the ecological conditions leading to disease spill-overs. this will not only benefit the health of humans and their domestic stock, but must surely also lead to benefits for the conservation and welfare of wild animals. effects of forest fragmentation on lyme disease risk population biology of infectious diseases: part i control of infectious diseases of wildlife in europe epidemiologie et lutte contre la rage en france et en ecology of infectious diseases in natural populations the dynamics of health in wild field vole populations: a haematological perspective poor condition and infection: a vicious circle in natural populations fox rabies. in ''the natural history of rabies.'' 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culling on badger meles meles spartial organisation: implications for the control of bovine tuberculosis key: cord- -oj v x authors: catala, m.; pino, d.; marchena, m.; palacios, p.; urdiales, t.; cardona, p.-j.; alonso, s.; lopez-codina, d.; prats, c.; alvarez lacalle, e. title: robust estimation of diagnostic rate and real incidence of covid- for european policymakers date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: oj v x policymakers need a clear and fast assessment of the real spread of the epidemic of covid- in each of their respective countries. standard measures of the situation provided by the governments include reported positive cases and total deaths. while total deaths immediately indicate that countries like italy and spain have the worst situation as of mid april , on its own, reported cases do not provide a correct picture of the situation. the reason is that different countries diagnose diversely and present very distinctive reported case fatality rate (cfr). the same levels of reported incidence and mortality might hide a very different underlying picture. here we present a straightforward and robust estimation of the diagnostic rate in each european country. from that estimation we obtain an uniform unbiased incidence of the epidemic. the method to obtain the diagnostic rate is transparent and empiric. the key assumption of the method is that the real cfr in europe of covid- is not strongly country-dependent. we show that this number is not expected to be biased due to demography nor the way total deaths are reported. the estimation protocol has a dynamic nature, and it has been giving converging numbers for diagnostic rates in all european countries as of mid april . from this diagnostic rate, policy makers can obtain an effective potential growth (epg) updated everyday providing an unbiased assessment of the countries with more potential to have an uncontrolled situation. the method developed will be used to track possible improvements on the diagnostic rate in european countries as the epidemic evolves. the evolution of the epidemic in europe has affected spain and italy more strongly than in other countries so far. this is clear from reported cases and fatalities in these however, they lack the recipe-type nature needed sometimes to direct a policy response. the focus of this paper is, thus, to introduce a method to compute the real diagnostic rate and the real incidence of covid- in each european country, testing that the key hypothesis of the method is fulfilled and that, if they were to be slightly off, they would affect all countries in the same direction. in other words, we provide a recipe for policymakers that we have tested to be correct, unbiased across countries and useful to make cross-country comparison provided the evolution and prognosis of the disease in a patient is not strongly dependent on socio-economic factors and only on age, sex and previous clinical history. we must recall here that the ability to determine the diagnostic ratio is essential to evaluate what the real number of infected people is. knowledge of this number is not only useful to visualize the full scope of the epidemic but also to properly estimate the number of people with probable short-term immunity. in this sense, our method can be added as an empirical take of other assessments about the real incidence of the disease and to study the possibility of developing herd immunity. a large number of real infected people would be a positive scenario for policymakers while a low number will be negative. it is thus very important to err on the side of caution in all our estimates giving always the less optimistic take. the basic structure of the paper is the following. first, we give a general overview of our framework in the methods section. then we discuss our key assumption: the real case fatality rate (cfr) in european countries experiencing a significative incidence will be roughly the same, given the similar structure of the population. if the real cfr were to be lower, or higher, it would affect all countries in the same way and would not affect most policy decision-making since it will move all countries in the same direction. we take this real cfr to be % and proceed to test that, effectively, there is a strong correlation between the day of reported deaths with the number of cases taken - days before. once a given value for the real cfr is taken, one must consider that people do not die immediately from the disease, as it takes roughly days after infection [ ] [ ] [ ] . in other words, the present values of the death toll can provide an estimation of the number of infected people days ago. knowing the number of infected people at present, not days in the past, is crucial. we attack this problem considering that people who become infected are usually diagnosed a few days after the onset of the symptoms, which can be to days after infection occurs. by comparing the number of people diagnosed on a certain date with our estimation of the real number of infected people, we can estimate what percentage of the cases are being diagnosed. we can calculate this for different countries and regions and test how this ratio has changed dynamically as the epidemic advanced. in the results section, we provide a full detailed description of how this fraction has become steady in the last weeks. we demonstrate that the percentage of diagnosis throughout the development of the epidemic has taken values that gradually converge for most countries. this gives a final clear picture showing the rate of diagnosis for each country. using this rate is straightforward to give a present-day estimate of the incidence given the number of reported infected people in each country as long as we can observe that the rate of diagnosis remains fairly constant. for policymakers, we have constructed an index named effective potential growth (epg) that combines this information with the growth rate of the epidemic to provide insight regarding which countries are, comparatively and in the short-term, in the most potentially complicated situation [ ] . framework of our methodology our analysis will be applied to european countries with a minimum of deaths on april so that we can guarantee a minimum statistical significance. the analyzed countries are: belgium, france, germany, italy, netherlands, portugal, spain, sweden, switzerlands and united kingdom. our two core assumptions are that the real cfr in all european countries is roughly the same and that reported data of death due covid- is uniform in all european countries under consideration. we will address these two hypothesis in the following sections. with these assumptions we need to carry out four steps, as indicated in fig. , to obtain the percentage of diagnosis. first, using a common reference cfr = % and, given the reported reported death count, we estimate the number of cases days ago. according to medical reports people die between and days after the development of the first symptoms [ ] . this time to death, ttd, after the development of the first symptoms will not be country-specific for demographic reasons. the estimated number of infected people with the disease at time t (see process in fig. this allows us to know to estimate the number of cases days ago. this value can be compared with the number of cases detected days ago, obtaining a diagnostic depending on the availability of tests, saturation of the health system and other external factors, countries have a great variability in the time of diagnosis delay. countries accumulate some delay that may arrive to days in the case that a country detected people as late as they were detected on death. this delay to detection (dd) due to lags in diagnosis corresponds to the time between the patient having the first symptoms and being reported by the health system. in fact, this time in some countries may vary throughout the course of the infection. therefore we cannot assume that the estimated and the reported are comparable and we need to know what the diagnostic time was for each of the countries studied. we can compare the reported deaths with the reported cases to find the maximal correlation, see process in fig. (a) , to estimate the dd, see process in fig. (a) . finally the ratio between reported cases at dd with the estimated cases, see below, provides an estimation for the percentage of diagnosis, see process in fig. (a) . note that the usual development of the reporting of a new case/death, see fig. (b), depends on the particular country under consideration, which determines dd. in fact, dd also includes a delay in reporting the diagnostic to death to official information systems. the cornerstone of our analysis is that the real cfr in all european countries will not be biased against any country in particular. we should point out immediately that we are not arguing that there are not important uncertainties in the real cfr, what we do claim and check in this methodology is that these uncertainties will not generate any biased against particular countries and should not affect policy decision. we take the cfr in of covid- in europe to be between . - % and we assume % to be the benchmark scenario. this value ( %) is the cfr observed in the initial stages of the south korea pandemic and the diamond princess cruise. in both cases, it was found to be around - . % and, in both, error margins came from different sources [ , ] . in south korea, the ability to test all the population in contact with infected people and the tracking of contagious chains was thorough, despite that, the reported cfr increased from initial values around . - . % to higher values around %. in the diamond princess cruise, cfr for confirmed cases was % but estimation of false negatives and the possibility that a fraction of the passengers never developed symptoms and was never tested put the cfr again around %. both south korea and the diamond princess cruise provide complementary evidence, one coming from a natural experiment and another from a country with the ability to perform half a million tests/day from the very beginning of the transmission chain [ ] . if we accept the two measurements of the cfr independent, the most likely interval of real cfr is between . and %. recent experimental results from random testing in the german city of gangelt [ ] and preliminary results from iceland [ , ] indicate the presence of a layer of people fully asymptomatic that are normally not detected. this group of people have passed the disease without any knowledge seems to be larger than previously thought. these preliminary studies point to a cfr of around . % in zones where the epidemics was not fully spread. we cannot disregard the possibility that, just as cfr inceased with time even in south korea, similar studies in countries with more cases, could have a real higher cfr. it is thus reasonable to consider cfr at % as an easy policy guiding principle and not to use the more positive scenario of . %. unbiased nature of cfr in europe there are three sources of possible biased cfr across countries. the disease affects more strongly elder people with comorbidity problems than healthy younger ones, and more men than women. in all european countries the male/female ratio is unbiased except for older people. this is precisely the group with higher mortality rate. it is thus very important to asses how the different demographic structure of european countries could affect our central benchmark [ ] . the same must be said about the relative prevalence of other comorbidity factors. we proceed to show that, with the data we have today: the demographic and comorbidity structure, none of these possible sources of bias can have anything but a small effect. to do so, we will do a comparison with the cfr of south korea on april , . %. table shows the demographic structure of south korea and the corresponding cfr for each analyzed age group reported on april . the first row shows the demographic structure according to eurostat, but the analysis has be performed by using only the three age groups shown in the second row: ≤ , − and ≥ years. this was done because for many countries reported cases and fatalities consider different age groups, and some countries even report this two figures using different age groups. the three age groups considered in the analysis were the only ones that includes all the analyzed countries. as can be observed in the to analyze what is the role played by the differences in demography in europe in the covid- cases and fatalities we have downloaded from eurostat the demographic distribution by age (see table ). we can readily asses that, when comparing with south korea, all the countries have a larger percentage of population above years ( % larger for italy) and larger median age except sweden and united kingdom, but the relative differences in each of the cohorts in between the european countries shown in the table is small. only italy presents a relevant larger than average ratio of people over . using this demographic data and assuming each european country presents the same cfr by age group as south korea on april , we have computed the cfr for each country. table shows the results of this analysis and the officially reported cfr by the different european countries on the same date. both values are presented relative to the cfr reported by south korea on april , . %. as can be observed in the first column, when demography is the only difference between countries, between the worst and best case of the relative cfr the differences april , / . cc-by-nc . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . having previously a very bad prognosis. we know this group is strongly affected by the virus [ ] . in blunt terms, we must examine the possibility that different countries are counting the raw number of dead people differently. before entering in the detail of the analysis, let us point out that two indications go against this possibility. first, health care systems in europe can have different april , / . cc-by-nc . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . resources in different countries with different focus and priorities, but they attend there is a single exception that we know of: belgium [ ] . belgium seems to be reporting unconfirmed cases from nursing homes without tests as due to covid- . it is quite clear that this includes a good number of people who, either, did not die from covid- or that covid- was not an important factor in the prognosis. therefore, we will include a reminder that belgium data is biased compared with other countries, being anywhere from % to % lower given the number of reported deaths from nursing homes compared with hospitals. there is a second argument regarding the treatment of the elder population in other countries. if large undercounting woul be the case, it should be noted in the mortality rate for people years and older, which is not observed in the countries where we have data. in this framework, spain becomes a key country. if spain were not to have an important undercounting is highly implausible to think that other countries would. we proceed to analyze the data of the national epidemiology center (instituto de salud march to april for the whole of spain, they see that, as expected, mortality is much higher than in previous years. an increase of % is observed. however, it is interesting to compare this with the data reported for covid- deaths. the reported deaths by covid- are roughly , depending on how you attribute deaths to a particular day in the calendar. on the other hand, the reported excess of deaths by the momo surveillance system is . we think that the assessment of around % underreporting can be taken indeed as a worst-case scenario for a highly impacted country. it seems reasonable to expect other countries to have underreported way below or slightly below this level. all the data point out right now, that the undercounting due to a different treatment of the very fragile population is highly unlikely across europe, and at most introduces changes in cfr around ± %. having shown that the real cfr should not present bias in european countries larger than %, we address now how to deal with the real sources of bias in the diagnostic rate for each country. to estimate dd we look for a correlation between the number of reported cases (see fig. a ) and the number of reported deaths (see fig. b ) [ , ] . to deal with noise effects we put a weighted moving average filter on the data of both cases and deaths. the correlation time between reported cases and reported deaths will be named as time from diagnosis to death (dtd), and: ttd = dd + dtd. ( ) correlation between reported cumulative cases and reported cumulative deaths exploring different delays between diagnose (reported) and death for germany (red), spain (green) and switzerland (green). (d) maximum correlation is marked with a red square for each country. % correlation interval can be seen with black bars. april , / . cc-by-nc . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . in fig. c we can see the correlation [ ] between reported cases and reported deaths assuming different dtd for germany, spain and switzerland. as you might expect, correlations have values close to . in most cases the correlation has a concave parabolic shape with a clearly defined maximum. we assume this maximum represents dtd for each country. the correlation interval is estimated as the points where the correlation is greater than % of the observed maximum. we decided to set a lower limit of days and a higher limit of days [ ] because we believe that time outside diagnostic rate by country as discussed in the methods, we use the same cfr = % in all european countries instead of making small corrections for demography. the bias due to demography was shown to be around - %, precisely the same order of magnitude we obtain for the possible bias in the counting of reported mortal cases. given that our aim is to provide a clear method for policymakers and that there is no data on how, or even if, both correlate, a common cfr allows us to homogenize the results with the clear limitation that we will obtain reasonable estimations and not exact results. the resulting picture is expected to be closer to reality than using purely reported data, but worse than correcting properly for age and diagnosis if the data of cfr for all age brackets and locations (nursery homes, hospitals, individual homes) were available, which is not the case. the estimation of the diagnostic rate is straightforward. from the cumulative number of deceased each day, and multiplying by ( % cfr) we get the cumulative number of people with symptoms days ago [ ] [ ] [ ] simply by rescaling and displacing backward in time the cumulated death curve of any country. to give an initial realistic and homogenous diagnostic rate we must establish how many days are needed as a bare minimum to be able to detect a patient from the onset of symptoms. first, the patient has to feel sufficiently sick and then contact the health service. from this contact, the doctor needs to be suspicious that the person has the disease and request a test. then, this test must be available, performed and the result received and annotated. it is clear that a bare minimum of one week is needed for this process. we use the name -days . cc-by-nc . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . symptoms and then days forward to be detectable/diagnosable. from this curve, we can obtain the rate between the cumulated number of people who had symptoms for or more days and the cumulated number of people detected days ago. it is thus clear that this homogenous analysis across countries could be performed assuming d-dr or d-dr and different cfr. it gives a proper first estimation of the situation. we argue, however, that there is indeed bias in the way people deal with the health care system in normal situations and, especially, under an epidemic. different countries and populations are in fact behaving very differently. we have observed that this is the case in the methods section checking the delay between diagnostic and death using time-displaced correlation analysis. this is the reason why we also define the delay to detection diagnostic rate (dd-dr) as the diagnostic rate computed using a time delay between the appearance of symptoms and detectability different for each country. we proceed to use fig. , with spain as an example, to explain the concept behind dd-dr. for spain, the maximum correlation between cumulated death curves and cumulated reported cases appears when cumulated deaths are displaced days backward. this suggests a dd of around two weeks ( − = days). this makes sense in a situation like the one in spain during march . the population receiving news that the health care system is under stress may decide to delay reporting of symptoms unless they are very serious. additionally, there is the possibility that tests are not available to people who report with symptoms to primary health care centers, and that the delay between the test, its positive result, and its record to official information systems is not negligible as well. it is thus important to correct for this bias in the estimation of the diagnostic rate. it is clearly not the same to have a time delay from symptom to the detection of days than . dd-dr can be computed from spain just like we did before for the d-dr using the same rescaling of the cumulated dead curve as before but using a displacement backward of days instead of days. fig. shows how the dd-dr is obtained in different countries depending on the delay between symptoms and detectability. countries with a lower dd, such as germany, have the same d-dr than dd precisely because they diagnose as early as realistically possible. we notice now that both d-dr and dd-dr can be tracked along time, as the epidemic advances we can check how these diagnostic rates changes. each new day we can look days back for the d-dr and compute the diagnostic rate. dd-dr can be tracked similarly. in fig. we show the evolution for both as a function of time for three selected countries. we observe that the dd-dr reaches a steady state after the initial stages of the disease while d-dr seems more affected by trends. this can be expected since dd-dr uses, precisely, the maximum of the correlation delay so it is expected to fluctuate less. the dd-dr is not only more stable but it also allows as to produce a proper assessment of the errors involved. the main one is the fact that the april , / . cc-by-nc . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . april as above million in spain and close to half million in germany. the table in fig. shows a list of the d-dr as of the beginning of mid-april of , and the dd-dr, which seems stable, together with the associated error. to interpret estimated cumulative cases and estimated attack rate we must take into account detection delay, because they are computed using the reported data. data updated on april . belgium data is biased due to reporting of unconfirmed death cases [ ] . best estimations might shift - % once the diagnostic rate is known, it is straightforward to establish a real incidence no longer affected by the presence of important differences in the time delays to diagnostic in different countries (see the table in fig. ). the level of diagnosis and the real incidence is indeed useful for policymakers since it gives a clear general picture. however, the policy response needed to improve the diagnostic rate is limited, in the short-term, by the ability to increase the production of pcr kits and other diagnostic tools. policymakers have more ability to affect immediately mobility patterns and social contact. in this sense, a key number for policymakers would be to have a reliable and robust estimation of the number of infected people in each country that can propagate the disease. providing an exact number is, right now, impossible. we can, however, produce an index of the effective potential growth using the dd-dr and the guidelines used by the ecdc to track the epidemic. even if the precise number of people with the disease were known, and the distribution of symptoms by sex and age was reported, there is no clear knowledge regarding the level of infectivity of the different type of person and symptoms. for instance, it is not known the days a person with mild symptoms can transmit the disease. the same can be said for people with serious symptoms. virus loads in the throat seem to be rather high across the board [ ] , but data on how this influence contagion is unclear. the only way to assess the situation is to use a general unbiased broad measure, which is indicative of the potential for infection. the ecdc uses the number of newly infected people in the last days [ ] . we use this same criterion. this number can only be obtained properly some days in the past, on the day we have a typical diagnosis. after that, we would need input from new data to properly compute how many people are diagnosed. so the number i is strictly a measure of the recent past, but good enough to give the proper picture that the system will face the following days. april , / . cc-by-nc . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . fig . schematics of the procedure to obtain incidence a , recovered and estimated cases using germany as an example. incidence of estimated cases (blue), contagious incidence (red) and total estimated recovered cases (green). blue shaded part is the number of cases used to compute the estimated contagious incidence. to interpret final number of total cumulative cases, recovered cumulative cases and estimated attack rate we must take into account detection delay, because they are computed using the reported data. similar figures for all countries are shown in si fig. we also consider those undetected cases which appear earlier than days as recovered r i . notice that here we use the word recovered lousily. it does not mean literally that all of them are fully recovered since most of them never fell ill to begin with, and some of them could not have neutralized tests yet, but that those infected and undetected for more than two weeks ago do not seem to pose a serious risk. a alone, however, does not give a full picture of the situation. it is not the same to have contagious per inhabitants when the number of contacts is high that when the number of contacts is low. it is important to take into account the level of spreading velocity of the epidemic related to the effective reproductive number (r t ). the effective reproductive number depends on multiple factors, from the properties of the virus itself to the number and type of contacts. those, again, depend on different social behavior and structure such as mobility, density or the typical size of the family unit sharing a house, to name a few. the only feasible way to estimate r t is using fits from seir models. complex seir models which include spatial and contact-processes have a large number of parameters which, due to the present lack of knowledge, are ( ) and epg.ρ ( ) is computed using the mean value for the last three days. epg: effective potential growth described in the text. to interpret table data we must take into account detection delay, because they are computed using the reported data. data updated on april . * belgium data is biased due to reporting of unconfirmed death cases [ ] . best estimations might shift - %. given the partial empiric nature of present r t , we prefer to take a fully empiric surrogate as a quantitative evaluation of the level of infections. we define an alternative reproductive number as the number of new cases detected today divided with the number of new cases detected five days ago as n t /n t− . however, the high fluctuations on this quantities imposes the use of averaged values over three days [ ] : where n t stand for new cases reported at day t. this rate is one if the number of new cases is constant. it will be below if new cases are decreasing and larger than if the april , / . cc-by-nc . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint number of cases is increasing. we take days as the key delay unit since this is roughly the time since infected people develop symptoms if they do develop them. there are still clear fluctuations on a day-to-day basis of this measure ρ t due to common delay and irregularities in reporting. most fluctuations can be eliminated by taking the average of ρ t during three daysρ ( ) which is normally enough to get a rather smooth measure. it is not uncommon to find still some fluctuations and one-week averages can be done if required. we propose the following day-to-day index epg: epg is just the multiplication of the growth rate of the diseaseρ ( cases is biased by diagnosis protocols and ratios in each country, as well as by the pool of asymptomatic cases. moreover, any attempt to improve diagnosis percentage requires an economic, infrastructural and logistical effort that is not always possible. in addition, this health system structure is a strong conditioning that limits the possible actions to carry out in this direction. the reported number of deaths, if uniformly and properly recorded, provides very relevant information as a first general overview. even in countries where there is a bias on death reporting, the effort that should be made to improve these data collection is much lower than the necessary effort to increase data about cases. the assumption of a common lethality, which has been situated around %, allows for using the cfr as an indicator of real incidence. current information on cfr is still not complete, since many countries do not report distribution of deaths by age or sex, neither provide covid- mortality outside hospitals. however, we argue that the kingdom ( , ), france ( , ) and belgium ( , ). if we estimate the cases that should have been diagnosed by that time, the ranking is lead by italy ( , , ) and followed by france ( , , ), spain ( , , ), united kingdom ( , , ), belgium* [ ] ( , ) and germany ( , ). thus, differences in diagnostic rate are absolutely significant when analyzing global situation in europe. countries like germany, portugal and switzerland would be diagnosing around % of cases, while belgium, france, sweden and united kingdom would be in the level of %. assessing the risk of countries to enter or remain in the epidemic growth phase is essential. in this sense, the epg index is a valuable tool for policy makers. a high epg in the situation where there is a high growth rate of the epidemic and large number of active cases is a clear situation of danger, while a very low epg because both the reported epg vs estimated real epg. different european countries in terms of the epg computed using the reported data on the attack rate vs the epg using our estimation of the real attach rate. the order of the different countries should be done from right to left (for the reported state of the index) and from top to bottom (for the estimated value of the index). we observe how the comparative situation of the different countries changes as of april . * belgium data is biased due to reporting of unconfirmed death cases [ ] . best estimations might shift - %. despiteρ ( ) is quite independent of the diagnostic rate, reported i directly depends on the level of diagnosis. thus, if epg is evaluated with reported data, it can provide a wrong picture of the situation. based on reported epg, the worst situation in europe at april would be for belgium, followed by spain, united kingdom, netherlands and portugal. if risk is evaluated with estimated epg, highest value would still correspond to belgium as well, but followed by sweden, united kingdom, spain, netherlands and italy. portugal is in much better position that its reported data suggest. actually, countries with similar reported epg like portugal, and netherlands have, in fact, totally different estimated epg, being the last country at significantly higher risk than the former . we have shown in the methods section that the basis for obtaining estimated i and a is not biased due to demographic differences and, right now, there is no indication that it is biased due to a different way of accounting for the cumulative dead toll of the epidemic. there is also no indication that comorbidity factors are largely different in different countries or that cfr is higher on some countries given that icu units and hospitals are not available for people that would need it, at least so far. if this were the case, under any scenario where the situation occurs, the epidemic in that country will have such a larger number of cases, attack rate and growth that the epg will be extremely high. the only real limitation is that the social and environmental issues could affect the prognosis of the infected. if living in a small house with other people infected could lead to worse prognosis than staying in a large house alone, a new analysis regarding the unbiased nature of the cfr would need to be done. it is important to indicate that not only i is unbiased, as analysed in previous sections, but thatρ ( ) is not biased as well. even though absolute reported cases is biased, as we have shown, ρ t deals with ratios and its evolution. as long as the diagnosis and recording of the people with disease follows roughly the same criteria along time in each country, ρ t is a good measure of the growth the epidemic. indeed, if evaluated diagnosis percentage is more or less constant in time, we can assume that ρ t correctly reveals tendencies in contagiousness. if a change in criteria in reporting the cases occurs (i. e., a large increase in the number of tests per day leading to an increase of cases due to more testing), ρ t will be temporally affected but will go back to be a good measure once the new criteria is established. in this case, epg will provide a wrong picture for a while as well, until stationary conditions in diagnosing and reporting are achieved again. there is another important point to address in order to guarantee that ρ t is a robust measure. as soon as we are estimating real number of cases, we can determine the associated ρ t . it is expected that both ρ t behave similarly but with a certain delay. this delay can be determined by translating both ρ t in time until error between both is minimized. we show this detailed analysis in the supplement material si file where we evaluate that both the reported ρ t and the inferred ρ t are indeed different but that follow the same type of evolution once the proper delay is accounted for si fig. . the third important outcome of this analysis is the estimation of recovered people. this is an important number to assess the possibility of herd immunity discussed as a possible exit strategy. the idea is that those that recover might have immunity and act as barrier in the transmission of the disease. a recent study from the fudan university at shangai [ ] has analyzed antibody titters of adult covid- recovered patients. the study is based in the detection in plasma of spike-binding antibody using rbd, s , and s proteins of sars-cov- using an elisa technique. it is also the first study that looks after neutralizing antibodies (nabs) specific for sars-cov- using a gold standard to evaluate the efficacy of vaccines against smallpox, polio and influenza viruses. the study highlights the correlation between the nab titters and spike-binding antibodies that were detected in patients from day - after the onset of the disease, remaining afterwards. middle and elderly age patients had higher titters compared with young age patients, in which in cases the titters were under the limit of detection. nab titters had a positive and negative correlation with c-reactive protein (crp) levels and lymphocyte counts, respectively. this indicates that the severity of the disease, in terms of inflammatory response (crp levels), usually worse in middle and elderly age, favors the increase of antibody titters. equally, the negative correlation with lymphocyte counts suggests an association between cellular and humoral response. therefore, it is possible that the immunity reached by young people, which were mostly asymptomatic, is residual. in that case, this sub-population would keep being carriers of covid- . serological studies that many countries are designing and carrying out should provide further information on post-infection immunity. even if the entire recovered population acquires middle-term immunity, current incidence situates european countries far from herd immunity. nevertheless, it is feasible that regions with highest affectation were closer to use herd immunity as a strategy for de-confinement. governments might wish to explore the possibility of local deconfinement. there are two possible limitations of this present study. it could be possible, in theory, that some countries present an intrinsically different cfr if they are able to isolate completely and significantly its elder population more than others. the epidemics real cfr is a measure of the case fatalities if all the population, or a representative sample of it, has become infected. if one country would effectively prevent all infections among all its elder population from contagious forever, it will certainly have a different cfr. right now, it is impossible to assess if this is indeed the case in different countries given the lack of reported cases and mortality rates by age and sex. we should notice however that, if this disaggregation were to be provided, we could proceed with exactly the same methodology but instead of using the whole country as a whole we would divide it into different age brackets and treat them separately. the second limitation is related to the first one but coming from a more structural perspective. a clear possibility is that countries under stress could be failing in providing the same medical support changing the cfr. we must notice that health care in european countries, even under stress, has been able to increase dramatically its number of health personnel, of beds and hospitalization in short notice [ , ] . italy and spain present some regions under stress but not the whole country [ ] . finally, one cannot disregard the possibility that complex mechanisms of mutations and repetitive exposure to the virus may change the prognosis depends on the type of residence and, hence on socio-economic factors, which are clearly different across countries. if any proof that a close environment not only increases the level of infections, which they obviously do, but also changes the disease evolution in the patient, one should again test that the uniform/unbiased cfr hypothesis holds with the proper knowledge at hand. to obtain dtd for each country and the corresponding evolution of the diagnostic rate. we also provide fore each country the evolution of recovered and the attack rate in the last days a . we also provide the demonstration thatρ ( ) is also unbiased showing the correlations between real and estimated growth rates. fig. series of figures showing the evolution of the estimated cases for different european countries. in blue, incidence of estimated cumulative cases. in green, estimated incidence of cumulative recovered cases. in red, estimated incidence of attack rate lasts days (a ). day is considered the first day where cumulative cases was over cases, it is different for each country. data extended till april . . cc-by-nc . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. (which was not certified by peer review) the copyright holder for this preprint this version posted may , . . growth rate and, in blue, reported cases growth rate. (b) the gorwth rate of estimated cases is displaced to find better match with the growth rate of reported cases. (c) error between estimated and reported growth rates using differents delays. minimum delay is marked and is the one used in (b). european centre for disease prevention and control. download today's data on the geographic distribution of covid- cases worldwide situación actual covid- ministerio della salute republica italiana. covid- , i casi in italia coronavirus testing: how are the hardest-hit countries responding? suppression of covid- outbreak in the municipality of vo transmission potential and severity of covid- in south korea correcting under-reported covid- case numbers: estimating the true scale of the pandemic using a delay-adjusted case fatality ratio to estimate under-reporting. centre for mathematical modelling of infectious diseases. london school for hygiene and tropical medicine analysis and prediction of covid- for eu-efta-uk and other countries. dpt. of physics. universitat politècnica de catalunya estimating clinical severity of covid- from the transmission dynamics in wuhan, china real estimates of mortality following covid- infection case-fatality estimates for covid- calculated by using a lag time for fatality clinical course and risk factors for mortality of adult inpatients with covid- in wuhan, china: a retrospective cohort study. the lancet estimating the asymptomatic proportion of coronavirus disease (covid- ) cases on board the diamond princess cruise ship estimating the infection and case fatality ratio for coronavirus disease (covid- ) using age-adjusted data from the outbreak on the diamond princess cruise ship central disease control headquarters. coronavirus disease- , republic of korea vorläufiges ergebnis und schlussfolgerungen der covid- case-cluster-study large scale testing of general population in iceland underway spread of sars-cov- in the icelandic population demographic science aids in understanding the spread and fatality rates of covid- neutralizing antibody responses to sars-cov- in a covid- recovered patient cohort and their implications chain safety and environment belgium government. the covid- figures: collection, verification and publication vigilancia de los excesos de mortalidad por todas las causas. momo world health organization. coronavirus disease (covid- ) situation reports correlation (in statistics virological assessment of hospitalized patients with covid- european centre for disease prevention and control. coronavirus disease (covid- ) in the eu/eea and the uk -eighthupdate el ministerio de sanidad amplía las medidas para el refuerzo de personal sanitario hospital fira salut ja està a disposició del sistema sanitari per si calen llits addicionals incidencia de la covid- en las camas uci en españa key: cord- -ogb xg authors: palomino, juan c.; rodríguez, juan g.; sebastian, raquel title: wage inequality and poverty effects of lockdown and social distancing in europe date: - - journal: eur econ rev doi: . /j.euroecorev. . sha: doc_id: cord_uid: ogb xg social distancing and lockdown measures taken to contain the spread of covid- may have distributional economic costs beyond the contraction of gdp. here we evaluate the capacity of individuals to work under a lockdown based on a lockdown working ability index which considers their teleworking capacity and whether their occupation is essential or closed. our analysis reveals substantial and uneven potential wage losses across the distribution all around europe and we consistently find that both poverty and wage inequality rise in all european countries. under four different scenarios ( months of lockdown and months of lockdown plus months of partial functioning of closed occupations at %, % and % of full capacity) we estimate for european countries an average increase in the headcount poverty index that goes from . to . percentage points and a mean loss rate for poor workers between % and . %. the average increase in the gini coefficient ranges between . % to . % depending on the scenario considered. decomposing overall wage inequality in europe, we find that lockdown and social distance measures produce a double process of divergence: both inequality within and between countries increase. the dramatic and unprecedented intensity of the shock due to the covid- pandemic has highlighted the importance of measuring the economic consequences of "social distancing". the lockdown measures implemented in many countries around the world will likely have a significant negative impact on their gdp. thus, the global economy is expected to shrink by % this year. asia will not have economic growth for the first time in years, the economies of the us and europe are projected to contract between % and % (imf, ) , and the global job losses are estimated to be over million (ilo, ) . however, the effect of the pandemic will not only take place at the aggregate level and is likely to have distributional implications (furceri et al., ) . the social distancing imposed by governments to limit the spread of the pandemic has caused an asymmetric effect on the labour market: discounting essential occupations like health services and food sales, only the jobs not closed by the lockdown that can be done from home ("teleworkable") will be not impeded. this asymmetry of the supply shock implies that the economic costs of social distancing could be significant, not only in terms of negative gdp growth rates but also in terms of higher wage inequality and poverty rates. in this paper, we analyse the potential effects of social distancing on wage inequality and poverty in absence of any compensating public policy across europe. recent studies have provided estimates of the supply shock caused by the emergency regulation imposed to contain the spread of covid- (dingel and neiman, ; hicks, ) . they have evaluated the possible economic consequences of social distancing -without considering the subsequent effects that may occur on the demand side-by calculating to what extent occupations can be performed from home (teleworking). mongey et al. ( ) find that workers with less ability to work from home have been indeed more unable to follow the 'stay at home advice' -using geolocation data-and that they have suffered higher unemployment increases. studies focused on the demand side have shown that, due to fear of infection, economic activity dropped and individuals changed their consumption habits even before lockdown was formally imposed (goolsbee and syverson, ) . in a more comprehensive analysis, del rio-chanona et al. ( ) provide quantitative predictions of supply and demand shocks associated with the covid- pandemic. these studies, mainly focused on the u.s. economy, have analysed the consequences of social distancing in the job market, at the occupation and industry levels, without delving into the study of poverty and inequality. we find two exceptions. first, irlacher and koch ( ) obtain a substantial wage premium (higher than %) in a mincer regression for german workers performing their job from home, and a lower share of teleworkable occupations in poorer german regions. second, brunori et al. ( ) study the short-term effects that two months of lockdown have had on the italian income distribution. by using a static microsimulation model, they find a non-negligible increase of poverty and inequality in italy. to estimate the impact of social distancing on wage inequality and poverty across europe, we concentrate on the legal restrictions (supply side) due to the closure of non-essential occupations and workers not being able to perform their activities at home during the lockdown period. additionally, we consider a range of scenarios with further partial closures of certain activities, caused by mandatory capacity limitations and by the change in individual consumption habits to prevent contagion. despite its relevance, our analysis does not consider additional indirect effects in supply (as shortages propagate through supply chains) and in demand (when the loss of labour income for some workers further reduces consumption). these effects are difficult to estimate at this early stage, and the legal and voluntary restrictions imposed to prevent the spread of the pandemic already provide a clear framework to study the consequences for inequality and poverty of having a particular productive structure. under a lockdown, the asymmetry of the restrictions may affect economies in a different way just because their productive structure is not the same: countries who are specialised in outdoor and non-essential activities like tourism will, in principle, suffer more from the lockdown and the capacity limitations. we thus restrict ourselves to measuring the potential impact on wage inequality and poverty of enforced and voluntary social distancing. in general, if occupations with higher wages are more teleworkable, we should observe an increase in wage inequality due to lockdown and the implementation of social distance measures within each country analysed. however, whether this happens, and the intensity of this change, will depend on the structure of the economy and the extent of essential and closed occupations under the lockdown. the wide set of european countries in our sample -with a variety of productive structures-will also allow us to test if different productive structures imply different potential effects on wage inequality and poverty under lockdown and social distancing. note that if that is the case, inequality would not only increase within countries, but also between nations, which could exacerbate the problem of cohesion in europe. the first step to measure the changes in wage inequality and poverty across europe due to lockdown is to calculate the index of teleworking at the occupational level. dingel and neiman ( ) found that percent of jobs in the united states can be done entirely from home. what is the share of occupations that allow teleworking in europe? following these authors, we use fifteen questions from the occupational information network (o*net) database such as, 'is the work done outdoors?' or, 'does it require significant physical activity?' to calculate the probability of teleworking for each occupation. we then use the european union labour force survey (eu-lfs) occupational structure to translate these probabilities into the european context. finally, we match the eu-lfs occupation teleworking index and the european union statistics on income and living conditions (eu-silc), which provides detailed information on wage at the individual level. after this process, we have for every worker in the eu-silc database the individual index of teleworking (according to her occupation) and the wage. a lockdown implies that some activities -like healthcare or food chain related jobs-will become essential while others will be closed. when the occupation is essential, workers will be not affected by lockdown regardless of their capacity to work from home. when a certain economic activity is closed -like hospitality-working is not at all possible. for the remaining economic activities, only teleworking is allowed. consequently, during the lockdown we need to adjust our index of teleworking for the workers whose occupation is essential or closed, to obtain an individual measure that summarizes the capacity of each worker to keep active under the lockdown. we will call this measure lockdown working ability (lwa) index. the next step is to estimate the wage loss due to the lockdown. because not all workers are able to perform their job at home and some activities are closed or limited (or less demanded to avoid infection), there are potential wage reductions for a significant part of the labour force. to simulate these wage losses, we consider four possible scenarios: two months of lockdown and two months of lockdown plus six months of only partial functioning of the closed activities at %, % and % of capacity. in addition, we consider in appendix e another three scenarios where partial functioning of closed occupations −at %, % and % of full capacity− lasts for nine months. with these proposals we intend to measure not only the effect of the lockdown, but also the impact of the de-escalation period imposed to contain the spread of covid- . although each european country may have followed slightly different lockdown and deescalation strategies, the core of the social distancing enforcing policies has been similar in most of them. for that reason, and to ensure that in our analysis differences across countries are mainly due to their productive structure, we simulate the same scenarios for all european countries. the last step is to measure the changes in wage inequality and poverty across countries, and the variation of wage inequality between and within-countries due to the lockdown. for this task, we first compute the lockdown incidence curve (lic), which represents the relative change in the wage of individuals ordered by centiles, and the related changes in the mean 'growth' rate of the poor (ravallion and chen, ) and headcount poverty index. then, we use the gini coefficient and the mean logarithmic deviation (mld) to calculate the changes in wage inequality. the first measure is a popular index of inequality which is widely used in the literature, while the second measure fulfils some properties which are necessary for our analysis of inequality decomposition. in particular, the mld is the only inequality index that is additively decomposable into a between-group and a within-group component (bourguignon, ; shorrocks, ) and has a path-independent decomposition (foster and shneyerov, ) . our results show that poverty increases for the headcount index and the mean loss rate of the poor in all countries for all simulations, although these changes vary greatly with the country and simulation under consideration. for the four simulated scenarios, we estimate for european countries an average increase in the headcount poverty index that goes from . to . percentage points and a mean loss rate for the poor that goes from % to . %. likewise, wage inequality increases for the gini coefficient in all countries for all simulations. for example, the average increase in the gini coefficient varies between . % and . % for europe as a whole. we thus find that poverty and inequality changes are sizeable in all countries and they increase with the duration of the lockdown. when we decompose overall inequality in europe, both within countries and between countries inequality increase, producing a double process of divergence in wage inequality in europe. but this increase in wage dispersion is not symmetric, and the within-countries inequality component increases more than the betweencountries inequality component. thus, the increase in the within-countries inequality component ranges from . % to . %, while the change in the between-countries inequality component goes from . % to . %, depending on the scenario under consideration. the observed increases in poverty and inequality are in general greater in eastern and southern countries than central and northern countries and are inversely related to the lwa index. to further understand these differences across european countries we need to analyse separately the three components of the lwa index: teleworking and essentiality (which increase the working ability of occupations during the lockdown), and closure (which implies a lower working ability during the lockdown and subsequent de-escalation periods). we observe that the occupational and industry structure of eastern european countries presents, in general, medium average levels of closure, but low average levels of essentiality and teleworking. on the contrary, northern and central european countries show large values of essentiality and teleworking, and low average levels of closure. meanwhile, workers in southern european countries have medium average levels of essentiality, rather low levels of teleworking and, especially, the highest average levels of closure, due to the large preponderance in their productive sector of activities that imply the agglomeration of large groups of people like recreation or hospitality. the rest of the paper is structured as follows. in section we present the lwa index and the methodology applied to calculate the wage losses and changes in inequality and poverty. in section we highlight the main results obtained for europe. finally, section concludes. to analyse the working ability of individuals during the lockdown, we need first to estimate occupational teleworking. for this task we use three different databases (see appendix a). first, we make use of teleworking information acquired using key attributes and characteristics of occupations from the american o*net database (dingel and neiman ) . second, we use the latest wave of eu-lfs ( release) -with detailed employment and occupational information for european countries-to accurately obtain occupational teleworking information for the european occupational categories. finally, this information is combined with the rich socioeconomic data -crucially, salaries-from the wave of eu-silc ( release). this strategy, which is explained in detail in appendix a, allows us to calculate the teleworking capacity of european workers. however, teleworking capacity is not the only determinant of workers ability to effectively work and keep their wage during the lockdown period. we need to consider that some having calculated the teleworking index and identified the essential and closed occupations, we construct the lockdown working ability (lwa) index. this measure summarizes the capacity of individuals to work under a lockdown taking into account not only the value of their occupation's teleworking index, but also if such occupation is essential ( ) or closed ( ). the central idea is that workers in essential occupations can work during the lockdown regardless of whether the occupation can be teleworked or not. on the contrary, workers in closed activities cannot work at all to the extent that their overall activity has been closed. in all remaining cases, working capacity will depend on the share of that occupation that can be teleworked. formally, the first step in constructing the lwa index requires to split the population of workers into three groups according to the occupation of each worker ∈ { , , … , }. if the individual has an occupation that is neither essential nor closed, the value of her index will be equal to the value of her index of teleworking, ∈ [ , ]. if the person has an essential job ( = ), we will compute the lwa index as is the essentiality score given to the occupation of the individual. thus, for partially essential occupations ( < < ), the non-essential share of the occupation ( − ) can work during lockdown only to the extent that it is teleworkable. finally, if the person has a job that is closed ( = ), we will calculate the lwa index as = ( − ) , where ∈ ( , ] is the close score given to the closed occupation of the individual. in partially closed occupations ( < < ), the non-closed share of the occupation ( − ) can work to the extend that is teleworkable. in summary, the lockdown working ability index is calculated as follows: for all ∈ { , , … , }. as shown in figure , the average lwa index varies significantly across european countries: our lwa index varies significantly not only by countries but also by gender, type of contract −permanent or temporary−, type of work −full or part time−, and level of education (see table ). according to their lwa index, women jobs are less affected by social distancing than men jobs in all european countries. interestingly, the biggest difference between both sexes is found in the nordic countries: norway ( points), denmark ( points the next step is to calculate the potential wage loss due to the lockdown for every individual in the population. following the facts observed in europe during the pandemic, we adopt four possible scenarios. as a lower bound, we assume a simple scenario: two months of lockdown (case i). here we suppose that occupations can be developed at full capacity ( %) after the lockdown. next, we assume two months of lockdown and six months of partial functioning of closed occupations at % (case ii), % (case iii) and % (case iv) of full capacity. the rationale for these scenarios is that governments may not allow a fully functioning of closed occupations after the lockdown to avoid a new outbreak of the virus and that individuals may voluntarily choose to stay home, which reduces their consumption, to avoid infection (goolsbee and syverson, ) . for example, some activities like arts, entertainment, recreation, restaurant, hotel and transportation are still under a large negative shock in production and consumption. consequently, to be more informative we have adopted a range of capacity constraints that goes from % to % of full capacity for a period of six months. in addition, given the uncertainty about the duration of legal and voluntary restrictions, in appendix d we have further replicated our results for another three scenarios where partial functioning of closed occupations −at %, % and % of full capacity− lasts for nine months instead of six. we simulate the same scenarios for all countries so that differences across countries are mainly due to their distinct productive structures, isolating our analysis from the influence of particular mitigation measures adopted by each european government. using the lwa index, we calculate the wage loss ( ) experienced by every individual during the lockdown according to the four simulated scenarios described. for the first case (i) the equation we estimate is the following: where − is the annual wage of individual in period − (before the lockdown) and represents the duration of the lockdown in annual terms, i.e., = . because measures the capacity of individual to work under a lockdown, the term − represents the incapacity of worker to perform their job under a lockdown. consequently, the wage loss the duration of the lockdown in many european countries has been approximately of two months. for example, the lockdown went from the rd of february to the th of may in italy, from the th of march to the th of may in spain, and from the th of march to the th of may in france. for these activities chetty et al. ( ) have found, as of july th , a decrease in spending compared to the pre-lockdown levels in the range of between . % and . % in the us. based on weekly credit card expenditure data, bbva research ( ) has found, as of july th , a negative interannual growth rate for these spending groups ranging between % and % in spain. experienced by workers under a lockdown is the proportion of annual wage they lose due to their inability to work during the shutdown period. for the cases (ii), (iii) and (iv) we apply the same equation ( ) unless the individual has a closed occupation, in which case, we need to additionally consider the wage loss due to the partial closure of %, % or % of their occupation for six (or nine) additional months. for these scenarios the relevant equation is: where = { = ≠ is the indicator function, represents the duration of the partial closure ( or ), and is the proportion of closure ( . for case (ii), . for case (iii) and . for case (iv)). to give the intuition of expressions ( ) and ( ), we provide an example. suppose we are interested in calculating the wage loss for three workers with different occupations under a lockdown of months and a posterior partial closure of some activities of % (keeping % of capacity) for months. the first worker is a physician, the second works as a clerk and the third one is a waiter. their pre-lockdown ( − ) annual salaries are − , − and − , respectively. the occupation of the first worker is essential with = so we apply equation ( ) to calculate her wage loss: = − · · = , i.e., the doctor does not lose earnings during the pandemic. the second worker has an administrative occupation that in our example is neither essential nor closed so her index is equal to the value of her index of teleworking, for example, . . then, according to equation ( ) her wage loss will be = − · · ( − . ) = − · . . finally, the occupation of the third worker is closed with = so the value of her index is . in this case, we apply equation ( ) closely related to the lic we have the mean loss rate for the poor, a measure that reveals the average wage loss for workers below the poverty line. let ( ) denote the headcount index defined as the proportion of workers whose salary is less than , where is the poverty line (set at % of the median wage). then, the mean loss rate for the poor ( ) is defined as the area under the lic up to the headcount index divided by the headcount measure, which can be expressed as: when < for all < ( ) one can conclude that the lockdown was poverty-augmenting. in addition to representing the lics for the set of european countries and compute the above unfortunately, the gini index is not additively decomposable into a between-group and a within-group component. its decomposition includes also a residual term which cannot be assigned to either component. for this reason, we use the mld index in the last part of our analysis, where we decompose the overall estimated change of inequality in europe into its between-countries and within-countries components. the mld belongs to the generalized entropy class, which is the only class of inequality indices that is additively decomposable into a between-group and a within-group component (bourguignon, and shorrocks, ) . moreover, the mld has a path-independent decomposition, so the result of the decomposition is independent of which component (between-group or within-group) is eliminated first (foster and shneyerov, ) . the mld ( ) is defined as: where is n-coordinated vector of ones. expression ( ) provides a breakdown of overall wage inequality into between-group and within-group terms. the between-group component ( , , … , ) is the level of wage inequality that would arise if each worker in a country enjoys the mean wage of the country, and the within-group component ∑ ( ) = is the weighted sum of wage inequalities within different countries. how large are the wage losses experienced by workers during the lockdown? are they evenly distributed? a visual approximation to the wage loss across the wage distribution can be achieved by looking at the lic curves. in figure we represent the lics for some european countries under two months of lockdown (case i). we highlight the lic for two extreme cases, romania, and cyprus, where the main wage losses take place at the upper and lower part of the wage distribution ( figure , panel a) and for germany, the uk and france (figure , panel b) , where wage losses tend to decrease with the percentile that the worker occupies. in our two-month lockdown scenario, wage losses are already sizeable and can be superior to % at some parts of the distribution in some countries, varying significantly with the centile of the workers. also, we find that the shape of the distribution of wage losses is similar for the rest of scenarios and the main difference lies only in the larger size of the wage drops. we formally capture the potential impact of social distancing on poverty with two measures: the mean loss rate for the poor ( ) and the changes in the headcount poverty index ( ). table (columns to ) shows the values of for the four simulations under consideration, which indicate that the lockdown is poverty-augmenting in the four scenarios for all european countries. under the most conservative scenario, a two-month lockdown, we find a mean loss rate for the poor of % for europe on average. if we consider an additional -month period of partial functioning of closed activities at % of full capacity (scenario iv), the mean loss rate for poor workers in europe increases up to % ( % if the period of partial functioning lasts for nine months). by countries, the highest loss rate for the poor is found in cyprus (between . % and . %), while the smallest one happens in romania (between . % and . %). the share of workers under the poverty line also increases significantly for the four simulated cases in all european countries (see table , columns - ). for europe on average, this increase varies between . percentage points under two months of lockdown and . percentage points if we consider an additional six months during which closed activities are partially functioning at %. these values imply that −in absence of compensating policies− the percentage of poor people in europe (those with earnings below % of the pre-lockdown median in the country) may substantially increase even if lockdown does not last long. if closed the contribution of growth and redistribution to poverty reduction over the period to in countries, including europe, is estimated in bluhm et al. ( ) . the relationship between the components of social mobility (growth, dispersion and exchange mobility) and inequality is estimated for countries of the european union in prieto et al. ( ) . the headcount poverty index gives additional relevant information about the consequences of the pandemic for poverty since this index is not necessarily consistent with the lic (essama-nssah and lambert, ). eastern and southern countries than central and northern countries. thus, we find that the highest increase in poverty according to the headcount poverty index is in croatia (between . and . points). on the contrary, the smallest percentage increase in poverty according to the headcount index happens in switzerland for case i ( . points) and denmark for cases ii ( . points), iii ( . points) and iv ( . points). if we look at the relationship between these changes in the headcount poverty index and the average lwa we find a negative correlation with = . for the first scenario ( figure , panel a) and = . for the fourth scenario ( figure c , panel a, in appendix c). when we consider the gini of lwa within countries instead of the average lwa the correlation becomes positive with = . for the first scenario (figure panel b) and = . for the fourth scenario ( figure c , panel b, in appendix c). the uneven distribution of potential wage losses across the distribution is prone to have an impact on wage inequality as well. european countries already have important differences in pre-lockdown inequality (table , pre-lockdown wage inequality ( = . ) . other countries with low levels of inequality before the lockdown are sweden, czechia, belgium and norway, all of them with a gini index slightly below . . on the other extreme of the spectrum, we find the countries with the highest level of pre-lockdown wage inequality: bulgaria, ireland, the uk and spain, all of them with a gini index above . . when comparing wage inequality after the lockdown with the baseline, it is observed that absolute and relative changes in inequality are sizeable and increase in all countries with the duration of the partial closure of some activities (table ) . for example, the relative increase of the gini coefficient, at scenario (i), ranges from . % (the netherlands) to . % (cyprus). at the more severe scenario (iv), relative increases in inequality range from . % in france to . % in cyprus (if the period of partial functioning lasts nine months these rates become % in france and % in cyprus). a scrutiny of the absolute changes in the gini coefficient −which makes the change scale independent of the previous level of inequality− reveals that cyprus is again the european country where inequality increases the most for the four simulation cases (between . and . gini points) followed by ireland (between . and . points). on the other hand, the smallest absolute change in inequality is found in norway ( . points) for scenario (i), in norway and france for scenarios (ii) ( . points) and (iii) ( . points), and in france and denmark for scenario (iv) ( . points). these variations in inequality also tend to be related with the lwa index. in figure given the link between the values of the lwa index and the increases in wage inequality (and poverty), it is not surprising that the observed changes are in general greater in eastern and southern countries than central and northern countries. as mentioned before, northern and central europe present large average levels of essentiality and teleworking, and low average levels of closure. meanwhile, eastern europe shows the opposite pattern, i.e., low average levels of essentiality and teleworking, and medium-high average levels of closure. the partial closure of activities that imply the agglomeration of large groups of people like tourism significantly penalises southern europe, whose countries tend to have the highest average closure scores. while some tertiary jobs like administrative, programmers and accountants are not affected by partial closures, others are strongly impacted by the measures to fight covid- and the fears to table . wage inequality changes in europe. note: cl , cl and cl refer to partial closure for months at %, % and % of full capacity, m is months, and ∆ a g and ∆ r g are the absolute and relative changes in the gini index. bootstrapped standard errors are in parenthesis. be infected: hospitality, restaurants, arts and entertainment. for this reason, the impact on inequality and poverty is also related to the share of these specific tertiary occupations. our findings show an increase in inequality for all european countries but, would inequality changes be different enough to increase inequality between countries? the answer is yes, although they are smaller than the inequality changes occurring within countries. in table we show the results of the decomposition of wage inequality for all european workers. our simulations show an increase in overall inequality in workers' salary of . % in europe ( . to . ) according to the mld index under a lockdown of two months. for the same scenario, the changes in the between-and within-countries inequality components are . % (from . to . ) and . % (from . to . ), respectively. for two months of lockdown and six months of partial functioning of closed activities at % of capacity (scenario iv) we find the following increases in inequality for europe on average: . % in overall inequality, . % in between-countries inequality and . % in within-countries inequality. when the period of partial functioning of closed activities at % of capacity lasts nine months the corresponding increases are: . % in total inequality, . % in between-countries inequality and . % in within-countries inequality. both components of overall inequality increase, but the within-countries inequality component increases significantly more than the between-countries inequality component in all scenarios. that is, cohesion between european countries decreases with the lockdown, though the main change in wage inequality happens within european countries. with the partial closure of some activities, changes get larger and the double process of wage divergence (between and within countries) deepens. the emergency measures adopted to contain the spread of covid- all around the world are largely based on social distancing and closure of high-risk productive activities. the paralysis of production imposed by the contention measures during the lockdown and the capacity limitations driven by official restrictions and by consumers precautionary behaviour will thus have an uneven impact on workers from different occupations and industries. our analysis reveals a sizable potential increase in poverty and inequality across europe. table . the between-and within-countries inequality components in europe. note: ∆ a is the absolute change in wage inequality and ∆ r (%) is the relative change in wage inequality. standard errors are in parenthesis. poverty will increase under our simulations in all countries. under the most conservative scenario, a lockdown of two months, we estimate a mean loss rate for the poor of % and an increase in the headcount index of . percentage points on average in europe, with the change ranging from . points (switzerland) to . points (croatia). likewise, wage inequality increases under a lockdown of two months, being the change in the gini coefficient equal to . % for europe on average, with changes ranging between . % (netherlands) and . % (cyprus). considering a more severe scenario with months of partial closure at % of full capacity after a two-month lockdown, we estimate a mean loss rate of . % for the poor workers in overall europe, a rise of . percentage points in the headcount poverty index and a gini increase of . % on average for europe. our results also highlight that lockdown measures are likely to worsen cohesion in europe both between countries and, especially, within countries. our simulations show that between-countries inequality will increase in europe between . % and . %, while within-countries inequality will increase between . % and . %. in general, we find a greater increase of both poverty and inequality in eastern and southern europe than in northern and central europe. workers tend to have a lower and more unequally distributed ability to work under the shutdown and social distancing in the economies of eastern and southern europe than in the northern and central european countries. these differences across european areas get larger with the severity of the measures to fight and prevent covid- infection. impacto de la covid- sobre el consumo en españa en tiempo real y alta definición poverty accounting decomposable income inequality measures distant and unequal. lockdown and inequalities in italy how did covid- stabilization policies affect spending and employment? a new realtime economic tracker based on private sector data supply and demand shocks in the covid- pandemic: an industry and occupation perspective how many jobs can be done at home? tax progression and inequality of income distribution measuring pro-poorness: a unifying approach with new results path independent inequality measures will covid- affect inequality? evidence from past pandemics decreto-legge febbraio , n. . retrieve at: www fear, lockdown, and diversion: comparing drivers of pandemic economic decline occupational exposure to social distancing: a preliminary analysis using o*net data ilo monitor nd edition: covid- and the world of work world economic outlook update working from home, wages, and regional inequality in the light of covid- what is pro-poor growth poverty equivalent growth rate which workers bear the burden of social distancing policies? a study on the relationship between economic inequality and mobility measuring pro-poor growth global inequality when unequal countries create unequal people the class of additively decomposable inequality measures an economic definition of poverty an axiomatic characterization of the watts index key: cord- -lv fy e authors: dávalos, alberto; henriques, rossana; latasa, maría jesús; laparra, moisés; coca, maría title: literature review of baseline information on non‐coding rna (ncrna) to support the risk assessment of ncrna‐based genetically modified plants for food and feed date: - - journal: nan doi: . /sp.efsa. .en- sha: doc_id: cord_uid: lv fy e this report is the outcome of an efsa procurement (np/efsa/gmo/ / ) reviewing relevant scientific information on ncrna and on rna interference(rnai) that could support the food and feed risk assessment of ncrna‐based genetically modified (gm) plants. information was retrieved through key words and key questions covering the stability and degradation of ncrnas after oral ingestion, the passage of ncrnas from food and feed to human and animal organs and tissues via the gastrointestinal tract and other barriers, as well as the potential effects on the gastrointestinal tract, the immune system or the entire organism.full description of the strategy used for the literature search and for studies selectionis provided and the number of retrieved publications is reported. this report is divided into four partsdiscussing the kinetics of exogenous ncrnas in humans and animals, with focus on ingested ncrnas (part ); the possible effects of ncrnas on the gastrointestinal tract (part ), systemically(part )and on the immune system (part ). this report suggests that some plant ncrnas (e.g mirnas and sirnas) show higher stability as compared to other ncrnas due to peculiar chemical characteristics ( ’‐o‐methylation at ’ end).however, ingested or administered ncrna must overcome many extracellular and cellular barriers to reach the intended target tissue or functional location in sufficient amount to exert any biological effect. literature data indicate that chemically unmodified and unformulated ncrnas exhibit very low stability in the gastrointestinal tract and in biological fluids and, in general, do not elicit major biological effects.this report also provides an overview of the rna content in plant‐derived foods and diets and discusses the controversies on the presence of dietary exogenous rnas in the biological fluids of humans and animals and their effects. finally, gaps in the scientific literature are highlighted and recommendations provided . introduction this scientific report is the result of a contract awarded by the european food safety authority (efsa) to the madrid institute of advanced studies (imdea)-food, under the contract title: "literature review of baseline information on non-coding rna (ncrna) that could support the food/feed risk assessment of ncrna-based gm plants" (contract number np/efsa/gmo/ / ). for the literature review of the available scientific information on foreign (exogenous) ncrnas that could support the risk assessment of ncrna-based gm plants, the following tasks were defined by efsa. based on available scientific literature (narrative review) to review the kinetics profile of foreign (exogenous) ncrna in humans and animals (experimental animals, livestock and pets). this to be based on information from research and development of ncrnas intended to be used as therapeutics and/or from research on biomarkers in humans (pharmaceuticals/medicine area) and focused primarily on the oral route of administration of ncrna, with detailed information on the absorption, distribution, metabolism and excretion of ncrna molecules. other routes of administration, however, not to be excluded. based on available scientific literature (narrative review), to review the effects (physiological, paraphysiological, and pathological) of foreign (exogenous) ncrnas on the gi tract and annex glands (e.g. liver, pancreas, salivary glands) in humans and animals (experimental animals, livestock and pets). this to be based on information from research and development of ncrna molecules intended to be used as therapeutics and/or from research on biomarkers in humans (pharmaceuticals/medicine area). based on available scientific literature (narrative review), to provide information on the possibility of systemic effects of foreign (exogenous) ncrnas in humans and animals (experimental animals, livestock and pets) with focus on gastrointestinal barrier to absorption, and other barriers as relevant (e.g. placenta). based on available scientific literature (narrative review), to assess the plausibility of effects on the immune system of humans and animals (experimental animals, livestock and pets) of foreign (exogenous) ncrnas. gm plants based on silencing approaches by rnai (through ncrna expression) are developed for food and feed purposes and assessed by efsa within the eu gmo regulatory framework. the team considered that pursuing the four tasks defined by efsa to support the risk assessment of gm plants in this regulatory context required gathering preparatory baseline information on ncrna and refinement of the scope of the tasks. therefore, further elaboration on the terms of reference provided by efsa has been conducted and it is described below. it should be emphasized that the available information on ncrnas which could be possibly relevant to food and feed safety, overlaps with and is barely distinguishable from the broader information on rnas. information on rna in general has therefore been included in the search as warranted; when possible, the relevance of distinguishing peculiar ncrnas features is described. the term "foreign ncrna" or "exogenous ncrna" used in this report refers to ncrna molecules to which humans/animals can be exposed through the diet or via therapeutic treatment. similarly, "foreign rna" or "exogenous rna" refers to rna molecules to which humans/animals can be exposed through the diet or via therapeutic treatment. part : kinetics of exogenous ncrna in humans and animals (efsa task ) efsa task : based on available scientific literature (narrative review) to review the kinetics profile of foreign (exogenous) ncrna in humans and animals (experimental animals, livestock and pets) . this to be based on information from research and development of ncrnas intended to be used as therapeutics and/or from research on biomarkers in humans (pharmaceuticals/medicine area) and to be focused primarily on the oral route of administration of ncrna, with detailed information on the absorption, distribution, metabolism and excretion of ncrna molecules. other routes of administration, however, not to be excluded. to address efsa task , preparatory baseline information is provided i) on general features of plant ncrnas including their function, movement in the plant, biogenesis and degradation (section . . ), ii) on their stability (i.e. for how long ncrna molecules retain their original structure, and how they resist degradation over time and in various conditions, both inside and outside the plant), and turnover, also in comparison with turnover of endogenous ncrna in animals (section . . ). if necessary, comparisons between the ncrnas class and other rna classes are made. general information on rnas used/intended for use as therapeutics (including ncrnas) is provided, expanding on the chemical modifications necessary to avoid degradation and to achieve a target effect after administration (section . . ). lessons from rna-based therapeutics as regards the pharmacokinetics of exogenous rna are discussed, with a focus on non chemically modified (naked)ncrnas. in addition, aspects of the pharmacodynamics of rnas as learnt by therapeutics are addressed (section . . ). a description of cellular uptake of rna (in general and with focus on ncrnas), and of the many barriers represented by human/animal cells after oral ingestion is addressed insection . . . further relevant information on dietary exposure to plant rnas (i.e. exposure to plant rnas following dietary consumption) by different diet types, and presentation of the gaps in the literature on dietary plant rna exposure is provided in section . . . efsa task : based on available scientific literature (narrative review), to review the effects (physiological, paraphysiological, pathological) of foreign (exogenous) ncrna on the gastrointestinal tract and annex glands (e.g. liver, pancreas, salivary gland) to address efsa task preparatory baseline information is provided on i) the gi tract barriers to ingested exogenous rnas, including ncrnas (section . . ); ii) experience in rna-based therapeutics for oral (gi) administration, with focus on delivery of naked (non chemically modified) ncrnas (section . . ). biological effects of dietary exogenous ncrnas (plant and nonplant origin) on the gi tract and its annex glands is presented in section . . . www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. to address efsa task , preparatory baseline information is provided on the molecular pathways relevant for the uptake of exogenous ncrnas by human and animal cells and the intracellular trafficking that follows, with description of the effects by exogenous ncrnas once they reach the specific subcellular location (section . . ). to exert systemic biological effects, in most cases the ingested exogenous rna must enter the systemic circulation to reach the target tissues. information is therefore provided on the landscape of exogenous rnas in biological fluids and tissues from humans and animals, with focus on ncrna when possible (section . . ). a description of possible systemic biological effects of dietary exogenous ncrnas is provided in section . . , with details on tissue barriers (i.e. placenta, brain) encountered. a detailed review is made of the contradictory information on the systemic effects exerted by plant-derived exogenous ncrnas. finally, studies on the safety and in general on risk assessment on ncrnas from gmos are reviewed (section . . ). efsa task : based on available scientific literature (narrative review), to assess the plausibility of effects on the immune system of humans and animals (experimental animals, livestock and pets) of foreign (exogenous) ncrna molecules. to address efsa task , preparatory baseline information is provided on ncrna mediated regulation of the immune system in humans and animals. the many pathways of sensing exogenous rnas, both at the endosomal compartment and in the cellular cytosol, and the mechanism of triggering the immune system are reviewed (section . . ). an overview is done of the plausibility of biological effects of exogenous plant ncrnas on the regulation and function of the immune system upon uptake by mammalian cells (section . . ). in addition, since the gut microbiota influences the immune system, a review on the possible effects of exogenous ncrnas as gut microbiota modulators was considered relevant and it is provided (section . . ). in the final section of this report, concluding remarks inform on the gaps existing in the scientific literature, and highlight areas needing further investigations to better understand and support the food and feed risk assessment of ncrna-based gm plants. an extensive search was done to identify as many studies as possible relevant to the literature review questions. to include all possibly relevant information multidisciplinary databases and information resources were explored. unpublished research reports ("grey literature") including dissertations, thesis or other scientific reports were also included, which were retrieved using general search engines (i.e. google). the following information sources, databases or search engines ( table ) were used for the literature review: pubmed (biomedical), world wide science (multidisciplinary), web of science (multidisciplinary), springerlink (multidisciplinary), scopus (multidisciplinary), scielo (multidisciplinary) and biorxiv (preprint biology). finally, the publications reporting the outcome of two efsa procurements aiming respectively at investigating and summarising the state of knowledge on the mode-of-action of dsrna and mirna pathways, the potential for non-target gene regulation by dsrna-derived sirnas or mirnas, the determination of sirna pools in plant tissues and the importance of individual sirnas for silencing ; and reviewing relevant scientific information on rna interference that could serve as baseline information for the environmental risk assessment of rnai-based gm plants ) were also used. www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the methodology used for this literature review process and report preparation followed three basic principles: i) methodological rigour and coherence in study retrieval and selection; ii) transparency; and iii) reproducibility. to ensure that these principles were implemented in this literature review process, the search methodology design, study selection and data and results presentation follow, whenever possible, most of the methods and techniques described in the efsa guidelines on application of systematic review methodology for food and feed safety assessments to support decision-making (efsa, ) . the following section describes the general key steps followed in the literature search methodology; detailed descriptions of the literature search methodology for each topic are also provided in this section. to find a comprehensive set of scientific literature related to exogenous ncrnas and their biological effects on a target organism, either local or systemically, key questions were identified and key words were used as search queries for each task section according to a pre-defined review process. to provide a fit-for-purpose literature search, the team defined key questions based on the efsa tender specifications and their interpretation as above described. to support this phase, the team identified key elements of the questions considering the pico approach of efsa ( ). the key elements identified include the population, in this case humans or animals (experimental animals, livestock and pets) as defined by the terms of reference of efsa. the intervention was identified as exogenous rnas (focusing on ncrnas). the comparator was identified as the control or reference group (not exposed to exogenous rnas) in most studies, whenever possible. the comparator could also be identified as the modified (either chemically or biologically) version of the exogenous rnas when referring to the stability or pharmacokinetic of the exogenous rna. the outcome is the biological effect (if any) as a response to the intervention. other specific key questions are determined for every part of the specific tasks defined by efsa. evidence is generally scarce for most of the topics covered by this scientific review, and in some cases the information was available from not directly related studies (i.e. pharmaceutical industry studies). the literature review was helpful in identifying both knowledge gaps and unexpectedly relevant studies or evidence not previously known to exist. these knowledge gaps are used to make informed proposals for future research designs (section . ). the key methodological steps in the literature search, selection and review process are shown in figure . three phases were defined in this process. the preparatory phase started with team discussions of topics to be included in the literature review. during this phase key elements addressing the efsa tender specifications and their interpretation as above described were identified and key questions were defined. the key questions aimed to to assist the literature search in order to produce a comprehensive overview of the topic that best fits tender specifications. the bibliography search and selection phase included several steps. the initial search in bibliographic databases and search engines was done using task-specific key words. the key words used for the search were derived from the efsa tender specifications and their interpretation (tasks - ) and were complemented with other key words identified in the preparatory phase. identified key words are detailed for each task in section . the final phase of review and final selection involved two additional steps, i.e. a detailed review of all relevant previously selected documents and a systematic evaluation of the final selected studies. a detailed review of the selected documents was done for studies with full text availability (either freely available or available for purchase) and, based on this, a final selection was made of full-text studies most relevant to answer the key questions for tasks ( - ) and apt for review in this report. a systematic evaluation of these final documents was done to develop a comprehensive overview of the topic. conflicting or contradictory results were reported in a neutral way and supported by references and the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. citations of all available evidence, both from the published and grey literature. retracted papers were carefully identified and noted. finally, each section of the narrative review was assessed by one reviewer. if more specific data was needed because of missing or novel information, additional bibliographic research was performed. additional key words that could directly address the specific data needed were identified and used to run a new specific search. the search among full-text studies was focused on obtaining only specific missing or required data (e.g. quantitative data on rnas levels in plants, earlier studies on exogenous rna exposure to mammalian cells). this new specific search generally represented less than % of the literature included in this report and it was performed only once for certain topics. finally, to reduce the risk of introducing bias into the literature review process and to assure reproducibility in the review methodology the following general approach was applied: when the search of the identified "key words" in different databases resulted in a myriad of duplicate documents, manual removal of these duplicates was performed in each section by each team expert; when relevant studies that might answer the identified key questions were identified after the first search run, these were used to further refine the literature search, for example refining the keywords; each selected full-text study to be included in the result section was manually examined by the team. to identify literature relevant to the topic, the methodology described in section . . . ( figure ) was applied, with adaptations. a multiple-step approach was followed based on the expertise of the team members who carried out research in the area. following a first search based on identified keywords, a careful selection of retrieved review studies was done, followed by a search of specific publications referred to in these reviews. additional specific key words were identified, and an additional search was run. this search retrieved a relatively small number of studies; therefore, team members reviewed all these to determine which were relevant to the topic. duplicates, irrelevant or erroneously assigned publications were removed. team members also cross-checked each others' findings, sharing discussing the results of their searches. the following areas were investigated. the purpose of this search was to provide general information on rnas function, movement in the plant and aspects of biogenesis and degradation, with a focus on ncrna (either small or long). by comparing animal and plant ncrna pathways, the aim was to identify similarities between animal and plant ncrna processing machineries that would indicate the possibility of ingested plant-derived ncrnas being processed in animals. in addition, this section was organized to avoid details that were outside the scope of this literature review in the topic of plant ncrna biology. this serves as an introductory chapter allowing easier integration of the knowledge presented in later chapters. the following combination of key words or phrases was used in some of the databases indicated above: plant small rnas; plant and sirna; plant and mirnas; plant and lncrnas; plant and circrnas; plant rna and movement. www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the initial search using the above-mentioned key words or phrases yielded a large number (≈ ) of documents ( table ) , most of which were not relevant to the topic. particularly, the databse web of science retrieved documents not directly related to the topic. to assist and refine the search, studies were further filtered by their relevance to the key questions identified: -what type of ncrnas are described in plants? -what are the main differences between mammalian and plant ncrnas? -what is the function of plant ncrnas? -what is known about the movement of plant ncrnas? title and abstracts were evaluated, and studies further refined using the key questions. detailed study of the most relevant full-text documents answering the key questions, retrieved documents which were finally selected as relevant to give an overview of plant ncrnas. search was then refined by using other key words including "half-life", "turnover", "fate" or "degradation". the identified studies were then manually evaluated by title and abstract and their number further limited due to their low relevance to the below identified key questions for this specific subchapter: -are plant ncrnas more stable than animal (mammalian) ncrnas? -if so, what makes a plant ncrna more stable than its animal (mammalian) counterpart? -why is it relevant to consider ncrna stability outside the organism of origin (plant)? -are certain types of plant ncrnas more or less stable than others? -what is known about the half-life of ncrnas? documents on the stability of rnas (not only ncrnas) were further screened for relevance to the question. most of the studies in the literature describe the stability of a specific class of intracellular rnas (i.e. mrnas). the stability of rnas other than ncrnas (e.g. mrna, trna), unless are used as model of general ncrna stability, is outside the scope of this literature review, since it is very unlikely they would be used as baseline information for food/feed risk assessment of ncrna-based gm plants. these keywords were specifically selected since they describe the kinetic profile of exogenous ncrnas in human and animals. the word "pharmacokinetics" was included in the search because it is a general term that retrieves data on absorption, distribution, metabolism or excretion of any molecule, particularly those intended for use as therapeutics (pharmaceutical/medicine areas). the keywords "increased intestinal permeability" or "increased plasma clearance" are intended to retrieve any study indicating changes in these parameters. although a large number of publications unrelated to ncrnas were expected, these keywords were chosen to initially explore the general aspects of physiological or pathological conditions associated with modification in these parameters. the search was further refined after this initial exploration. the search of the above-mentioned key words on different databases yielded ≈ . entries ( table ) . a quick exploration of titles and abstracts suggested that (as expected) a number were unrelated to or were not useful to appropriately address the topic. it is worth noting that the search in the previous section (section . . . .) indicated that the chemical modifications generally introduced for rna-based therapeutics are normally absent in nature. scientific data that could serve as a baseline to support risk assessment of ncrna-based gm plants should contain primarily information on ncrnas resembling those present in the nature or in gm plants. therefore, studies on naked exogenous rnas were analysed to prepare this narrative review. to promote comparisons between natural and synthetic rnas, a very few examples of slightly chemically modified rnas were included. heavily chemically modified rnas were not considered due to their unlikely existence in nature. only in vivo studies were considered, both in animals (mammals) or humans. studies were further evaluated for their relevance to answer the key questions identified for this section: -what are the pharmacokinetic properties of rna-based therapeutics? -what routes of administration have been used for rna-based therapeutics? -what is it known about the pharmacodynamic properties of rna-based therapeutics? -do disease conditions modify the pharmacokinetic properties of rnas when administered to mammals? the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. after detailed evaluation of the full-text reports, documents meeting the described criteria were finally selected for review. understanding whether and how exogenous rnas are taken up by cells and what barriers are encountered is relevant to study the possible absorption and systemic exposure to ncrnas introduced by food and feed. this topic is reviewed in this dedicated section and excluded from the section on the pharmacokinetics of exogenous rnas (section . . . . above). to search and select the relevant studies concerning the uptake of exogenous rnas in mammals, the following combination of key words was used in some of the databases listed above: rna and cellular uptake; ncrna and cellular uptake; mirna and cellular uptake; lncrna and cellular uptake. these key words were used to screen for studies where "cellular uptake" was mentioned in the title or abstract for any type of rnas. it was also decided to include specific types of ncrnas including "mirna" or "lncrna" to expand the search to literatures where the more general term "rna" was not necessarily used. the initial search of scientific documents using the above keywords yielded ≈ documents ( table ) . these were explored by their relevance (title and abstract), and only studies addressing naked ncrnas were considered for full text analysis, since highly chemically modified rnas are not common in nature. to address the reference terms for task- of this procurement, and after expert discussion for this section (preparatory phase) key questions were identified and the studies then further filtered for their relevance to the these: -are exogenous rnas taken up by mammalian cells? -how does exogenous rnas uptake in mammalian cells occur? -what are the biological barriers to consumed rnas? -what are the biological barriers for the exogenous rnas after absorption and entering into the mammalian circulatory system? only studies where exogenous rnas were evaluated were included in the literature review. after detailed evaluation of full-text reports and expert judgement on their relevance to efsa task , documents were finally selected for review. to select relevant studies for plant rna exposure, the following combination of keywords were used: plant and rna content; plant and mirna content; plant and sirna content; plant and lncrna content; plant and dsrna content; rna intake and vegetarians; vegetables/fruit intake and vegetarians. these specific keywords were used because the wording "rna (ncrna, mirna, sirna, dsrna, lncrna) content" would yield most of the documents related to quantities of rnas in a specific substrate. these were combined with "plant" to retrieve only those studies where plants were used to evaluate rna content. it was also decided to use "rna intake", "vegetable intake" or "fruit intake" to specifically refer to consumption, and to combine this with "vegetarian" to search only for studies where comparisons are made between vegetarian and other types of diets. alternative wordings including "rna amount" or "vegan" were used to search for alternative information. using the above keywords, ≈ studies were retrieved (table ) during the initial search. to more appropriately cover this section and to refine the search, the following key questions were identified during the preparatory phase: -what is the general amount of rnas naturally occurring in edible plants? -what percentage of total plant rnas are ncrnas, mirnas, sirnas, lncrnas or dsrnas? -does exposure to rnas change in different diet types? -do changes in rnase activity, due to pathologies or human polymorphisms, influence dietary exposure? titles and abstracts were evaluated, and studies further filtered using the key questions above. the search was also refined to focus on those studies where quantitative rnas data were mentioned. only in vivo exposure studies were used (either epidemiological or experimental). common types of diets were only considered (i.e. vegetarian, vegan or omnivorous). for rna exposure by different diet types, studies where comparative analysis of at least two different diets (one of which was the general population diet type) were mainly considered. in all cases, full-text studies were evaluated to screen for relevance to answer the identified key questions. a detailed study of the full-text documents resulted in the identification of studies most relevant to address this specific topic of task- , and which were reviewed for this section of the report. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. to identify literature relevant to the topic, the methodology described in section . . . (figure ) was applied. during the preparatory phase key questions were identified to assist in the literature search. the document search was first done using specific key words and then refined according to the key questions. detailed analysis of the full-text documents was done by team members with expertise in the area. search and analysis results were discussed collectively among team members. the following areas were investigated: in this subsection, there is a general description of the biological barriers within the gi tract that exogenous rnas would need to overcome to exert a biological effect, either locally or systemically. to select relevant studies, the following combination of key words or phrases were used: "gastrointestinal tract anatomy"; "gastrointestinal tract physiology"; intestinal transport pathway; physicochemical properties of rnas. an exploration of titles and abstract of the yielded entries (table ) suggested that a large number of documents were unrelated to the topic. only one key question was identified in the preparatory phase: -what are the biological barriers within the gastrointestinal tract to ingested exogenous rnas? the search for relevant studies addressing this key question was based mainly on team member judgement. detailed study of the most relevant full-text documents answering the key question retrieved documents which were finally selected as relevant to this section of the report. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. this search was intended to collect data from rna-based therapeutics employing oral administration (and other gi routes) which could support the search on the possible biological effects of plant dietary exogenous rna and ncrnas on the gi tract and its annex glands. the initial search was done using the following key words on the different databases: ncrna oral delivery; rna oral delivery; sirna oral delivery; aptamer and oral; sirna gastrointestinal delivery; mirna gastrointestinal delivery; oral exosome rna; extracellular vesicle and gastrointestinal. these key words were selected to search for any documents related to rnas and oral administration. any information on oral administration was considered relevant. to expand the search, different terms referring to different types of ncrnas (mirna, aptamer, sirna or others), and a synonym for "administration", i.e. "delivery", were included. the initial search using the above key words or phrases yielded a small number (≈ ) of documents ( table ). the different databases generated similar number amounts of publications, most of which were duplicates. using the key word "lncrnas" in combination with the other key words did not yield any information relevant to the topic. indeed, just the search using "lncrna exosome" identified fewer than documents, but none was relevant to oral delivery or gi administration. the wording "dsrna and oral" produced ≈ documents, but none of them was relevant to mammalian organisms. the documents were selected by relevance, evaluating title and abstract. in this section, only studies in animals (mammals) or humans were considered. when replacing the word "delivery" with "administration", ≈ additional documents were identified, but almost % were unrelated to the review topic. since most of the documents in the literature describe minimally modified rnas for oral use, all chemically modified rnas were considered in this section. these can provide baseline information to support food/feed risk assessment of ncrna-based gm plants. to assist and refine the search, studies were further filtered for their relevance to the key question identified: is the oral route of administration relevant for rna-based therapeutic development? -what types of formulation (delivery vehicles) are tested for rna-based therapeutics for the oral route of administration or other gastrointestinal tract routes of administration? the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. -do exogenous ncrnas (naked rnas) resist the conditions of the gastrointestinal tract environment? -could ncrnas-based therapeutics -delivered by oral administration or other gi tract routes of administration-exert a biological effect in the gi tract or its annex glands? -are there any natural vehicles for oral administration of exogenous ncrnas? from the initial ≈ entries identified using the above key words and after filtering for relevance to the key question with evaluation of titles and abstracts and detailed study of the full-text reports, documents were selected for review in this section. twenty ( ) of these documents reports selected examples of local effects of nucleic acids-based drugs (mainly exogenous rnas) within the gi tract, using oral administration. since efsa required a literature review also including livestock species, an additional specific search was run for birds and fish. this search was intended to compile information on administration of exogenous ncrnas in either fish or birds, regardless of administration method, exerting any biological effects in vivo. the initial search was done using the following key words on the different databases: exogenous rna and fish; exogenous rna and bird; rna oral delivery and fish; rna oral delivery and bird; ncrna administration and fish; ncrna administration and bird. to expand the search, different terms referring to different types of ncrnas were used (sirna, mirna, circrna, or dsrna). the initial search using the key words or phrases produced a small number (≈ ) of documents (table ) in most of the databases. to assist and refine the search, only one key question was identified during the preparatory phase for this subsection: -are exogenous ncrnas also used to exert rnai effects in fish or birds? from the initial ≈ entries identified using the above key words, and after filtering for relevance to the key question, and evaluating titles and abstracts and detailed study of the full-text reports, documents were selected for the review in this subsection. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. this chapter provides baseline information supporting food/feed risk assessment of ncrnas-based gm plants. in the effort to cover the widest amount of literature containing relevant scientific data on exogenous ncrnas and their possible biological effects, the following key words or phrases were used in the search: dietary plant non coding rnas; food plant non coding rnas; exogenous plant non coding rnas; diet plant micrornas; food plant mirnas; breast milk exosomes; breast milk mirna; breast milk ncrnas; breast milk lncrnas; breast milk rna; dietary exogenous rna. these key words were selected to cover most, if not all, available studies on the topic ( table ) . the search of other common examples of exogenous ncrnas (non-plant origin) consumed by oral intake was focused on breast milk ncrnas, due to their relevance to human nutrition (table ) . almost all documents yielded by the three databases (pubmed, scopus or web of science) were duplicates. in this section, studies reporting local effects (in the gi tract or its annex glands when consuming exogenous ncrnas) were preferentially (but not exclusively) evaluated. other databases including the preprint server for biology (biorxiv), scielo or search engines (i.e. google) were also used. the initial search in some databases using the above key words yielded a number of entries (table and ) not exceeding ≈ documents. because this specific topic is especially salient to future understanding of the possible food/feed risk assessment of ncrnas-based gm plants, all the identified studies were further evaluated by title and abstract and their relevance to answering the identified key questions: does the literature describe the effect of exogenous plant-origin ncrnas when consumed orally? -does the literature describe negative results or contradictory results regarding the possible biological effect of ncrnas when consumed orally? -are ncrna stability properties under gi tract conditions normally evaluated? -are exogenous ncrnas evaluated quantitatively? -are exogenous ncrnas consumed in an amount sufficient to exert a local biological effect?are there other common examples of exogenous ncrnas (other than plant-origin) that could exert a biological effect on the gi tract and its annex glands when ingested orally? local effects (in the gi tract or its annex glands) are mainly described in this section. the possible mechanisms of plant exogenous ncrnas cellular uptake, their intracellular trafficking and systemic biological effects (including plasma levels) are reviewed in another section (section . .). of the initial ≈ documents retrieved using the above key words, and after filtering for relevance to the key questions by evaluating titles and abstracts, and detailed study of the full-text reports, documents were selected for the review in this subsection. from these, documents were selected for plant-origin ncrnas and for non-plant origin (breast milk) ncrnas. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. in line with the methodology described in section . . . . (figure ), a multiple-step approach based on the expertise of the team members responsible for this section was applied to identify literature relevant to the possible systemic effects of exogenous ncrnas. the document search was first done using specific key words in the databases and refined based on the appropriate key questions. detailed analysis of the full-text documents was done by team members and the results discussed collectively. the below areas were investigated. the search objective was to obtain information on basic molecular cellular uptake pathways of exogenous ncrna, including proteins involved in this process (if any). a review was also done on several other aspects of intracellular trafficking, once an exogenous ncrna is internalized, and how it exerts a biological effect. tissue barriers to exogenous ncrna function were also investigated. the content of this section deepens on molecular pathways previously described in section . . . to assist the search the following key words were used in the different databases: exogenous ncrna and mechanism and absorption; ncrna and mechanism and absorption; sidt (sid transmembrane family member ) and ncrna; sidt (sid transmembrane family member ) and ncrna; ncrna and uptake; intracellular trafficking and ncrna. these key words and terms were selected to identify any document related to the mechanisms of uptake and intracellular trafficking of ncrnas exerting a biological effect. the search was expanded by adding and combining different terms for exogenous ncrnas (see table ), including the general term "rna", www.efsa.europa.eu/publications efsa supporting publication : en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. or specific ncrnas such as "mirna". the initial search using the above key words or combinations thereof produced a moderate number of documents (table ). the different databases generated similar amounts of documents, most of which were duplicates. the largest proportion of the references was obtained using the key wording "ncrna and uptake" (≈ ). however, most of these had no relation to this specific section. the search was refined by filtering for relevance to answering the identified key questions: is there any clear transport of exogenous ncrnas into mammalian cells? -are orthologous of environmental rnas transporters in lower organisms (i.e. sid family of proteins) involved in the mammalian exogenous ncrna mechanism of uptake? -what molecular mechanisms are involved in exogenous ncrnas trafficking inside the cell? -what is the cellular fate of exogenous ncrnas once inside the cell? -what are the specific tissue barriers to exogenous ncrna function? very few documents evaluate the molecular mechanism of exogenous ncrnas uptake, intracellular trafficking and function. from the initial ≈ documents retrieved using the above key words, and after evaluation of their titles and abstracts and filtering for relevance to the key question, less than documents initially seemed relevant to the topic and were finally selected for review after detailed study of the full-text reports. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. this subsection is intended to retrieve available information on the presence of exogenous rnas in human and animal biological fluids (i.e. blood, plasma, serum or any other biological fluids) and tissues. the search focused on the presence of dietary plant-derived exogenous ncrnas, due to their relevance to the overall literature review. to cover the largest possible portion of the relevant literature, the following key words were used in the search: exogenous rna and human plasma; exogenous rna and serum; plant ncrna and human tissue; plant lncrna and human plasma; plant mirna and blood; plant mirna and tissue; plant rna and tissue. to expand the search, the above key words were used in different combinations for the same search (i.e. blood, serum or plasma for biological fluids) ( table ). the general wording "rna" was also incorporated into some searches. specific types of ncrnas were also added, including "mirna" or "lncrna", to avoid missing documents possibly related to these specific molecule types. the search using these key words or phrases in different databases yielded more than ≈ . publications (table ) . a quick exploration of the titles and abstracts suggested that only few studies were related to the review topic. only one key question was identified as relevant which was related to the presence or absence of evidence of exogenous ncrnas in biological fluids and tissues. since these findings are important for the risk assessment of ncrna-based gm plants, the key question was presented in its positive and negative forms as follow: are there studies describing the presence of exogenous rnas in the biological fluids and tissues of humans and/or animals? -are there studies contradicting the presence of exogenous rnas in the biological fluids and tissues of humans and/or animals? after a detailed assessment of the full-text, documents were finally selected for review. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. this search primarily aimed to gather information relevant to understand the possibility of systemic effects of dietary exogenous ncrnas after intake (oral administration) by humans and/or animals (mammals). information on the possible passage through specific biological barriers (i.e. placenta or brain) was also searched. the search was guided by the following key words to identify studies providing any data on the possibility of systemic effects of dietary exogenous ncrnas: plant ncrna and diet; plant exogenous rna and diet; plant rna and diet, plant environmental rnai and diet; plant rna and placenta; plant rna and brain; dietary rna and cross kingdom. these key words were used to guarantee that every single document potentially providinginformation on the possibility of systemic effects of exogenous ncrnas was found. to widen the search, other wording such as "environmental rnai", "cross kingdom" or "mobile rnas" were used. although the wording "systemic effects" was not used in the key words during the search, the above key words were used to retrieve any documents in which a biological effect (local or systemic) might be reported. different wording for the term ncrnas was also used (including lncrna, circular rna, mirna, or dsrna). the initial search in some databases using the above key words produced a small number of entries (table ) . the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. approximately ≈ documents were initially identified using the above key words (table ) . after filtering for their relevance to the key question by detailed assessment of the full-text reports, documents were finally selected for review. within this specific subsection, the selected documents are mainly grouped as studies that "support the evidence" or studies that "contradict the evidence" of systemic biological effects of dietary exogenous ncrnas in humans and animals (mammals). although not specifically included in the scope of this literature review, search for toxicological effects of dietary exposure to exogenous ncrnas in humans or animals is relevant to risk assessment. this search was aimed at identifying information on possible toxicological effects of plant-derived exogenous ncrnas following food/feed consumption. the initial search was conducted using the following key words in the different databases: the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. few documents were initially retrieved and after a quick evaluation of titles and abstracts, it became evident that very little information is available in this area. therefore, different terms were used to refer to exogenous ncrnas, including circular rna, lncrna and mirnas. the search using the above key words yielded a small number of documents (table ) . only in vivo studies were considered, both in animals (mammals) and humans (if available), with focus on dietary plant-origin exogenous ncrnas. the key questions identified to refine the literature search for this specific topic were: -is there evidence of any toxicological effects of plant-derived dietary exogenous ncrnas? -are the toxicological effects related to rnai-mediated interaction with the human or animal genome? -are there possible interactions related to unintended rnai-mediated gene regulation? approximately documents were identified using the above key words. after filtering for relevance to the key questions, and assessing the full-text reports, only documents were finally selected for review. a systematic and comprehensive review and collection of the literature relevant to the topic was performed with the aid of the team members' expertise. an extensive search using key words was initially done on multidisciplinary databases to avoid publication bias, followed by a grey literature search using general search engines. the resulting documents were screened by title and abstract to determine relevance to the topic and refined using the identified key questions. after full-text analysis of the documents, irrelevant documents were eliminated. all stages of the screening process were independently assessed and, in case of uncertainty, discussed with other experts to avoid personal biases. the methodology followed the key steps described in section . . . . (figure ) and a total of papers were identified, investigating the areas below described. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the final goal of this search was to identify relevant information on ncrnas-mediated processes associated with innate and adaptive immune responses. this section serves as an introduction to the studies presented later, and it is based on the results of the seach described in the next paragraph. sixty-four ( ) documents were reviewed for this section. the search was aimed primarily at gathering state-of-the-art knowledge on the possible biological effects of exogenous (plant) ncrnas on the regulation and function of the immune system. to cover this specific section, a combination of the following key words was used in the different databases: exogenous rna and immune and human/mice/rat; exogenous plant rna/mirna/sirna/lncrna/dsrna/transgenic rna and immune function/inmmunity and human/mice/rat; genetically modified plants and immune function/immunity and human/mice/rat; exogenous plant rna/mirna/sirna/lncrna/dsrna and primary cells and lymphocyte/monocyte/macrophage; exogenous rna/genetically modified plants and immune function/immunity and zebrafish. the key words were designed to identify all publications that use in vivo, including non-mammalian animal models, and in vitro ncrnas testing to evaluate the influence on tolerance processes as well as immune cell function. the search also included publications on in silico methods. the different key words were entered one by one into the search field 'topic' of the databases ( table ). the search was designed to identify documents including methods that could use non-mammalian animal models (i.e. zebrafish embryos). all search terms within one key word were combined by the operator 'or' and 'and' copied in a second or third search field. 'review' and 'meeting' documents were excluded. a search for grey literature was done in general search engines (i.e. google) using the different key words in the 'search' function of the respective websites. all the information on in vivo studies, using mammalian and/or non-mammalian organisms, and in silico studies was collected. these key words were chosen because this section aimed to describe the possible effects that exogenous ncrnas, specifically of plant dietary origin, might exert on the immune system of humans and animals. the search yielded ≈ entries (table ) . publications were excluded if one of the following criteria was met: the biological effect could not be unambiguosly attributed to ncrnas; the publication addressed immunodeficiency or autoimmunity; the publication reported no immunity (innate or adaptive)-associated endpoint or it is not specific for immune function and/or homeostasis evaluation; evaluation of exposure of and/or effect on cells other than immune cells (e.g. epithelial cells); the study was unable to measure processes related to immune function and/or homeostasis; the study reported data on potentially immunocompetent cells not fully differentiated as mature immune cell; no ncrnabased plant was tested in the publication; and the publication did not address plant-derived exogenous ncrna. the unexpectedly low number of publications retrieved using the key words proved a challenge in the selection process. for example, selection for in vivo studies produced only articles after title/abstract screening. to expand the number of documents, additional key word searches were included and less conservative criteria were set for the title/abstract screening. retrieval of grey literature was also based on information containing "preliminary results" or on-going projects that could possibly answer the key questions. www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. studies were further evaluated for their relevance to answering the key questions. the key questions identified using the above key words were: -which test methods or approaches are available to investigate the possible role of plant-derived exogenous ncrnas on the immune system of humans or animals? -what human or animal immune response(s), innate or adaptive, are possibly associated with administration of plant-derived exogenous ncrnas? -which mediators in humans and animals have been identified and/or ascribed to immune response(s) possibly mediated by administration of plant-derived ncrnas? -which innate immunity mediators (i.e. chemokines, cytokines) have been identified in humans and animals as associated to effects of plant-derived ncrnas? -which human or animal cells (i.e. lymphocytes, macrophages, myeloid) are possibly involved in immune responses mediated by plant-derived ncrnas? after the selection of the full-text documents, publications were excluded if not providing sufficient information (i.e. endpoint, exposure time), if investigating a mechanism other than immune-related processes, or if describing an operational procedure or guideline rather than a research study. from the initial ≈ documents identified using the above key words, and after filtering for the relevance to the key questions and detailed study of the full-text reports, documents were finally selected for review in the three sections of part (efsa task ). of these, documents were selected for review in this section. gut microbiota is important for the function of the immune system, therefore the purpose of this additional search was to provide general information on the effect of dietary exogenous ncrnas on the gut microbiota and the possible consequent secondary modulation of human or animal immune systems. this would serve as an introduction to identifying information gaps and experimental needs for future studies relevant to food/feed risk assessment of ncrna-based plants. selected full-text studies were analysed for their relevance to answering the following key question: -is there evidence for influence of exogenous plant ncrnas on the gut microbiota related to the immune system? five documents were finally selected for review in this section. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. results are presented narratively. when appropriate, tables and figures are included to allow better understanding of the topic covered by the literature review. some sections of this review provide only limited amount of quantitative analysis due to the nature of the available evidence and the key questions identified for each subchapter. at the end of each subchapter, a short summary presents the general points identified in the reviewed literature. gaps in the literature and open questions identified in the field are presented in a subchapter of certain sections. these gaps in the literature may serve as suggestions for further research in each topic. part : kinetics of exogenous ncrnas in humans and animals (efsa task ) ncrnas are transcripts that are not translated into any functional peptides or proteins. these transcripts include structural (transfer, ribosomal, small nuclear, and small nucleolar rnas) and regulatory ncrnas. the latter comprise both small and long ncrnas that can regulate gene expression by acting on chromatin, transcription, rna processing, rna stability and translation. this part of the report introduces general features of regulatory ncrnas including their function and aspects of their biogenesis and degradation. it also establishes possible comparisons between animal and plant ncrna pathways. this can be considered background information to clarify the possibility of plant ncrna biological effect in humans and animals that ingest food/feed from ncrna-expressing gm plants. in addition, ncrna movement inside the plant is also described. specific details on the structural ncrnas and the broad topic of plant ncrna biology are outside the scope of this literature review. following an extensive literature search as described above and based on the team expert decisions on the topic, this section is a review of scientific documents. small ncrnas (srnas) srnas are common to both plants and animals. two major classes of srnas are found in eukaryotes: micrornas (mirnas) and small interfering rnas (sirnas). they function as regulators of endogenous genes or as defenders from invasive nucleic acids. they are characterized by the double-stranded nature of their precursors, and differ from the less abundant piwi-interacting rnas (pirnas) which derive from fragmentation of single-stranded rnas (juliano et al., ) , and are primarily found in animals; they have not been described in plants. pirnas exert their functions in germ and stem cells through interaction with piwi-proteins (juliano et al., ) . by contrast, both mirnas and sirnas are bound to argonaute (ago) proteins, where they identify 'target' rnas by base-pairing interactions, and act in both somatic and germ cells. all srnas in plants are modified at the ' -terminus by '-o-methylation, including mirnas and sirnas, which lack this modification in animals. '-o-methylation is essential to conferring stability and protection from ' uridylation and degradation (borges and martienssen, , ) . mirnas and sirnas are distinguished by their origin and biogenesis (figure ) . mirnas are derived from processing of single-stranded precursors with a hairpin structure, whereas sirnas are generated from long, fully complementary double-stranded rna (dsrna) precursors (carthew and sontheimer, the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. endonucleases (dcls) that excise rna precursors into short double-stranded fragments, - nucleotide (nt) long, with -nt ' overhangs; and argonaute proteins (agos) that engage these duplexes and support their silencing activies based on target complementarity (carthew and sontheimer, ). further details on sirnas and mirnas are provided below. sirnas several different classes of sirnas with specialized biological roles have been identified in plants. the most abundant class consists of heterochromatic sirnas (hetsirnas), which are usually - nt long and originate from the repetitive and intergenic regions in the chromosome and transposable elements. hetsirna are crucial in dna methylation and chromatin modification through a process known as rnadirected dna methylation (daxinger et al., ) . hetsirnas are processed by dcl nuclease and preferentially loaded into ago . mature ago -sirna complex can interact with complementary noncoding nascent rna polymerase triggering the recruitment of dna methyltransferase (domains rearranged methyltransferse ) to silence target loci at the transcriptional level via dna methylation and repressive chromatin modifications (vaucheret, ; lee and carroll, ) . these hetsirnas may account for approximately half of a plant's total mass of srnas. another class of sirnas are naturalantisense transcript sirnas (natsirnas) which can be generated from dsrna precursors through hybridization of independently-transcribed complementary rna strands (vaucheret, ) . in addition, secondary sirnas generated as a "secondary effect" of mirna-mediated target cleavage are found in plants (axtell, ; vaucheret, ) . the mirna-mediated cleaved target is occasionally used by rna-dependent rna polymerase (rdr) to produce secondary sirnas, which can either give rise to a phased set of sirnas (phasirnas) or trans-acting sirnas (tasirnas) with the ability to target genes different from their loci of origin. this secondary pool of sirnas can greatly amplify and sustain a systemic silencing throughout the organism. recognizable rdr-encoding genes are present in the genome of many rnai-competent eukaryotes, with the notable exceptions of insect and vertebrate species in which these secondary sirnas are absent (carthew and sontheimer, ). the lack of these secondary sirnas might have a positive effect on specificity (as sirna amplification can lead to the silencing of multiple transcripts, specifically if they share a highly conserved sequence or a common exon) and a negative effect on the amplification of rna silencing in these species. plants have more diversified and specialized sirna-based pathways than other organisms, which are thought to contribute to plant plasticity . it is generally accepted that these pathways evolved as a cellular defence mechanism against rna viruses and transposable elements, which were later adapted to regulate the expression of endogenous genes. this is consistent with the fact that most small rna classes have a recognized role in defence responses, as well as in epigenetic regulation, and that plants have larger and repetitive genomes (borges and martienssen, ) . the diversification of srna-directed silencing pathways in plants occurred through the expansion of the rdr-polymerases, dcl and ago proteins. rdr genes are found in rna viruses, plants, fungi, protists and some animals, but are absent in flies, mice and humans; this is consistent with the fact that the vast majority of srnas in humans are mirnas (kurzynska-kokorniak et al., ) . absence of rdr activity may also justify the lack of intercellular spreading of rna silencing in vertebrates, whereas systemic silencing is a phenomenon widely reported in plants and nematodes. dicer or dicer-like proteins in plants constitute a four-member gene family, whereas vertebrates have only one member (mukherjee et al., ) . the diversification of agos has resulted in development of distinct gene-silencing processes based on differential ago affinities to small-rna duplexes (borges and martienssen, ) . www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. mirnas mirnas are well-studied srnas deeply conserved over long evolutionary distances, even between the plant and animal kingdoms. however, there are substantial differences between these two kingdoms with regard to mirna biogenesis, and mechanism and scope of mirna-mediated gene regulation (jones-rhoades et al., ) . detailed pathway comparisons and species information are described elsewhere . transcription of mirnas is typically performed by rna polymerase ii, and transcripts are capped and polyadenylated. in animals, most mirnas are derived from longer hairpin transcripts (pri-mirna) by the consecutive processing by the rnase iii-like enzymes drosha and dicer, whereas in plants only dicer (particularly dcl ) is responsible for mirna processing. thus, the biogenesis of most mirnas in plants occurs in the nucleus, whereas in animals it requires the sequential cleavage of pri-mirna in the nucleus and then in the cytoplasm by distinct rnaseiii enzymes (rogers and chen, ) . once processed, one strand of the hairpin duplex is loaded into an ago protein to form the core of the mirna-induced silencing complexes (miriscs). miriscs silence the expression of target genes predominantly at the post-transcriptional level (ptgs). the targets to be silenced are recognised through base-pairing interactions between the loaded mirna and mrna target, which contains a partially or fully complementary sequence (huntzinger and izaurralde, ) . plant mirnas recognize fully or nearly complementary binding sites, which are generally located within the open reading frames (orfs) of the mrna target. of importance is that mirna nt - are usually engaged in base pairing, which allows target cleavage by ago proteins (between nt and ). by contrast, animal mirnas recognize partially complementary binding sites, which are generally located in ' utrs. in both plants and animals, complementarity to the ' end of the mirna (the 'seed' sequence, containing nt - ) is a major determinant in target recognition and is sufficient to trigger silencing. for most mirna-binding sites complementarity is limited to the seed sequence (seed-matched sites) or to the seed sequence plus mirna nucleotide . however, in some rare cases complementarity to the ' region of the mirna might contribute to target recognition, particularly when the mrna has a weak seed match. even for these sites, however, mirna nucleotides - generally bulge out, preventing endonucleolytic cleavage by agos. in both animals and plants the mirna ' terminal nucleotide is buried in the mid domain of agos and is not available for pairing with the target (huntzinger and izaurralde, ) . therefore, there is an important difference in complementarity between plant and animal mirnas and their targets; this is less extensive in animals than in plants. in both animals and plants mirnas can move from cell-tocell. while in mammalian cells mirnas and other types of rnas can be transferred through secretory vesicles (ruvkun, ; valadi et al., ) , in plants they mostly use a different mechanism (kobayashi and zambryski, ) (see section . . . . for details). however, recent evidences also suggest that plants can send small rnas in extracellular vesicles to pathogens to silence virulence genes (cai et al., ) . regarding the silencing mechanism, mirnas were initially thought to inhibit translation in animals and to predominantly promote target endonucleolytic cleavage in plants. however, recent evidence has changed this view by showing that mirnas can trigger translational repression and mrna destabilization in both kingdoms. in both plants and animals, the current evidence suggests that target mrna degradation provides a major contribution to silencing by mirnas (huntzinger and izaurralde, the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. next generation sequencing approaches have revealed a new transcriptional landscape in which many novel lncrnas have been identified. these are transcripts longer than bp without any clear protein coding potential. lncrnas are transcribed either from intergenic regions (lincrnas), introns (incrnas) or the opposite strand to protein-coding genes or other lncrnas; they are thus natural antisense transcripts (nats) (ariel et al., ; chekanova, ; shafiq et al., ; yamada, ) . as a confirmation of their widespread nature and possible biological relevance, lncrnas have been identified in plants, fungi and animals. in fact, lncrnas are involved in diverse biological processes across the eukaryotes, ranging from regulation of mating type in yeast to the pluripotency of embryonic stem cells in mammals (zofall et al., ; flynn and chang, ) . in plants, lncrnas play key roles in flowering time regulation, gene silencing, root organogenesis, seedling photomorphogenesis, and reproduction (ariel et al., ; chekanova, ; shafiq et al., ; yamada, ) . although plant lncrna biological characterization is still limited, detailed analysis of over arabidopsis thaliana transcriptome data sets identified , putative lncrnas, including approximately , nats and over lincrnas. both in plants an mammalians (jin et al., ; liu, j et al., ) these lncrnas do not show association with small rnas and are also expressed at lower levels ( -fold to fold less) than mrnas. plant nat-lncrnas can overlap completely ( %) or have complementary sequences in the ' or ' regions of mrnas. they accumulate in a tissue-specific manner and many are modulated in respond to biotic or abiotic stresses, suggesting fine-tuning regulatory roles wang et al., ; ben amor et al., ; xin et al., ) . nat-lncrnas also accumulate under specific environmental conditions such as light exposure, and their expression overlaps with accumulation of histone acetylation marks, suggesting some transcription regulation effect . in plants there are other types of lncrnas. such is the case of the lncrnas involved in: ) the rna-dependent dna methylation silencing pathway (rddm); and ) the lncrnas generated from phasloci which serve as precursors of -nt and -nt secondary phased phasirnas in many plant genomes (zhai et al., ; fei et al., ; zheng et al., ) . however, the function of most plant lncrnas is still unknown (yamada, ; shafiq et al., ; ariel et al., ) . the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. many lncrnas in mammals and yeast originate from specific genomic locations such as the regions around transcription start sites (tsss), enhancer regions, intron splicing sites, and transcription termination sites. most of the studied lncrnas are expressed around the tss, including exosomesensitive yeast cryptic unstable transcripts (cuts) and stable un-annotated transcripts (suts) (xu et al., ) , and upstream antisense rnas (uarnas) (flynn et al., ) , among others. many mammalian non-polyadenylated lncrnas also correspond to divergently transcribed, exosome-sensitive ernas mapped to enhancer regions (andersson et al., ) ; so far plant ernas have not been reported. although most plant lncrnas are transcribed by rna pol ii, there are two plant-specific rna polymerases, pol iv and pol v, that also produce lncrnas (wierzbicki et al., ; li et al., ; yamada, ) . like many yeast and mammalian lncrnas, most plant lncrnas are polyadenylated, although non-polyadenylated lncrnas do exist (heo and sung, ; shin and chekanova, ; kim and sung, ; andersson et al., ) . in fact, hundreds of non-polyadenylated lncrnas induced by specific abiotic stresses were identified in arabidopsis . the expression of lncrna is regulated at transcriptional level and in combination with the pathways involved in their biogenesis, ' end processing and degradation. the main '- ' exoribonuclease complex may regulate lncrna levels since various groups of polyadenylated ncrnas were originally identified in arabidopsis exosome mutants (chekanova et al., ) . rna exosome is an evolutionary conserved cellular rna processing/degradation complex (lange and gagliardi, ). some of these lncrnas originate from the tsss of protein-coding genes, resembling cuts, or they overlap with the ' ends of protein-coding transcripts and extend into the first intron (chekanova et al., ) . gene expression can be regulated by lncrnas (either in cis or in trans) by sequence complementarity or homology with other rnas or dna, and/or by their structure; lncrnas can thus form specific scaffolds or platforms for assembly of specific complexes (chekanova, ) . most lncrnas regulate transcription. in animals this can be achieved by: ) modulation of transcription factor dna-binding activity; ) control of rna pol ii pausing; or ) recruitment of chromatin modellers, which will ultimately affect chromatin topology and nuclear organization (bonasio and shiekhattar, ) . in plants, there are two lncrnas (hid , apolo) that associate with chromatin, promote loop formation, and modulate transcription ), (ariel et al., (figure ). some lncrnas act at the post-transcriptional and translational level (chekanova, ; jabnoune et al., ) . specifically, lncrnas can act as "decoys" or mimics by blocking certain rnas and/or dnas to access their protein regulators (e.g. ips ) (franco-zorrilla et al., ; huang et al., ) . bioinformatics analyses have also identified other putative mirna target mimics in animals that act similarly to plant mirna sponges (chekanova, ) . another plant lncrna (asco) also acts as a decoy of nuclear speckle rna binding proteins leading to different alternative splicing events and consequent altered root development (bardou et al., ) . other lncrnas with biological function are the enhancerrnas (ernas), described in yeast and mammals, but still unknown in plants. these ernas can act in cis as scaffolds to recruit co-activators and thus promote chromosome looping between enhancer and promoter regions; they can interact with other lncrnas to form specific chromosome structures to control gene expression. ernas are regulated by the exosome which can affect either rna synthesis or degradation in these regions (pefanis et al. ) . the exosome can also protect ernas from genomic instability by resolving r loops, which are stable rna-dna triplexes naturally formed during transcription, which can persist in regions that have divergent transcription, eventually becoming deleterious (skourti-stathaki and proudfoot, the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. action of ernas and exosome can control gene expression and nuclear organization in these enhancer regions. interactions between lncrna and chromatin modifiers can be either dependent or independent of srnas. in animals they are srna-independent and occur via the trithorax complex (h k trimethylation) ; or prc (polycomb repressive complex , that regulates h k methylation) (tsai et al., ) . in plants they are -srna-dependent, with the rna-dependent dna methylation pathway (rddm) (matzke and mosher, ) (see below).further details on specific aspects are provided below. the rddm pathway seems to be a plant-specific pathway that relies on lncrnas transcribed from pol iv that will produce nt sirnas. in parallel, lncrnas produced by pol v act as a scaffold that can then recruit the sirna-ago complex by sequence complementarity (matzke and mosher, ) . however, pol v-dependent lncrnas are difficult to identify probably due to their low accumulation rates. nevertheless, the few identified lncrna are non-polyadenylated and can be either tri-phosphorylated or capped at their ' end (wierzbicki et al., ) . in addition, pol v also seems to collaborate with pol ii to promote rddm (zheng et al., ) . another group of regulatory ncrnas that are pol iv -rdr dependent were identified at intergenic regions overlapping mostly with transposons or sequence repeats. these ncrnas are also non-polyadenylated and at their ' end have a monophosphate instead of ' tri-phosphate or cap (zheng et al., ) . the arabidopsis exosome is involved in metabolism or processing of these lncrnas generated by pol iv, v and also some from pol ii (chekanova, ) . also worth mentioning is that the exosome is constituted by different subunits that are functionally diverse and can affect metabolism of smrnas and dna methylation (chekanova, ) . lncrnas and regulation of flowering some of the best characterized plant lncrnas are those involved in regulation of flowering upon "vernalisation" (the need of plants to experience a period of cold -winter-to enable flowering in spring). therefore, transgenic plants expressing any of these lncrnas may have biotechnological value. to assess the future relevance of these lncrnas it is important to understand the molecular mechanisms behind their function, since they are quite different from those of srnas (sirnas and mirnas).the best studied lncrnas in this process are: ) coldair (heo and sung, ) ; ) coldwrap (kim and sung, ); ) coolair (swiezewski et al., ) and asl (shin and chekanova, ) . all these lncrnas modulate the flclocus, which encodes a transcription factor able to repress flowering (berry and dean, ) . interestingly, these four lncrnas originate from different regions within the flclocus (promoter, first intron, and ' end antisense) suggesting a function in cis. they all contribute to prevent flc transcription by recruiting the prc silencing complex and the accumulation of repressive chromatin marks (crevillén and dean, ; csorba et al., ; rosa et al., ; sun et al., sza; swiezewski et al., ; liu, f et al., ) . of interest is that certain aspects of this regulation are similar to that of the mammalian lncrnas hotair and xist. it has been proposed that some of these lncrnas could directly bind to prc components, but findings showing that mammalian prc can bind very strongly to unrelated rnas (davidovich et al., ) question the need for specific lncrnas in this recruitment step. the study of components of the arabidopsis exosome (rrp l and rrp l ) revealed their role in coolair and asl expression or processing, although this seemed to occur independently of the exosome core complex. both rrp ls could process the ' end of coolair and promote asl accumulation, similarly to xist in humans (shin and chekanova, ; ciaudo et al., the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. regulate the levels of different lncrnas and probably act in different mechanisms to silence flc (chekanova, ) . lncrnas and r-loop formation coolair transcript levels are decreased due to r-loop formation at its promoter (sun et al., b) , which could promote exosome recruitment through the ' end processing pathway (chekanova, ) . this function is also seen in mammals where the exosome also resolves/degrades deleterious r loops (pefanis et al.) . surprisingly, in mutants affected in r loop formation, coolair may accumulate together with flc, suggesting that this regulation is still not fully understood (chekanova, ) . lncrnas and nuclear architecture animal lncrnas are involved in tethering rna, dna and proteins, and thus affect nuclear d structure (engreitz et al., ; quinodoz and guttman, ; hacisuleyman et al., ) . in plants, extensive evidence supports a similar role, either within the rddm pathway (moissiard et al., ) , in the regulation of flowering time (hepworth and dean, ) or in auxin signalling and root development (ariel et al., ; ariel et al., ) . plant and animal lncrnas can be intergenic, intronic or natural antisense transcripts, depending on their location in the genome. although only a few lncrnas have been characterized in detail, they are known to have different modes of action, ranging from chromatin modifications including prc recruitment, to promotion of translation, mirna target mimicry, hijacking splicing factors and formation of chromatin loops. adapted from (ariel et al., ) . not all processes occur simultaneously. different types of lncrnas are shown schematically. circrnas earlier studies of rnas structure proved that viroids (existing as uncoated rna molecules and are known to infect plants) have circular rna (circrna) molecules. the nature of covalently closed circular rna molecules was determined due to: i) the inability to phosphorylate at the '-terminus; ii) resistance to metaperiodate oxidation or borohydride reduction of the '-terminal ribose; or iii) resistance to venom phosphodiesterase degradation (sanger et al., ) the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. al., ), plants (daros and flores, ) , and mammals (nigro et al., ) including humans memczak et al., ) , and across the eukaryotic spectrum. circular rnas' lack of coding potential was identified early on by the inability of circular mrna (kozak, ) or rna (konarska et al., ) to assemble/adhere to eukaryotic ribosomes. circular rnas are also unable to be translated in plants extracts . as with other ncrnas, circrnas in plants exhibit tissue-specific expression, have a considerable number of isoforms, alternative backsplicing (canonical and noncanonical) and alternative circularization patterns (lu et al., ; sun et al., ; wang et al., ; darbani et al., ) . the biogenesis of circrnas is a conserved feature in animal and plant cells. transcribed by rna polymerase ii and backsplicing reactions of pre-messenger rnas, they occur in the spliceosome and require a repeated sequence and rna-binding proteins. spliceosome formation is initiated by sequential assembly of small nuclear ribonucleoproteins onto a specific region of the pre-mrna downstream of the ' donor splice site (dinucleotide gt) and upstream of the ' acceptor site (dnucleotide ag). thus, exonic or intronic circrnas are generated depending on the initial small nuclear ribonucleoprotein binding site (lee et al., ; sun et al., ) . although their mechanisms of action in plants are still unclear, some studies in rice suggest that they do not imply any significant enrichment (as occurring in humans) for mirna target sites, and that circular rna and its linear form may act as a negative regulator of its parental gene (lu et al., ) . other studies have shown that fluctuations in circrnas do not correlate with the levels of their parentalloci encoded linear transcripts (darbani et al., ) . some circrnas have also been shown to contain putative mirna binding sites lu et al., ) and have been identified as mirna sponges (zuo et al., ) . by binding to mirnas, and consequently repress their function, circrnas act as mirna sponge to regulate the response to stress . recent data suggest that circrnas in plants may affect plant response to abiotic and biotic stress. for example, circrnas may be involved in chilling injury in tomato (zuo et al., ) , or may respond to imbalances in iron and zinc (darbani et al., ) . in another study, tan and colleagues (tan et al., ) showed that overexpression of a tomato circrna derived from phytoene synthase (psy ), reduced psy mrna abundance, and lycopene and β-carotene accumulation. this was likely due to the continuous highly expressed circrna and/or the low abundance of linear rna from the overexpression vector. similar results were reported for another circrna derived from the phytoene desaturase gene (tan et al., ) . their role in developmental processes has also been established (cheng et al., ) , including their possible role in different aspects of development and senescence in arabidopsis cheng et al., ) . other biological processes such as photosynthesis (dou et al., ) or mitochondrion function (darbani et al., ) have also been proposed as involving circrnas. circrnas are a popular topic in animal research because of their potential as post-transcriptional regulators (memczak et al., ; piwecka et al., ; guarnerio et al., ; hansen et al., ; ashwal-fluss et al., ) , their recognized function as mirna sponges, as sponges for rna-binding proteins or their competition with linear splicing; and their role as diagnostic markers (zhao et al., a; zhao et al., b) . indeed, circrnas have also been found in biological fluids including saliva (bahn et al., ) , seminal fluid (dong et al., ) and plasma . research in plants is just emerging and functional studies are still lacking. it is unknown if this type of novel ncrnas could resist gastrointestinal tract conditions, due to their expected high stability, and have a biological effect on an organism. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. ncrnas can regulate many biological processes through their impact on gene expression by acting on chromatin structure and affecting transcription, rna processing, rna stability and translation. among these, srnas are produced by cleavage of dsrnas intermediates, either from hairpin precursors (mirnas) or from the synthesis of dsrnas by rdrs (sirnas); most lncrnas regulate gene expression without being processed. despite size and biogenesis differences, most ncrnas share sequence-specific inhibitory functions. important differences are present between plants and other organisms in srnadirected silencing pathways, which are highly diversified and specialized in plants through the expansion of rdr, dcl and ago proteins that mediate srnas biogenesis and function. the vast majority of small regulatory rnas in humans and animals are mirnas. in contrast to plants, no evidence for systemic spreading of rna silencing in humans and vertebrates have been found. although very few plant lncrnas have been studied in detail, they seem to share with their animals' counterparts the ability to recruit chromatin modifiers and thus regulate gene expression. in the same context, very few studies are available on plant circrnas. their functional characterization will bring novel insights into their possible role as a novel class of noncoding regulators. when considering the potential presence of plant ncrnas in food or feed it is important to determine the mobility potential of these molecules inside the source plant (figure ). there are several comprehensive reviews (chitwood and timmermans, ; dunoyer et al., ) addressing sirna/mirna movement in detail. it has been hypothesiezed that srnas may move throughout the plant. although the initial model would suggest non-cell autonomous (one whose action extends beyong the cell producing the signal) rna silencing by srnas, other intermediates could also account for this signal: dsrnas produced by rdr activity (plant specific), or fold back rna acting as silencing trigger (dunoyer et al., ; dunoyer et al., ; himber et al., and smith et al., ) . these intermediates could also diffuse from one cell to another and be processed again by dcl , which has been associated with nt sirna movement (dunoyer et al., ) . in detail, secondary sirnas processed from dsrna by dcl could move cell-tocell to propagate silencing by signal amplification. it has been shown that dcl expression from phloem companion cells is critical to the short-term spread of the silencing effect of a sirna duplex. this was reported in a study in which the viral suppressor p was found to be expressed in phloem companion cells and able to bind to nt small dsrna (vargason et al., ) . dcl -dependent sirna movement has also been shown to occur within certain organs, such as leaves, to create gradients (chitwood et al., ; levine et al., ) . other dcls-dependent sirnas are involved in sirna movement and these can also be sorted into different risc complexes (montgomery et al., ; mi et al., ) . other examples of mobile sirnas could be the ta-sirnas and dcl -dependent nt sirnas, also acting through ago . dcl processing of long dsrna substrates generated by pol iv (a plant-specific rna polymerase) and rdr (another plant-specific protein) leads to production of nt sirnas that are incorporated into ago and will transcriptionally silence transposons (chapman and carrington, ; mi et al., ) . in fact, sirnas derived from transposons or methylated regions in the genome have been shown to be associated with chromatin silencing. these -nt mobile sirnas could then promote epigenetic changes transmissible to following generations. when transgenes are expressed in plants, the generated dsrnas can be processed by dcl and dcl . another dcl, dcl , processes dsrnas into nt sirnas which can induce gene silencing of viral origin or from transgene expression (deleris et al., ) . sirnas may also function as mobile signals able to promote epigenetic modifications. grafting experiments with roots from dcl dcl dcl mutants and shoots expressing gfp sirnas could the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. indicate that sirnas to nt long move throughout the plant, but still unclear if in free or proteinbound form (i.e. bound to ago or other proteins) (molnar et al., ) . within the plant, sirnas are known to move in a source-to-sink direction and into growing meristems (molnar et al., ; palauqui et al., ; schwab et al., ) . their accumulation in reproductive cells could promote epigenetic changes in subsequent generations. in fact, slotkin et al. (slotkin et al., ) showed that dcl dependent sirnas generated by transposon activation in the pollen vegetative nucleus can silence transposons in pollen sperm cells. these cells are then able to transmit genetic material to the following generation. a similar effect may occur if maternally-derived sirnas in the endosperm moved to the embryo and influence genome integrity or even lead to the creation of new epialleles (molnar et al., ) . similar findings of sirnas silencing transposable elements in the animal germ line have also been reported in an animal cell line and tetrahymena (malone and hannon, ). considering that plant development relies more on positional effects than cell lineage, the role of mirnas in the control of several developmental processes has to be strictly regulated at the mobility level (chitwood et al., ; levine et al., ) . for example, as small rnas diffuse into regions of low mrna (targets) expression, it eliminates target molecules therein, but cannot affect regions of high mrna levels (levine et al., ) . as mentioned previously, dcl processes imperfect hairpins into nt mirnas, with a ' u nucleotide incorporated into ago , and cleavage the mrna targets. ago can also incorporate these mirnas (brodersen et al., ) . while sirna movement is better understood, this is not the case for mirna movement. mirnas can be isolated from phloem sap (mir , mir , mir ) but it is unclear if they can leave the phloem (buhtz et al., ; pant et al., ; yoo et al., ) . some reports support both endogenous mirna and artificial mirna movement (chitwood et al., ; schwab et al., ) . it has been shown that mirnas can move from below the shoot apical meristem into the meristematic layers (chitwood et al., ) , and, similarly to sirna movement, from the phloem towards the root meristems (molnar et al., ) . although initial evidence suggested sirna mobility, with only dcl responsible for this, it is now accepted that almost every known rnai pathway in plants has non-cell autonomous activity. movement has been shown for srnas ) of viral origin; ) induced by transgene expression; ) ta-sirnas; ) mirnas and ) repeat-associated sirnas (dunoyer et al., ; carlsbecker et al., ; chitwood et al., ; molnar et al., ; slotkin et al., ) . however, it is still unclear if srnas can move in a free state or associated with specific proteins, and in a single or double strand form. whereas some reports suggest that the srna molecule movement happens in a duplex form, the strand bias found in the srnas present in dcl dcl dcl grafted roots suggests that single strand sirnas can also diffuse (molnar et al., ) . unlike in animals, in which sirnas can move from cell-to-cell through secretory vesicles (ruvkun, ; valadi et al., ) , in plants the size of small rna would allow cellto-cell movement through plasmodesmata channels connecting different plant cells (kobayashi and zambryski, ) . this movement seems to occur in a source-to-sink direction, and could be regulated in some tissues through the formation of sirnas-ago complexes (molnar et al., ; schwach et al., ) . www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. plant small rnas (srnas) can move either cell-to-cell or long distance. cell-to-cell movement occurs through plasmodesmata (green arrows) that allow spreading of srnas from the cell where they are generated to - neighbouring cells (a). this cellto-cell movement can be extended over the surrounding cells by signal amplification (b). in this case, srna targets will be converted into new dsrnas by the combined action of dcls and rdrs. several grafting experiments have demonstrated that - nt sirnas can move over long distances in the plant (c). whether they move as ssrna, dsrna or associated with proteins is still unclear. adapted from (dunoyer, p. et al., ) , (melnyk et al., ) and (molnar et al., ) . in terms of mirna mobility, it is still unclear if all mirnas, or only a subset, can diffuse. different mobility could be associated with different tissues/structures, and/or the size and stability of the passenger strand mirna (a.k.a. mirna*, star strand). in addition, mirna movement regulation could involve subcellular compartmentalization (by nuclear export proteins such as hasty (park et al., ) ) and sequestration via ago proteins (e.g. mir / can be sequestered by ago and affect shoot apical meristem differentiation (liu et al., ; tucker et al., ) ). most of the information is available for srnas since they represent the most studied class of ncrna to date. in addition to the role of srna movement when considering ingestion of transgenic plants expressing altered levels of srnas, it is also important to address the issue of their biogenesis and turnover to assess their temporal availability when ingested. this is especially important since transgenic plants expressing rna interference constructs will accumulate the different srna intermediary forms (pri-mirnas, pre-mirnas, mature mirnas, drnas, sirnas), which may also be processed outside the plant when ingested. several reviews published in high-impact journals (rogers and rogers and chen, ; or scientific reports the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. view of the processes of mirna maturation and degradation in plants. it is important to consider the homology between the mirna biogenesis pathways in plants and animals as well as mirna stability. this knowledge allows understanding ) whether any aspect of animal metabolism or other biological processes could be modulated by plant mirnas; and ) if mirnas can survive for long periods after ingestion. as typical pol ii transcripts, pri-mirnas are stabilized by addition of a ' -methylguanosine cap (xie et al., ) and a ' polyadenylated tail (jones-rhoades and bartel, ; zhang et al., ) . mirna genes are also subjected to transcription factor regulation and can be alternatively spliced. the size of these transcripts varies and their processing normally occurs in a base-to-loop direction (rogers and . in plants, there are multiple dcl endonucleases that possess dexd/h-box rna helicase, duf , paz, tandem rnaseiii, and dsrna-binding domains. dcls cleave the dsrna precursor generating a -nucleotide ' overhang (margis et al., ) . processing of the pri-mirna requires at least two catalytic cycles to free the mirna:mirna* duplex. in animals, this requires sequential cleavage of the pri-mirna in the nucleus first and then in the cytoplasm by different rnase iii enzymes. the biogenesis of most mirnas in arabidopsis requires dcl (park et al., ; reinhart et al., ) , although other dcls may be involved, but are not essential (xie et al., ; gasciolli et al., ) . in contrast to animals, all plant mirna processing steps occur in the nucleus (papp et al., ) . dcl activity will result in mature mirnas of - nucleotides (nt), although variations in size can occur (rogers and chen, ) . confirming its fundamental biological relevance, dcl mutants are lethal (golden et al., ) . other dcl proteins that function in different srna biogenesis pathways might also have minor roles in mirna biogenesis. for instance, several mirna genes are partially processed into - -nucleotide mature mirnas species whose accumulation depends on dcl (vazquez et al., ) . in the absence of dcl , mature -nucleotide mirnas can accumulate due to dcl activity. in contrast, processing of mir , mir , and mir depends primarily on dcl , probably as a consequence of highly complementary fold-backs in these pri-mirnas (ben amor et al., ; rajagopalan et al., ) . dicer cleavage of animal pri-mirnas is facilitated by the action of dsrnabinding domain (dsrbd) proteins (drbs) (jiang et al., ; parker et al., ) . in arabidopsis, there are five drbs that have been shown to bind dsrna in vitro (hiraguri et al., ) . the best studied member of this family is hyl (hyponastic leaves ) which is involved in pri-mirna processing, but also in pri-mirna intron splicing (laubinger et al., ; szarzynska et al., ) . hyl seems to increase the accuracy of dcl processing of most, but not all, pri-mirnas, suggesting that other mechanisms may also be involved (liu, c et al., ) . from the same family, drb and drb are also involved in pri-mirna processing, by associating with dcl (rogers and . another protein involved in mirna biogenesis is serrate (se), which can interact with hyl and dcl (lobbes et al., ; machida et al., ) . like hyl , se also seems to increase dcl pri-mirna cleavage efficiency (dong et al., ) . of interest is that these two proteins (hyl and se) can be regulated by phosphorylation. although hyl phosphorylation can decrease its activity (manavella et al., ) , the biological relevance of this modification in se is not known (rogers and . another regulator, dawdle (ddl), a phosphothreonine-binding forkhead associated (fha) domain protein, seems to stabilize pri-mirnas, possibly by direct binding (yu et al., ) . like se and hyl , ddl can also interact with dcl , but does not seem to be involved in pri-mirna processing. ddl most likely facilitates dcl access or recognition of pri-mirnas. in addition, ddl can recruit phosphorylated se or hyl to the pri-mirna. the human homologue of ddl, snip , seems, among other functions, to be involved in mirna biogenesis and interacts with drosha (yu et al., ) . ddl could therefore be an evolutionarily www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. conserved factor in mirna biogenesis. other examples of conserved srna biogenesis machinery include ago, dicer, hen and exportin (yu et al., ) . two components form the cap structure of rna pol ii transcripts in plants: cap-binding proteins (cbps) and . there is substantial overlap among cbp , abh /cbp , and se mutants in terms of: ) the specific pri-mirnas affected; ) the specific mrna splicing defects observed, and ) a bias for accumulation of first introns (laubinger et al., ) . this connection between pri-mirna processing and pre-mrna splicing seems to converge at se and cbps. in fact, splicing defects are also observed in cbp and abh /cbp in plants. in animals, the cap and its associated cap-binding complex are essential for the correct splicing of the first intron (lewis et al., ) . the connection between pri-mirna processing and splicing also converges in tough (tgh), a g-patch domain rna-binding protein. the tgh mutant accumulates pri-mirnas in vivo and has been shown to affect processing of pri-mirnas into -nucleotide mature mirnas (ren et al., ) . tgh may have either a structural or a regulatory role in pri-mirna processing. tgh binds both pri-and pre-mirnas in vivo, but this probably occurs via association with the loop structure since tgh binds ssrna but not dsrna in vitro (ren et al., ) . tgh paralogs have been described in metazoans (calderon-villalobos et al., ) , but their roles in mirna biogenesis are still unknown. a human tgh paralog is reported in spliceosomal preparations (jurica et al., ) . a conserved connection may exist between tgh and splicing and possibly pri-mirna processing (rogers and chen, ). mirna processing seems to occur in specific subnuclear regions. hyl , se and dcl can make pairwise interactions that occur in certain subnuclear particles (fang and spector, ) . tgh can also interact with dcl , hyl , and se in subnuclear foci (ren et al., ) . these foci seem to be zones of functional pri-mirna processing (fang and spector, ; fujioka et al., ; manavella et al., ) . some of these components (dcl , hyl , se, tgh) also co-localize with components of the splicing machinery, further strengthening the connection between the two processes (calderon-villalobos et al., ; fang and spector, ; fujioka et al., ) . processed mirna:mirna* duplex will leave the nucleus to carry out its function in the cytoplasm. several components have been implicated in this transport and function and are described below. nucleo-cytoplasmic transport hasty (hst) is a member of the importin-ß family of arabidopsis that accumulates in the nuclear periphery (bollman et al., ) . hst is a paralog of human exportin- involved in pre-mirna transport. hst can participate in dcl -dependent mirna:mirna* duplex export (zeng and cullen, ) . but this is probably not the sole mechanism for mirna nucleocytoplasmic transport, since passive diffusion via the nuclear pore may also occur (jacob et al., ; park et al., ) . ema /sad is another importin-ß family protein involved in mirna function. ema is an orthologue of human importin- which facilitates nuclear import of ago and reduces its association with target mrnas (weinmann et al., ) . it is still unclear if ema ) sequesters mirnas; ) is involved in transport of other risc loading factors or; ) affects loading of certain mirnas into risc (rogers and . the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. after leaving the nucleus the mirna:mirna* duplex is loaded into ago, a risc component. the arabidopsis ago family consists of members that associate with small rnas possessing specific ' nucleotides (mi et al., ; takeda et al., ) . in plants, the slicer activity of agos has been shown for ago , ago , ago , ago , and ago (rogers and . ago seems to have a predominant role since it preferably associates with 'u small rnas (most common ' nucleotide in mirnas) (mi et al., ; takeda et al., ; vaucheret et al., ) . however, other agos can also associate with specific mirnas depending on mirna length or even their accumulation in different tissues (ebhardt et al., ; rogers, kestrel and chen, xuemei, ) , suggesting a certain flexibility in this association. in plants, mirna guide strands generally accumulate in higher levels than mirna* strands, although the exact mechanism for strand selection is still not fully understood (rogers and chen, ). hyl (drb) seems to be involved in this process (eamens et al., ; manavella et al., ) . in animals, loading of sirnas into risc requires the cytosolic drb protein loq, but loading of mirnas into risc requires dicer rather than loqs (liu et al., ; liu, x et al., ) , suggesting different mechanisms in plants and animals. hyl is predominantly nuclear, therefore its association with the mirna:mirna* duplex suggests that ) hyl may be transported to the cytoplasm together with the processed mirna duplex; or ) loading may also occur in the nucleus. in addition, other risc loading mechanisms can exist (eamens et al., ). in animals, sirna passenger strand unwinding requires ago slicer activity; mirnas duplexes are unwound by a slicing-independent mechanism (matranga et al., ) . in plants, ago slicer activity removes sirna passenger strand but this function is not required for mirna passenger strand removal (carbonell et al., ; iki et al., ) . as occurs in animals, plant mirnas use an alternative slicerindependent unwinding mechanism. ago loading seems to require heat shock protein (hsp ) too (iki et al., ) . in addition, via hsp , ago is able to interact with squint (sqn) and the phosphatase pp (iki et al., ) . these interactions seem to regulate ago loading and consequently risc activity. ago phosphorylation has been detected in arabidopsis (de la fuente van bentem et al., ) in manner similar to human ago . apparently, phosphorylated ago exhibits reduced srna binding (rüdel et al., ) . phosphoregulation by pp or other proteins may modulate arabidopsis ago loading capacity. unlike in animals, in which mirna biogenesis occurs in the nucleus and cytoplasm, the mirnas in plants are generated in the nucleus in a mostly dcl- -dependent manner. studies confirm that several of the major components are evolutionary conserved, such as the dicer, ago, hen and exportin paralogs involved in nucleocytoplasmic transport of the mirna:mirna* duplex. however, shared pathway components among the different species do not reflect totally identical processes, such as, loading of mirnas into risc which occurs differently in plants and animals. one interesting common aspect remains the pri-mirna processing and pre-mrna splicing which seems to occur both in plants and animals. www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. following a systematic literature search as described in section . . . and following the methodology described in section . . . ., a total of publications were selected as relevant to the stability and turn-over of non-coding rnas. of these, only four analyse ncrnas stability at a genome-wide scale, and one (enuka et al., ) simultaneously addresses the stability of several circrnas. using these studies, the stability of small ncrnas, lncrnas and circrnas can be compared. while earlier studies report the degradation of plant nucleic acids when exposed to hard conditions (i.e. cooking), six recent studies suggest that some ncrnas (i.e. mirnas) from plants can resist these conditions. turnover of ncrna molecules in plants and mammals depends both on their stability and degradation rate and it is described that chemical modifications can increase stability of different types of srnas (e.g. sirnas, mirnas). in mammalian cells, artificially introduced -o-methyl groups can stabilize sirnas without affecting their rna interference activity (czauderna et al., ) . considerable work has been done to address the issue of mirna modifications and how these affect their stability. among these, '-o-methylation has been identified as relevant. plant mirnas have a naturally occurring methyl group on the ' nucleotide ribose ( figure ). in this case, methylation does not require guide rnas, since hen (hua enhancer ) can methylate mirna:mirna* duplexes. this hen -dependent '-omethylation on the ' terminal ribose is a mg + dependent methylation mechanism that will ultimately stabilize mirnas (abe et al., ; molnar et al., ; yu et al., ; yang et al., ) .the hen mg +-dependent methylation mechanism relies on its two dsrbds binding the substrate duplex, and the la motif-containing domain interacting with the ' end of the substrate strand (huang et al., ). this methylation step occurs after dcl processing creates the mirna:mirna* duplex but before duplex www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. unwinding and selection of the mirna guide strand. however, it is still not clear whether hen can methylate free mirnas or can bind hyl , dcl and ago , although it has been shown to interact with hyl and dcl . in animals, the hen homolog has a different structure since it does not possess the dsrbd and la motif-containing domains. therefore, the animal hen homolog methylates singlestranded srnas associated with ago or piwi proteins (saito et al., ; horwich et al., ) . the degradation of ncrna molecules in plants and mammals is mediated by ' nucleotidyl transferases and exoribonucleases. the arabidopsis hen suppressor (heso ) belongs to the dna polymerase β gene family (zhao et al., ; ren et al., ) . in other organisms, heso putative homologs are ribonucleotidyl transferases that are able to add specific nucleotides to the ' end of different rnas (martin and keller, ) . arabidopsis heso can also add nontemplated nucleotides to the ' end of unmethylated mirnas in vitro and exhibits a preference for uridine (ren et al., ; zhao et al., ) . analysis of hen and heso mutants indicates that this ' oligo uridylation triggers degradation of mirnas (ren et al., ; zhao et al., ) . the arabidopsis family of small rna degrading nucleases (sdns) of '- ' exonucleases has similarity to the yeast rex exonucleases. sdn has '- ' exonuclease activity versus short ssrnas but is not active against longer or double-stranded substrates (ramachandran and chen, ) . in plants, ' truncated mirnas can be modified by the addition of ' oligonucleotide tails (see above) (ibrahim, fadia et al., ; li, j et al., ; lu et al., ). in addition, these ' truncated mirnas also seem to associate with ago . this interaction delays their degradation and allow addition of the ' oligonucleotide tail leading to their degradation at a later stage (zhao et al., ) . whereas '-o-methylation may protect against ' oligouridylation by heso (see above) and consequent targeting to degradation, this modification would not protect against sdn . it is possible that sdn can promote mirna ' truncation and then facilitate subsequent ' polyuridylation by heso . however, after ' polyuridylation sdn would be unable to degrade these tailed mirnas, suggesting the existence of other nucleases (ramachandran and chen, ) .for instance, the addition of ' polyuridylated tails in mirnas seems to attract exosome-mediated degradation in algae (ibrahim, f. et al., ) , but in arabidopsis this regulation has not been addressed in detail (rogers and rogers, kestrel and chen, xuemei, ) . another possible mechanism would include loss of ago association and protection of mirnas with long ' tails due to heso activity. in addition, ' polyuridylation may act as a nucleases recruitment platform, thus promoting mirna degradation. nevertheless, ' polyuridylation and nuclease processing seem to be interconnected events that will ultimately result in degradation of the tagged mirnas. the overall turn-over of rnas is mainly described using half-life studies in both plants and animals. most of the available reports assessing ncrna half-life focus on the role of ' truncation, ' uridylation and consequent degradation (see above) as the main processes regulating mirna turnover in plants. however, a recent report identifies the enzymes (sdn and sdn exonucleases) responsible for ' truncation in vivo and its relationship with ' tailing (uridylation). also of importance, this report shows the opposite role of ago and ago in mir / degradation by sdns, specifically sdn . ago promotes mir degradation by sdn , an unexpected role for a plant ago which has been associated with a protecting role in mirna stability (yu et al., ) . this function of ago (enhancing mir degradation) keeps mir out of the stem cell niche in the shoot apical meristem (where ago is not expressed). accumulation of ectopic mir in the shoot apical meristem fails to maintain the stem cell population. this kind of mechanism is thus needed to clear mir since this particular mirna can move between cell layers (see mirna movement section, above). another report evaluates the activity of nucleotidyl transferases in ' uridylation of mirnas and in vitro assays indicated a fast enzymatic process (tu et al., ) . also, xrn - nucleases that affect mir homeostasis in c. elegans do not www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. seem to process mature plant mirs but rather the by-products of pri-mirna processing (ji, lijuan and chen, xuemei, ) . in summary, there are no specific studies on mirna half-life in plants. the reports mentioned here evaluate mirnas levels at steady state in different mutants, for example, when either the exonucleases and/or nucleotidyl transferases involved in mirna turnover are deleted. the half-life of mirnas in mammals has been addressed in studies of dicer ablation in mouse embryonic fibroblasts showing that mirnas are highly stable inside the cell. indeed, turnover assays revealed that the average mirna half-life is h (i.e. ≈ days) (figure ). although some mirnas have a shorter half-life, these data generally indicate that mirnas are much more stable (≈ x more) than messenger rnas. (gantier et al., ) . several features have been described that might explain this high half-life for mirnas. for instance, partial pairing of mirna and the mrna target site generally produces translational repression, while extensive pairing of mirna and mrna produce a mrna cleavage (pasquinelli, ) . in the former case the mirna remains stable, but in the latter it degrades. further studies in mammalian cells have also shown that argonaute proteins stabilize mature mirnas in a slicing-independent manner, increasing mature mirna stability (winter and diederichs, ). for specific mirnas, it has been found that certain rna-binding proteins (i.e. roquin) can enhance mature mirna- stability by reducing its ' end uridylation (srivastava et al., ) . while intracellular levels of mirnas are mostly affected by cell division (ghosh et al., ; gantier et al., ) , mirna activity and turnover can also be controlled by subcellular distribution of microribonucleoproteins; that is, polysome sequestration contributes to an increase of cellular mirna levels, but without an increase in mirna activity (ghosh et al., ) . plant small rna duplexes can be '-o-methylated at their ' terminal ribose by hen , loaded into ago and protected from uridylation and degradation by '- ' exonucleases. ago -bound unprotected small rnas, however, are uridylated by heso and degraded by sdn . adapted from (borges and martienssen, ) and (ji and chen, ) . the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. widespread variation of mirna decay in slow-dividing primary macrophages. bone marrow derived macrophages from when analysing lncrnas stability in mammals at a genome-wide scale, lncrnas half-life generally varies over a wide time range, comparable to, although on average less than, that of mrnas (clark et al., ) . in a study of all transcripts, including mammals, it was found that the range of half-lifes for ncrnas (all types) was similar to that of the mrna half-life (tani et al., ) . indeed, a large number of ncrnas transcripts have a half-life of less than h (tani et al., ) , which agrees with the < h half-life for hundreds of unstable lncrnas (clark et al., ) . however, although there are lncrnas with extreme stability (half-life > h) they are still much less stable than mirnas (gantier et al., ) (see above), but more stable than protein-coding rnas (wang, l et al., ) . circrnas have a covalently closed loop structure with neither '- ' end nor a polyadenylated tail, which confers high resistance to rna exonuclease or rnase r treatment when compared to that of their linear sequence counterparts (memczak et al., ) . indeed, earlier studies of the structure of rnas from viroids, which were found to be circular, suggested that the nature of covalently closed circular rna molecules confers certain properties including resistance to metaperiodate oxidation or borohydride reduction of the '-terminal ribose; the inability to phosphorylate at the '-terminus; or resistance to venom phosphodiesterase degradation (sanger et al., ) . in mammals, some studies have evaluated the stability of circrnas compared to that of their linear isoform derived from the same host gene. for example, circrnas were found to be less abundant and dynamic than their counterparts (enuka et al., ) . calculating the half-life of circrnas and their linear counterparts, enuka et al., showed that the media half-life of circrnas of mammary cells ( . - . h) was at least . times longer than the median half-life of their linear counterparts ( . - . h). this suggests that cirrnas are generally more stable and static compared to linear species (enuka et al., ) (figure frame a). in a stability study on some circrnas it was found that while the associated linear transcripts exhibited a half-life of < h, the circular rna isoforms were highly stable, with transcript half-life exceeding h (jeck et al., ) . in plants, covalently circularized exogenous rna incubated with wheat embryo cell extract in vitro have shown that the circular rnas exhibited considerable stability compared to the linear version (makino et the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. al. , ) . while linear rna degraded within the first few minutes, its circular counterpart was stable for more than min in vitro (figure frame b). by contrast, circularization dramatically reduced the translation capability of the rna, which was restored to some extent after linearization. plant srnas are naturally methylated at their ' nucleotide ribose. this modification depends on hen and protects srna from degradation. unmethylated or truncated srnas can be uridylated by heso and thus be targeted for degradation by '- ' exonucleases of the sdn family. although ago binding to mirnas was believed to protect them from degradation, recent reports suggest that ago in particular may promote mirna degradation by sdns. differently from the plant situation, animal srnas are not methylated. a) mcf a cells were metabolically labelled using su, for or h. the rna was then extracted, biotinylated and purified on streptavidin magnetic beads. flow-through rna was also collected. next, rna was reverse transcribed and quantified using highthroughput real time pcr (fluidigm). the half-lives of circrnas and their corresponding linear counterparts are shown. halflife values were calculated from two samples, labelled with su for or h and then averaged. all data were corrected for any bias introduced due to low uridine (short length) of rna species (see 'materials and methods' section). the circrnas (red dots) and their linear counterparts (blue dots) were sorted from high to low according to the difference between their half-lifes. error bars represent standard errors. source: from (enuka et al., ) . b) stability of circular rnas in the wheat germ cell-free translation system. a denaturing polyacrylamide gel separating the synthesized circular and linear rnas with a (gaaa) sequence after incubation with the wheat embryo extract for the indicated time periods. untreated rna is in lane n. positions of circular, linear, and ribosomal rnas are indicated as c, l, and rrna, respectively, on the right. source: from . compelling evidence supports the significant contribution of srnas to communication between hosts and some eukaryotic pathogens, pests, parasites, or symbiotic microorganisms. (baulcombe, ; (ghag et al., ; koch et al., ; vega-arreguín et al., ; helber et al., ) . srnas or dsrnas, efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. for example, can be transferred from plant to pests such as insects that eat leaves or nematodes that infect roots. in fact, transgenic plants can be created that express dsrna homologous to essential genes of insect pests or nematodes and thus control them (baum et al., ; fairbairn et al., ; mao et al., ) . thus, this silencing transfer mechanism is very relevant for the food and feed risk assessment of ncrna gmo plants, highlighting the need to evaluate the stability of ncrnas outside the plant. earlier studies suggested that food nucleic acid content was hydrolysed when cooked (colling and wolfram, ) . most of ncrnas, including srnas and lncrnas have been discovered in the last years, and thus new data has been added to the body of knowledge. in a study of srna stability under cooking conditions, plant mirnas were detected after cooking although at significantly lower levels when compared to fresh plant tissues (see table ) (zhang, lin et al., ) . differences were observed for mirnas from different food/feed sources (rice, cabbage, wheat or potato), ranging from nearly undetectable values to almost % persistance of mirnas after cooking. levels of mirnas were monitored with a stem-loop quantitative reverse transcription polymerase chain reaction (qrt-pcr) assay using the u snrna for normalization, a commonly used housekeeping gene in mirnas analyses. the same study found that most of the plant mirnas and mammalian mirnas could survive under acidic conditions that mimic the gastric acidic environment. of note is that the degradation rate of mammalian mirnas under acidic conditions was similar to that of their synthetic form (without '-omethylated ' ends), whereas plant mirnas had a much slower degradation rate compared with their synthetic form (without '-o-methylated ' ends), suggesting that methylation had a protective effect on the stability of plant mirnas (table ) . another study reported that endogenous mirnas can be detected by qpcr analysis in rice and soybean plant materials after storage, processing and cooking (philip et al., ) . for processing and cooking, soybeans were soaked in rnase-free water with . % w/v nahco overnight at ºc, then separated from the soaking liquid, rinsed in fresh rnase-free water, and then boiled in rnase-free water for www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. min until they became soft in texture. by constrast, a synthetic mirna showed a significantly higher susceptibility to simulated food processing conditions as compared to plant mirnas, as well as high molecular weight rnas (total rna). the synthetic mirna used for comparisons was the cel-lin- , which is a caenorhabditis elegans mirna. the synthetic mirna was synthesized without '-o-methylated ' ends (philip et al., ) . these results suggest that the methylation and the small size of plant mirnas make them more resistant to degradation than synthetic (not chemically modified) mirnas. in addition, the study showed significant plant mirna stability in a simulated digestion system for min prior to absorption or transport into the blood stream. however, this study provided most of the quantitative data from q-pcr analysis without normalization, and data should be considered as relative compared to the treated control in the experiments. studying the stability of plant srna in vitro, liang and colleagues exposed total rna ( µg), isolated from brassica oleracea, to freshly drawn serum from mice. in serum, a large amount of srnas survived after h of incubation at ºc, but after h only about % of the rna could be detected. the authors also exposed the plant rna molecules to fecal suspensions at ºc. after h incubation significant amounts of srnas were still detectable, but after h rna molecules were undetectable . suggesting that srnas were more resistant to degradation in the presence of serum suspension than in fecal suspensions. these results were produced by gel electrophoresis of srnas, which is known to be less sensitive than qrt-pcr or rna sequencing methods. this study also reports that, when orally ingested, plant mir (chosen because it is the most highly enriched plant mirna from b. oleracea) can pass intact through the gastrointestinal tract in mice (a maximum of . % recovered from the stomach of some individuals), and can be detected in the blood, liver, spleen and kidney. levels of mir were monitored by qrt-pcr using a taqman probe, which can distinguish differences of one nucleotide. since only one mirna was evaluated, the above findings cannot be generalized to other mirnas. other studies have reported mirnas to be stable in watermelon and mixed fruit juices produced by simple extraction without additives . plant mirnas were detected by stem-loop qrt-pcr using taqman probes and some were validated by northern-blot analysis. to reduce the nonspecificity signal of qrt-pcr, no-template controls were used. yang and colleagues studied artificial in vitro digestion systems that simulate mammalian gastric and intestinal conditions, and found that mir was between - fold more stable than other mirnas . this effect was observed for cabbage extracts when compared to either the 'o-methylated or the non-modified form. detection was also done by qrt-pcr. the same research group reported recently that several plant mirnas, including mir , were degraded during storage at ºc in cabbage extracts obtained by mechanical maceration . degradation of these mirnas correlated with that of s rrna, whereas mir accumulation was increased over -fold after hours of storage, and then gradually decreased over a span of days. mirna (mir ) increasing in macerated tissues (rather than degrading) is uncommon for most mirnas. indeed, the authors indicated that, in terms of genesis and amplification, mir is an atypical mirna, which could explain the disparity in serum detection with respect to canonical plant mirnas. thus, mir was shown to be derived from s rrna and processed in a dcl -independent manner, meaning that when s rrna is degraded mir is produced. this study presents the possibility that certain ncrnas can have non canonical features for synthesis or stability, and should be evaluated individually. in another recent study, the in vitro stability of curcuma longa mirna (clo-mir ) was assessed. when exposed to foetal bovine serum (fbs) for up to h, the mir exhibited high resistance . in summary, food nucleic acid content are generally hydrolysed during processing, with some studies suggesting that mirnas from plants may be more resistant to degradation than synthetic or animal www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. mirnas due to endogenous modifications ( '-o-methylated ' ends). however, these findings are not consistent with mirna stability studies in the animal gastrointestinal tract, and further research is required. although there are many studies on plant ncrnas stability (see . . . .), these mainly focus on ncrna stability within the plant itself. mammalian ncrna stability differs from that of plants because lack the natural '-o-methylation at the ' ends occurring in plant ncrnas, mostly in mirnas (yu et al., ) . very few reports were identified in which plant ncrna stability is investigated in a mammalian organism ) (see sections . . and . .) . understanding the molecular basis that confers plant ncrna stability is of vital importance and needs to be further evaluated to better inform on the relevance of plant ncrna for food and feed risk assessment. for instance, modifications plant mirnas not occurring in mammals, such as the methyl group located on the ' nucleotide ribose, could hypothetically have an impact on plant mirnas stability in mammalian systems due to the lack of the appropriate enzymes for their recognition and degradation, or due to increased stability to mammalian rnases . should this be the case, plant mirnas could manifest a much longer than expected half-life in mammalian organisms when compared to mammalian mirna, which could increase the probability of encountering target molecules. the literature contains descriptions of possible in vitro interaction of plant mirnas with mammalian mirna silencing complexes which may trigger target repression chin et al., ) . however, this hypothesis needs to be experimentally validated since there are gaps in the literature. moreover, it needs to be determined if sufficient levels of ingested plant mirnas reach a target cell to determine an effect in mammalian cells. for instance, in mammalian studies it has been suggested that the threshold for target gene regulation is between and copies of mammalian mirna per cell . accumulation of plant exogenous ncrnas has not yet been reported, but recent next generation sequencing studies have reported a plethora of plant mirnas and other rnas in circulating in the human organism (see section . ). the biological significance of their presence is still unknown. it remains unknown if under certain pathological conditions (i.e. compromised intestinal permeability or renal function) exposure to these plant ncrnas could change. all these gaps in the literature would need to be experimentally validated. to determine the half-life of plant ncrnas in mammalian cells would require experiments utilizing labelled plant ncrnas, first transfected into mammalian cells (in vitro studies) and then orally administered to experimental animals (in vivo studies). the markers used for these experiments could be radioactive probes or labelling molecules of very low molecular weight, such as biotin or fluorescein, to prevent distorting the actual size of the plant ncrna under study. due to their small size, this is important when studying mirnas stability. there is evidence that certain plant ncrnas can induce silencing in some eukaryotic pathogens, pests, parasites or symbiotic microorganisms as a defence strategy. in return, various pathogens develop mechanisms to evade this defence system, proving the existence of a two-way sirnas traffic between pathogens and their plant hosts. plant ncrnas stability outside the plant and in animal systems may play a role in cross-kingdom effects but has received limited attention. the resistance of plant ncrna to the gastrointestinal environment or processing has been partly investigated but more research would be needed. efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. following the methodology described in the section . . . and based on a literature search as described in section . . . . , a total of documents were selected as relevant to describing the state of the art in the field of ncrnas used/intended for use as therapeutics in humans (pharmaceuticals/medicine area). this constitutes valuable preparatory material on the pharmacokinetics (and pharmacodynamics) of ncrna, as required by task . of these publications at least indicate that to ensure the biological activity of exogenous rnas for therapeutic requires numerous chemical modifications and delivery methods. although composed mainly of only four basic building blocks, rnas form simple to very complex structures. its intrinsic base-pair property, enhanced by the ability of the extra hydroxyl groups in the ribose sugar to form hydrogen bonds, are the foundations of its diverse structure and functions (lu et al., ) . rna can fold into various secondary structures including stem, loop, bugle, pseudoknots, gquadruplex and kissing hairpin. it can perform a wide range of biological functions, ranging from regulation of gene expression at various levels to catalysing chemical reactions which closely depend on the ribonucleotide chain's spatial structure. other rnas, such as ribozymes (doudna and cech, ) , can form tertiary structures with catalytic activity. certain rnas can also undergo transformation between alternative structures, depending for instance on binding to specific ligands or environmental changes, as is the case of riboswitches (mironov et al., ) . riboswitches are noncoding rna structures located within mrnas that bind endogenous ligands to regulate gene expression or rna splicing events (cheah et al., ) . the sirna mechanism of action involves dicing of endogenous dsrnas from longer rnas transcripts by dicer and loading into the protein complex of the rna-induced silencing complex (risc). argonaute (ago) proteins are the catalytic core of the risc complex in plants and animals (tolia and joshua-tor, ) . one strand (the passenger strand) is discarded and the guide strand is paired to a complementary mrna sequence via the risc complex. gene silencing can be achieved mainly through post-transcriptional gene silencing (ptgs) or transcriptional gene silencing (tgs). in ptgs mechanisms, the mrna undergoing translation can be sequence-specific cleaved by the risc complex and degraded when the target mrna is perfectly complementary to the sirna. when the interaction is partial or only limited complementarity exists, translational repression and rna degradation occurs, which is a mechanism exerted by mirnas. the mechanism of action for mirna differs somewhat. the mirnas are endogenous encoded singlestranded rnas of about nt long that are important post-transcriptional regulators of gene expression by pairing through a sequence-specific complementary binding to the ' utr of the target mrna. this is usually mediated by a sequence of - nucleotides, known as the seed region, at the ' end of the mature small rna (bartel, ). however, mirnas can also bind to other mrna sites including ' utr or protein coding exons (forman and coller, ) . depending on the degree of sequence complementarity, the interaction will result in mrna degradation and/or translational repression (ameres et al., ; bartel, ) , with destabilization of target mrna being the predominant efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. mechanism to reduce protein output exerted by mammalian micrornas (guo et al., ) . most animal mirnas are transcribed by rna polymerase ii into a long primary transcript (pri-mirna) containing a stem-loop structure. the pri-mirna is then processed by a multiprotein complex containing the nuclear rnase iii drosha into a ≈ nt long hairpin-shape precursor (pre-mirna) and exported to the cytoplasm via an exportin- and ran-gtp-dependent mechanism. the cytoplasmic rnase iii dicer further processes the pre-mirna by removing the terminal loop to produce a small rna duplex, containing the functional microrna and the passenger (*) strand (mirna/mirna*). the duplex mirna is then separated and the functional mirna incorporated into ago proteins within the rna-induced silencing complex (risc), guiding it to the target mrna to induce its translational repression or mrna destabilization (krol et al., ; bartel, ). in , zamecnik first reported the use of an exogenous synthetic oligodeoxynucleotide complementary to the rous sarcoma virus s rna as a potent inhibitor of protein translation (stephenson and zamecnik, ) and virus replication (zamecnik and stephenson, ) . since then, the ability to use exogenous (synthetic) agents to control gene expression has revolutionized many aspects of biological research and catalysed the development of a new promising class of exogenous molecules to treat human diseases. however, despite the obvious promise of this approach, progress has been slow because of the need to overcome myriad technical hurdles, particularly those related to the pharmacokinetics, pharmacodynamics and delivery of antisense molecules. indeed, as most antisense-based potential therapies have not yet produced significant clinical results, very few rnabased or dna-based antisense drugs have been approved for clinical use. in , the u.s. food and drug administration (fda) approved the first aso-based therapeutics for the treatment of a human disease. in , fomivirsen (marketed as vitravene) was approved for clinical use by the european medicines agency (ema) for the european market for treatment of cytomegalovirus (cmv) retinitis in immunocompromised patients. fomivirsen is a synthetic -mer aso (dna) that binds complementary to the sequence of the mrna encoding the ie protein from cmv. the first rna-based therapeutic approved for clinical use was pegaptanib (marketed as macugen), which was approved by both the fda (in ) and ema (in ), for treatment of neovascular (wet) age-related macular degeneration. pegaptanib is also the only approved single strand rna aptamer drug, and it targets the vascular endothelial growth factor as an antagonist. in both cases, fomivirsen and pegaptanib, the route of administration is intravitreal injection, and the unique properties of this route provide benefits in terms of pharmacokinetic and immunological response. in , the fda (but not the ema) approved the second rna-based therapeutic, mipomersen (marketed as kynamro) for treatment of homozygous familial hypercholesterolemia. mipomersen is a dna-rna aso that binds to the mrna of apolipoprotein b (apob), thus reducing apob protein and concomitantly ldl cholesterol levels. antisense oligos like fomivirsen (dna) and mipomersen (dna-rna) act by binding to a complementary sequence of a mrna, thus preventing production of its encoded protein. nusinersen (marketed as spinraza) approved by the fda (in ) and ema (in ) as an orphan-drug for treatment of spinal muscular atrophy is a rnabased aso that increases the production of the full-length survival motor neuron (smn ) protein. eteplirsen ( the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. (bobbin and rossi, ) and many others, which suggests that exogenous rnas can be delivered to humans and used to modify gene function, protein accumulation and treat human diseases. the ability of nucleic acids, both dna and rna, to form duplexes by base complementarity, has been used to develop oligonucleotide-based drugs for gene silencing. in principle, an oligonucleotide binds a target rna through watson-crick base pairing and exerts gene silencing through different mechanisms (kole et al., ) . a single strand antisense oligonucleotide (aso) binds to the mrna, which is also single-stranded, and is translated into proteins. different molecular mechanisms to either block gene expression or degrade the rna duplex formed by watson-crick base pairing have been described. cleavage of the rna strand of dna•rna hybrids is predominantly mediated by the enzyme rnase h (walder and walder, ) , which is an abundant enzyme in both the nucleus and cytoplasm of eukaryotic cells (wu et al., ; ten asbroek et al., ) . asos can also bind rna and block ("steric blockers") gene expression rather than facilitating rna cleavage (dias et al., ) . also considered to be within the aso family are peptide nucleic acids (pnas) or phosphorodiamidate morpholino oligomers (pmos or "morpholinos"). their modification usually acts through this mechanism (michel et al., ) , which in some cases exerts high efficacy in vivo (iversen et al., ; kole et al., ) . exogenous rna molecules or asos can alternatively target and bind a splicing site on the target pre-mrna and thus modify its exon content (mayeda et al., ; skordis et al., ) , which results in the production of alternative splicing products with potential applications in exon skipping therapy (goemans et al., ) . because many of these asos contain dna with multiple modifications, they are not discussed here in detail. the present literature review focuses mainly on ncrna and particularly on exogenous ncrnas. the discovery of rna interference (rnai) (fire et al., ) increased the interest in using exogenous chemically-synthesised rnas for silencing genes in mammals, and promoted their potential usage in human therapeutics (song et al., ) . to achieve therapeutic results, rnai can be administered via delivery to the cell of small exogenous rna duplexes -including short interfering rnas (sirnas) (zuckerman and davis, ), mirnas mimics (bouchie, ), short hairpin rnas and dicer substrate rnas (dsirnas) (foster et al., ; kim et al., ) -to the cell for further processing into an rna interfering silencing complex. although double-stranded rnas have been widely used for sirna therapy, single-stranded sirna (ss-sirna) is capable of activating the rnai pathway and exerting rnai effects (holen et al., ; lima et al., ; prakash et al., ) . moreover, singlestranded mirna mimics have also been shown to activate the mirna pathway (chorn et al., ) . in principle, then, both dsrnas and ss-sirnas can trigger gene silencing of complementary messenger rna sequences (holen et al., ) . other small rnas with imperfect matches have also been described to repress mrna translation (saxena et al., ) . for mirna therapeutics, different pharmacological tools have been developed which involved either inhibiting or enhancing mirna function (van rooij and olson, ; janssen et al., ) . approaches to inhibiting mirna function include small-molecule inhibitors; mirna masking; mirna sponges; and aso, such as anti-mirs, locked-nucleic acids (lna), or cholesterol-modified antagomirs (krutzfeldt et al., ) . the latter's complementarity allows them to bind to mirnas, inducing duplex formation or mirna degradation. the lna therapeutic approach successfully led to the first mirna-based clinical trials for the treatment of hepatitis c (miravirsen; santaris pharma, denmark) with promising results (janssen et al., ) . there are also approaches that can be utilized to enhance mirnas function called "mimic". these include small-molecule activators of mirna expression and mirna mimics, which are www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. exogenous mirnas delivered by several methods aiming to repress the function of their endogenous targets. they are also referred to in the literature as "mirna replacement therapy". as proof of concept, a synthetic version of mir- a (mrx , mirna therapeutics), delivered using a liposomal delivery formulation, was the first mirna to enter a clinical trial for cancer (beg et al., ) . the literature also describes other types of rnas that can exert physiological functions similar to that of mirnas and sirnas. for example, a -nt long guide hairpin rna (ghrna), that might not require ago or dicer, has been shown to exert mirna or sirna activity in vivo (ohno et al., ) . this novel rna interference technology may act as a novel platform for rna-based therapy. indeed, systemic or local injection of the ghr-form mirna- a (ghr- a) suppressed tumour growth in a mouse model of ras-induced lung cancer. understanding the manifold mechanisms of rna function requires detailed knowledge of the rna tertiary structure (magnus et al., ) , which can lead to therapeutic development (digiusto et al., ; strobel et al., ) . indeed, different experimental (lu et al., ; weinreb et al., ) and computational methods (magnus et al., ) have been developed to predict rnas structure and interactions. the capacity of rna to fold in various ways can generate unique three dimensional secondary structures capable of specific molecular recognition of their target cognate (zhou and rossi, ; tuerk and gold, ) . aptamers are short single-strand rna molecules ( - nt) with a defined structure that can specifically bind to a molecular target via tertiary structures. aptamers can be rationally designed using selex technology (tuerk et al., ) , and can therefore be incorporated into chemically modified rnas with high nuclease resistance properties suitable for animal and clinical studies (sullenger and nair, ). unlike other ncrnas therapeutics, aptamers can target soluble extracellular proteins or extracellular domains of cell-surface receptors. the latter characteristic is unique and can be used to deliver sirnas, mirnas or other compounds to target tissues, binding them to a cell-specific aptamer (zhou and rossi, ) . aptamer therapeutics have rapidly progressed to clinical studies, and some are already in phase clinical trials (sullenger and nair, ; zhou and rossi, ) . the unique properties of aptamers led to development of the first therapeutic aptamer and the first rna-based drug approved for clinical use, pegaptanib. however, aptamer and other ncrnas therapeutics are not free of adverse effects (lincoff et al., ) . when compared to sirna, dsirna was found to be comparable in potency and duration of effect. however, dsirna was found to be more immunostimulatory when compared to the shorter sirnas (foster et al., ) . preclinical studies suggest that when administered in lipid nanoparticles (lnp) dsirnas can exert biological effects in different mouse models with tumours of diverse origin (ganesh et al., ) . although preliminary data in nonhuman primates suggest acceptable tolerance for this specific lnp formulation (ganesh et al., ) , the dsirna therapeutic arena still needs to demonstrate its efficacy and safety in humans. catalytic rnas, the so-called ribozymes, are another family of rnas for which therapeutics have been actively developed (kobayashi et al., ) , reaching advanced clinical trials (burnett and rossi, ; mitsuyasu et al., ), but this review will not focus on this type of rna molecules. unmodified nucleic acids have been shown to possess limited stability in biological media and are subject to rapid enzymatic-mediated degradation (braasch et al., ; . there are three major classes of intracellular rna-degrading enzymes (houseley and tollervey, ) (a.k.a. ribonucleases or rnases): (i) endonucleases acting within the rna chain, ii) ' exonucleases and iii) ' exonucleases degrading rna from the ' or ' end respectively. very large amounts of nuclear rnas efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. are rapidly degraded (brandhorst and mcconkey, ) , this being considered either as translation noise (struhl, ) or part of the biological function of ncrnas. instead, most cellular rnas are modified to resist to exonuclease degradation (ren et al., ) . most genomes encode a plethora of rnases, often with overlapping activities, making redundancy a general feature of rna degradation systems, presumably with the goal of enhancing the overall efficiency and robustness of these degradation pathways (houseley and tollervey, ). normal physiological degradation of endogenous rna molecules or degradation of endogenous rna molecules with defects in processing, folding or assembly are not addressed in this literature review. abundant rnase activity has been described in different tissues, including human body fluids (blank et al., ) (weickmann and glitz, ) and with differential catalytic properties (leimoni et al., ) . different tissues contribute to body fluid rnases (neuwelt et al., ) , supporting the cellular defence system against, for instance, viruses (barrangou et al, )(gupta et al., . the mammalian ribonuclease a family seems to contribute to this host defence by exerting antimicrobial activity (harder and schroder, ; dyer and rosenberg, ) . rnase , for instance, is a rnase a superfamily member with potent ribonuclease activity that is secreted by epithelial tissues including skin, respiratory tract, genitourinary tract, and the gut (harder and schroder, ) . the abundant rnase a produced in humans very probably functions to reduce rna contamination, whether endogenous or exogenous, preventing entry into unwanted rna-processing pathways (houseley and tollervey, ). as above described, unmodified rnas (naked) are generally unstable in biological systems, due to the large amount of ubiquitously expressed nucleases. thus, several chemical modifications have been tested for exogenous rna developed for in vivo therapies to increase resistance to nucleases, enhance binding affinity, facilitate cellular uptake, improve the pharmacokinetic and pharmacodynamics profiles, and reduce immunological response or toxicity (wan and seth, ; bennett et al., ) . there are several sites of an exogenous ncrna molecule susceptible to chemical modification ( figure ) without interfering with its ability for base-paring or enhancing its drug-like properties. the several sites on rnas that can be modified include the base, the sugars, and the backbone. this allows them to conjugate with a wide variety of molecules. different chemical modifications have been tested in the context of developing rna-based therapeutics aimed at increasing the pharmacological properties of exogenous rnas and allowing their successful use in different pathologies/targeted therapies. the following section briefly reviews the types of rna modifications included in different studies in the available literature. nucleobase modifications can affect base-paring properties, binding affinity or specificity for the target mrna, as well as exert adverse effects due to competition with natural nucleotide pools or interfere with polymerases (wan and seth, ). replacement of adenosine in the sirna guide strand with adenosine analogues has been found to modify immunomodulatory activity. watson-crick face-localized n-ethylpiperidine triazole modification has been found to be less reactive than the hoogsteen edge modification, and the ' end was more effective at blocking cytokine production than those placed at the ' end (valenzuela et al., ) . c- substitution in pyrimidines in the rna guide strand increase thermal stability of sirna duplexes. indeed, smaller -methyl group substitutions resulted in better sirna activity (terrazas and kool, ). major groove modifications have also been found to increase the serum stability of sirnas (terrazas and kool, ). -position purine modifications, where the major groove edge (i.e. the hoogsteen face) is modified, have also been tested. thiazole modifications of guide strand sirna at position is well tolerated in sirnas, but -ethynyl at this location reduces efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. potency (ibarra-soza et al., ) . several other nucleobase modifications have been evaluated to modify thermal stability of the sirna duplex, hydrogen bonding and sterics, or off-target effects (peacock et al., ) . although not exactly a base modification, movement of the nucleobase from c- to another position on the ribose (i.e. isonucleosides) in sirnas has been evaluated (zhang, j et al., ) . while maintaining binding capacity to rna and stability toward nucleases, passenger strand modifications with isonucleoside at the ' or ' terminals can retain the silencing activity and minimize the passenger strand specific off-target effect (zhang, j et al., ) . chiral inversion of the natural d-forms (spiegelmers) of rna has also been tested in aptamers, leading to their clinical evaluation (ludwig et al., ) . increase nuclease resistance modify plasma protein binding prevent rapid renal excretion improve pharmacokinetics enhance rna-binding affinity enhance thermal stability increase nuclease resistance enhance rna-binding affinity enhance cellular uptake reduce renal filtration enhace delivery to certain tissues modulate protein binding enhance specific cellular uptake increase possibility of formulations for specific tissues/mucosal mebranes ability to traverse biological barriers using extracellular vesicles ´to d irection sites susceptible for modification on an rna molecule. exogenous rnas for therapeutic use can be subjected to different chemical modifications without interference on their base-pairing ability and enhancing their drug-like properties. conjugates at the ´-end or the ´-end may have the same objectives. earlier studies showed that the '-oh of the rna ribose sugar is not required for sirna activity (chiu and rana, ) and '-modifications influence the sugar to adopt a '-endo sugar pucker due to the gauche effects between the o '-and '-substitutes, improving affinity properties (egli et al., ) . therefore, the furanose ring structure at the ' position of rnas has been extensively modified to efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. achieve pharmacological properties including increasing nuclease resistance, stability and efficacy (egli et al., ) . the '-o-methyl ( '-ome) or '-o-methoxyethyl ( '-moe) modifications increase resistance to nucleases and duplex melting temperature (lamond and sproat, ; prakash and bhat, ) , and have been widely used to develop mirnas (meister et al., ) , sirnas (judge et al., ; choung et al., ) or aptamer-based (burmeister et al., ) therapies. for example, '-ome in sirna has been shown to selectively protect the particularly vulnerable '-end of the guide strand against exonucleolytic degradation in human blood serum (hoerter and walter, ) . moreover, '-ome is a naturally occurring modification of rna found in trna and other small rnas. '-ome has also been used to modulate rna splicing ( toxicity related to lna modifications has also been described (swayze et al., ) . although sugar-modified oligonucleotides or exogenous rnas, including sirnas or antimirs, are more effective rna inhibitors than their unmodified counterparts, they are still susceptible to serum in addition, the phosphorothioate linkage promotes plasma protein binding (srinivasan et al., ; wan and seth, ; sands et al., ) . this increases pharmacokinetic benefits by reducing rapid renal clearance (sands et al., ) and facilitating tissue delivery, binding to cell surface proteins and cellular uptake (miller et al., ; overhoff and sczakiel, the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. binding affinity (tm) for its target complementary nucleic acids (≈ . ºc/linkage) (freier and altmann, ) including mrnas and mirnas (krutzfeldt et al., ) . thus, the use of a mixture of phosphodiester and phosphorothioate bonds may be preferred over all phosphorothioate bonds for in vivo applications (ghosh et al., ; krutzfeldt et al., ) . also, the phosphorothioathe linkage confers chirality at phosphorus and might be relevant for sirna activity (jahns et al., ) . although initial studies have shown exogenous homologous rna uptake by cell suspension, there was very rapid degradation after uptake (shanmugam and bhargava, ) . duplex rnas may be more stable than single-stranded rnas, even in the absence of phosphorothioate modifications (braasch et al., ) . indeed, '-ome hairpin-designed antimir oligonucleotides do not seem to require phosphorothioate modification to exhibit in vitro activity (lennox and behlke, ). minimally-modified phosphodiester aso has been reported to have less (brigui et al., ) , similar or superior activity than its complete phosphorothioate counterparts (uhlmann et al., ) , but sugar modifications in phosphodiester backbones can compensate the loss of activity (monia et al., ) . some adverse effects related to phosphorothioate modifications, compared to their phosphodiester backbone counterparts, have been described in aso and sirnas, including nonspecific protein binding (brown et al., ) , the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. (dsrna). alpha-tocopherol conjugated sirnas can be delivered to liver tissue (nishina et al., ) or the brain when administered locally (uno et al., ) . other lipid conjugates of rnas have been evaluated including bile acids, long-chain fatty acids, and medium-chain fatty acids (murakami et al., ; lorenz et al., ; wolfrum et al., ) . conjugation with specific peptides has also targeted sirnas to specific tissues (hsu and mitragotri, ; yamayoshi et al., ) . another approach includes conjugation with high-molecular-weight polyethylene glycol (peg) to overcome renal filtration. this strategy was used in rna aptamers either approved (macugen) or in advanced clinical trials for ocular disorders (zhou and rossi, ; drolet et al., ) . the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. rna delivery to a target tissue for appropriate and specific cellular uptake is currently a main hurdle in rna therapeutics. several synthetic non-viral methods for rnas delivery have been developed for local and systemic rna therapeutics ( multiple studies have suggested that exosomes, naturally-released extracellular vesicles, can deliver rnas (mrnas, mirnas, lncrnas) to recipient cells (zhang, l et al., ; . due to their ability to traverse biological barriers, the use of exosomes or exosome-mimetic nanovesicles for rna delivery is an open field for future rna therapeutics (lunavat et al., ; zhou et al., ) . indeed, preclinical studies suggest that exosome-mediated delivery of rnas is feasible in vivo (didiot et al., ; el-andaloussi et al., ) . different rnas have been found in milk exosomes from different species, including humans zhou, q et al., ) , but whether these vesicle-protected exogenous rnas are bioavailable is still in debate (zempleni et al., ) . it is important to note that most of the above described chemical modifications are not found in nature and have been developed generally to increase the drug-like properties of rnas and other nucleotides. to the best of our knowledge there is no evidence of clinical trials using unconjugated naked or unmodified exogenous rna for human use with any administration route. efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. early studies suggested that rnas were poor drug candidates due to their relatively high instability, relatively short half-life in vivo, immunomodulatory effects and the many hurdles to their cellular absorption caused by their negative charge. however, several improvements in stabilization chemistry have been identified in recent decades. indeed, a number of chemical modifications and conjugation strategies have improved their rna-binding affinity, in vivo nuclease stability and pharmacokinetic and pharmacodynamic properties. an exogenous rna can be amenable for chemical modification without interfering with its base-pairing ability. several parts of the exogenous rna molecule can be chemically modified including the nucleobase, the backbone or the ribose (sugar). rnas can also be conjugated with a variety of molecules which can exert different biological properties. in addition to these, chemical and biological approaches can be applied to deliver exogenous rna to cells or specific tissues. although many aspects are still to be fully understood in the field of chemical modification of exogenous rnas for therapeutic use, several exogenous rnas (sirnas, aptamers, mirnas, etc.) have entered advanced clinical trials for human use. adams the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. following a systematic literature search as described in section . . . and . . . ., a total of papers were reviewed on the pharmacokinetics of foreign exogenous ncrnas used/intended to be used as therapeutics (hereafter referred as exogenous ncrnas). this section focuses on the pharmacokinetics of exogenous ncrnas (naked, i.e. nonchemically modified or minimally chemically modified) other than those consumed orally from plants; these will be covered in chapter . . moreover, only in vivo studies, in mammals and humans, are reviewed here. of the documents included here, were used for review of ncrna pharmacokinetics in disease and other conditions, which may be relevant to risk assessment considerations. although not specifically requested by the mandate, information on pharmacodynamics aspects of ncrnas was considered. pharmacodynamics is the study of a drug's effect on an organism, meaning its biological effects and other aspects of its xenobiotic action, (efficacy, potency, and toxicity) and this is considered relevant for ncrnas (see . ). pharmacokinetics is dedicated to determining the fate of substances administered to a living organism. it describes the trajectory of a xenobiotic (in this case exogenous ncrna) after delivery into an organism, and encompasses from the movement of administration, to its movement through the organism, to its complete elimination; in other words, absorption, distribution, metabolism and excretion. a drug's pharmacokinetics depends on an individual's physiology (patient-related factors, i.e. renal function, sex, age) or pathology (i.e. renal failure, hepatic failure), and drug chemical properties. pharmacokinetic relevant parameters include: dose, the amount of drug administered at a single point in time; dosing interval, the time between drug dose administration; c max , the peak plasma concentration of a drug post administration; t max , the time to reach c max ; elimination half-life, the time required for the drug concentration to reach half its original value; area under the curve (auc), the integral of the concentration-time curve (after a single dose or in steady state); and clearance, the volume of plasma from which a substance is completely removed per unit time. pharmacokinetics modelling is done using noncompartmental or compartmental methods; the former estimate exposure to a drug by estimating the auc of a concentration-time graph, and the latter estimate the concentration-time graph using different kinetic models. considering an organism as a homogeneous compartment (monocompartmental model) implies that a drug is distributed equally throughout the entire body (tissues and fluids). therefore, blood plasma drug concentrations are a true reflection of drug concentration in other fluids or tissues, and drug elimination is directly proportional to its concentration in the organism. not all tissues in an organism receive the same blood supply and different drugs have different characteristics (i.e. variation in drug passage through natural barriers, such as the brain blood barrier, due to a drug's biochemical properties), meaning other models are sometimes needed. the two-compartmental model assumes that there is a compartment where distribution is more rapid (central compartment) and another compartment www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. (peripheral compartment) consisting of organs with a lower relative blood flow, which consequently exhibits a slower drug distribution rate. saturation of the enzymes responsible for drug metabolisation, the presence of active transmembrane transport mechanisms independent of drug plasma concentration, and many other factors not taken into account by the two-compartmental model, require the use of multi-compartmental models. very few studies have focused on the pharmacokinetic profile of exogenous rnas (naked or unmodified), with most of the knowledge on the absorption, distribution, metabolism and excretion profile of ncrnas coming from the development of antisense oligonucleotides therapeutics (dirin and winkler, ) . scaling dosing from preclinical animal studies to humans (i.e. based on body weight or surface area) is challenging and apparently depends on the mechanism of clearance. for example, the pharmacokinetics of a naked sirna formulated into polymer-based nanoparticles in mice, rats, monkeys and humans exhibited blood c max shortly after intravenous administration and rapid elimination across all species in correlation with the body weight (zuckerman et al., ) . the pharmacokinetic profile investigated during the development of exogenous rna therapeutics has shown to be influenced by the route of administration, as described below. a comprehensive summary of available information of in vivo studies and related pharmacokinetics of exogenous rnas is provided in table . most studies on exogenous ncrnas have focused on iv administration. studies in mice have shown that naked unconjugated sirna ( µg/kg) injected intravenously disappears from peripheral blood min after administration, restricting the likelihood of accumulation to peripheral organs (gao et al., ). similar results were found following iv administration of naked sirna ( i labelled), which distributed to the kidney and liver within the first minute, peaking in the kidney, liver and bladder within the first minutes (braasch et al., ) . although distribution of sirna ( i labelled) decreased markedly after h, it persisted in the kidney and liver up to h, and lower concentrations were observed in the lung, spleen and heart (braasch et al., ) . in rats, sirnas were distributed to the kidney and excreted into the urine one hour after injection (van de water et al., ) . exogenous mirna iv administration has also been tested in animal models and entered human clinical trials. in this case (mirna mimics), they are either chemically modified dsrnas formulated in different nanoparticles (xue et al., ) to preferentially target a tissue (trang et al., ), or incorporated into naturally circulating extracellular vesicles (bala et al., ) . for example, mirnas mir- and let- b formulated in a neutral lipid emulsion have been iv administered to mice and have shown to exert a biological effect (kasinski et al., ) . the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. administration site for less than one day, while in a chitosan-formulated version it persisted for up to days (ma et al., ) . peptide-modified and chemically modified sirnas applied topically to the skin have been found to exert a local biological effect, as compared to the sirna alone (hsu and mitragotri, ) . mirnas formulated with transfection agents or nanocomplexes have also been injected intradermally (srivastava et al., ) , or subcutaneously (urgard et al., ) , and demonstrated to exert local biological effects. intranasal administration of sirnas in rats for brain targeting has resulted in very low levels of radioactive sirnas in the brain or plasma when naked sirnas were administered compared to a formulated version (perez et al., ) . mirnas administered by intranasal injection using transfection reagents reached the dorsal root ganglia and the olfactory bulb (cheng et al., ) and, using another nanovector delivery system through intranasal administration (as drops), it reached the brain, exerting a local biological effect (zhuang et al., ) . due to ncrnas susceptibility to degradation, very few studies have focused on exogenous ncrna administration in the gi tract. non chemically modified naked sirna administered by oral gavage to female mice ( µg in total), was found intact and at high levels (analyzed by northern blot and by quantitative pcr analysis) and hours after dosing in the stomach, small intestine and colon when formulated with chitosan nanoparticles (ballarin-gonzalez et al., ) . by contrast only low sirnas levels were observed in the stomach, intestine and colon of mice hour after oral administration of the non formulated naked sirna. at this timepoint intact sirna was almost exclusively present in the stomach while partial degradation products were observed in the proximal and distal regions of the small intestines. further reduction of sirna levels was observed hours after administration, with traces only found in the colon (ballarin-gonzalez et al., ) . orally administered sirnas, formulated using specific vehicles including thioketal nanoparticles ( µg kg - d - the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. counterparts were present in all epithelial cells (martirosyan et al., ) . oral administration of exogenous exosomes containing mirnas of bovine origin has been shown to attenuate arthritis in mice models. bovine milk derived extracellular vesicles were administered daily by oral gavage starting at week till week after birth and arthritis was induced by collagen gavage. animals receiving exosomes ameliorated the clinical condition, although no clear mechanisms for this effect have been described . enteral (large intestine) sirna administration for therapeutic gene silencing targeting the liver (via the lymphatic route) has also been described, but using chemically modified and formulated sirnas (murakami et al., ) . enzyme-and ph-responsive microencapsulated nanogels are also being developed for oral sirna delivery, as reported in recent in vitro tests (knipe et al., ) . naked sirna formulated in a water-in-oil microemulsion has been administered via the rectal mucosa to mice along days ( - µg sirna/kg, administrations) and has reached the brain where exerted a biological effect through downregulation of the prion protein expression (lehmann et al., ) . prion infected mice presented an improvement of some neuropathological conditions after receiving the formulated sirna following the rectal mucosa route of administration. being the eye the target for the first rna-based therapy, sirna intraocular administration has been investigated. intravitreally injected naked sirna (slightly modified, dtdt) distributed throughout the eye (vitreous, iris, retina, retinal pigment epithelium and sclera) of rabbits when administered at mg/eye, and the pattern of ocular distribution was similar in male and female rabbits ( . conjugated or formulated sirnas have also been tested in animal models (zhang et al., ; janout et al., ) . intravitreal injection of mirnas has been described in the literature using either naked mirnas in mice , rats (qin et al., ) or using transfection reagent in rats (mcarthur et al., ) or exosomes in rats (mead and tomarev, ) . some studies have evaluated ip administration of naked sirnas (slightly modified, dtdt) in animal models (wilcox et al., ; shimizu et al., ) . the half-life of ip administered naked sirna (dtdt) was found to be ≈ times lower than the formulated version (perepelyuk et al., ) . sirna (dtdt) was also quickly excreted into the urine compared to a formulated version (shimizu et al., ) . in a comparison of a formulated (complexed) and naked sirna after ip administration in mice it was found that after min the formulated version was detectable in muscle, kidney, liver and tumour tissue, while the naked sirna was completely absent (urban-klein et al., ) . while not a common administration option, chemically modified sirnas formulated in ph-responsive nanoparticle complexes have been administered in the submandibular gland of mice (by retroductal injection) and found to exert rnai effects (arany et al., ) . the monitoring of the distribution of the chemically modified sirna h post-administration indicated that the formulated sirna was internalized www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. in the cytoplasm of duct cells via the endosomal pathway. at h post-administration, the signal was concentrated in the acinar compartment as well. after h, ≈ % of the submandibular gland cells were positive for the sirna. the delivery of the sirna formulated in nanoparticles was more efficient compared to that of naked sirna (arany et al., ) . in another study, iv administration of chemically modified sirna ( . mg/kg) was shown to accumulate remarkably in the submandibular gland just min after injection, and had a half-life in this tissue of ≈ . h, but did not have a rnai biological effect . in contrast, direct local injection of liposome-encapsulated sirna ( . mg/kg) into the submandibular gland was shown to have strong gene-silencing effects, while naked sirna injection ( mg/kg) did not . few studies have focused on this administration route. senn and colleagues compared central nervous system administration of sirna vs. aso (senn et al., ) . compared to phosphothioate and 'methoxyethyl protected aso (up to µg), naked (slightly-modified, dtdt) sirna ( µg) administered to rats via icv was not found in any brain region, including the area around the injection site. in contrast, h after aso injection it was distributed in the thalamus and hypothalamus. the naked sirna clearly exhibited very poor distribution and uptake in the brain compared to the aso. although aso exhibited a capacity to reach different brain structures, its administration at a total dose of µg over two days produced no rnai effect. in the same context, icv administration of sirna (up to µg/rat/day) on two consecutive days did not exert rnai effects. however, in another study, a sirna chemically modified to allow for delivery in the absence of transfection reagents (accell sirna) was administered via the icv route at µg/rat. the sirna was incorporated into different types of neurons, although not glia, where it exerted a biological effect in regions like the cortex, the caudate subregion of the striatum and in the pyramidal cells of the hippocampal ca subregion (nakajima et al., ) . in a different study, a mirnas mimic (chemically modified double-stranded rnas) administered by icv ( pmol/g body weight, mixed with cationic lipid dotap) was found to increase forty fold the level of this mirna in the brain (stary et al., ) . several chemically modified mirnas (agomirs) were also found to be active when administered via this route in rats (huang et al., ; ge et al., ) . a double-stranded modified pre-mirnas was also reported as active in rats (davis et al., ), but a naked mirna (double-stranded -bp rna) injected by icv ( pmol) in this study was unable to increase its levels or exert a biological effect, suggesting that the naked mirna had a higher susceptibility to rnase degradation (davis et al., ) . lipid-complexed naked sirna is reported to be absorbed by mouse epithelial and lamina propria cells of the vagina and ectocervix (palliser et al., ) . however, uncomplexed and unmodified sirnas were immediately degraded (< min) when exposed to genital wash fluid (wu et al., ) , suggesting high rnase activity in the cervicovaginal secretion. also, slightly modified but still naked sirna (dtdt) was unable to exert a clear biological effect (zhang et al., ) . in summary, different administration routes have been tested during ncrna therapeutics development. the administration route used for exogenous ncrna delivery clearly influences pharmacokinetic profile of chemically synthesized ncrna. no matter the administration route, it seems that naked and unformulated rnas rapidly degrade and exert very limited or no biological activity. very few studies have evaluated the distribution profile of administered naked oligonucleotides in organs and tissues or compared it to the distribution profile of their chemically modified counterparts. using the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. imaging techniques in baboons (positron emission tomography -pet -and [ f]-labelling) it was shown that naked oligonucleotides are poor drug candidates due to limited distribution into organs and tissues and very rapid elimination (tavitian et al., ) . compared to '-o-methyl or phosphorothioatemodified oligonucleotides, naked oligonucleotides were found in the heart, liver and kidney and, to a lower extent, in other tissues min after iv injection. at that timepoint, naked oligonucleotides were already found in the bladder suggesting that naked oligonucleotides are very rapidly eliminated compared to '-o-methyl or phosphorothioate ones. moreover, one hour after injection, almost all naked oligonucleotides were found in the bladder or residually in the kidney, while the phosphorothioate oligonucleotides were found largely in the liver and kidney, and the '-o-methyl ones were in an intermediate distribution (tavitian et al., ) . kinetics studies with radiolabelled oligonucleotides also showed a dramatic reduction of naked oligonucleotides in the plasma within the first min, while '-o-methyl and phosphorothioate-modified ones showed a much slower reduction during the first min. naked sirna have been reported to distribute to the kidneys and bladder of mice min after iv administration (braasch et al., ; naeye et al., ) . intravenous administration of mg kg - partially-modified unconjugated sirna (partial phosphorothioate backbone and '-o-methyl sugar modification) or cholesterol-modified sirna in mice resulted in a broader tissue distribution (liver, heart, kidney, adipose and lung) of the cholesterol-conjugated sirna h after injection (soutschek et al., ) . in another study, naked sirna (free sirna) iv administered (retro-orbitally) was found in the kidney min after injection (naeye et al., ) . ip administration of synthetic sirna formulated with low molecular weight polyethylenimine resulted in its presence in muscle, liver, kidney and tumour tissues min after injection, while the naked counterpart was completely absent (urban-klein et al., ) . in rats, chemically modified naked sirnas were distributed in a similar manner to those of '-o-methyl or '-f modified sirnas (viel et al., ) , even though the circulation time of the '-f was increased. in other studies with rats, iv injection of sirna (chelated to indium- ) resulted mainly in renal distribution hour after exposure (van de water et al., ) . some liver distribution of sirnas have been also reported in mice, when using either the two phosphodiester rna strands duplexes or one phosphodiester strand and one phosphorotioate strand sirna duplex (braasch et al., ) . orallyadministered naked sirnas ( µg sirna) were found to be present in the stomach, proximal and distal small intestine and colon tissues hour after administration but at very low levels, further lowering hours after administration (ballarin-gonzalez et al., ) . the same sirna formulated with chitosan nanoparticles was found to distribute systemically and reach the liver, spleen and kidney (ballarin-gonzalez et al., ) . in an experimental model of arthritis in mice, iv administered naked sirna ( µg) was found to distribute to arthritic joints immediately after injection, but disappeared within hours, compared to that of specific formulations (komano et al., ) . overall, these finding suggest that naked sirna, although distributing at very low levels and with short residence times compared to other formulated counterpart versions, does distribute to the gi tract when administered orally or other tissues when administered systemically. www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. formulation type (i.e. cationic lipids, neutral lipids, or others) can drive preferential distribution to specific tissues. as described for certain mirnas, a neutral lipid emulsions (nle) formulation has been shown to distribute to the lung after iv injection ( µg) (trang et al., ) . the lack of cationic lipids in the nle seems to bypass some of the shortcomings that can be attributed to the charge, including formation of aggregates in biofluids, filtration by the liver, adherence to the endothelium or uptake by scavenging macrophages. the result is excellent distribution to lung tissues (trang et al., ) . extracellular vesicles (exosomes)-formulated synthetic mir- was found to distribute to liver, adipose, lung, muscle and kidney tissues min after iv injection (bala et al., ) . administration of exosomeloaded mir- resulted in its detection by isolated hepatocytes and liver mononuclear cells, suggesting cellular uptake (bala et al., ) . in summary, naked oligonucleotides, including ncrnas, rapidly distribute to the liver, lung and kidney after iv injection. within the first minutes they are found in the bladder, suggesting rapid renal elimination. chemical modifications or formulations increase their distribution to other tissues or delay clearance. exosome-loaded rnas have recently been shown to be an alternative tool for distribution to different tissues. ncrnas by oral administration seem to distribute to the gi tract or, if formulated, systemically. there are few studies which have evaluated the metabolism of exogenous rnas administered to animals or humans, most of which only investigate their degradation. naked sirnas seem to be rapidly degraded when administered iv (viel et al., ) , decreasing by ≈ % within the first min, although certain chemical modifications (i.e. 'f, 'ome) can slightly increase this time. stability of naked sirna was reduced (< min) compared to that of other chemically modified or formulated sirna when administered by iv injection in mice (gao et al., ) . indeed, incubation of naked sirna with plasma or foetal bovine serum causes a very rapidly degradation of the genetic material (jiang et al., ; braasch et al., ; urban-klein et al., ) . unmodified aptamers in general are also very rapidly degraded (≈ min) (zhou and rossi, ) . when naked sirna is administered as formulated (i.e. nanoparticles) it was shown that the renal filtration barrier can separate the sirna from its carrier (naeye et al., ; zuckerman et al., ) . in the eye, sirnas are degraded by endonucleases without preferences for one side of the duplex (as observed also for chemically modified sirnas) (beverly et al., ) . early in vitro studies also showed that exogenous rnas taken up by cells are rapidly degraded (shanmugam and bhargava, ) , and exogenous rna injected into the blood is rapidly degraded to nucleosides, ribose phosphate and free bases (sved, ). the renal filtration barrier owns an effective size cut-off of ≈ Å (batsford et al., ; bohrer et al., ) , but is also influenced by net molecular charge discrimination (brenner et al., ) . this is supposed to facilitate rapid renal clearance of small molecules and free sirna (zuckerman et al., ; naeye et al., ) since aso, sirnas and aptamers are commonly smaller than this cut-off (huang et al., ) . by comparing naked sirnas and ribose-modified sirnas ( '-o-methyl or 'f) using pet scans in rats and mice, the kidney was identified as the main organ for sirna elimination (viel et al., ) . however, it is worth noting that also the liver participates in sirna elimination. in other studies, sirna appeared to accumulate in the bladder shortly after administration (hatanaka et al., ) . this was also the case for modified sirnas (lna) compared to their versions conjugated with peg administered in mice, in which their levels peaked in the bladder min post injection and they were found in the urine as soon as min after injection (iversen et al., ) . efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. some studies have described rapid elimination of naked sirna and renal excretion after iv injection at doses including µg (jiang et al., ) and µg/kg (gao et al., ) in mice or rats (van de water et al., ) . other studies using dynamic imaging techniques or radiolabelled sirnas have confirmed that most of the naked sirna (free sirna) administered to mice was found in the bladder after just min up to min, faster than the formulated counterparts (naeye et al., ; zuckerman et al., ) . the literature mostly addresses chemically modified rnas (dyke et al., ) in clinical trials (zhou and rossi, ) . the small size of rna aptamers renders them susceptible to rapid clearance by renal filtration which has driven efforts to increase their size by conjugation for therapeutic use (healy et al., ; haruta et al., ) . overall this indicates that naked exogenous small rnas are rapidly cleared from systemic circulation and excreted in the urine. iv injection of a cocktail of four different plant-based small rnas (mir , mir a, mir a, and mir ; pmol each) showed them to be rapidly cleared from circulation (< min) . the exception was mir , which peaked min after injection compared to the other mirnas administered at equal dosages, and then dramatically decreased at min. when compared to other small rnas, anomalous or atypical stability and genesis were proposed for this specific mirna. efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. although scarcely described, there may be several conditions in which, theoretically, the pharmacokinetic properties of exogenous rnas could be modified. increased intestinal permeability has been described in different conditions including consumption of nonsteroidal anti-inflammatory drugs, celiac disease, crohn's disease or other intestinal inflammatory disease (oman et al., ; sundqvist et al., ; michielan and d'inca, ) . for instance, absorption may be altered in cases where vasculature permeability of the gut is increased, as it has been observed in murine models of ageassociated inflammation (jeong et al., ) . increased intestinal permeability ("leaky gut") has been described in alcoholism, contributing to the extra-intestinal tissue damage caused by alcohol (bjarnason et al., ) , including liver damage (keshavarzian et al., ) . similarly, leaky gut has been linked to liver steatosis in obese patients ( theoretically, leaky gut may increase the possibilities of passively absorbing larger amounts of exogenous rnas with consequent release into the circulatory system. however, very few studies are available in which sirnas, aptamers, mirnas or other exogenous ncrnas have been tested under these conditions. absorption of naked sirna formulated into nanoparticles following intrarectal administration was tested in mice models of dextran sulphate sodium (dss) induced colonic inflammation. uptake of sirna (observed mainly in epithelial cells, lamina propria lymphocytes, and cells from the mesenteric lymph nodes including dendritic cells and t cells) was increased in mice suffering from acute colitis compared to uptake (in the same cell types) under healthy conditions (frede et al., ) , suggesting that inflammatory conditions may alter ncrna absorption. although there are no studies using naked unmodified exogenous ncrnas, this topic deserves further attention. acute kidney injury or chronic kidney disease are important public health issues. the kidneys play a key role in elimination of xenobiotics and metabolism products and thus renal clearance should be considered in drug discovery and drug interaction research (feng et al., ) . very few studies have focused on the clearance of exogenous ncrnas in pathological renal conditions. for example, plasma clearance of chemically modified sirna administered by iv injection ( mg/kg) was slightly reduced in renally impaired rats than in normal animals (thompson et al., ) . although sirna concentration in residual kidney tissue of partially nephrectomised animals was slightly greater than the mean in normal animals, the differences did not result in appreciably greater sirna distribution into other organs (thompson et al., ) . in general, similar results, but using asos, were observed in rats after cisplatin-induced renal injury (masarjian et al., ) . also in a study of an aso against mirnas, it was recently reported that in end stage renal disease patients, only relatively modest increases in aso plasma levels were observed when compared to control subjects (http://regulusrx.com/wpcontent/uploads/ / / -aasld-rg- -pk-and-safety-in-esrd-vs-normal.pdf). also using asos, distribution within the kidney was found to be altered in a mice model with chronic kidney disease (ckd) compared to controls (donner et al., ) . moreover, concentration of `-moe asos in the liver of mice with ckd was elevated as compared to mice without ckd, suggesting that a reduction of renal function and aso excretion can potentially influence the systemic delivery of asos (donner et al., ). www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. liver disease may also lead to alterations in drug pharmacokinetics, and thus the potential effects of hepatic impairment are considered in drug development (allen et al., ) . although exogenous ncrnas have not been analysed quantitatively, one study showed that chemically modified and formulated sirnas in lipid nanoparticles were found to be distributed and similarly accumulated in pathological or normal liver tissue from mice (yu et al., ) . other physiological, pathological or lifestyle factors, such as aging (cohen, ) and exercise activity (van baak, ) , may influence the pharmacokinetics of exogenous molecules. however, no information is yet available regarding exogenous rnas and their behaviour under these conditions. pharmacodynamics refers to the relationship between the concentration of a drug at the site of action and the resulting effect, including the time course and intensity of therapeutic and adverse effects. there is scarce data on the pharmacodynamics of naked unmodified exogenous ncrnas. some information is available on chemically modified rnas (or nuclease-stabilized rnas, the latter being outside the scope of this literature review). for example, in a mouse model of viral hepatitis the administration (hydrodynamic injection) of naked or chemically modified (methyl-modified) sirnas was associated with clear dose-related effects of various rnai constructs (peng et al., ) . in this study the naked sirnas had the highest inhibitory effects, but these rapidly declined (shorter-lasting inhibitory effects); in contrast, the methyl-modified sirna duplexes were more stable inside cells and exerted their effect over a longer time period (peng et al., ) . non-compartmental analysis of rna therapeutics is used in most preclinical studies when analysing pharmacokinetic and pharmacodynamic parameters. for example, a non-compartmental model was used to estimate the properties of intravitreal injection of slightly chemically modified sirna (dtdt) (dejneka et al., ) . also, in a mirna mimic therapy by iv administration of a liposomal formulation, a non-compartmental model was used to evaluate different parameters (beg et al., ) . systemic delivery of naked sirna has generally failed to produce significant biological effects (gene silencing). no biological effects have been reported when naked sirna ( µg/eye) was administered by intravitreous injection in rats . few studies report gene silencing effects following local administration, although some of the tested rnas were slightly chemically modified (i.e. dtdt overhangs). intrathracheal administration of naked (but slightly modified, dtdt) sirna was effective through gene silencing effects in a mouse model of haemorrhagic shock and sepsis (perl et al., ) . in non-human primates, naked (but slightly modified, dtdt) and unformulated sirna administered intranasally was effective in severe acute respiratory syndrome sars coronavirus infection reducing viral load and host symptoms (li et al., ) . it looks that naked ncrnas require proper formulation to attain efficacy. naked sirna administered intranasally to mice was effective in reducing viral replication when complexed with lipofectamine (fulton et al., ) . naked sirna formulated in ph-triggered nanoparticles exerted gene silencing effects when administered by iv injection in mice ( mg/kg) (kolli et al., ) . a formulation of naked sirnas using cyclodextrin-containing polycations and transferrin, as a targeting ligand for delivery to transferrin receptor-expressing tumor cells, also showed gene silencing effects when administered by iv injection ( . mg/kg) in a mouse model (hu-lieskovan et al., ) . when administered orally efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. ( µg/kg) to a mice model of thioglycollate-elicited inflammation, a formulated sirna reached macrophages in the peritoneum, spleen, liver and lung, exerting a sirna effect (aouadi et al., ) . a liposome-formulated naked mirna iv administered twice a week was efficacious in clinical trials (beg et al., ) . a cationic liposome formulated dicer-substrate sirna (dsirna) (a solid dsirna core in an envelope of cationic lipids and cholesterol) was found to be efficacious in mice when administered by iv injection ( mg/kg) (ganesh et al., ) . preliminary results within this formulation demonstrated a tolerability profile in non-human primates upon repeated administration (ganesh et al., ) . when comparing the naked sirna and to a highly formulated sirna in cyclodextrin-containing polycations with surface peg and transferrin as the targeting ligand, only the formulated version ( . mg/kg iv) was effective in reducing the expression of the ews-fli gene and tumour engraftment, while the naked version exerted no biological effect (hu-lieskovan et al., ) . similarly, in a mouse model of concanavalin a-induced hepatitis a reduction of hepatic injury by liver-specific induction of rna interference was observed upon iv administration a of galactose-conjugated liposome nanoparticles (gal-liponp) bearing naked sirna, while no effects were noted following administration of the unformulated naked sirna (jiang et al., ) . in a collagen-induced arthritis mouse model and using systemic administration, naked sirna (dtdt) was used against tnf-α, and was completely unable to repress tnf-α mrna expression, whereas a formulated version repressed it (komano et al., ) . in a mouse tumour model, intraperitoneal injection of naked sirna exerted no rnai effects, while its complex-formulated version did (urban-klein et al., ) . while developing a model to study sirna pharmacodynamics, intramuscular administration ( µg) of naked sirna (chemically stable) in mice showed no effects without use of electroporation (stevenson et al., ) . there are several ongoing clinical trials using sirnas, rna aptamers, mirnas and other noncoding rnas intended for use in humans to treat diseases including cancer, cardiovascular disease, inflammation, ocular disease and others (zuckerman and davis, ; li and rana, ; bobbin and rossi, ; zhou and rossi, ; sullenger and nair, ) . most of these ncrnas are chemically modified or formulated. adverse effects were identified in some of these studies. the delivery component of the formulation (i.e. liposomes), rather than the naked sirna component, was primarily responsible for the adverse effects observed in different preclinical toxicity studies (zuckerman et al., ) . for the mir- a mimic, adverse effects in humans have been reported including acute renal injury, back pain and fatigue, all likely attributed to the liposomal carrier rather than the mirna (beg et al., ) . liposomerelated toxicities are considered due to activation of the complement and have been well characterized with the most frequent symptoms observed being flushing, rash, dyspnoea, chest pain, back pain and subjective distress (szebeni et al., ) . in non-human primates, repeated iv administration of escalating doses of a non chemically modified sirna led to increased il- and inf-gamma levels, as well as kidney and liver toxicity at the highest dose tested ( mg/kg) (heidel et al., ) . in several clinical trials on rna therapeutics, dexamethasone premedication was required to reduce infusion-related adverse effects or reactions (i.e. hypersensitivity, flushing, oedema, etc.) (beg et al., ) . for some sirnas used in the clinic, a predosing hydration protocol has been used to protect the kidneys (zuckerman et al., ), thus reducing in humans the toxicities observed in animals. there are also animal studies in which no toxic effects are reported. for instance, when highly formulated naked sirnas were administered by iv route at . mg/kg, no apparent liver or kidney toxicity, immunogenicity-related toxicity or organ damage were detected in a murine model of metastatic ewing's sarcoma (hu-lieskovan et al., ) . no histological toxic effects were seen in liver, kidney, lung, heart and spleen in the same model following repeated treatment with the same formulation the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. (twice weekly for weeks) (hu-lieskovan et al., ) . the same results were obtained for the naked sirna administered at the same dose and for the same duration (hu-lieskovan et al., ) . in addition, when administered daily via oral gavage ( µg sirna/kg) for days in mice, sirna nanoparticles in galactose modified trimethyl-cysteine conjugates were reported to have no histological toxicity in major organs (brain, heart, kidney, lung, liver or spleen) (han et al., ) . since there are so few studies in the current literature on the pharmacokinetics of naked/unmodified rna molecules, this section included studies of other, minimally-modified, exogenous rnas to compare their pharmacokinetic and pharmacodynamic properties to that of naked rnas, when possible. the findings suggest that naked or unmodified rnas are rapidly cleared from circulation and have been reported in the kidney and urine immediately after administration (generally intravenously). in general, no major gene silencing effects have been observed for naked rnas when compared to chemically modified or formulated ones. this lack of efficacy has probably discouraged studies investigating the toxicity and safety of naked unmodified rnas. while limited studies have reported minor biological effects of certain naked rnas in animal models (very much dependent on the route of administration), the lack of studies in humans prevents scientists from understanding the kinetic profile of exogenous rnas and developing effective therapeutics. finally, scientific literature on toxicological or safety studies provides no evidence for concern about the systemic toxicity of naked exogenous small rnas under normal physiological conditions. allen the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. following a systematic literature search as described in section . . . . and following the methodology described in the section . . . , a total of papers were selected as relevant to reviewing the topic of rnas uptake. information on naked (non chemically modified) exogenous rnas is mainly presented in this section. of these publications, support the uptake of exogenous ncrnas. one study reports no intestinal absorption of rna (baintner and toth, ) , and three others suggest that rna is degraded before absorption (loretz et al., ; o'neill et al., ; sonoda and tatibana, ) . uptake of plant exogenous rnas, after oral intake is not included here, and will be reviewed in chapter . ; and uptake of exogenous rnas from the pharmaceutical/medicinal area is described in section . . . herein, uptake refers to passage of the molecule (in this case a ncrna) through the cellular membrane as a part of the absorption process. an important feature for the possible biological effect that exogenous rnas, including ncrnas, may have within an organism is their cellular uptake. because of their size and negative charges, exogenous rnas cannot easily passively cross cell membranes (see below). however, earlier studies suggested that exogenous rnas, either homologous (shanmugam and bhargava, ; galand and ledoux, ; sirdeshmukh and bhargava, ) or heterologous (i.e. from e. coli, yeast or others) (natural academy of sciences, ; herrera et al., ; galand et al., ) , could be taken up by mammalian cells (bensch and king, ) . intact, rna may be present inside the cell but is reported to degrade rapidly in some studies (herrera et al., ; shanmugam and bhargava, ) . extracellular ribonuclease activity seems to inhibit uptake (shanmugam and bhargava, ) or function (niu et al., ) of exogenous rnas. uptake of exogenous rna is apparently not due to altered membrane permeability, impaired cell viability, or ribonuclease degradation of the macromolecule during incubation experiments (herrera et al., ) . radioautographic evidences (radioactive assays) have shown the uptake of macromolecular rnas by normal and neoplastic mouse cells, but degradation was also shown to occur prior to or after rna uptake (schwarz and rieke, ) . although intracellular degradation may occur, early studies also described biological actions in response to uptake of exogenous rnas (niu et al., ; niu et al., ; ; esposito, ) , suggesting that once taken up by a cell, rna could eventually exert a biological effect. studies on other types of cellular uptake of exogenous rnas, such as transfer rnas, either homologous or heterologous (depending on donor rna and recipient cell type) have been conducted (gallagher et al., ; sakai and cohen, ; crooke et al., ) . however, as noted in rna therapeutics development (see section . . . ), the large number of barriers encountered by rnas from intake to cell uptake needs to be considered when evaluating possible exogenous rna bioavailability (figure ). the gi tract offers a large surface area potentially relevant for ncrnas cellular uptake and following systemic release, since it contains a large variety of cell types at different maturational levels (e.g. stem cells in the crypts of lieberkühn) located only a few microns from an extensive capillary network (lozier et al., ) . however several barriers exist that could prevent ncrnas uptake by mammalian cells (o'neill et al., ) . for example, in a study conducted in neonatal pigs, no intestinal absorption of rna was observed (baintner and toth, ), suggesting possible rna degradation or an inability to cross the intestinal barrier. gastrointestinal barriers can be distinguished in extracellular and cellular barriers and are presented below. efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. orally ingested exogenous ncrna would have to pass different barriers in the human gi tract. for clarity, only the general barriers are shown. gi barriers to the uptake of orally ingested rnas. the first environment encountered by any orally ingested rna is the oral cavity, where lytic enzymes such as amylase or lipase are secreted in the saliva, and the ph ranges from . to . . the stomach is a more acidic environment (ph . - . ) (dressman et al., ) , and nucleic acids are known to be denatured and depurinated (hydrolysis of nucleic acids to release purine bases) over time in acidic gastric fluid (loretz et al., ) . the gastroenteric fluid flow and peristaltic activity in the gi tract could reduce the contact time between ncrnas and the epithelial layer, therefore diminishing the opportunities for cellular uptake, while nuclease enzymes present in the gi lumen could degrade nucleic acids before any cellular uptake (o'neill et al., ) . indeed, purines and pyrimidines in nucleic acids are believed to be absorbed mainly in the form of nucleosides (sonoda and tatibana, ) . the gut flora, predominant in the distal ileum and in the large intestine, produces a range of enzymes that could degrade ncrnas as well. the gi tract is lined by a viscous sticky layer of mucus entrapping foreign particles before reaching the underlying epithelium (figure ) . this "trapping" effect could be mediated by mucus composition, which confers a net negative charge to the mucous layer. since nucleic acids present a net negative charge as well, this mucus could represent an electrostatic barrier difficult to overcome by rnas, and particularly ncrna, if not properly delivered or chemically modified. furthermore, mucus turnover efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. (secretion, shedding and discard) happens in a relatively short time period ( - min) (lehr, ), preventing rnas from traversing the mucous layer and to reach the underlying epithelium. in conditions such as ulcerative colitis this mucus layer can be reduced or missing in areas of acute inflammation (pullan et al., ) , which would allow nucleic acids to reach the epithelial cell layer. another barrier is the glycocalyx, a glycoprotein and polysaccharide layer ( - nm thick) associated with the apical cellular membrane of enterocytes (frey et al., ) . this layer prevents the access of certain viruses, bacteria and particles (such as rnas) to the underlying plasma membrane by acting as a size-selective diffusional barrier. the intestinal mucosa is structured as a three-layer barrier: a single layer of epithelial cells, the lamina propria ( furthermore, the epithelial cells (enterocytes), which represent approximately % of the epithelium, have a short lifetime of - days, and are continuously shed and replaced (quastler and sherman, ; jung et al., ) . the enterocyte apical membranes are characterized by a high number of microvilli, structures approximately mm long and nm wide (johnson, ) . since endocytosis occurs primarily at the base of microvilli (jung et al., ) , particles with a diameter greater than nm could not be efficiently endocytosed (uduehi et al., ) . m-cells (cells dedicated to trans-cellular transport and associated to intestinal lymphoid follicles) are fewer in number than enterocytes, but, due to their role in the transport of foreign material from the lumen to the immune cells of the lamina propria, are characterized by a high endocytic activity. moreover, m-cells have microfolds in their apical membrane, and are covered by a reduced glycocalyx and mucous layer. this determines a low breakdown capacity (e.g. low alkaline phosphatase activity, and a lower number of lysosomes) so intracellular degradation of ncrnas could be spared. conversely the capillary network underlying the m-cells is less dense and less permeable (o'neill et al., ) . although the literature has described these cellular barriers when studying several molecules, there is little specific data (o'neill et al., ) on the relevance of these to exogenous rna molecules. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. assuming that ncrnas could overcome the gi barriers and reach the circulatory system, they would still encounter other biological barriers before reaching the appropriate machinery to exert any effects. significant degradation occurs of sirna duplexes (non chemically modified) in min when they are incubated in foetal calf or human sera (haupenthal et al., ; urban-klein et al., ) . sirna in human plasma is rapidly degraded, with nearly % degraded within min (layzer et al., ) . nuclease activity in plasma and tissues is relevant to degrading rnas, including ncrnas. the major activity in plasma is a ' exonuclease, but cleavage of internucleotide bonds can also take place (juliano et al., ) . encapsulation methods, which protect rnas from degradation, include exosomes, microvesicles and apoptotic bodies, all of which are extracellular vesicles distinguished by size, biogenesis and cargos (ela et al., ; van niel et al., ) . exosomes, for instance, contain various species of rna, conferring them protection against degradation and providing them a vehicle for cellular uptake of rnas by endocytosis (cui et al., ). the reticuloendothelial system (res) is comprised of phagocytic cells, including circulating monocytes and tissue macrophages, whose physiological function is to clear the body of foreign pathogens, remove cellular debris generated by tissue remodelling, and clear cells that have undergone apoptosis. additionally, the res plays an important role in uptake and clearance of individual "free" oligonucleotides. phagocytic cells of the res, particularly the abundant kupffer cells in the liver and splenic macrophages, express a number of cell surface receptors, including integrins and scavenger receptors, that are potentially involved in uptake, although the role of scavenger receptors in uptake of free oligonucleotides is somewhat controversial (juliano, ). following internalization of an oligonucleotide, the phagosome fuses with lysosomal compartments, where the contents are subjected to enzymatic degradation by proteases and hydrolases that operate efficiently in the low-ph lysosomal environment. tissues macrophages are most abundant in the liver and spleen, and both these tissues receive high blood flow (juliano et al., ) . the capillary lumen is surrounded by a layer of endothelial cells interlinked by adherence junctions (vecadherin containing) and by tight junctions (occluding and claudin-containing) and tightly adherent to the underlying extracellular matrix. this structure forms a barrier between blood and the parenchymal space. molecules in the blood can be transported across the endothelial barrier by two routes. paracellular transport occurs through the junctions between cells and is limited to molecules of approximately nm diameter or less; only micrornas would be able to transit this barrier by this method. caveolar-mediated transcytosis carries large proteins across the endothelium within vesicles of about nm. in some tissues, such as liver and spleen, there are gaps or fenestrations of - nm diameter between the endothelial cells, which allow egress of larger molecules (juliano, ). endothelial permeability is also increased in sites of inflammation and in some tumours, in which increased permeability is associated with fenestrations between the endothelial cells that result from rapid and disorganized angiogenesis in the tumour. this is known as the epr effect (enhanced permeability and retention), although some argue that this is mostly due to poor lymph flow in the area the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. and many minutes to several hours for the elimination phase; (ii) oligonucleotides are accumulated in most tissues, particularly in kidney and liver, but not in the central nervous system (probably due to the blood brain barrier); (iii) the major route of elimination is via the kidneys; and (iv) although the most detailed studies have been performed in rodents, the pharmacokinetic behaviour of the modified oligonucleotides in humans is similar to that observed in lower animals (juliano et al., ) . the pharmacokinetics and biodistribution of sirna duplexes is similar to that of single-stranded antisense molecules, with the highest uptake in the kidney followed by the liver (juliano et al., ). sirna and uncharged oligonucleotides do not bind extensively to plasma proteins, and are thus cleared by the kidney much more readily than modified oligonucleotides and tend to accumulate at lower levels in tissues (juliano, ) . typically, molecules with sizes of - nm or less can be ultrafiltered by the kidney; micrornas could therefore be rapidly excreted by the renal route, unlike long ncrnas. in fact, sirna molecules intravenously injected can be visualized in renal filtrate within seconds of injection (molitoris et al., ) . plasma sirna concentrations are reported to decline by more than % within min (relative to levels at min post dose) and decline by more than % within h (thompson et al., ) . only about - % of the iv administered dose is absorbed by tissues and the majority of this uptake occurs in the kidney (thompson et al., ) , most of it being cleared from tissues within h of administration . to reach their target, ncrnas must enter mammalian cells and arrive at the appropriate subcellular location (the nucleus or the cytoplasm, depending where the target molecules are located). like other large, polar and charged biological macromolecules, ncrnas are internalized by endocytosis and then trafficked through multiple membrane-bound intracellular compartments. the ability of ncrnas to reach their targets depends on both cellular internalisation (uptake) and intracellular trafficking. (figure ). there are five major classes of endocytosis: (i) the clathrin-coated pit pathway; (ii) the caveolar pathway; (iii) the noncaveolar, clathrin-independent pathways (clic pathways); (iv) phagocytosis (mainly taking place in "professional phagocytes" such as macrophages and granulocytes); and (v) macropinocytosis (in which internalized macromolecules are simply dissolved in the ambient medium) (juliano et al., ). non-coding rnas (ncrnas) can enter cells via different endocytic pathways, including micropinocytosis. internalized ncrnas can traffic from early endosomes (ee) to late endosomes (le) and to lysosomes. ncrnas must escape endosomal organelles to reach the cytosol and nucleus and act on the target molecules. some proteins may direct ncrna to non-productive pathways. the micropinocytosis pathway may represent a non-productive pathway for naked ncrnas. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the first four classes often involve a cell surface receptor and are collectively termed receptor-mediated endocytosis. these pathways are mainly utilized in cellular delivery of oligonucleotides. in general, receptor-mediated endocytosis includes three major steps. (i) receptor binding and internalization represents the primary barrier for oligonucleotide transport; the ligand-receptor binding determines which target cells and tissues oligonucleotides are delivered to. (ii) sequential intracellular trafficking leads oligonucleotides into a variety of low ph endomembrane compartments, including early/sorting endosomes, late endosomes/multivesicular bodies, and lysosomes. in some cases, receptors/ligands can traffic to the golgi complex. in many instances, receptor and ligand are dissociated in the low ph endosome environment. vesicular trafficking can prevent ncrnas from reaching their targets, for example, by sorting to secretory or lysosomal vesicles which may lead to export of ncrnas out of cells or degradation in the lysosomes. (iii) ncrnas must exit from the endosome to reach the site of action in the cytoplasm or nucleus. endosomal trapping represents an important barrier for ncrnas final functional objective. on the other hand, oligonucleotides are able to continuously shuttle between the nucleus and the cytoplasm mediated by nuclear pore structures, and do not require classical nuclear localization signals (juliano et al., ). the available data clearly describe several barriers to be considered when evaluating exogenous rna absorption, circulation, cellular uptake or intracellular trafficking. exogenous rna must overcome numerous biological barriers to reach its intended target within mammalian cells. considering oral administration as the most relevant for exogenous plant ncrnas, the first major limiting step to exogenous rna bioavailability is the gi tract itself. the gi tract encompasses both extracellular and cellular barriers. the extracellular barriers include the presence of enzymes in the lumen, such as amylase or nucleases, a harsh environment with ph ranging from . to . , and a net negative charged mucous layer with a very rapid turnover ( - min). the cellular barriers include a single layer of epithelial cells, the lamina propria and the muscularis mucosa, constituting the threelayer intestinal mucosa barrier. ncrnas could cross the epithelium between cells but this is limited by the presence of tight junctions. pore size in the human intestine would prevent passage of all ncrnas other than mirnas, the smallest ncrnas. ncrnas could also cross this epithelium through transcytosis by traversing the cells; this mechanism implies exposure to intracellular nucleases, recycling of ncrnas back to the lumen and nuclear uptake. many biological barriers and environmental conditions between the gi epithelium and the target tissue are then encountered by ingested rnas. ncrnas would be exposed to nucleases both in the plasma and in the target tissues. once inside the circulatory system, these rnas would be subjected to distribution and elimination, accumulating mostly in the liver and kidney. due to their small size, mirnas are especially rapidly cleared by the kidney, unlike long ncrnas. therefore, a very small percentage of rnas could actually be absorbed by tissues. furthermore, ncrnas would need to escape the reticuloendothelial system, the function of which is to clear the organism of foreign molecules. then rnas must cross the vascular endothelial barrier to reach the target tissue. for ncrnas to exert their functions they must enter the target cells. rna cellular uptake is achieved by endocytosis after which they would enter the intracellular trafficking system through multiple membrane-bound compartments. ncrnas would need to exit these compartments to reach their functional sublocation within the cell, be it the cytosol or the nucleus, while simultaneously escape degradation in the lysosomes. efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. seeds, corn kernels and rice grains, all of which are common food and feed, and numerous endogenous plant srnas are reported to have perfect complementarity to genes in humans, and other animals . however studies in mammals and humans with sirna indicates that the exposure levels required to produce regulatory effects exceed the levels achieved following ingestion . all animal and plant-related foods and feeds contain naturally occurring coding (i.e. mrnas) and ncrnas. early (srivastava, ; lassek and montag, ) and more recent studies ) have estimated that the total rna content in plants is about mg/g plant tissue. plants contain an array of ncrnas, including highly abundant transfer rnas (trnas) and ribosomal rnas (rrnas), single-stranded antisense rnas, and the mirnas and sirnas that trigger rnai and their precursor dsrnas . long dsrnas from non-plant, exogenous sources are particularly common in plants, including plants used for food and feed, due to infection from rna-containing viruses. animalderived foods are generally richer in rna than plant-derived foods, and are likely to contribute significantly to overall rna consumption (jonas et al., ) . of the mg/g tissue of total rna content in plants, relative percentages (by weight) of the major rna forms are approximately % rrna, - % mrna, and - % trna, with srnas accounting for less than % of total rna in plants . srnas are present at levels of up to . µg/g of conventional soybean grain (average = . µg/g grain) and comparable amounts are present in the grains of conventional corn and rice . in tobacco plants engineered to overexpress a dsrna under the control of a constitutive promoter, sirnas levels in leaves reached about . % of total rna (chau, b. l. and lee, k. a., ) . total exposure to construct-derived srnas from a putative biotech soybean product was estimated to be µg/kg/day in the general population and µg/kg/day for children aged six and under . using these same assumptions for rna expression levels in sweet corn, consumption of corn at the highest possible rate would result in an estimated intake of µg/kg/day in the general population and µg/kg/day for children aged in the absence of encapsulation or chemical stabilization to prevent degradation or without the addition of penetration enhancers, the absorption of rna -including sirnas -across the gi tract is described as negligible (akhtar, ; jain, ) . one study suggests that activity could be possible for certain highly expressed plant mirnas following food intake (zhang et al., a) . this study reports that in mice fed a diet consisting entirely of cooked rice (i.e. human equivalent of about kg/day of cooked rice) , several rice mirnas were detectable in mouse serum and liver. some of the results presented in this study were contested in later studies (chen et al., ) . petrick et al. clearly stated the unlikelihood of achieving such high concentrations of mature plant mirnas in the serum, plasma and organs of humans or animals via food intake since the high levels of rice administered the experimental animals did not reflect anticipated dietary exposure levels. in another study, it was concluded that plant mirnas identified in animal srna sequencing data can originate from artefacts of the sequencing process (zhang et al., b) . even in the case of -mer oligonucleotides modified to increase their stability, oral bioavailability in rats was only . % (nicklin et al., ) and intestinal absorption was less than % in a model that bypasses the gastric acidic environment. most of the labelled oligonucleotide was associated with the luminal epithelial cell membrane and very little efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. was localized intracellularly (khatsenko et al., ) . regarding the possibility of dietary sirnas and other dsrnas effects on gastrointestinal tissues, studies with radiolabelled or immunostained dna oligonucleotides demonstrate that these are primarily located extracellularly within the intestinal tract (lumen and luminal wall) (khatsenko et al., ; nicklin et al., ) , and are thus presumed to be minimally absorbed (negligible bioavailability, < %). it has also been observed that absorption decreased as the oligonucleotide length increased (from to nt) (khatsenko et al., ) . the only published quantitative data on srnas content in plant-derived foods ) (detailed above) indicate that . µg of srna is obtained per gram of soybean, rice seeds or corn kernels. as total rna content in plant-derived foods appears to be around mg/g tissue (lassek and montag, ), srnas could therefore represent . - . % of plant seed materials. the amount of sirnas in plants genetically modified to carry hairpin transgenes is estimated at about ng sirna/ µg total rna/g tobacco leaf tissue (chau, bess l. and lee, kevin aw, ) . the resulting estimated human dietary exposure to plant small rnas from rna-based biotech crops was derived from this (table ). . (b) assuming % of the specific food is consumed from a biotech crop, with total rna levels of ≈ mg/g grain and . % of these small rnas derived from the transgene. for details see . these doses are significantly lower than those required to elicit adverse effects in rodents or monkeys ( or mg/kg of chemically-stabilized sirna, respectively) by iv administration (thompson et al., ) . moreover, high mirna concentration is required to target suppression, and levels below copies/cell mirnas have little regulatory capacity (mullokandov et al., ) . according to snow et al., dietary plant mirnas are present at less than one copy per cell in target organs assuming reliable quantification . in another study the calculated dietary exposure in humans of a specific dsrna (dvsnf ) was estimated to be around . x - µg/kg/day in the us population (petrick et al., b) . the same study reported that mg dvsnf /kg/day in a -day repeated dose oral (gavage) toxicity study in mice did not produce any toxic effects (petrick et al., a) . levels for this specific dsrna in this biotech crop were reported to be ≈ . x - µg/g grain tissue on a dry weight basis for corn food, and ≈ . x - µg/g in the whole plant on a dry weight basis . of note is that there is little information on the impact of intended rnai in gm plants on the coding and non-conding plant rna. knowledge of which other rnas are modified, or how other rnas are compensated or desregulated in the gm plant needs to be experimentally evaluated for each specific case. different studies have evaluated the amount of plant-derived foods consumed by the general population. in a study of cohort of women in the uk, the difference between the mean intake of overall fruit and vegetables consumption was three times greater in high consumers vs. low consumers (pollard et al., ) ; vegans and vegetarians were the highest consumers of fruit and vegetables. in an efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. evaluation of the flemish population it was observed that vegans, while having the lowest total energy intake compared to omnivores, have the highest total fruit and total vegetables consumption. the intake of fruit, nuts and/or legume about twice in vegans, using either the healthy eating index (clarys et al., ) or total mean intake (clarys et al., ) . in the epic cohort study, which includes ten european countries, mean daily consumption was between and g/day vegetables and between and g/day fruit (agudo et al., ) . although not homogeneous among countries and centres, the ratio between the highest and lowest consumers was around . for vegetables and for fruit (agudo et al., ) . countries in southern europe had the highest vegetables and fruit consumption. for example, consumption of vegetables and fruit in healthy adults in spain (epic data), considered higher than most european countries and the usa, was g ( . servings) of vegetables and g ( . servings) of fruit (agudo et al., ) , equal to ≈ g/d vegetables and fruit. in the united states (us) the total vegetable and fruit intake has increased slightly (up to ≈ . servings/day) in all consumer categories from the late s (krebs-smith and kantor, ) and remained stable over the last decade (rehm et al., ) . actual quantities, however, are around . servings/day (rehm et al., ) , that is . for fruit and . for vegetables (eaton et al., ) . in cup-equivalents, this is . cup equivalents for vegetables and . cup equivalent for fruit (moore and thompson, ) . mean total vegetable intake in the us for vegetarians is reported to be around g/day and that for non-vegetarians as g/day. total fruit intake was around g/day for vegetarians and g/day for non-vegetarians (haddad and tanzman, ) . in other words, there is less than two-fold difference in the amount of fruit and vegetables consumed between vegetarians and non-vegetarians in the general us population. in a specific population (i.e. the adventist health study) in the us, daily mean consumption of fruits was higher (≈ g/day) for vegans than for other types of vegetarians or non-vegetarians (≈ g/day). the same trend was observed for daily mean consumption of fruit which was higher in vegans (≈ g/day) than other vegetarians or non-vegetarians (≈ g/day) (orlich et al., ). based on these data, dietary exposure to rnas from plants can in general be assumed to be higher in vegans and vegetarians than omnivores. although total rna content in plant-derived foods varies (lassek and montag, ), a person consuming an estimated daily dose of total fruit and vegetables ≈ g/day would theoretically ingest mg of total rnas assuming about mg/g of plant tissue. this value agrees with the dietary rna intake, which typically ranges from . - g/person/day (jain, ) . considering that srnas represent up to . % of plant seed materials ), estimated daily intake in the general population would be around . mg srnas. since vegans and vegetarians normally consume higher amounts of vegetables and fruits (see above), they could be exposed to an average of three times more plant exogenous rna ( . mg/day). no pharmacokinetics studies were identified in these groups. changes in rnase activity can occur in physiological or pathological conditions, this possibly impacting exposure after dietary intake. for instance, increased serum rnase activity has been observed in pancreatic carcinoma, pancreatitis, and renal failure (peterson, the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. et al., ) . human argonaute (ago- ) is a catalytic core component of rnai machinery. ago- mediated gene silencing has been proposed to be linked to the mitogen activated proteine kinase signalling pathway (zeng et al., ) , which is activated in response to cellular stress. since various kinds of stress on human cells induce formation of stress granules, ago- can be recruited to these granules, modifying its intracellular localization and the efficacy of rnai activity (detzer et al., ) . ago- is thus regulated at both the transcriptional and post-translational levels, suggesting that ago- levels can influence cellular mirna activity (adams et al., ). it has also been proposed that inactivation of mitochondria could lead to a strong decrease in mirna-mediated rnai efficiency, and, to a lesser extent, to sirna-mediated rnai (huang et al., ) . due to the interaction of p-bodies with mitochondria, reduced ago- activity could be due to changes in location of endogenous ago- from pbodies (huang et al., ) . association of ago- with cytoplasmic rna granules is known to regulate the translational repression activity of the protein, but phosphorylation of certain residues may prevent this inhibition, allowing the protein to remain active in the cytoplasm (lopez-orozco et al., ) . overall, it seems clear that ago- activity can be modified by cellular stress conditions. however, whether this can influence the function or the exposure to dietary ncrnas remains unknown and would benefit from further research. most studies have been focused on mirnas since these ncrnas were the first to be described. these studies have consequently extended to transgenic dsrnas as these have increasingly used to induce gene silencing. as mentioned previously, little is known about dietary exposure to other ncrnas. given the role of lncrnas in plant physiology, it is important to better understand human and animal exposure to these ncrnas. further research is needed on how transgenic ncrnas affect expression levels of other ncrnas, and other rnas such as mrnas. alteration of the expression levels of certain ncrnas within the plant may modify the levels of other ncrnas due to putative compensatory circuits and codifying rnas. this in turn could lead to changes in protein and enzymatic content, with putative consequent alteration of the modified plant's nutritional value. controlling for these changes would require complete transcriptome sequencing, and comparison of rna levels in unmodified to those of the altered plant . another point to consider is whether special diets (e.g. vegetarian or vegan) modulate ncrnas uptake throughout the gi tract, possibly leading to increased exposure. humans and animals have been continuously exposed to naturally-occurring plant rnas. estimated amounts of ingested plant ncrnas contrast with the much higher doses administered to experimental animals and in clinical trials to humans. although some reports on the presence of plant mirnas in serum, plasma and organs of humans or animals do not agree, the extremely low reported concentrations prevent them from being functional, even in the case of rnas modified artificially to augment their bioavailability. humans following special diets, such as vegetarians and vegans, have high intake of plant rnas, but their increased estimated exposure to such rnas is barely on average fold greater than in omnivorous diets. there are no specific studies determining the effects of increased dietary exposure to plant ncrnas. adams bd, claffey kp, white ba. . argonaute- expression is regulated by epidermal growth factor receptor and mitogen-activated protein kinase signaling and correlates with a transformed phenotype in breast cancer cells. endocrinology , - . efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. to exert a biological effect, exogenous ncrnas need to overcome the physical and biological barriers present in both the gi tract and the circulatory system, reach the target tissue, and enter the appropriate intracellular pathway at sufficient dose. this section describes the obstacles and challenges encountered by exogenous ncrnas following oral intake. following an extensive literature search, and based on expert judgement in the topic, publications were reviewed for this section. rna molecules are large, hydrophilic and negatively charged. even the smallest rna molecules (mirnas) have a high molecular weight (∼ kd) and their net charge is negative (akhtar and benter, a; akhtar and benter, b) . due to these physicochemical properties, rna molecules are highly water soluble with low membrane permeability, thus falling into class iii of the biopharmaceutical classification system (fda, ; amidon et al., ) . like many other class iii compounds, they are not absorbed at all or only to a very small degree. being highly charged species, rna molecules resist partitioning across lipid bilayers, and higher molecular weights effectively restrict their movement by function of the tight junctions present in the gi tract epithelium (tillman et al., ) . in vitro and in vivo preclinical studies have assessed the use of medium chain fatty acids ( - carbon atoms), as well as bile salts, as absorption enhancers to cross the intestinal mucosa (gonzalez ferreiro et al., ; tillman et al., ; raoof et al., ) . orally introduced rna molecules encounter relevant obstacles in the gi tract, these precluding their absorption and activity. in general oligonucleotides are rapidly degraded in the harsh biological milieu of the acidic stomach and enzyme-rich gi tract, as described previously, and show poor transcytosis across the gut (akhtar, ) . in humans the ph in the gi lumen can vary from in the stomach to in the intestine (gamboa and leong, ) . exposure to these ph values can cause ph-induced oxidation or de-amination of rna, resulting in loss of activity (pridgen et al., ) . in addition, nucleases present in the gi lumen for digestion of biological molecules enzymatically degrade rna molecules too (gamboa and leong, ; kriegel et al., ) . the presence of a rich bacterial population, mostly in the intestines (the human microbiome project consortium, ) may also contribute to degradation of these molecules, as bacterial degradation of ribonucleic acids through the secretion of extracellular nucleases has been early described (nishimura, ; eaves and jeffries, ) . finally, if the above-mentioned obstacles are overcome, rna molecules still face the intestinal extrinsic and intrinsic barriers (see also . . ). the extrinsic barrier consists of the mucus layer covering the epithelial cells. this is a complex hydrogel material composed of proteins, carbohydrates, lipids, salts, antibodies, bacteria, and cellular debris (o'neill et al., ; pridgen et al., ) . it consists of loosely and firmly adherent layers that vary in thickness along the gi tract and can fluctuate based on diet. the mucus is secreted by cells presenting a rapid cell turnover, approximately - days in the small intestine in humans (gamboa and leong, ; o'neill et al., ; atuma et al., ) . among other functions, the mucus layer protects www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. epithelial surfaces by trapping pathogens and foreign particulates, and rapidly clearing them. penetration of this mucus barrier is necessary to reach the absorptive epithelial cells. several mucuspenetrating and mucoadhesive materials can be used to enable penetration of molecule to the mucus layers, increasing residence time and contact of the delivered molecules to the epithelium (pridgen et al., ) . the intrinsic barrier consists of the epithelial cell monolayer, constituting the greatest barrier to material from the intestinal lumen to the lamina propria and bloodstream. cells maintain this barrier by forming tight junctions, whose permeability can be modulated through specific combinations of different proteins (o'neill et al., ) . there are several possible pathways across the intrinsic (epithelial) barrier (figure ).  the transcellular pathway passes through the apical and basolateral cell membranes, as well as the cytoplasm. this pathway is very restrictive to the passive flow of hydrophilic solutes such as rna molecules, because of the lipid bilayer membrane and its impermeability to large molecules. transport mechanisms for this pathway can be passive for hydrophobic molecules, or active with membrane pumps for specific molecules such as ions.  the paracellular pathway is the major passive permeation pathway and allows diffusion of small molecules in the space between epithelial cells. the tight junctions regulate permeability of this pathway based on molecules' size and charge.  finally, transcytosis is an active transport pathway that relies on molecule-specific receptors guiding the molecule through the cell without entering degradation pathways. because of their large hydrodynamic size, macromolecules such as rnas are restricted to this pathway (pridgen et al., ) . the m cell transcytosis pathway is the most extensively studied for oral delivery of rnas. this pathway, which is used to transport antigens across the epithelium for immune surveillance, is attractive because m cells lack mucus secretion and have a sparse glycocalyx, among other features. however m cells are closely associated with immune cells in the lamina propria and peyer's patches, such as dendritic cells and macrophages, this from one side limiting the ability of the delivered molecule to reach the bloodstream and on the other side increasing the risk of triggering an immune response (florence, ; kriegel et al., ; pridgen et al., ) . moreover, m cells only make up a small percentage ( - %) of the non-absorptive epithelium in humans. the surface properties of m cells, as well as the number of peyer's patches vary among species, which could make difficult extrapolating animal model data to humans (pridgen et al., ) . an additional obstacle is represented by the immune system, which is intimately associated with the epithelium. numerous types of immune cells patrol the lamina propria, including t cells, macrophages and dendritic cells. if the ingested rna molecules reach the bloodstream, they must also evade the mononuclear phagocytic system (reischl and zimmer, ; pridgen et al., ) . the presence of endogenous and luminal nucleases, together with gi ph conditions, in general determine rapid degradation and low rna molecules' biological half-life (bhavsar and amiji, ) the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the paracellular pathway allows diffusion of molecules in the space between epithelial cells and is regulated by intercellular tight junctions. the transcellular pathway passes through the apical and basolateral cell membranes and cytoplasm. it is restricted to hydrophobic molecules or molecules transported by membrane pumps. the transcytosis pathway is an active transport system that relies on molecule-specific receptors guiding the molecule through the cell escaping degradation pathways. transcytosis pathways are found in both epithelial and m cells. from pridgen et al, . transepithelial transport: paracellular, transcellular and transcytosis pathways. first-pass elimination occurs when a compound is metabolised between its site of administration and the site of sampling for drug concentration measurement. this greatly affects the bioavailability of orally administered compounds since the concentration of the active substance reaching systemic circulation is decreased. the liver is usually assumed to be the major site of first-pass metabolism of an orally administered drug, but other sites are the gi tract, blood, vascular endothelium and lungs (pond and tozer, ) . in fact, the obstacles previously described for rna molecule absorption in the gi tract can be considered a first-pass effect. rna molecules reaching the circulatory system must avoid filtration by the kidneys, accumulation in non-target reticuloendothelial system (res) of the liver, kidney, lungs and spleen, degradation by endogenous nucleases, aggregation with proteins in the serum, and uptake by non-targeted cells such as phagocytes (kriegel et al., ; reischl and zimmer, ). once cellular uptake has taken place in the potential target tissue (surmounting the obstacle of crossing the vascular walls and the lipid bilayer cellular membranes), nucleic acids have to undergo efficient intracellular trafficking to ultimately produce the intended pharmacological effects. this includes endosomal transportation (escaping endolysosomal degradation), cytoplasm release, and efficient incorporation into the relevant intracellular pathways such as risc loading (for sirna, mrna, and mirna) or nuclear uptake (for lncrna) kriegel et al., ; pouton and seymour, ) . www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. rna molecules are large, hydrophilic and negatively-charged molecules. to achieve effect via the oral route, these molecules need to overcome the physical and biological barriers present in both the gi tract, where they encounter variable and harsh conditions along with degrading nucleases, and the circulatory system. rna molecules must reach the target tissue, escaping endo-lysosomal degradation, and enter the appropriate intracellular pathway in sufficient doses to exert their effects. www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. following a literature search as described in section . . . . and based on the methodology described in section . . . , a total of documents were selected as relevant to a general review of rna-based therapeutics administered by gi route. of these documents, at least report the use of specific vehicles for gi administration of exogenous ncrnas (i.e. nanoparticles). six of these documents simultaneously evaluated formulated exogenous rnas and their naked version, and reported either rapid degradation under gi conditions, or reduced or absence of biological effect of the naked version compared to the formulated one (table ) . twenty ( ) of these documents provide specific examples of local effects of rna-based drugs administered orally. in addition, papers were selected as relevant to inform on administration of exogenous ncrnas in either fish or birds and possibly exerting biological effects. orally delivered rna has been used as a dietary supplement or empirical therapeutic agent since s (jain, ). for example, yeast rna supplementation as a source of dietary ribonucleic acids has been used in different animals (choudhury et al., ; jha et al., ) including mammals (sukumar et al., ; heaf and davies, ) and in human studies (gianotti et al., ; tepaske et al., ) . in human studies, up to g per day (in divided doses) of exogenous rnas have been administered (for - days), producing an increase in plasmatic uric acid levels (zollner and grobner, ; zollner, ) . the absence of attempts to study the gi absorption of pure rna in these studies suggests that its effects, if any, could be related to rna molecules as a source of nucleotides. indeed, nucleotide supplementation in rats has been shown to affect the healing of ulcerative conditions, ameliorating indomethacin-induced ileitis and aggravating the severity of dextran sulfate sodium (dss)-induced colitis (sukumar et al., , sukumar et al., . also, a study of body fluid composition of rats orally administered yeast rna, mixtures of its constituent nucleosides or its constituent bases and ribose was performed (heaf and davies, ) . the authors showed that ingestion of rna increased intestinal levels of ribose, inorganic phosphate, uridine, pseudouridine, uracil, inosine or uric acid. the effect of orally administered mixed nucleosides on blood and urine composition was similar to that of rna, while some differences were noted for some equivalent mixture of free bases (heaf and davies, ) . this study showed that the dietary rna-phosphate passed to the urine from the gut, suggesting that most of the rna-ribose was probably metabolized (heaf and davies, ) . it has been shown in a murine model of staphylococcus aureus infection that the oral administration of rna was ineffective while the intraperitoneal nucleoside-nucleotide mixture was more effective in maintaining host resistance against bacterial infection (adjei et al., ) . supplementation with nucleotides has also been shown to exert biological effects, including expression regulation of certain genes (sanchez-pozo and gil, ; gil, ) , although the mechanisms of action are still unknown. it is generally assumed that nucleotides are not essential nutrients, but under certain conditions (i.e. low dietary intake, tissue needs increased or stress) dietary nucleotides may play a role as conditionally essential nutrients (jung and batal, ; carver and walker, ) . in this context, dietary supplementation with yeast rna promoted ulcer healing in a rat model of indomethacin-induced ulcerative ileitis (sukumar et al., ) . similar effects were observed when nucleosides and nucleotides were administered intravenously (veerabagu et al., ) . in both cases, the possible mechanism of action was partly due to increased cell proliferation in the damaged tissue (sukumar et al., ; veerabagu et al., ) . similar effects were observed when rna was administered orally or its constituents (nucleotides) were injected iv, suggesting that the effects are due to the absorbed nucleotides. dietary nucleotide supplementation in formula-fed infants efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. has been shown to improve gut microbiota composition (singhal et al., ) . in studies on certain fish model, supplementation with dietary rna at a range of concentrations ( . - . % of nucleic acids in the diet compared to the non-suplemented control group) enhances immunological response (choudhury et al., ; jha et al., ) . in human studies, postoperative enteral diet supplementation with rna, arginine and omega- fatty acids has been shown to modulate the postoperative immune response after surgery for upper gastrointestinal cancer (senkal et al., ) , overcoming more rapidly the immunologic depression after surgical trauma (kemen et al., ) . also in humans, oral supplementation with rna (from yeast) during days before surgery improved host defence in patients undergoing cardiac surgery (tepaske et al., ) or gi cancer surgery (gianotti et al., ) . in both cases, rna supplementation was also accompanied with arginine and omega- fatty acids administration. in summary, the specific contribution of orally administered rna to these effects cannot be ascertained. the instability of orally administered rna molecules and the presence of a large array of rnaases within the gi tract (see sections . . and . . ) suggest low absorption, if any. exposure to a highly active enzymatic environment in the gi tract, extreme ph conditions, and the existence of a mucosal epithelial barrier are the main challenges for oral delivery of rna therapeutics (martirosyan et al., ) . although a considerable amount of food rna is ingested, which varies widely between individuals but is typically in the range . - g/person/day, it is assumed it is degraded and absorbed in minimal amount (jain, ) . however, further data are required to document the extent to which this conclusion pertains to rna molecules with complex structures. to overcome the multiple barriers encountered by exogenous rnas for oral delivery, several strategies in the laboratory or clinical trials have been developed (see . . . below). the oral administration route is advantageous because it increases patient compliance and comfort over injection, provides for simple, repeatable administration, and offers a large surface area for absorption (forbes and peppas, ) . rna delivery may also benefit from advances in the oral delivery of aso for animal (raoof et al., ; raoof et al., ; . indeed, polymeric nanoparticles containing alginate, dextran, chitosan, polyethylenimine, polyethylene glycol, polylactide, yeast derived β-glucans and/or several other natural or chemically synthesized molecules have been tested by the pharmaceutical/medicine field. the literature describes several strategies for the oral delivery of rna oligonucleotides. the target is to stabilize and protect rnas during gi transit, enable cellular uptake in the intestine, and in some cases, promote endosomal escape and increase biological action. thioketal nanoparticles (tkns) were formulated from a polymer, poly-( , -phenyleneacetone dimethylene thioketal) that degrades selectively in response to reactive oxygen species ( the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. (plga) nanoparticles were also tested in the same study, but with no efficacy in diminishing tnfα expression. to obtain efficient mucus transportation, targeted cellular uptake and endosomal/lysosomal escape, different nanoparticles have been tested ( nanoparticles of galactosylated trimethyl chitosan-cysteine containing sirna against map k were tested and found to target activated macrophages in colonic tissue (zhang et al., b) . compared to nanoparticle-protected sirnas, naked sirnas were completely degraded by the intestinal fluids. when administered orally ( µg sirna/kg) for days, nanoparticles improved the dss-induced model of colitis in mice (zhang et al., b) . other ternary polymeric nanoparticles formed by thiolated trimethyl chitosan with tripolyphosphate have been tested in mice and found to efficiently deliver sirnas to the intestine and other tissues when administered orally (zhang et al., a the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. ( %, % and %) were also evaluated in trimethyl chitosan-cysteine conjugate nanoparticles . in a mouse model of ulcerative colitis, the mannose density of % was the most effective for sirna knockdown of tnfα following oral administration ( µg/kg) of '-o-methyl-modified tnfαspecific sirna duplex . the use of supramolecular self-assembly nanoparticles (ssnps) -which contain oleyl trimethyl chitosan, poly(γ-( -(((piperidin- -yl)ethyl)amino)methyl)benzyl-lglutamate), oleyl-peg-mannose, oleyl-peg-cysteamine, and sodium tripolyphosphate -with specific functions of mucoadhesion, transepithelial permeation, membrane penetration and active targeting have also been tested ( however, not all nanoparticle types can efficiently deliver rnas. in a study evaluating the ability of unassisted epithelial entry of nucleic acids as a consequence of nanocarrier contact with mucus, antisense oligonucleotides but not sirna (naked) were shown to be delivered to mouse intestine when formulated with nanocarriers composed by chitosan (martirosyan et al., ). multi-compartmental formulations -consisting of one or more internal compartments surrounded by protective external compartments -have been developed to surmount the many barriers to oral rna delivery (kriegel et al., ; . one example is the "nanoparticles in the microspheres oral system" (nimos), a solid-in-solid multi-compartmental system (kriegel et al., ) . gelatin nanoparticles entrapped in poly(epsilon-caprolactone) microspheres were prepared carrying tnfα sirna (slightly chemically modified dtdt) to treat dss-induced colitis. oral administration of sirna ( . mg/kg) to female balb/c mice decreased colonic tnfα expression and supressed the expression of proinflammatory cytokines (kriegel, c and amiji, m, ) . when combined with sirna against cyclin d (naked sirna, also at . mg/kg/day), the silencing effects were more potent than with tnfα sirna alone (kriegel c and amiji mm, ) . dectin- recognizes beta- , and beta- , linked glucans rich particles and intact yeast and is particularly expressed on the monocyte/macrophage and neutrophil lineage (herre et al., ) . β- , -d-glucan has been used to deliver exogenous rna targeting macrophages (aouadi et al., ). delivery of β- , -d-glucan sirna particles containing µg/kg unmodified sirna by oral gavage to mice for days reduced map k expression in different macrophages from different tissues and protected them from lipopolysaccharide-induced lethality (aouadi et al., the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the cd gene was the target of the antigen-presenting dendritic cells and oral administration of the recombinant yeast. short hairpin rna delivered in lactobacillus casei have also been tested to target the intestine (kuwahara et al., ) . other organisms, including attenuated salmonella typhi, were used as vectors to deliver rnas, even ribozymes (bai et al., ) or sirna (jiang et al., ) , using oral administration in mice to target either local or systemic effects (bai et al., ; jiang et al., ) . the the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. extracellular vesicles are a heterogeneous family of vesicles delimited by membranes. these can be classified by their size as i) exosomes ( - nm in diameter), which originate from the endosomal system, ii) microvesicles ( - nm) formed by budding out of the plasma membrane, and iii) apoptotic bodies (> nm) formed by blebbing of the plasma membrane during apoptosis (reviewed in yanez-mo et al., ; colombo et al., ) . increasing evidence suggests that extracellular vesiclemediated communication can take place in vivo, but the nature of the extracellular vesicles involved in these effects remains to be clarified (reviewed in tkach and thery, ; pitt et al., ) . exosomes, which are formed in multivesicular compartments, are secreted when these compartments fuse with the plasma membrane (kowal et al., ; abels and breakefield, ) . exosomes have been shown to contain or transport a myriad of molecules including proteins, carbohydrates, lipids, and a variety of genetic material including dna, mrna and ncrnas (reviewed in choi et al., ; abels and breakefield, ) . exosomes are also described as participating in intercellular communication (costa-silva et al., ; pironti et al., ; nojima et al., ) , delivering their parental cell-derived molecular cargo to a recipient cell. some of these processes are thought to be mediated by ncrnas, including mirnas thomou et al., ) . detailed information on how all these biological processes occur is outside the scope of this review. given the above characteristics of exosomes, these vesicles hold promise as delivery vehicles for therapeutics (munagala et al., ; van der meel et al., ) . indeed, exosomes have been used to deliver sirna to different tissues when administered systemically, including the brain (alvarez-erviti et al., ; cooper et al., ; didiot et al., ) . mirnas have also been delivered via exosomes to different tissues including xenograft breast cancer tissue (ohno et al., ), brain and other tissues when administered systemically or locally (zhang, d et al., ) . interestingly, oral administration of exosomes or extracellular vesicles has also been described in animal models agrawal et al., ; oliveira et al., ; oliveira et al., ) . however, their biological effects were not related to transport of ncrnas. some extracellular vesicles have been shown to resist digestion under in vitro simulated gi conditions (benmoussa et al., ; , particularly those from bovine milk (vashisht et al., ) . moreover, milk exosomes seem to exhibit cross-species tolerance and are not described to induce adverse immune and inflammatory responses (munagala et al., ) . however, whether exosomes or other extracellular vesicles resist the harsh in vivo conditions of the gi tract and the digestive process remains poorly described and needs to be studied (tomé-carneiro et al., ) . exosome-like nanoparticles from plants have been isolated from different species including ginger, grape, grapefruit and carrot (mu et al., ; ju et al., ; zhang et al., ) . some authors indicated that the approximate amount of exosome-like nanoparticles in these edible plants were reported to be ≈ mg/ g edible plant (mu et al., ) , while other authors reported only ≈ mg/kg ginger . most of these studies reported the presence of rnas, either small or large, including ncrnas (mu et al., ; zhang et al., ; ju et al., ) . the amount of rna present in these exosome-like nanoparticles was reported to be ≈ µg rna/ mg exosomelike nanoparticles for grape and grapefruit, and ≈ µg rna/ mg exosome-like nanoparticles for ginger and carrot (mu et al., ) . the in vivo biological effects of these nanovesicles have been assessed. oral delivery of ginger nanovesicles during days at a dose of . mg/day was found to target the colon, to be taken up by epithelial cells and macrophages, and to reduce dss-induced colitis in mice . treatment for weeks with the same dose also exhibited antiinflammatory activity in a mouse model of chronic colitis . ginger-derived lipid vehicles have also been generated from ginger lipids and loaded with sirna-cd (zhang m et al., www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. ). oral administration of these ginger-derived lipid vehicles targets the colon tissue and they are effective in treatment of induced ulcerative colitis in mice. grapefruit exosomes at a dose of mg/kg per day for days also ameliorated the dss-induced colitis targeting intestinal macrophages . when administered orally at a dose of mg of nanovesicles/day, grape exosomes-like nanoparticles were also found to protect mice from a dssinduced model of colitis by induction of intestinal stem cells (ju et al., ) . local intestinal macrophages and stem cells from the small and large intestine were also found to be the target of these exosome-like nanoparticles when administered by oral gavage ( mg/day) (mu et al., ) . even though mirnas were reported to be present in these exosome-like nanovesicles (ju et al., ; mu et al., ; zhang et al., ) , the biological effects of these ncrnas were not evaluated. whether these mirnas resist the gi tract conditions remains unknown. a main hurdle in rna-based therapeutics is the successful delivery of naked rna (non-chemically modified) molecules to specific (target) tissues in the gi tract. the literature contains few studies in which exogenous rnas are administered in vivo using routes of administration other than oral, and most are in the field of inflammatory bowel disease. an amphiphilic cationic cyclodextrin (cd) vector was developed for a complex sirna against tnfα, where the complex was concentrated to µg/ µl in % glucose (mccarthy et al., ) . cd.tnfα.sirna was found to be efficient against dss-induced colitis in c bl/ mice treated twice (day and post-dss) by intrarectal administration with the test solution ( µl). cd.sirna administration reduced tnfα and il- expression as compared to the non-silencing sirna control or the naked tnfα sirna (not delivered in formulated cyclodextrin). enteral delivery of sirnas for systemic effect was also tested. a nuclease resistance and chemically modified sirna against apolipoprotein b was conjugated with α-tocopherol and was administered ( mg/kg) as lipid nanoparticles in the large intestine of mice. the lipid nanoparticle was composed of mixed micelles comprised of linoleic acid and peg- hydrogenated castor oil. the sirna efficiently reached the liver and other tissues using the chylomicron-mediated pathway via the lymphatic route (murakami et al., ) . delivery of sirna into hepatocytes was markedly reduced when the sirna was not bound to α-tocopherol, indicating that conjugation with α-tocopherol was essential to this delivery system using this route. a calcium phosphate (cap) core coated with sirnas and encapsulated in poly(d,l-lactide-co-glycolide acid) (plga) nanoparticles containing an outer layer of polyethyleneimine (pei) was developed for intrarectal delivery (frede et al., ) . in a dss-induced model of colonic inflammation, intrarectal administration ( µg/day) of nanoparticles containing ng sirna against either tnfα, ip- or kc from day to after % dss treatment, ameliorated the colitis. indeed, while intestinal epithelial cells, dendritic cells, macrophages and t cells could uptake the tnfα sirna nanoparticles, reduction of tnfα was only detected in epithelial cells and t cells (frede et al., ) . the stability of sirna within the cap/plga/pei nanoparticles against enzymatic nucleases under colonic conditions was evaluated. while the sirna-loaded cap/plga/pei nanoparticles were detectable after h incubation at ºc with colonic fluid or homogenate, the same sirnas not formulated in nanoparticles was not detected and was assumed to be degraded. even after exposure to colonic homogenate for h at ºc, the nanoparticles retained biological activity. since the supernatant of the homogenate subsequently incubated with a cell line, still reduced the expression of tnfα by % (frede et al., ) , suggesting that sirna-loaded nanoparticles retain its biological activity compared to that of nacked sirnas (not formulated). www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. other alternatives for rna delivery have been tested. ultrasound-mediated rna delivery has been efficiently achieved in mice (schoellhammer et al., ) . chemically modified sirna against tnfα ( ng sirna in µl water) was delivered in the rectum using ultrasound exposure to khz for . s and after a -day treatment tnfα was efficiently repressed in colonic tissue of a mouse model of induced colitis. in the same study, a mrna (≈ kda) complexed in lipid nanoparticles (lnps) was administered in the colon ( µl lnps solution) with ultrasound and found to be successfully translated locally in the colonic tissue (schoellhammer et al., ) . oral administration of exogenous rnas to exert a biological effect, either locally or systemically, is successful if combined with specific delivery technologies, such as vehicles developed for pharmaceutical/medical purposes. nanoparticles have been designed to release their cargo in specific regions of the gi tract (i.e. where inflammation occurs) or to resist the gi conditions. in contrast to other routes of administration, where normally highly chemically modified rnas are used, oral delivery normally involves use of naked rna or minimally modified rnas formulated with complex delivery vehicles. achieving the local desired effect (i.e. targeting inflammation in the colon) has contributed to the development of delivery technologies and the usage of other routes of administration different than the oral as described above. however, the available literature also suggests that naked or unmodified exogenous rnas are rapidly degraded when exposed to the gi conditions without incorporation into delivery vehicles. extracellular vesicles are natural lipid particles released by many cell types with the potential to mediate cell-to-cell communication. these extracellular vesicles have a tremendous potential as delivery vehicles for therapeutics. however, there are still very few studies evaluating their resistance to the harsh conditions of the gi tract following oral administration. knowledge of their biological effects as transporters of exogenous ncrnas is still very limited. nucleic acid-based therapeutics have the potential to treat numerous diseases by correcting abnormal expression of specific genes. as indicated previously, the oral route of drug administration poses serious delivery challenges due to the gi degrading environment, as well as the need to overcome other barriers preventing nucleic acid delivery. because of these obstacles, efforts in developing oligonucleotide-based therapeutics using the oral route have focused on local gastrointestinal delivery, i.e. directly delivering the drug to the target tissue for localized effects, and, therefore, increasing local bioavailability and maintaining doses low while diminishing possible effects to non-target tissues kriegel et al., ) . crohn's disease and ulcerative colitis are the two principal forms of ibd, characterised by being a chronic relapsing inflammatory condition of the gi tract. the pathogenesis of ibd is dependent on the interaction between local immune and environmental factors in genetically-susceptible individuals (o'neill et al., ) . inflammation in crohn's disease is characterised by high production of the cytokines interferon (ifn)-gamma, il- and tnf-alpha, representing a t helper (th )-th response. in ulcerative colitis, the immune response is characterised by th and an atypical th , with high production of il- , il- and il- (o'neill et al., ) . conventional treatment consists of antiinflammatory and immune-suppressive drugs but, while some medications are effective to combat inflammation in the acute phase, they are ineffective in maintaining remission due to toxicity, dependency and higher relapse rates (kriegel, c and amiji, m, ) . the goal for ibd treatment is local delivery of therapeutics to intestinal immune cells. initial approaches involved il- supplementation www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. due to its huge potential in blocking proinflammatory cytokines and inflammatory tissue damage. but human clinical trials have not proven that systemic il- supplementation can help prevent or improve the ibd symptoms, probably due to il- low mucosal levels . separate studies have used gene therapy to increase il- mucosal availability by engineering il- plasmid dna, and achieving more success in rodent models (barbara et al., ; steidler et al., ; lindsay et al., ; bhavsar and amiji, ) . deregulation of tumour necrosis factor (tnf)-alpha production, a proinflammatory cytokine primarily produced by activated macrophages along with other cell types, is associated with the onset and progression of a number of inflammatory diseases (such as ibd, rheumatoid arthritis), alzheimer's disease and psoriasis . the harmful effects of tnf-alpha are thought to be mainly mediated by activation of nf-kappab, therefore regulating expression of over genes. systemic blockage of tnf-alpha is accompanied by several side effects that can be avoided if local inhibition occurs. rnai-mediated local suppression of tnf-alpha can be beneficial for overcoming many of these side effects. different groups have developed various delivery systems for sirna molecules targeting tnf-alpha expression in the gi tract, assaying them both on cells and several rodent models for ibd (wilson et al., ; kriegel, c and amiji, m, ; kriegel, c and amiji, mm, ; laroui et al., ; yin et al., ; he et al., b, a; he et al., ) . even though therapeutic rnai development is in its infancy, current approaches and tools for delivery of these biomolecules via the oral route are improving. in fact, recent reports describe the effects of an oral smad antisense oligonucleotide in a randomized, placebo-controlled, double-blind, phase clinical trial in patients with crohn's disease (monteleone et al., ; monteleone et al., ) . although in this instance the biomolecule used is an antisense oligonucleotide and not a sirna, the design of the oral formulation could be very similar. iron is an essential metal required for numerous physiological functions, but in excess it is a well-defined risk factor in the pathogenesis of several diseases, including cardiovascular and neurodegenerative diseases. since there is no recognised active pathway of iron excretion, disposal of excess iron from the body is the primary therapeutic goal of treating patients with iron overload. use of iron chelators is limited due to nonspecific distribution in non-target tissues, which results in a number of serious side effects and toxicity . another way to treat iron excess is by limiting absorption of exogenous iron. the divalent metal transporter (dmt ) protein plays a well-established role in iron absorption. the primary site of dmt activity is the intestinal epithelium. dmt expression is regulated in response to body iron levels, with expression enhanced when iron stores are low and, conversely, reduced when iron stores are high. oral delivery of sirna-encapsulated nimos to selectively suppress intestinal dmt could decrease intestinal uptake of dietary iron, thereby mitigating iron overload and preventing iron-mediated toxicity . celiac disease is caused by a t-cell mediated immune response in the small intestine against deamidated cereal gluten peptides modified by the enzyme transglutaminase (tg ). the only way to prevent celiac symptoms is strict adherence to a gluten-free diet. the pathophysiology of celiac disease involves a combination of environmental, genetic and immunological factors. among several immune mediators, enzyme tissue tg and the proinflammatory cytokine interleukin- (il- ) have emerged as key players in promoting inflammatory responses against dietary gluten. given their important role in the pathophysiology of celiac disease, sirna mediated silencing of intestinal tg and il- (i.e. by oral administration using nimos) may result in neutralizing proinflammatory effects and could therefore alleviate disease symptoms . efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. colorectal cancer is one of the most common forms of cancer, and rnai therapy has great potential in the treatment of local intestinal cancers. however, rnai therapy faces major challenges since the genotypic footprints of the various cancers differ widely, even from individual to individual. depending on the tumour, several different sets of genes may be misexpressed, making it extremely difficult to determine a common/unique "culprit" gene. moreover, mice and rats do not generally spontaneously develop colon cancer, so induction of tumour growth is required. but these developed animal models do not reflect the genetic changes observed in human tumours (o'neill et al., ) . genetic mouse models that spontaneously develop intestinal cancer have also been developed. mutations in the apc gene are associated with development of colon cancer as well as a condition called familial adenomatous polyposis (fap). fap is an inherited condition characterised by development of numerous polyps in the colon. some of these polyps can subsequently become malignant adenocarcinomas (o'neill et al., ) . regarding fap and human gene therapy, one study explored oral delivery of escherichia coli expressing shrna against beta-catenin, and found significant silencing activity in healthy mice (xiang et al., ) . the gi tract provides the possibility of acting at systemic level by targeting local resident cells, such as m cells. m cells can take up encapsulated biomolecules to be phagocytosed by macrophages. in fact, aouadi et al. reported that glucan encapsulated sirna particles (gerps) loaded with sirna were phagocytosed by the underlying gut-associated lymphatic tissue (galt) macrophages and then translocated to distal organs of the reticuloendothelial system such as the liver, spleen and lung (aouadi et al., ) . another report claims functional cd shrna expression delivered into dendritic cells in mice by oral administration of recombinant yeast (xu et al., ) . in this manner, the targeted cells located in the intestine can undergo sirna-mediated gene silencing and migrate into tissues throughout the body. when successfully administered, rnai is a very useful therapeutic strategy for efficiently modulating gene expression. orally administered local rnai therapeutics have great potential due to the increasing bioavailability of the therapeutic biomolecule at the targeted tissue or cell type, coupled with simultaneous evasion of systemic side effects and immunogenic reactions. most advanced studies are focused on inflammatory bowel disease, although other potential diseases are being explored. the current main challenge is development of suitable encapsulation methods to protect the rna molecules from the harsh and varying conditions of the gi tract. the literature on rna delivery in fish and birds is scarce, with some examples of dietary delivery of exogenous rnas. in birds, most of the studies have been done using central nervous system administration of sirnas. for example, in the zebra finch bird, sirnas (three - nt long sequences against tyrosine kinase b, trkb) administration via direct injection into the brain was effective in reducing the volume of the high vocal centre by diminishing the robust nucleus of the arcopallium and the relative number of cells within it (beach et al., ) . in another study, direct administration of sirnas in the brain of white-crowned sparrows reduced resting time, spontaneous production of complex vocalizations, and stimulated brief agonistic vocalizations (ubuka et al., ; ubuka et al., ) . in the japanese quail, central administration of the same sirnas against gonadotropin-inhibitory hormone the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. sirnas for rna interference in chicken embryos has also been used to target marek's disease virus in vivo (chen et al., ). in fish models, systemic exosome-mediated sirna delivery to zebrafish resulted in effective crossing of the brain blood barrier and localization in the brain in vivo (yang t et al., ) . exosome-delivered sirnas decreased fluorescence-labelled tumour cells in the brain more than fourfold by inhibiting vascular endothelial growth factor (vegf) in a xenograft zebra fish brain tumour model (yang t et al., ) . these results suggested that brain endothelial exosomes could be used to deliver exogenous sirnas to the target site in the brain for treatment of brain cancer. the proposed higher delivery efficiency, lower immunogenicity, and better compatibility than existing foreign rna carriers have promoted the possible use of exosomes for exogenous rna delivery (mei et al., ) . several other studies have evaluated systemic delivery of sirnas to fish animal models, including delivery to the heart using peg-pla nanoparticles (diao et al., ) or using a neutralized non-charged polyethyleniminebased system . the literature describes other examples of sirna delivery into zebrafish embryos to inhibit specific gene function (gruber et al., ) . in the context of other types of ncrnas, direct injection of mir- a mimic rnas at the one-cell stage of zebrafish embryos resulted in inhibition of formation of the caudal vein plexus by means of targeting the endothelial cell bone morphogenic protein smad signalling (icli et al., ) . also in the zebrafish model, injection of a mir- mimic into embryos at the one-cell stage to deliver mir- ubiquitously resulted in protection against sterile inflammation (hsu et al., ) . dsrna microinjection administration to zebrafish embryos ( - cell stage) at a concentration of - pg rna/embryo resulted in rna interference, an effect that was diminished when ssrna was used (wargelius et al., ) . in cultured european sea bass, intramuscular injection ( weeks, µg of dsrna per dose) followed by in vivo electrically mediated dsrna delivery resulted in a reduction of myostatin gene expression (terova et al., ) . delivery of exogenous rnas has also been tested in lower aquatic organisms. in the crustacean red claw crayfish cherax quadricarinatus, systemic injection of sequence specific dsrnas against viral protein b of macrobrachium rosenbergii nodavirus (mrnv) was conducive to rnai effects that were able to functionally prevent and reduce mortality in infected individuals (hayakijkosol and owens, ) . dsrnas (intramuscular injection) have been used to induce antiviral immunity in invertebrates as they can also recognize dsrna as a virus-associated molecular pattern, resulting in activation of an innate antiviral response (robalino et al., ) . oral administration of encapsulated dsrnas (inactivated bacteria expressing dsrna) in the prawn macrobrachium rosenbergii fed twice daily at a rate of % total biomass for days resulted in their protection against white tail diseases caused by mrnv (naveen kumar and karunasagar, ) . dsrnas designed against the capsid and b genes of mrnv reduced the mortality of prawns by ≈ % due to sequence specific-mediated silencing of the viral genes (naveen kumar and karunasagar, ) . injection of viral protein -targeted dsrna (≥ µg dsrna/g shrimp body weight) into shrimp resulted in a high level of nt sirnas mapping across the entire white spot syndrome virus genome after virus challenge, which resulted in protection against the virus (nilsen et al., ) . in planarians, ingestion of bacterially-expressed dsrnas or direct microinjection into the animals can inhibit gene expression through rna interference (newmark et al., ) . efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. mongersen, an oral smad antisense oligonucleotide, and crohn's disease. n engl j med , - . mu j, zhuang x, wang q, jiang h, deng zb, wang b, et al. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. - . zollner n. . purine and pyrimidine metabolism. proc nutr soc , - . zollner n, grobner w. . influence of oral ribonucleic acid on orotaciduria due to allopurinol administration. z gesamte exp med , - . following a literature search as described in section . . . and based on the methodology described in section . . . , a total of documents were selected as relevant to reviewing the effect of dietary exogenous ncrnas on the gi tract and annex glands. very few studies have focused on the gi tract and their annex glands. four documents specifically evaluated the resistance of plant-derived srnas to simulated in vitro digestion system, ex vivo digestion system or in vivo digestion samples (philip et al., ; yang, et al., a; and reported that the percentage of unaltered mirnas after digestion might be below . %. due to their relevance in early human nutrition, literature on breast milk ncrnas was chosen to describe the biological effects of dietary exogenous ncrnas of non-plant origin, for which full-text reports were studied and reviewed. zhang et al. were the first to report on the transfer of mirnas from food plants to the mammalian circulatory system causing effects on recipient cells . this study reported the detection by high-throughput sequencing of plant srnas in human serum, as well as in tissues of the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. humans, mice and calves. the plant mirnas were distinguished from human mirnas by their '-omethyl modification, which made them resistant to periodate whereas the human mirnas having free ' hydroxyl were periodate sensitive. it was then assumed that the identified plant mirnas were coming from the food uptake. two rice mirnas named osa-mir a and osa-mir a were particularly abundant in human serum samples, since rice is common in human diets. additional in vitro experiments showed that these plant mirnas can endure conditions that mimic the acidic environment of the gut, suggesting that plant mirnas can survive being eaten and digested, pass through the mammalian gi tract and reach target organs. the authors also showed that diet-derived plant mir a can target the low density lipoprotein receptor adapter protein (ldlrap ) mrna based on sequence complementarity. this mirna was able to reduce the ldlrap protein level in the blood and liver of mice fed rice, and eventually increase their plasma low-density lipoproteins (ldl) content. this publication raised questions about whether food-derived srnas could play an active role in human/animal health. however, it was controversial and its results were questioned due to nutritional imbalances in the test-diet studies or lack of rnai-mediated modulation of ldlrap protein levels in mouse liver (witwer and hirschi, ; chen et al., ; . scientists from the monsanto and miragen companies were unable to reproduce these findings . measuring ldlrap protein levels in the liver of mice fed with mir a-containing rice diets by elisa instead of western-blot analysis, dickinson and colleagues did not detect any protein change modulated by mirna a-diets. although changes in the ldl plasma levels were indeed detected, the authors concluded that ldl increases resulted from nutritional imbalances rather than from a mir consumption mediated effect. additionally, plant mirnas including the highly abundant and stable osa-mir a were not detected in liver and plasma samples of mice fed under different rice-based regimes by srna sequencing with the hiseq illumina system . other independent research groups failed to detect dietary plant mirnas in plasma and tissues when conducting controlled feeding studies in humans and in animal models using different detection methods, such as real time quantitative pcr (rt-qpcr), droplet digital pcr (ddpcr) and next generation sequencing (ngs) analysis . these studies provide contradicting evidence of mirnas dietary absorption. in all the attempts, plant mirnas were detected at substantial levels in the diets but were undetectable in animal fluids and tissues. snow and colleagues studied three plant mirna species (mir a, mir a, and mir a) highly abundant in fruits routinely ingested by a group of ten healthy athletes. examination of the athletes' plasma by rt-qpcr analysis using taqman probes revealed high levels of endogenous mirnas but not the three dietderived plant mirnas . the same results were obtained with mice fed vegetarian diets rich in these mirnas. furthermore, using a mir null mutant mouse they showed that none of the dietary mirna including mirna was detected in blood and body tissues, suggesting that dietderived mirnas were not absorbed or maintained in mouse body fluids or organs. another group reported negative results on dietary mirna absorption in non-human primates . using qrt-pcr analysis and the same timeframe of as in zhang's report a, plant-specific mirnas were undetectable in tissues from macaques given a vegetarian diet. another feeding study in humans showed no significant differences in the levels of the plant mir a and mir in the plasma after consumption of broccoli sprouts highly rich in these two mirnas (baier et al., ) . more recently, a different research group using by ngs analysis failed to detect plant mirnas in the plasma of healthy volunteers that normally consume olive oil . the possibility that the plant rnas detected in human tissues and body fluids in some studies were contaminants remains a serious concern . by searching the composition of srna-seq data generated by the zhang's team from amphioxus animals, www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. it was found that the amphioxus data contained rice mirnas as previously reported for human serum. given that amphibia have an exclusive algae-based diet, this strongly suggests that the rice mirnas in both studies are the result of samples contamination . spurious detection of such sequences could arise due to contamination during sample handling, library preparation or sequencing, or result from errors in data analysis. since next-generation sequencing is very sensitive, just a few molecules could cause a false positive detection. more recently a comprehensive meta-study surveying for the presence of cross-species mirnas in sequencing data sets from various human tissues and body fluids indicated that xenomirs (cross-species mirnas) are technical artefacts rather than the results of dietary intake . a bioinformatics study identified plant mirnas in human and porcine milk exosomes . by searching publicly available ngs datasets, the authors identified several plant mirna species, most of them belonging to evolutionarily conserved and highly abundant mirna families. the target prediction suggests that these mirnas may interact with mrnas coding several transcription factors, protein receptors, transporters and immune-related proteins, thus potentially influencing biological functions in humans. of note is that the plant mirna profiles found in mammalian breast milk were similar to the composition in human blood presented by zhang and colleagues . liang and colleagues reported positive results on the absorption of food-derived mirnas from feeding studies in mice using cabbage (brassica oleracea) . they showed that mir , the most abundant mirna in cabbage, was detected in blood and various organs after cabbage consumption. however, there was no absolute quantification of mir or a description of an endogenous control for normalization of qrt-pcr data. a more recent study by zhang's group described how plant mirna from honeysuckle (lonicera japonica) inhibited influenza a virus replication in mice when entering through the gi tract, and they hypothesized that this mirna might be the active component in the traditional chinese medicine based on honeysuckle herb to treat influenza infection . the authors demonstrated that mirna is highly stable in honeysuckle decoction by solexa sequencing and northern blot analysis. they also showed that mir levels increased in mouse peripheral blood and lung following continuous honeysuckle decoction intake (drinking) or gavage feeding, as measured by qrt-pcr using taqman mirna probes. additionally, luciferase reporter assays and in vivo experiments showed that mir was able to target the influenza a virus, and consequently inhibited its replication and reduced mice mortality. the same authors reported that plant mirnas can be found in human umbilical cord blood and amniotic fluid, transferred from mother to the foetus, and that these influence foetal development and health . they showed that mir content increased in the maternal plasma and foetal liver of mice fed honeysuckle. a fluorescently labelled sirna was used to trace the transplacental transmission through feeding. furthermore, they showed that after feeding mice with different amounts of sirnas targeting the alphafetoprotein mrnas the levels of this protein were down regulated in foetal tissues. these results suggest that dietary sirnas delivered from the mother to the foetus could regulate foetal gene expression. more recently, the anti-proliferative effect of a plant mirnas on breast tumours was demonstrated in feeding experiments in mice (chin et al., ) . the plant mir , particularly abundant in broccoli, was found in sera from women, and its levels were inversely correlated with breast cancer morbidity and progression. in human sera, mir was detected in extracellular vesicles by qrt-pcr using a taqman probe. transfection of breast cancer cells with a synthetic mimic mir was shown to reduce their proliferative growth by targeting the transcription factor mrnas. more importantly, oral administration of synthetic mir significantly inhibited the growth of xerograph breast tumours in mice. these results agree with a previous report in which oral administration of a cocktail of three www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. tumour suppressor mirnas that were '-o-methylated to mimic plant mirnas reduced colon tumour burden . additional research is needed to address discrepancies among different studies and verify whether plant srnas derived from diet are absorbed at functional levels in animal species. most of the available data suggests either that gastrointestinal absorption of dietary plant mirnas does not occur in healthy consumers or mirnas levels in blood and tissues of consumers are too low to have a biological impact. however, the biological activity of food mirnas may be more dependent on their accumulation in exosomes than on their abundance per se, because of their reported low concentration. development of sensitive sensors to help to detect the functionality of low-level absorbed mirnas would facilitate assessment of the effects of dietary mirnas in consumers. future studies are needed to establish how exogenous ncrnas absorption and tissue distribution occurs, as well as to address their bioavailability and their biological function. reported that increased mir was detected in sera and urine after consumption of particular foods (i.e. honeysuckle), and disappeared h after the honeysuckle was removed from the diet; this supports the idea that these small rnas were of dietary origin. the role of kidney damage in mirna retention in the animals was also evaluated. using two models of acute renal failure (baliga et al., ) , i.e. the cisplatin (a chemotherapeutic agent) and the glycerol-induced models, it was observed that only mice receiving cisplatin exhibited measurable levels of dietary mirnas in sera and urine. this suggests that kidney damage alone did not lead to enhanced mirna retention in the mouse circulatory system . of interest is that cisplatin treatment, but not glycerol treatment or honeysuckle feeding, disrupted the organization of small intestine epithelial cells as shown by histological analysis. in this study, yang et al. used droplet pcr to demonstrate that the amplified products were likely to be specific and not due, for example, to nonspecific amplification of endogenous rnas . whether particular foods and/or alterations in intestinal permeability could improve the capacity to absorb small rnas from the diet or influence the biological effect within the gi tract, annex glands and systemically is poorly described, but still relevant for risk assessment of exogenous ncrnas in a subset of population. no studies specifically report the biological effects of dietary exogenous plant ncrna on the gi tract. only a few studies (described above) report the possible biological effects in their annex glands, including the liver . for gastrointestinal tissues or the liver, liang et al. reported that icr mice (a strain of albino mice) fed plant total rna ( - µg) extracted from brassica oleracea showed detectable levels of mir in the stomach (up to ≈ copies), intestinal (up to ≈ copies) and faecal content (up to ≈ copies), with the maximum amount present at to h after feeding . estimation of the proportion of orally administered rna that survived digestion suggested that the stomach contained . - . %, the intestines . - . % and the faeces . - . %, while the blood contained about . - . % and the spleen about . - . % . the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. which was similar to that of their synthetic form (without '-o-methylated ' ends) . whereas plant mirnas had a much slower degradation rate compared to their synthetic form without '-o-methylated ' ends. it is important to note that extreme ph conditions are not the only physicochemical or biological condition within the gi tract (see section . . . . for details). using a simulated human digestion system in vitro, philip et al. evaluated the resistance of soybean and rice mirnas to simulated gastric fluid early stage digestion (philip et al., ) . the study found continual survivability of plant mirnas in an in vitro simulated digestion system for over min without any significant decrease in their levels (philip et al., ) . mirna levels have been analysed qualitatively by real-time pcr using taqman mirna assays. stability of the plant-derived small rna mirna was also assessed using an artificial in vitro digestion system that simulates mammalian gastric and intestinal conditions . yang et al. compared three samples which contained a) ml cabbage extract with pmol plant-derived mir (measured by qrt-pcr), and pmol spiked-in synthetic '-o-methylated mir a and artificial mirna termed c ; b) ml phosphate buffered saline (pbs) with pmol each of synthetic '-o-methylated mir , mir a and c ; and c) ml pbs with pmol each of synthetic mir , mir a and c without the '-o-methylation. time course analysis during both gastric and intestinal phase digestion showed that most mirnas displayed modest resistance in the acidic gastric environment (up to min), while in the intestinal phase the levels of all mirnas were drastically reduced after five minutes (figure ) . regardless of synthesis origin and '-o-methylation, the digestive stability of mir was up to -fold higher than that of the other mirnas. the most stable form of mir was the plant-derived form in the cabbage extract, since . % survived, compared to . % for the '-o-methylated form, and . % for the non-modified form. similar results were observed when pmol of synthetic '-o-methylated mir , mir a and c were digested in vitro using the ex vivo intestinal fluids ( figure ) . indeed, mir appeared to be more stable after -hour digestion than the other tested mirnas. finally, by directly measuring mir levels in vivo in the small intestines of mice fed the plant-based diet (daily dietary mir intake pmol) it was observed that this particular mirna reached level more than -fold higher than those observed in the animals consuming a single pmol dose of the synthetic mir . mir a, a mirna also present in the plant-based diets, was also detected in the small intestines at a level times lower than that of mir . using transgenic arabidopsis lines expressing the artificial mirna amir-rice, the murine mirna mmu-mir- a or their controls, yang et al. studied gastrointestinal digestion and bioavailability of transgenic mirnas (yang et al., b) . levels of transgenic mirnas in plants (qrt-pcr quantification) were similar ( . and . fmol/g of fresh weight for amir-rice and mmu-mir- a, respectively) to that of srna mir ( - fmol/g in fresh shoot tissue). however, in diets stored at room temperature, the abundance of amir-rice and mmu-mir- a decreased gradually by ≈ -fold (≈ fmol/g diet) while that of the srna mir increased ≈ fold after h. in vitro gastric and intestinal simulated digestion of the synthetic forms of the plantbased transgenic mirnas showed that amir-rice and mir had similar digestive stability, while mmu-mir- a was significantly less stable. the surviving percentage in the intestinal phase after min was . % for mmu-mir- a compared to ≈ . % for mir or amir-rice (figure , frame a). the surviving percentage in the in vivo assays (gavage-fed pmol of synthetic srnas) further decreased the stability of the mirnas, reducing their level to . % for amir-rice and . % for mir (figure , frame b) . however, the levels of mirnas in the small intestines were higher for mir ( fmol/mouse) than amir-rice ( . fmol/mouse), while mmu-mir- a levels were undistinguishable from the host mouse mirna (figure , frame c). amir-rice was not found in serum (neither are mirnas reported in previous studies and present in transgenic plants) while mir was found only in the serum ( . fm, or . x copies per mouse) of mice fed the plant transgenic diets (yang, j. et al., b) www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. altogether, these results suggest that the plant-derived mir might be more stable than other plantbased small rnas. whether this exceptional stability could be generalized to other ncrnas is unknown. however, recent evidence, including some studies evaluating mir and mir , suggests that there may be some misidentified alternative mirnas . indeed, several other rarer but consistently mapped plant mirnas also have % or near % matches to human transcripts or genomic sequences (i.e. rrna), and some do not map to plant genomes at all, suggesting artefactual results of plant mirnas in mammalian sequencing datasets. this emphasizes the need for rigorous filtering strategies when assessing possible dietary exogenous mirnas . indeed, the small rna mir is derived from s rrna and does not undergo canonical mirna processing, as it does not depend on dcl (yang, j. et al., a) . mir is also inefficiently assembled into the risc complex in human cells and modestly regulates gene expression. this suggests that this exogenous srna is different from the canonical plant-based mirnas (yang, et al., b) . these results highlight the need to evaluate other ncrnas as they could originate from precise processing at the ' or ' end of mature or precursor rnas generating other abundant small rnas (lee et al., ) . also, the rna degradome is a crucial component of the total cellular rna pool of plants (nowacka et al., ) . they can be stable degradation intermediates present in a high copy number, or they can derive from various rna species including trna, rrna, mrna and snrna (addo-quaye et al., ; nowacka et al., ) . among the few examples in the literature of possible biological effects of exogenous non-plant origin ncrnas, milk has been studied because it constitutes a rich source of secreted mirnas. human breast milk is an essential source of nutrition for new-borns. in addition to its nutritional function, breast milk contains myriad biologically-active components that influence the development of the the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. more than % of the unique mirnas identified in porcine milk exosomes (gu et al., ) or % in other studies (chen et al., ) . in human exosomes, the ten most abundant mirnas account for % of the unique mirnas identified . lncrnas previously reported as being important for developmental processes were found in human breast milk extracellular vesicles (karlsson et al., ) . maternal factors such as the mother's diet seem to influence mirna expression in the breast milk fat globule (munch et al., ) or exosomes (sun j, ) . maternal weight (xi et al., ) also seems to influence mirna expression. certain mirnas found in breast milk have also been found in placenta tissue (munch et al., ) , although their physiological role is unknown. other studies have shown that colostrum mirnas (early lactation period) are more abundant than in the later lactation period (gu et al., ; xi et al., ; sun et al., ) . bovine milk mirnas isolated from the colostrum whey fraction were more abundant than in the mature milk whey fraction (izumi et al., ) . immunerelated mirnas expression is reported to be higher in colostrum than mature milk (izumi et al., ; na et al., ) . in vitro studies of human milk mirnas using either rnase digestion, freeze-thaw cycles, low ph or other gi conditions have shown that these mirnas are relatively highly stable (kosaka et al., ; liao et al., ) . similarly high stability of mirnas was found when using one or all of the above mentioned hard conditions for milk from cow (hata et al., ; izumi et al., ), pig (gu et al., or panda . indeed, it has been proposed that rna in milk is present in microvesicles and is protected from rnases and the other gi conditions by surrounding membranes (hata et al., ; gu et al., ; . however, technological conditions such as pasteurization (golan-gerstl et al., ) have been shown to reduce mirnas abundance in cow and goat milk by up to ≈ %, or up to ≈ % for certain mirnas (howard et al., ) , and in human exosome (personal communication, faseb conference ). microwave heating also reduces the amount of certain mirnas (howard et al., ) , while ultrasonication influences exosome membrane integrity and thus increases cargo instability (sun et al., ) . among dairy products, mirnas concentrations varied considerably, but were generally lower than the concentration in pasteurized whole milk (howard et al., ) . the exception was fresh cheese ("queso fresco") dip, which contained higher concentrations of mirnas than observed in pasteurized milk (table ). in vitro studies using mirnas isolated from breast milk exosomes, either human or bovine, were found (liao et al., ) , and crl , k or lim (golan-gerstl et al., ) . when subjected to proteinase k treatment or when the vesicle structure was destroyed, uptake or transport of these exosomal mirnas was compromised (kusuma et al., ; wolf et al., ; sun et al., ) . different studies have attempted to quantify the amount of rnas present in breast milk. rna content in microvesicles isolated from cow milk has been estimated to be ≈ ng for colostrum and ≈ for mature milk when isolated from ml sample (hata et al., ) . in human milk, total rna was estimated to be ≈ ng/ cells of the milk cell fraction and ≈ ng/µl fat from milk fat (alsaweed et al., c) . exogenous plant mirnas have also been found in panda milk exosomes , the whole milk or exosomes of human breast milk the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. et al., ). however, levels of mirnas or total rnas were very low (golan-gerstl et al., ; alsaweed et al., c) compared to that of breast milk. indeed, alsaweed et al. reported that the total amount of rna found in bovine milk-based infant formula was ≈ . ng/µl formula ( human mirnas detected) as compared to . ng/µl for a soy-based formula ( human mirnas detected) (alsaweed et al., c). in terms of possible breast milk mirna absorption, plasma levels of mirnas were found to be higher in colostrum-fed piglets compared to mature milk-fed piglets (gu et al., ) . in humans, plasma mirnas (mir- b and mir- c) increased when different doses of cow milk were consumed (baier et al., ) . in contrast to the studies above, the literature also describes several studies that show either different or contradictory results regarding uptake of exogenous rnas from breast milk. auerbach et al. re-analysed a study in which mir- b- p and mir- c- p were found in plasma after bovine milk ingestion by healthy humans (baier et al., ) and found no significant altered mirna levels after milk ingestion (auerbach et al., ) . these results were confirmed by qpcr and rna-sequencing. several technical issues including low mir- b expression, use of control mirnas (normalization), and variation level may have contributed to these discrepancies (auerbach et al., ) . piglets fed cow milk for weeks showed very low levels of cow-specific mirnas in the bloodstream, which were compared to the levels measured from animals fed maize, which also showed similar counts of cow-specific sequences , suggesting a lack of transfer of exogenous small rnas. using a transgenic mouse model that overexpresses mir- b precursor in the mammary epithelial cells, and thus increases levels of mir- b in milk (≈ -fold), no differences were found in blood, liver, small intestine, kidney or lung tissues of pups from these dams when compared to pups from wild type dams (laubier et al., ) . interestingly, the level of mir- b was ≈ -fold higher in the stomach of mice consuming the transgenic milk. oral ingestion of . µg and . µg of total rna from porcine milk exosomes administered daily during three weeks to mice, significalty increased villus height and crypt depth of the duodenum and jejunum relative to the control group, suggesting an effect improving the development of the gi tract in mice . using genetic knock-out (ko) mouse models for mirnas mir- and mir- c/ , title et al., evaluated the uptake of maternal-milk derived mirnas using the foster mother offspring exchange model to prevent the confounding effects of mirnas derived from tissues of suckling offspring . wild type (wt) offspring consuming mir- ko milk at day exhibited basal levels of mir- in milk from the stomach, which likely comes from stomach epithelial cells. at day of suckling, it was confirmed that most mir- /mir- c came from the milk itself rather than the offspring www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. genotype, because there was no significant difference between wt pups and ko pups receiving wt milk. evaluating different parts of the gi tract (jejunum, ileum and colon), no evidence of uptake was observed because there was no measurable increase in mir- levels in enterocytes of ko pups receiving wt milk compared to ko milk. moreover, no changes in plasma, liver or spleen levels of mir- were observed in ko pups receiving wt or ko milk . similar effects were observed at day of treatment and upon evaluating mir- c on day . when the fate of milkderived mirnas downstream of the stomach was analysed, the levels of mir- in the intestinal content of ko pups receiving wt milk was seen to dramatically decrease compared to that of the stomach contents. these data suggest that milk mirnas are being degraded by the digestive system (since no evidence of uptake was observed). indeed, when the spike in cell-mir- was tested, it was degraded more rapidly than milk mir- , suggesting that milk mirnas exhibited a certain degree of resistance (perhaps because of their exosome content). but less than % of mir- copies remained after h incubation of intestinal content, suggesting a possible digestive enzymatic degradation of the milk-derived (exosomes) mirnas . observational studies also suggest that human breast milk mirnas are not related to preventing atopic dermatitis in infancy months postpartum . in general, rna contamination has been proposed as the main source of controversy in mirnas studies reported in the field of breast milk (bagci and allmer, ) . the literature contains very few studies on the biological effects of dietary exogenous ncrnas in the gi tract and its annex glands. indeed, these few studies only report a biological effect in the liver. the available literature suggests that dietary exogenous plant small ncrnas seem to resist the harsh conditions of the gi tract. however, the stability of plant mirnas under gi conditions in vitro, ex vivo or in vivo is low. moreover, these studies suggest that the percentage of surviving mirnas in the gi tract in the best-case scenario is around ≈ %, although this depends on the specific small ncrnas evaluated. several studies report the presence of dietary exogenous mirnas in breast milk of different mammals, including humans. some protected mirnas (i.e. transported in extracelluar vesicles) have been found to be stable under in vitro degradative conditions. while some evidence suggests a possible absortion through oral feeding, other studies indicates a very limited biovailability or lack of accumulation within the gi tract or its annex glands. recent evidence supports the significant contribution of srnas to communication between hosts and some eukaryotic pathogens, pests, parasites, or symbiotic microorganisms. this silencing transfer phenomenon is very relevant to food and feed risk assessment of ncrna gm plants. although this section (efsa task ) will focus on studies related to the possible trafficking of ncrnas between plants and humans and animals, studies supporting the movement of rna-silencing signals between plants and pathogens are also reviewed. virus-induced gene silencing represents the model of gene silencing in the host by exogenous sirnas. in this case, the sirna specificity determinant is derived from viral rnas. typically, with a discrete length of nt they form perfect duplexes which are produced in plants by dcl and dcl . virusinduced gene silencing is a very effective defence system, and, consistently with the normal dynamics of host-pathogen interactions, all viruses encode silencing suppressors as a counter defence (baulcombe, ) . www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. cross-kingdom rnai implies that a translocation of gene silencing signals occurs between hosts and these organisms. this implies a two-way traffic of sirnas between pathogens and their plant hosts. several studies report that rnas produced in a host plant can be transferred to a fungus or oomycete, symbiont or pathogen, to induce rna silencing in the interacting microorganism (ghag et al., ; koch et al., ; vega-arreguín et al., ; helber et al., ) . transgenic plants carrying a rnai construct, usually with a sense-intron-antisense palindromic structure that produces dsrnas and sirnas, can induce silencing of the targeted transcripts in the interacting organism. furthermore, treatment of fungal conidia with dsrnas of essential genes led to fungal growth inhibition, demonstrating rna interference from environmental silencing signals (koch et al., ) . conversely, the rna produced in a fungus can affect a host plant's defence system (weiberg et al., ) . remarkably, scientists have developed an effective disease control strategy, called host-induced gene silencing (higs), by generating transgenic plants that express exogenous rnai triggers to successfully silence essential genes in pathogens and pests (weiberg et al., ) . in this context, transgenic plants expressing dsrnas or hairpin rnas targeting vital fungal genes of fusarium sp. developed resistance to these important phytopathogens (ghag et al., ; koch et al., ) . small rnas or dsrnas can also be transferred from plant to pests, such as insects eating leaves or nematodes infecting roots. indeed, transgenic plants that express dsrna homologous to essential genes of insect pests or nematodes became resistant to these specific parasites through the silencing activity produced within the target organism when srnas expressed from the plant transgenes is consumed (baum et al., ; fairbairn et al., ; mao et al., ) . many aspects of these cross-kingdom rna interference phenomena are still poorly understood. one of them is how these rna molecules 'travel,' sometimes over long distances through diverse cellular boundaries between plants and interacting organisms. these silencing signals may utilize conserved cell-to-cell as well as systemic rnai pathways present in plants and animals, and may also use organismspecific pathways. rna-protective factors such as agos, other rna-binding proteins, or encapsulation into extracellular vesicles likely play important roles in protecting mobile rnas against degradation during transport (weiberg et al., ; lefebvre and lecuyer, ) . another important question is how these srnas use the target cell rna interference machinery to convey the silencing effect. based on current knowledge, rna-mediated gene silencing seems to be an ubiquitous phenomenon that exists in almost all eukaryotes, and which always follows the principle of complementary nucleotide base-pairing between regulatory srna and mrna sequences (weiberg et al., ) . despite the tremendous differences present in the structural features of regulatory rnas, and the completely unrelated or highly divergent rna gene-silencing mechanisms and pathways that have evolved in diverse organisms, complementary sequence matches seem to be sufficient enough to trigger cross-kingdom gene silencing, as exemplified by plants-parasites, bacteria-to-worms and other lower organism interactions (weiberg et al., ) . following a literature search as described in section . . . and based on the methodology described in the section . . . , a total of documents were selected for review of the topic molecular mechanisms of exogenous ncrna uptake and function. the uptake of exogenous ncrnas is described from the molecular mechanisms point of view (i.e. the presence of a specific transporter, if any). efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. although plant-derived exogenous ncrnas are the main topic of this subsection, some other examples of general molecular mechanisms of exogenous ncrnas uptake, intracellular trafficking and function are included as relevant to understanding the possible fate of plant-derived exogenous ncrnas. other general aspects of rnas uptake are reviewed in chapter . . , and the uptake of exogenous rnas from the pharmaceutical/medicinal area is covered in section . . . as previously mentioned in section . . , cellular uptake of ncrnas presents great challenges due to the physicochemical properties of these molecules. their size and negative charges prevent their easy entry into mammalian cells. cell membranes pose a major barrier to ncrnas entrance since entry into cells is tightly controlled and regulated (mcerlean et al., ) . the anionic lipophilic bilayer prevents entry of macromolecular anionic nucleic acids in their naked form, both restricting their binding to and passive diffusion across these lipophilic cell membranes . furthermore, even though their internalization may depend on the endocytic pathway, endosomal entrapment and lysosomal degradation can be major issues because of the reduced accessibility of ncrnas to their sites of action (nucleus, cytoplasm or mitochondria) (won et al., ; . many different carrier molecules have been developed to achieve the entrance of ncrnas (mostly mirnas and dsrnas) into target cells (bolhassani, ; lindgren and langel, ; mao et al., ; ragelle et al., ; rudzinski and aminabhavi, ; shum and rossi, ) . exogenous ncrnas must be carried by polymers, synthetic cationic lipids or cell-penetrating peptides neutralizing the negative charges of the nucleic acids. most of these cationic lipids tend to form liposomes when dispersed in an aqueous phase, such as blood. endocytosis has been suggested as the main pathway for this nucleic acids-cationic lipid complex internalization by the cell (el ouahabi et al., ) . most reports suggest that so-called cell-penetrating peptides (cpps) bind initially to negatively-charged proteoglycans at the cell surface and are internalized into endosomes (juliano et al., ) . there are multiple pathways of endocytosis (see also section . . ). i. the clathrin-coated pit pathway is the archetype of endocytosis pathways. cell surface receptors and their associated ligands interact with adapter proteins and accessory factors, clustering the receptors into specialized membrane areas subtended by a network of clathrin triskelions. the clathrin network is invaginated by means of membrane curvature promoting specialized proteins, giving rise to a clathrin-coated endosome. this endosome quickly uncoats, generating an uncoated vesicle that will begin its intracellular journey (juliano et al., ) . ii. the caveolar pathway implies the presence of small cell membrane invaginations rich in cholesterol and sphingolipids containing caveolin . cavins, which coat proteins helping to stabilize caveolar structures, are present in these invaginations. there is controversy as to whether caveolae generate independent intracellular vesicles or whether they remain as tubular structures linked to the plasma membrane; however, some data suggest that caveolae generate vesicles able to participate in intracellular traffic, generally presenting a smaller size than other forms of endocytotic vesicles (juliano et al., ) . iii. a number of clathrin-and caveolin-independent pathways have been described. these pathways are often defined in terms of the morphologies of the vesicles they generate or in terms of the cargo that is preferentially internalized. for instance, the flotillin pathway involves the presence of flotillin-rich membrane microdomains, where flotillins are membrane-inserted proteins that may be involved in ordering lipid domains and subsequent endocytosis, similar efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. to caveolin (juliano et al., ) . another such clathrin-and caveolin-independent pathway is the clic/geec pathway, which seems to be particularly important for fluid phase endocytosis. clic/geec stands for clathrin and dynamin independent carriers/gpi-ap enriched early endosomal compartments. this pathway gives rise to high-volume tubular endosomes rich in gpi-proteins and that typically contain fluid phase markers such as dextrans (juliano et al., ) . additional clathrin-and caveolin-independent pathways exist, some involving dynaminmediated disjunction of vesicles from the plasma membrane (juliano et al., ) . iv. macropinocytosis is a process by which cell protrusions pinch off large volumes of extracellular fluid and is therefore an important pathway in fluid phase endocytosis. it is also involved in internalization of clustered, activated receptor tyrosine kinases. v. phagocytosis and entosis are large-volume internalization mechanisms as well. they come into play in specialized cells or unusual circumstances but do not play much of a role in oligonucleotides processing in most cell types (juliano et al., ) . vi. the actin cytoskeleton plays an important role in most of the endocytotic processes described above, although certain arenaviruses enter cells by a pathway independent of clathrin, caveolin, dynamin and actin. it has been reported that phosphorothioate antisense oligonucleotides seem to enter the cells by this pathway as well (juliano et al., ) . there have been many attempts to identify endogenous receptors for antisense or sirna molecules. however, no direct evidence for their involvement in oligonucleotide trafficking has been provided (juliano et al., ) . integrins of the beta- subclass as well as scavenger receptors have been suggested as candidates, but this is controversial. toll-like receptor (tlr) family members seem to be the most convincing examples of cellular receptors for oligonucleotides; tlr binds dna having cpg motifs, tlrs / binds single-stranded rna, while tlr binds double-stranded rna. although these tlrs are usually found within endosomes rather than at the cell surface, in some cases they seem to be able to assist in accumulation of oligonucleotides by cells (juliano et al., ). one interesting candidate as a receptor for oligonucleotides is the mammalian homolog of the doublestranded rna (dsrna) transport protein sid- found in caenorhabditis elegans (feinberg and hunter, ) . the human homolog of this protein, sidt , has been described as facilitating rapid contactdependent intercellular small rna transfer (elhassan et al., ) and as selectively binding long doublestranded rna (li, w et al., ) . another member of the sid transmembrane family, sidt , has been shown to take up extracellular double-stranded rna in drosophila s cells (mcewan et al., ) via endocytosis, although in mammalian cells this protein has been located in lysosomal membranes (jialin et al., ) . in fact, some authors have proposed that sidt could mediate cellular rna degradation inside lysosomes through a novel type of autophagy called rnautophagy (aizawa et al., ) . another research group recently reported that sidt is required to transport internalized dsrna from endocytic compartments into the cytoplasm for immune activation (nguyen et al., ), while others have described that this protein mediates "gymnosis", facilitating uptake of naked single-stranded oligonucleotides into living cells (takahashi et al., ) . a more recent report indicates that both sidt and sidt not only do not transport rna, but are involved in cholesterol transport (mendez-acevedo et al., ) . once an oligonucleotide has entered a cell in an endosome, it encounters a complex maze of intracellular pathways leading to multiple destinations and regulated by an intricate protein machinery. key subcellular membrane bound compartments include early and recycling endosomes, late www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. endosomes/multi-vesicular bodies, lysosomes, the golgi apparatus and the endoplasmic reticulum. intracellular trafficking is not a random process, but rather a carefully orchestrated choreography that allows the cells to transport endogenous and exogenous materials to the most appropriate places. for nucleotides to exert their function, they need to leave the membrane bound compartments and access the cytosol and/or nucleus (juliano et al., ) . knowledge as to how endocytotic cargos are delivered to subcellular compartments is still partial. many of the internalization pathways previously described converge at the stage of early endosomes, raising the question of how the differentially internalized components traffic to different destinations. certain evidences suggest that membrane domains originating from different internalization pathways maintain their identity within early endosomes, thus providing the means for specific sorting and trafficking to distinct downstream destinations (juliano et al., ) . all membrane traffic proceeds through the same basic steps: i) a coated vesicle is pinched off from a larger donor membrane compartment; ii) the vesicle uncoats, allowing display of the tethering and fusion proteins; iii) the vesicle is carried to its destination along "tracks" provided by actin-or tubulin-based cytoskeletal structures; iv) the vesicle recognizes its target membrane compartment using tethering proteins and then utilizes snare proteins to complete the fusion process and deliver membrane and contents to the target compartment (juliano et al., ) (figure ). ncrnas must escape from the endomembrane compartments and reach the cytosol to exert their function. as mentioned above, intracellular trafficking involves a highly dynamic flux of membrane vesicles engaged in a multitude of fusion and disjunction events. there are a few key points in these processes to be considered when designing non-viral oligonucleotide delivery strategies: ) fusion involves localized stress on the fusion partners, including formation of non-bilayer lipid domains; ) non-bilayer regions of membranes can be much leakier than bilayer regions; ) many enveloped viruses fuse with cells via specialized membrane interacting proteins that, while differing in sequence, act in a manner similar to cellular snare proteins; in many cases these proteins can also induce increments in membrane permeability. therefore, there is an intrinsic relationship between the fusion events inherent to intracellular trafficking and transient leakage of vesicular contents. thus, the innate activity of oligonucleotides taken up by cells is likely due to a modest amount of continuous leakage from endomembrane compartments spontaneously occurring during intracellular trafficking (juliano et al., ) . some ncrnas function is exerted in the nucleus, although nuclear entry may not be the rate-limiting step for oligonucleotide action. studies have shown that oligonucleotides, particularly those with phosphorothioate backbones, are able to continuously shuttle between the nucleus and cytoplasm. this is an active process mediated by nuclear pore structures but does not require classic nuclear localization signals. for phosphodiester oligonucleotides, both passive diffusion and active transport have been described as nuclear entry mechanisms (juliano et al., ) . in terms of the uptake and trafficking of "free" or "naked" oligonucleotides, the co-existence of both productive and non-productive uptake routes has been suggested. the non-productive pathway seems to involve trafficking to lysosomes, while the pathway that results in rnase h-dependent antisense effects eventually leads to interaction with cellular pre-mrna (koller et al., ) . other studies have involved so-called "gymnotic" uptake of antisense oligonucleotides modified with lna (locked nucleic acid) moieties (stein et al., ; zhang et al., ) . subcellular distribution studies surprisingly suggested that the deoxy lna compounds became associated with p-bodies that are usually thought to be sirna action sites (stein et al., ) . unlike the study performed with phosphorothioate www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. oligonucleotides in which the concentrations used were in the nanomolar range, the antisense effects of "naked" lna required micromolar concentrations (juliano et al., ). as described in section . . , exogenous ncrnas must overcome a series of barriers to be functional at systemic level. in addition to the extracellular barriers specifically related to oral absorption, defensive humoral and cellular barriers efficiently prevent the intrusion of exogenous entities into the organism (jeong et al., ) . systemically-administered nucleic acids are rapidly degraded by nucleases in a few minutes and, even in nucleic acid drug therapy using positively-charged polyplexes, they readily interact with serum components to form larger aggregates, resulting in rapid clearance by res or phagocytes (jeong et al., ) . if the nucleic acids survive in the blood stream, they must still reach the appropriate tissues and enter the target cells. in systemic nucleic acid therapy, continuous endothelial walls in the microvasculatures are one of the main barriers limiting the access of oligonucleotides to target cells. in certain pathological cases, such as cancer, the presence of leaky vasculatures in the vicinity of highly penetrating solid tumours allows nanoparticulates to penetrate and accumulate in tumours due to the enhanced permeation and retention effect (jeong et al., ) . caveolae-mediated transcytosis has been considered one of the important mechanisms of macromolecules transport across the endothelium (schnitzer, ) . another possible limiting barrier is the extracellular matrix consisting of various proteoglycans, which are proteins covalently cross-linked with carboxylic or sulphated glycosaminoglycans (gags) (ruponen et al., ) . since gags, such as heparin sulphate, chondroitin sulphate and hyaluronic acid, are polyanions, they may repel the negatively-charged oligonucleotides, affecting mobility of the nucleic acids in the tissue extracellular matrix and therefore limiting their access to target cells. in the case that oligonucleotides reach the target cell membrane, they must enter the cell. the first barrier is the anionic plasma membrane itself, and the second is cellular uptake via endocytosis (see section . . ). without a specific ligand-receptor interaction mechanism, the entrance of ncrnas results in a lack of cell specificity (jeong et al., ) . once taken up by cells, the nucleic acids are localized within the endosomal compartments, where the ph rapidly drops to about by the action of membrane-bound atp-driven proton pumps. the endosomes mature to lysosomes where the oligonucleotides can be degraded by various enzymes (jeong et al., ) . as previously indicated, any surviving nucleic acids must then escape from the endosome to exert their functional effects. in nucleic acid drug therapy, certain endosomal disruptive agents, such as lysomotropic chloroquine, fusogenic peptides and ph-sensitive neutral lipid, are used to facilitate or promote endosomal escape through membrane disruption (jeong et al., ) . after endosomal escape, ncrnas should move through the cytoplasm to encounter their molecular targets, or to the peri-nuclear space where nuclear translocation takes place if the nucleus is the target compartment. passive diffusion of high-molecular weight macromolecules is limited in the cytoplasm. a complex network of microfilaments and microtubules, highly concentrated proteins and various subcellular organelles provide the cytoplasm with a high fluid phase viscosity and mesh-like structure with to Å pores from, making the diffusion process size-dependent (jeong et al., ) . in fact, microinjection of plasmid dna into the cytoplasm showed that normal dna was practically immobile in the cytoplasmic space (dowty et al., ) . however, dna molecules with less than base pairs seem to undergo free diffusion in the cytoplasm (lukacs et al., ) , suggesting that small oligonucleotides (i.e. mirnas) do not face limitations regarding their intracellular trafficking, contrary to large ncrnas (i.e. lncrnas) which might encounter more restrictions reaching the peri-nuclear space. finally, some ncrnas need to enter the nucleus to be functional. the nuclear envelope consists of a double-layer membrane with nuclear pores. the nuclear pore complex allows free diffusion of ions and small molecules, but restricts passage of macromolecules with molecular efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. weights greater than kda, unless accompanied by a nuclear localization signal peptide (nls) (jeong et al., ) , which is not the case with lncrnas oligonucleotides are initially accumulated in an endomembrane compartment (the donor compartment, e.g. early endosomes) and are then trafficked by shuttle vesicles to various other endomembrane compartments (the recipient compartment, e.g. the trans-golgi). the first step ( ) involves disjunction ('pinching off') of a shuttle vesicle under the influence of a coat protein as well as other accessory proteins. at this stage there are non-bilayer regions at the junction between the membranes of the donor compartment and the shuttle vesicle. this provides an opportunity for some oligonucleotides to escape into the cytosol. step involves uncoating of the vesicle; and rab proteins can contribute to this step. step comprises movement of the shuttle vesicle toward its destination along cytoskeletal tracks. motor proteins such as various myosins (for the actin system) or dyneins or kinesins (for the microtubular system) propel the vesicle. rab proteins are involved in forming the appropriate linkages to the cytoskeleton. step entails recognition of the recipient ('target') compartment by the shuttle vesicle. tether proteins work with rab proteins to provide interaction specificity while v-snare proteins in the vesicle membrane interact with t-snare proteins in the recipient compartment membrane to provide firm bridging, as well as contributing to specificity. in step the snare proteins undergo major conformational changes, and, with the assistance of accessory proteins, trigger fusion of the shuttle vesicle membrane with the membrane of the recipient compartment. at this stage non-bilayer regions exist at the junction between shuttle and recipient membranes potentially allowing oligonucleotide escape. reprint with permission from (r l juliano, x ming, o nakagawa. cellular uptake and intracellular trafficking of antisense and sirna oligonucleotides. bioconjugate chemistry : - ). copyrigth ( ) american chemical society. figure : proposed mechanism of vesicular trafficking of oligonucleotides some research has been done on cellular uptake, trafficking and tissue distribution of oligonucleotides without specific targeting or carrier mechanisms. in terms of tissue distribution, certain cell types exhibit a preferential in vivo uptake, particularly kidney proximal tubule cells and liver kupffer cells for phosphorothioate (ps) antisense compounds and sirna (juliano et al., ) . however, only one report has studied systemic level uptake and function of orally administered mirnas . these authors studied intestinal uptake of maternal milk-derived mirnas by new-born mice employing genetically-modified models to distinguish endogenous mirnas from milk-derived exogenous mirnas. their analysis of the intestinal epithelium, blood, liver and spleen revealed no evidence for mirna www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. uptake, even though milk in lactating mothers has been shown to be a particularly rich source of secreted mirnas. sequencing and microarray analysis of mirnas in the milk of various mammalian species has led to the discovery of hundreds of mirnas in cow, pig, rat and human milk, which are derived from exosomes and cellular components. furthermore, their study revealed rapid degradation of milk mirna in intestinal fluid, indicating that mirnas in the milk play a nutritional rather than a generegulatory role in new-born mice . there are certain tissues containing special physical structures that can function as additional barriers to ncrnas entry. such is the case of the central nervous system (cns), where delivery of nucleic acids is particularly challenging because of its anatomical and physiological complexities. the cns is protected by the blood-brain barrier (bbb), which consists of tightly joined capillary endothelial cells, restricting access of large molecules into the brain. the cns extends to include the spinal cord, which is protected by the blood-cerebrospinal fluid barrier (bcsfb), made up of choroid plexus epithelial cells limiting the free diffusion of molecules into the cerebrospinal fluid (csf). the bbb and the bcsfb express numerous transporters and receptors which contribute to the transfer of essential nutrients such as glucose and amino acids into the cns. within the cns, different cell types exist including neurons, astrocytes and other glial cells. entry of nucleic acids into neurons is notoriously difficult for unclear reasons, although it is likely related to their post-mitotic nature, as well as the complex structures and intricacies of neuronal networking (o'mahony et al., ) . for instance, variances in the uptake of sirna lipoplexes (liposome and nucleic acid complexes) were reported at different parts of the neuron structure, with greater uptake efficiency at the cell soma compared to the neuritis, which may occur because of different membrane compositions in these domains (o'mahony et al., ) . circumventing the bbb is only possible by direct administration of ncrnas to a target region of the brain or by administration to the spinal cord where trans-synaptic retrograde transport to the brain is possible. some delivery systems exploit the receptor-mediated uptake of molecules such as transferrin, lactoferrin and insulin receptors by attaching a ligand for the receptor to the non-viral delivery system, or promoting a transient mechanical disruption of the bbb (o' mahony et al., ) . in certain neurological diseases, and in particular in brain cancers, anatomical and physiological changes occur that can impact bbb integrity and the cns accessibility. delivery to the csf to bypass the bbb presents its own obstacles, including rapid clearance from the csf, the bcsfb and limited diffusion from the csf into the brain parenchyma (o'mahony et al., ) (figure ) . another specialized barrier is the placenta. the placenta serves as the interface between the maternal and foetal circulatory systems, and regulates the transfer of oxygen, nutrients and waste products. when exogenous substances are present in the maternal bloodstream, the extent to which these affect the foetus is determined by transport and biotransformation processes in the placental barrier. this barrier is formed by fusion of the outer blastocyst trophoblast cells facing the uterine epithelium into multinucleated syncytiotrophoblast. proliferation of syncytiotrophoblast is mediated by the fusion of precursor cytotrophoblast cells. maternal blood in the intervillous space and foetal blood in the villous capillaries are separated by a continuous layer of syncytiotrophoblast, a discontinuous layer of cytotrophoblast cells, basal lamina, connective tissue, and foetal endothelial cells (al-enazy et al., ) . the placenta undergoes several changes as pregnancy progresses. overall, the placental barrier becomes thinner over time, the distance separating the maternal circulation from the foetal circulation decreasing from - µm in the second month to only - µm at term. passive diffusion across the placenta is favoured for lipophilic substances, allowing them to cross the phospholipid bilayer. smaller compounds tend to cross the placenta more readily, with compounds having a molecular weight less than da crossing it most easily (al-enazy et al., ) . since nucleic acids are polarized, negatively charged molecules, it is unlikely they would be able to cross the placenta by passive diffusion. nevertheless, the placental syncytiotrophoblast is capable of efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. endocytosis (al-enazy et al., ) . no reports refer to exogenous ncrnas entering the placental barrier. however, the presence of placental mirnas has been described in the blood circulation of pregnant women. these mirnas have been reported to be actively secreted into microvesicles and exosomes. these specific exosomal mirnas may be important for embryo implantation, playing a significant role in the intercellular communication pathways that potentially contribute to placentation and development of maternal-foetal vascular exchange (mitchell et al., ) . since exosomes can cross the blood brain barrier, meaning they can cross several layers of cells, it has been speculated that exosomes could directly move from maternal to foetal tissues, and vice-versa, during pregnancy (record, ). the the molecular mechanisms of exogenous ncrna cellular uptake have been inferred from studies performed when developing strategies for nucleic acid delivery in therapeutics. few studies have been done with "naked" oligonucleotides, and even in these cases, the oligonucleotides were chemically modified. these molecular mechanisms of cellular uptake include the different types of endocytosis mechanisms known to date. the existence of receptor-mediated uptake of oligonucleotides through the sidt and sidt proteins has also been described, although their function has been contradicted since they have been described as cholesterol transporters. among the numerous challenges ncrnas must overcome once taken-up by the cell, escaping the endosomal compartments is the most relevant and significant, although reaching their site of action is also crucial to exerting their function. specialized barriers such as the blood-brain barrier or the placental barrier present further obstacles to overcome. www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. zhang y, qu z, kim s, shi v, liao b, kraft p, et al. . down-modulation of cancer targets using locked nucleic acid (lna)-based antisense oligonucleotides without transfection. gene ther , - . zhou g, zhou y, chen x. following a literature search as described in section . . . . and based on the methodology in section . . . ., a total documents were selected for reviewing the landscape of exogenous rnas in biological fluids of human and animals. using public databases of rna sequencing datasets, several documents report the application of in silico bioinformatics analysis (table ) clearly describing the widespread presence of exogenous ncrnas in biological fluids(from multiple sources, including diet) wang et al., ; beatty et al., ; yeri et al., ; . however, four documents suggest that exogenous ncrnas present in the public datasets may represent contamination from the laboratory environment since their abundance is extremely low, or they are of non-dietary origin, or could originate from technical artefacts or contamination bagci and allmer, ) . five documents report the development of bioinformatics resources to either predict transportable mirnas, predict functional analysis, or list and compare the exogenous mirnas found in samples chiang et al., ; zhang et al., ; zheng et al., ) . the generalized use of highly parallelized next generation (nexgen) sequencing technologies has promoted the use of sequencing to study complex biological systems, including human and animal biological fluids and tissues. initial studies evaluating the possible presence of exogenous ncrnas in biological fluids were done in silico using public databases of rna-seq results. several studies report the presence of exogenous rnas (i.e. dietary plant ncrnas) in the biological fluids of humans and animals. initial studies of the srnas spectra in human plasma showed that a small fraction of sequence reads from plasma mapped to human mirnas (≈ . %) or human transcripts and human genome sequences (≈ %) . this fraction increased by up to ≈ % when a higher sequence mismatch tolerance was allowed (under two mismatch allowance). surprisingly, a significant number of the unmapped reads aligned with various bacterial and fungal sequences (i.e. ribosomal rnas and trnas). also, many processed reads mapped to common food items, the most abundant being rna sequences from corn (zea mays) and rice (oryza sativa). compared to data from a serum sample from a chinese individual, the sequence abundance between corn and rice was reversed , suggesting the influence of dietary habits. other sequences from other foods in human plasma samples included soybean (glycine max), tomato (solanum lycopersicum), grape (vitis vinifera) and others. including exogenous mirnas, various common household insects were identified . treating plasma samples with dnase, protease, triton x- , and additional rnase reduced the total number of reads compared to the addition of rnase alone, suggesting that some of the exogenous rna molecules are probably associated to circulating protein and/or lipid complexes . in the same study, a lower percentage of exogenous sequences was observed in mouse plasma samples compared to human samples . another study evaluating a larger number of human plasma samples (n= young male athletes) reported exogenous srnas from different bacteria, but very few srnas were consistently detected in all samples (yeri et al., ) . in a study on public srna datasets from various tissues from mammals, chicken and insects, plant mirnas were found in out of datasets analysed, with mir- being extremely over-represented efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. . in datasets ( humans, mice, pig and others) the plant mirnas reads ranged from ≈ . - . % of the total animal mirna reads. insects fed with plant foods containing mir- , did not show accumulation of mir- when evaluated by northern-blot. however, in a very few samples sequencing data detected plant mirnas not present in the food source, including mir- , in addition to mir- at a moderate read, suggesting that plant mirnas observed in some public srna databases may be artefactual and due to sequencing methodology . plant mir- a from monocots was also found to be the most abundant exogenous plant mirna, in most cases, in other srna sequencing datasets studies with levels typically < % of human mirnas . but when analysis was performed in the absence of any known sources of plant contamination, plant mirnas were not detected. this suggests that precaution should be taken to prevent cross-contamination of the samples due to the high sensitivity of next-generation sequencing methods . other human public srna sequencing datasets have also been questioned for the presence of exogenous plant mirnas . of analysed human plasma samples, plant mirnas were found when compared to the genome of different plant model organisms (arabidopsis thaliana, triticum aestivum, oryza sativa, zea mays, and brachypodium distachyon). at least one read was present in selected samples. peu-mir- , found in all samples, is a singular mirna found in selected samples with more than counts. mir- is found in zea mays and conserved in fruits and vegetables including melon, sorghum, tomato, tea and oil palm. other highly-expressed plant mirnas found in up to hundreds of copies in selected samples include peu-mir- (found in samples), peu-mir- (found in samples), and tae-mir- (found in samples) . plant mir- (found in samples) and mir- a (found in samples) were detected in relatively small amounts (less than copies). using bioinformatics tools, the same study showed that seed sequences of mir- and mir- are similar to hsa-mir- and mir- - p for the former and has-mir- - p for the latter, and thus potentially target several gene targets of their human mirna counterparts . other studies have also reported in mouse plasma samples the presence of exogenous rnas matching exogenous species including bacteria, fungi, viridiplantae and food items . these included mir- d and mir- a (corn or rice as the most likely source), and mirnas from worms and insects . abundant non-human rnas were found in other healthy individuals human plasma samples (n= ) assigned to metazoan, bacteria or fungi (beatty et al., ) . several exogenous srnas were found that could potentially derive from food items including fragaria vesca, medicago truncatula, lotus japonicus, glycine max, arabidopsis thaliana, and solanum lycopersicum (beatty et al., ) . twelve public srna sequencing databases were analysed for the presence of plant exogenous mirnas. breast milk exosomes from humans ( samples, . million reads) and pigs ( samples, . million reads) were analysed. although plant mirna species belonging to mirna families ( samples) were found, they showed low abundance (< read counts for the highest expressed mirna). zma-mir- a, zma-mir- a, ath-mir- a, ath-mir- b and ptc-mir- d showed the highest abundance levels, while in humans only samples contained plant mirnas from mirna families (< read counts, for the highest expressed mirna, average of samples). ath-mir- a, pab-mir- , pc-mir- a, bdi-mir- , aly-mir- d, osa-mir- b. and zma-mir- a showed the highest abundance level in human samples . however, bagci and allmer reanalysed the data from lukasik and zielenkiewicz study and found that exogenous plant-derived mirnas in milk are likely to be contamination (bagci and allmer, ) . in a later observational study, lukasik et al. evaluated plant mirnas mir- a, mir- a, mir- a, mir- a and mir- a and reported the presence of plant mirnas in human (n= ) breast milk, both in whole milk and exosomes . out of mirnas, only were found in exosomes (mir- a and mir- a). while the human has-mir- a was found in all samples in a concentration range of . - pm for whole milk efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. and . - . pm in exosomes fractions, plant-derived mirna mir a was found in all samples at a concentration of - fm (≈ -fold higher than other plant mirnas) . compared to endogenous mirnas, the levels of exogenous plant mirnas were rather low and varied significantly between the samples . kang et al. analysed public sequencing srnas datasets from various human tissues and body fluids (n= ), including from serum and plasma samples, from exosomes, from cerebrospinal fluids, from liver, from blood cells and from brain . of note is that exogenous mirnas (xenomirs) were only found in % of the tissue samples (from total samples), with several studies being completely void of any xenomirs. xenomirs were absent from most human tissues samples, or present at very low abundances (with a median of . read counts). no enrichment of xenomirs was found in tissues relevant to dietary intake (i.e. hepatocytes and blood cells). by contrast, out of human body fluids datasets, xenomirs were found in % of samples, although at very low levels (median of read counts). even though the blood brain barrier separates the cerebrospinal fluid from the bloodstream, xenomirs were found in comparable quantities in both , suggesting no depletion (as expected by the presence of the barrier) of xenomirs in body fluids separated from the main bloodstream. moreover, the lack of co-occurrence of xenomirs in cerebrospinal fluid and serum of the same individuals, do not support that xenomirs detected in cerebrospinal fluid samples have entered through the bloodstream, and thus do not support a dietary origin of xenomirs. the most common xenomirs in body fluid samples belonged to the clades for rodents ( %), dicots ( %), insects ( %) and euphyllophytes ( %), suggesting that xenomirs composition does not reflect human food sources. from billion sequence reads (all samples), xenomirs were in low abundance, comprising only . % of the total reads ( , ). the same study reported a controlled feeding trial in rats and piglets (see section . . below for details). shu et al. ( ) used bioinformatics tools to characterize the structural characteristics of exogenous mirnas that can contribute to cross-species transportation and thus be transferred into human circulation . thirty-four thousand ( . ) mirnas sequences were compiled from species (from five kingdoms including animal, plantae, fungi, protista and viruses), including dietary mirnas ( mirnas) from types of common food species such as cow milk, breast milk, tomato, grape, and apple. using a support vector machine (smv)-based feature elimination strategy, features based on sequence, structure, and physicochemical properties were analysed to distinguish human circulating mirnas from the remaining sequences. predictions were made for features (categorized into groups) to better predict transportable mirnas in human circulation. known human plasma mirnas were ranked among the top of the list (dominated by animalia origin). only dietary mirnas were ranked among the top mirnas, five of which have a sequence identical to that in humans (bta-mir- b, bta-mir- , bta-mir- , gga-mir- a- p, gga-mir- b- p) . while only one mirna from plantae ranked among the top mirnas, plantae mirnas ranked in the top as possibly transportable , suggesting that animal-borne mirnas may be subject to more significant absorption in humans than in plant mirnas . it is important to note that this study of evaluated , animalia mirnas and plantae mirnas, whether this discrepancy in the original number of evaluated mirnas may have biased the smv-based classifiers to mammalian transportable mirnas was not addressed. in the same context, a database called "dietary mirna database" that describes different dietary mirna parameters and functional analysis has been described and is freely available. additional databases for exogenous mirna discovery using rna-seq data are also available in the scientific community . using bioinformatics tools (i.e. rnahybrid database), other studies have also predicted the potential function of dietary mirnas in the human body . in a bioinformatics study of plant mirnas, some similar functions between human and plant target genes were identified, such as ion transport and stress response . efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. analysing public srna sequencing datasets from humans ( ), mice ( ), pig ( ), chicken ( ), fly ( ), c. elegans ( ) and many other nematodes, protozoa or prokaryota, zheng et al. developed a database (exo-mirexplorer) for exploring and comparatively analysing exogenous mirnas (zheng et al., ) . two hundred and thirty-seven ( ) plant-derived mirnas were detected in sequencing samples. common mirna families included mir- , mir- , mir- , mir- and mir- . mir- a was the most frequently found mirna, and was detected in human samples from different studies (zheng et al., ) . the average abundance of mir- a was , . reads per million. in total, mir- a was found in samples from multiple species. mir- a is another frequently observed exogenous mirna from plants, and was detected in samples with an average abundance of . read per million (zheng et al., ) . regarding other biological fluids, exogenous rnas of bacterial origin were found in (urine) and (saliva) samples from young male athletes analysed for srna-seq (yeri et al., ) . plant small rna mir- was detected in urine of mice fed honeysuckle decoction (yang et al., b) . mir- a has been also detected in serum when honeysuckle was supplemented with pmol synthetic mir- a. mir- and mir- a were detected in the urine of mice that had received cisplatin treatment (cisplatin administration produces acute renal failure) and consumed a honeysuckle decoction alone or supplemented with synthetic mir- a. the increased detection of plant small rnas was attributed to disruption of the organization of small intestine epithelial cells produced by cisplatin treatment or longterm consumption (pre-fed) of honeysuckle (yang et al., b) , by mechanisms as yet unknown. mir- levels in urine reached ≈ fm. in tissues, the overall percentage of exogenous sequences in human lung tissue (normal lung rna samples), was found to be less than % . in spermatozoa samples, using public srna sequences datasets, tosar et al. found high amounts of exogenous sequences in three samples . however, in their own analysis of three spermatozoa samples they found no detectable traces of any plant mirnas, suggesting that exogenous mirnas present in the public datasets may have been caused by laboratory environment contamination . in terms of the possible passage of plant mirnas through the brain-blood barrier, kang et al. analysed samples from cerebrospinal fluid (csf) (from one study), which is separated from the bloodstream by the brain-blood barrier, and brain samples (from studies) . in general, xenomirs were absent from most of the human tissue samples, and, when present, they were found at very low abundance ( % of analysed tissues). although the brain-blood barrier would relatively protect the brain from dietary molecules, xenomirs were present in similar fractions in the exposed liver and in the relatively protected brain samples , suggesting, according to the authors, a nondietary origin . xenomirs were present in more body fluids samples (≈ % of samples) than tissue samples (≈ % of samples) and were identified in comparable proportions in the serum and cfs. this study does not provide support for the dietary origin of xenomirs (including plant exogenous rnas) meaning it may originate from technical artefacts . efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. in addition to mirnas reported by the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. very few studies have reported the influence of dietary or physiological factors on levels of exogenous ncrnas in biological fluids. in a small study (n= per group), wang and colleagues compared some abundant exogenous mirnas species identified in plasma samples from healthy subjects, colorectal cancer and ulcerative colitis patients . although ulcerative colitis patients apparently had increased levels of circulating mirnas of certain species (i.e. from a bantam family), a large variability of mirnas levels were observed among subjects of the three groups , suggesting that health/disease status may influence the presence of circulating mirnas in human plasma. tissue injury produced by excessive consumption of common pharmaceutical drugs can also influence exogenous rnas levels in plasma . wang et al. showed that acetaminophen overdose in a mouse model produced liver injury, and an increase of rna concentration in plasma compared to the control group was observed (from ≈ to ≈ ng/ml of total rna h after treatment). plasma ncrnas levels were affected by acetaminophen overdose, both endogenous and exogenous. plantderived exogenous mirnas levels were reduced, and this drop in plant rna was related to the observation that mice lose appetite after acetaminophen injection , suggesting the influence of diet on plasma plant-derived mirna levels. levels of exogenous rnas can be affected in case of drug-induced kidney damage. sera and urine from mice treated with cisplatin, a chemotherapeutic agent also producing small intestines epithelial cells disruption, showed increased plant exogenous ncrnas (i.e. plant mirnas), not observed in untreated mice (yang et al., b) . the effect was not observed in a glycerol-induced model of acute renal failure (which does not affect small intestine organization). detection levels of exogenous plant mirnas, including mir- and mir- a, were below background levels in mice pre-fed with chow diet devoid of honeysuckle. however, when mice were pre-fed with honeysuckle and honeysuckle was administered (alone or mir- a-supplemented) dose-dependent levels of mir- a was detected in serum and elevated serum levels of mir- were observed (yang et al., b) . the study shows that long-term honeysuckle feeding potentiates absorption of dietary mirnas through a mechanism that likely differs from cisplatin treatment, suggesting that particular diets can modify the capacity to absorb small rnas, and that altered or damaged guts resulting from illness and/or therapeutic treatment may enhance dietary rna uptake (yang et al., b) . while comparing the presence of exogenous srnas in three human plasma samples, beatty et al. reported that the largest proportion of reads matching plant sequences were found from one individual who reported following a vegetarian diet (beatty et al., ) , suggesting that the diet can influence levels of exogenous srnas of food origin. in another study comparing human breast milk for mirnas derived from plants, no significant differences were observed for exosomal breast milk plant-derived mirnas (mir- a and mir- a) between vegetarian or non-vegetarian volunteers . in the whole milk an increase in plant mirna mir- a was observed in non-vegetarian versus vegetarian diet volunteers . although some studies (see above) have reported the presence of exogenous rnas in human and animal circulation and tissues, very few have focused on quantitative data relevant to risk assessment. wang et al. estimated the concentration of rna in plasma to be < . pm based on an average of ng/ml of rna in plasma with an average of nt in length and a selected rna (i.e. mirna) sequence representing less than . % of the total rna population. after days feeding of herbals, yang et al. reported an increase of mir- in the sera of mice from fm ( -fold higher than control chow-fed animals) following ginseng intake to fm ( -fold higher than control chow-fed animals) following honeysuckle intake . the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the lack of enrichment of plant-derived small ncrnas in samples to be analysed by rna-sequencing (i.e. through periodate oxidation) might also influence the low enrichment of these exogenous sequences in biological fluids. however, although '-o-methyl modification of synthetic rna oligonucleotides '-ends can affect ligation activity of the t ligase in srna library preparation and may potentially result in srna sequencing bias and under-representation of srna with '-o -methyl '-ends in quantification experiments (munafo and robb, ) , others suggest a broad capability to detect plant mirnas during the sequencing procedure of small rna library construction . the available literature clearly suggests the widespread presence of exogenous rnas from multiple species, including plants, microbiota and many others, in human and animal biological fluids. evidence for plant-origin ncrnas, possibly relevant to risk assessment of ncrna-based gm, suggests that plantderived mirnas can be found in the biological fluids of humans and animals. whether these sequences are directly derived from dietary sources has not been clearly demonstrated. their overall low abundance, lack of enrichment in tissues most exposed to dietary changes, and their high variation between samples suggest that some may have originated as technical artefacts or contamination, as indicated by some authors. although few reports were available in this area, recent evidence also suggests that the levels of exogenous plant ncrnas in biological fluids may be influenced by diet, subject physiological or pathological conditions, or certain therapeutic treatments. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. following a literature search as described in section . . . . and based on the methodology described in the section . . . ., a total of papers were selected as being relevant to review of the systemic effects of dietary exogenous ncrnas. very few of these studies ( ) provide supporting evidence (in favour) that dietary exogenous ncrnas can be detected in biological samples and/or exert a biological effect after dietary ingestion. by contrast, studies provide contradicting evidence (against) that dietary exogenous ncrnas are either bioavailable or exert a systemic biological effect. one study provides evidence of the possible transfer of small ncrnas through the mammalian placenta . in addition, following a literature search as described in section . . . . and based on the methodology described in the section . . . , papers were selected as relevant to review toxicological effects of dietary exposure to exogenous ncrnas in humans or animals. the first report of systemic effects of dietary exogenous mirna in animals was published in by . plant mirnas were detected in human chinese healthy donors, among which mir- a and mir- a were highly expressed. these mirnas were also detected in calves and mice. plant mirna levels were relatively lower, but at a similar concentration range as endogenous mammalian mirnas present in serum. periodate oxidation, which oxidize mirnas but not plant mirnas (which are '-o-methyl modified), confirmed that these mirnas were genuine plant mirnas as most mammalian mirnas in human sera were oxidized and failed to be sequenced by solexa . the plant mirnas mir- a and mir- a were also detected in different mice tissues, while plant mir- a was not, although it was present in the sera. both mir- a and mir- a were primarily detected in microvesicles in c bl/ j mouse plasma. foods were found to contain the plant mirnas mir- a, mir- a and mir- a, including a chow mice diet at a concentration of . , . and . fmol/g, respectively. fresh rice contained between to -fold higher amounts than the chow diet. plant mirnas were detected in other foods including chinese cabbage, wheat and potato. plant mirnas were found to be stable in cooked foods. plant mir- a and mir- a were higher in the sera and liver of mice after h feeding with fresh rice. mir- a also increased in the stomach and small intestines, but not in the kidney, of mice fed a chow diet or fresh rice. mice fed with total rna extracted from rice or synthetic mir- a (methylated) showed an increased level of mir- a in serum and liver h after feeding. in vitro stability assays suggested that plant mirnas had a much slower degradation rate than their synthetic form (without '-o-methylated 'ends), suggesting that methylation has a protective effect on the stability of plant mirna . of note efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. is that mir- a targets the liver-enriched gene low-density lipoprotein receptor adapter protein (ldlrap ), which plays a role in facilitating the removal of ldl from the circulatory system (garcia et al., ) . direct binding of mir- a to exon of ldlrap , which is in the orf (open reading frame), was demonstrated by luciferase reported assays. transfection of both pre-mir- a and mature mir- a resulted in decreased ldlrap protein expression (but not that of mrna) in hepg cells. moreover, transfection of the intestinal epithelial caco- cell line with mir- a produced microvesicles with plant mir- a which can be taken up by hepg and reduce ldlrap protein expression. mir- a seemed to be associated to ago protein and other cells like t cells which were also able to package mir- a in microvesicles and then deliver mir- a into hepg . injection (iv) of antisense oligonucleotide against mir- a into mice during fresh-rice feeding reduced mir- a levels in the liver and reversed ldlrap repression. mice were finally fed with fresh rice for days to evaluate the physiological function of food-enriched in plant mir- a. serum and liver mir- a levels were induced already at day of feeding and the liver ldlrap protein was repressed after , and days. consequently, ldl levels in mouse plasma were significantly elevated on days and after fresh rice feeding. administration of an antisense oligonucleotide against mir- a reduced plasma levels of mir- a, repressed reduction of ldlrap in the liver and blocked the rice-induced elevation of plasma ldl levels, suggesting that elevation of fresh-rice derived mir- a in the mouse liver specifically decreased liver ldlrap expression and that this caused elevated ldl levels in mouse plasma (figure ). this study provides the first overall evidence of cross-kingdom regulation of gene expression through dietary plant mirnas. the authors further tested a mature mammalian mir- added to the chow diet for mice, and reported an increase in serum and liver levels of mir- after three days , leading to downregulation of liver c-myc expression . the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. by analysing western human sera samples from female stage ii-iii breast cancer patients, chin et al. detected multiple plant mirnas from arabidopsis thaliana and soybean (glycine max), but with sparse counts (chin et al., ) . among the detected plant mirnas, ath-mir- a, gma-mir- a- p and gma-mir- e- p had the highest counts (≤ read counts) and were over times more abundant than other plant mirnas. circulating mir- was more abundant in female healthy donors (n= ) than breast cancer patients (n= ) as analysed by qrt-pcr. moreover, circulating mir- was less abundant in patients who relapsed with metastatic disease, suggesting that human serum mir- levels inversely correlate with breast cancer incidence and progression (chin et al., ) . mir- in human serum was detected in an extracellular vesicle-enriched serum fraction obtained by ultracentrifugation and was resistant to sodium periodate oxidation, suggesting its plant origin due to '-o-methylation of the ' end (which makes plant mirnas more resistant to periodate oxidation). in situ hybridization confirmed the presence of mir- in tumour samples, suggesting that mir- in human serum is capable of entering breast tissue (chin et al., ) . the authors analysed several plant food items for the presence of mir- and found that broccoli is particularly rich in this mirna and most of this mirna was still present after cooking (boiling for min, > % remained stable, compared to fresh). in vitro studies showed that synthetic mir- (both double-stranded and '-o-methylated) reduced cell proliferation in breast cancer cell lines but not the non-cancerous breast cell line. extracellular vesicles from healthy human donors also have a similar effect. luciferase assays determined that mir- targets transcription factor (tcf ) and binds to two different sites within the ' utr. tcf is a transcription factor of the wnt signalling pathway and is upregulated in breast cancer. in vitro assays also showed that mir- inhibits breast cancer cell growth by targeting tcf . oral gavage of synthetic '-o-methylated mir- ( mg/kg mir- or scramble control oligos, daily) reduced tumour growth from day of treatment until the end of xenograph experiments ( weeks) (figure ) , an effect that was lost when tumour cells stably overexpress tcf without the ' utr (chin et al., ) . mir- was detected in xenografts of tumours by in situ hybridization analysis. overall, these experiments suggest that plant mirnas can inhibit cancer growth in mammals. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. liang et al. evaluated and quantified plant mirnas in human plasma after consuming a single dose of watermelon juice or a mixture of fruits (liang et al., b) . using taqman probe-based qrt-pcr assays and appropriate non-template controls, sixteen plant mirnas including ath-mir- a, mir- a, mir- a, mir- a, mir- a, mir- a, mir- a, mir- a, mir- a, mir- a, mir- a, mir- a, mir- a, mir- , mir- and mir- were evaluated and a standard curve generated for quantification. mirnas concentration ranged from . pm for mir- a to , . pm for mir- in watermelon juice, and from . pmol/kg for mir- a to , . pmol/kg for mir- in the fruit mixture. watermelon juice was orally administered to nine healthy male adult volunteers and plasma samples were taken . , , , and h after drinking . l of juice. different mirnas followed different peak time or appearance kinetics in plasma after administration (liang et al., b) . using six plant mirnas ( table ) that showed typical kinetic absorption curves, the total amount of plant mirnas absorbed by the body was calculated. using the plant mirnas concentration in watermelon juice and the areas under the plasma concentration-vs-time curves (auc) of plant mirnas, the total uptake amount, total absorption amount and absorption rate for each plant mirna was calculated (table ) . absorption rate ranged from . % to . %. mir- a, mir- a and mir- a were found almost exclusively encapsulated in microvesicles. mirnas were validated in watermelon juice or plasma by northern blotting. using the same protocol, a fruit eating study was performed. each subject consumed an equal portion of watermelon, apple, banana, orange, grape, mango and cantaloupe ( . kg in total). while mirnas concentration varied, the kinetics of plant mirnas after fruit administration appeared similar to the kinetics after juice administration (liang et al., b) . the authors also suggest that the real concentrations of plant mirnas may be underestimated and, for instance, the real concentration of mir- a in the watermelon juice might be six times higher and its plasma levels four times the previously calculated (table ) concentration (liang et al., b) . adapted from (liang et al., b) in a comment (correspondence) to the editor, the results of above article were criticized as being either a result of technical artefacts or a contamination problem, due to the presence of mir- in monocots and not in the dicot watermelon (witwer, ) . in a reply letter, the original authors indicated that mir- also exists in dicots and that the mirnas display a dynamic physiological kinetic absorption curve in human plasma, supporting the concept of uptake of food-derived mirnas by humans and animals (liang et al., a) . efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. analysing mir- by qrt-pcr in dried flowers and herbs used in chinese medicines or as tea, yang et al. reported different levels in different products including: sophora (≈ fmol/g), honeysuckle (≈ fmol/g), chamomile (≈ fmol/g), blue mallow (≈ fmol/g), ginseng flowers (≈ fmol/g) and as low as . fmol/g in hibiscus . herbal feeding of male mice for days increased plasma circulating levels of mir- compared to that of chow-fed animals. honeysuckle increased -fold ( fm) in sera levels, while chamomile increased -fold ( fm), sophora -fold ( fm), lavender -fold ( fm), blue mallow increased -fold ( fm), ginseng increased -fold ( fm) and no significant changes were detected in hibiscus ( fm). post-feeding urine levels of mir- ranged from -fold higher ( fm) for honeysuckle, and -fold higher ( fm) for chamomile and to -fold higher ( fm) for lavender . however, when chemically synthesized mir- ( '-o-methylated, plant-specific) was administered after single dose feeding of pmols, mice serum levels were elevated roughly -fold within min after gavage but decreased to background levels ≈ h after gavage . mir- was not associated with ago in sera and remained in the unbound fraction. clearance of mir- and other plant-derived synthetic mirnas (mir a, mir a and mir ) were analysed in mice by tail vein injection of fmols. sera collected from mice ( min, min, h, h and h post injection) showed that most mirnas were rapidly cleared from circulation. compared to the other mirnas administered at equal doses, mir- levels were substantially higher at min after injection but after h the apparent clearance of all srnas tested was complete . in the same study, gut microbiota did not seem to influence mir- absorption despite its -fold increase in their microbiome titre after honeysuckle consumption. honeysuckle feeding of mice was also found not to influence gut permeability, discarding its possible absorption due to changes in gut permeability . rapeseed (brassica campestris) bee pollen was evaluated for the possible absorption of plant mirnas in male mice after pollen oral feeding ( g/kg). mirnas were analysed by srna sequencing after or h in mice serum ( µl). from ≈ m clean reads, only reads were plant mirnas, which belonged to mirnas. of the plant mirnas, bna-mir- a ( reads) and bna-mir- ( reads) showed the highest levels in mouse blood and were mapped to the rapeseed genome. the rest of the plant mirnas had ≤ reads, of which mirnas had ≤ reads. both mirnas, mir- a and mir- , were also detected in the rapeseed bee pollen as evaluated by qrt-pcr, with higher abundance for mir- a. serum levels of mir- a after h of rapeseed bee pollen ingestion versus a control were ≈ pm for treated animals while in chow diet fed animals it was ≈ . pm, suggesting that plant mirnas from rapeseed bee pollen can be absorbed by mice by oral ingestion and detected in systemic circulation . evaluated herbal medicine honeysuckle mirnas before and after decoction (boiling for min, ≈ . g/ml water). from ≈ m reads, ≈ . m reads of mirnas were obtained, from which osa-mir- g, peu-mir- and peu-mir- contained more than . copy numbers. most mirnas were degraded by decoction (i.e. mir- g degraded from . to reads), while mir- remained high ( . reads) reaching % of total mirnas in the final honeysuckle decoction. concentration analysis of mir- showed a reduction from . pmol/g in honeysuckle to . pmol/ml in decoction when analysed by qrt-pcr, or from ≈ . pmol/g to . pmol/ml when assessed by northern-blot . compared to other plant mirnas, the apparent high resistance to boiling of mir- was also supported by its high resistance to rnase treatment, which was dependent on its unique sequence and high gc content. oral administration (n= per group) of a single dose ( µl) of honeysuckle decoction (mir- concentration ≈ . pmol/ml) resulted in a maximum peak level in mice plasma and lung tissues h after ingestion (i.e. plasma levels increased from . to . fm). most plasma mir- was found in cell-derived microvesicles and associated efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. to the ago complex. continuous drinking of decoction during three days ( . pmol/ml, ≈ ml/day) increased plasma levels from . to fm, while no changes were observed in the control mice (from . to . fm). levels of mir- in lung and liver tissues increased from to -fold, while the intestines and kidneys remained unchanged. synthetic mir- administration ( nmol/ml, µl given once per mouse, n= per group) resulted in an increase at peak ( h) from . to . fm in plasma and also an increase at peak ( h) in lung tissue, suggesting overall that atypical dietary mirna mir- is absorbed and delivered to tissues . studying its possible biological function, zhou et al. found influenza virus h n encoded mrnas targets of mir- , i.e. virus-encoded proteins pb and ns , which are relevant for viral replication (zanin et al., ) . luciferase activity assays confirmed that mir- binds to pb and ns , which is lost by point-mutation of mir- binding sites of the individual viral gene sequences. in vitro h n replication was reduced by synthetic mir- or total rna extracted from honeysuckle decoction. the inhibitory effect was abolished by co-transfection with anti-mir- antagomir. moreover, no effects in viral replication were observed when pb and ns mutant virus were used with synthetic mir- or honeysuckle decoction rnas . in vivo administration of honeysuckle decoction or synthetic mir- ( . nmol per day) by gavage one day before and during days after inoculation reduced weight loss and viral titre in lung tissues. these results were abolished when the anti-mir- antagomir was used concomitantly. moreover, weight loss and high viral titre were not affected when pb and ns mutant virus were used, and synthetic mir- or honeysuckle decoction administered . the authors also showed similar effects for synthetic mir- or honeysuckle decoction with other influenza viruses, including h n and h n , tested in vitro and in vivo (figure ) . the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. regarding this particular mirna, yang et al. observed a different bioavailability of plant-derived mir- vs. its synthetic '-o-methylated counterpart . oral administration of cabbage extract containing . pmol of mir- per µl gavage volume was compared with a -fold higher dose of synthetic '-o-methylated mir- ( pmoles per mouse). after gavage, mice plasma levels of cabbage mir- reached an approximately -fold increase (≈ fm) versus that of synthetic mir- (≈ fm), suggesting that mir- derived from plants is more bioavailable than synthetic rnas . whether this observation can be generalized to other dietary plant mirnas is unknown. the authors also showed that mir- was not present in exosome vesicles that were proteinase-k resistant in circulation . du et al. evaluated the presence of srnas in hong jing tian (hjt, rhodiola crenulata), which is a chinese herbal medicine. male c bl/ j mice were fed with hjt rnas by gavage for three days ( µg/kg per day) and lung tissue collected at , , or h (n= mice per time). the srna sequences of lung tissues (top eight ranked) that were also present in the plant hjt had ≈ k read counts . one of the small rna (hjt-srna-m ) was effective in reducing in vitro transfected gene expression vectors for α-sma, fibronectin and collagen type i α (col a ) in a tgf-β induced fibrosis model. although the type of srna is not clearly described, hjt-srna-m targets the ' utr of these three genes. using the hjt-srna-m agomir (cholesterol-conjugated srna), by intratracheal administration ( mg/kg in µl of saline) (n= mice per group), the authors showed that fibrosis was reduced in a bleomycin-induced fibrosis mouse model . due to the relevance of liposomes in delivering rna molecules, the authors evaluated the possible formation of liposomes during decoction (boiling for min) with plant lipids. they found that phosphocholine lipid molecules were also present in the decoction and suggested that liposomes may be formed that might facilitate the entry of rnas from plants to human cells and tissues; this was not tested . in another study, mlotshwa et al., described oral administration of a cocktail of tumour suppressor mirnas (normally downregulated in colon cancer) and a reduction of tumour burden in the apc min/+ mouse (c bl/ j) model of colon cancer . mir- a, mir- and mir- were chemically synthesized with the same mice sequence, but with a methyl group on the ' position of the ribose at the ' end, which is a chemical characteristic of plant mirnas (see section . . ). for days (starting at weeks of age), mice (n= per group) received either total plant rna spiked with the mix of the three mirnas, total plant rna alone (from a. thaliana) or water. oral dose was µg of total plant rna alone or plus . to . µg (corresponding to . to . nmole) of each species of tumour suppressor mirnas. tumour burden was dramatically reduced in mice receiving the mirna mixture. moreover, mice receiving the plant rna alone also showed a reduced tumour burden, although it was not statistically significant. after periodate oxidation, intestinal levels of mir- a were -fold higher than that of the group receiving water alone . mir- and mir- intestinal levels failed to be detected due to high background of endogenous mice mirnas after periodate oxidation. although the study does not directly evaluate the mirnas produced by the plant, mirnas designed to mimic small rnas produced in plants seemed to be taken up by the digestive tract of apc min/+ mice upon ingestion and exert a biological effect . oral ingestion of total rna isolated from brassica oleracea ( µg) resulted in the detection of plant mir- (the most highly enriched exogenous plant mirna in b. oleracea) in blood of icr mice between h and h after feeding . mir- was also detected in the spleen ( . - . % recovered of orally administered mir- ), and merely detected in the liver or kidney. no evidence of sex differences was reported . at lower doses of total rna ( µg or µg), no mir- was detected in the blood of mice . efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. using pigs as experimental animals, luo et al. evaluated the detection of plant-derived mirnas in the serum and tissues after feeding fresh maize to pigs (sus scrofa). jinhua female pigs consumed a fresh maize diet and water ad libitum for days (n= ), and blood and different tissue samples were obtained for plant-derived mirna analysis. eighteen maize mirnas were evaluated ( were detected) in serum and tissues, and exhibited relatively low abundance in the pancreas and longissimus dorsi muscle tissue . periodate oxidation treatment of the samples confirmed the detection of zma-mir- a- p, zma-mir- e- p, zma-mir- a- p, zma-mir- a- p, and zma-mir- a- p (while the rest were completely degraded), suggesting that these mirnas are bona fide plant mirnas (figure ). in a separate experiment, female pigs were fed one meal with fresh maize ( kg per pig) after overnight fasting. serum was collected at different times (i.e. , , , , , and h). after h, pigs were provided with fresh maize diet ad libitum, and serum collected at , , and days and tissue collected after sacrifice at days. the serum levels of some of these mirnas (mir- a- p, mir- a- p, mir- e- p, mir- a- p and mir- a- p) gradually increased after one feeding of fresh meal, reaching peak values at or hours within the first hours and maintaining stable detectable levels (by qrt-pcr) in the serum following days of access to fresh maize. the changes in mirna expression at all measured time-points in most cases were modest (≈ to -fold increase). the five mirnas were primarily present in serum exosomes (≈ . % of concentration in the serum) ( figure ). using bioinformatics, potential targets were determined within the porcine mrnas for zma-mir- a- p. using in vitro luciferase assays with porcine kidney cells, the authors showed that zma-mir- a- p binds to three of these potential target genes including cspg , otx and plagl , an interaction lost when their mutant constructions were used, suggesting overall that dietetically-absorbed mirnas can target endogenous porcine mrnas . the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. several of the above-mentioned studies have reported the presence of dietary plant mirnas in circulation associated to extracellular vesicles or exosomes (chin et al., ; liang et al., b; . this could be relevant for their possible biological effects, as extracellular vesicles can deliver a cargo to several tissues, including crossing the blood-brain barrier, and other biological barriers such as the placenta (matsumoto et al., ; yang, t et al., ; jayabalan et al., ) . their presence in microvesicles could modify their function, for example, it has been described that in some cases delivered naked mirnas cannot exert the same effects as those exerted by delivered exosomal mirnas (thomou et al., ) . also, exosomes as therapeutic drug carriers and delivery vehicles are gaining relevance in therapeutics (aryani and denecke, ; ha et al., ) . recently, it has been described that edible plant-derived exosome-like nanoparticles can be isolated from these plants (ju et al., ) , can transport ncrnas (i.e. mirnas) (mu et al., ) and can exert a biological effect (raimondo et al., ) , highlighting their role as therapeutic nanoparticles . it is still unknown if these exosome-like nanoparticles contribute to the resistance of ncrnas in the harsh conditions of the gi tract and to the absorption of plant ncrnas or the formation of exosomes circulating in biological fluids. regarding other biological barriers, only one study has reported on the transfer of small ncrnas through the placenta. li et al. found that exogenous plant mirnas were consistently detected in the umbilical cord blood and amniotic fluid of healthy chinese pregnant woman . this belongs to foetal circulation system, suggesting that mirnas may transfer through the placenta to the foetal side. three hours after gavage feeding of synthesized mature mirnas of the influenza virus to pregnant mice (at last -day pregnant and with mature placenta) increased mirnas levels were detected in the maternal plasma. an increase of exogenous mirnas levels in the foetal liver was also observed . to determine if mature plant mirnas can pass through the placenta and efficiently enter foetal organs, the traditional chinese herbal honeysuckle was tested. mice were gavage fed with . ml honeysuckle decoction ( . pmol/mice of mir- ) and maternal plasma and foetal liver collected h after gavage feeding. plant mir- levels were elevated ≈ . -fold in the maternal plasma (reaching fm), ≈ -fold in the placenta and ≈ . -fold in the foetuses, suggesting that plant mirna mir- in honeysuckle can efficiently transfer through the placenta to enter the foetal liver (figure ) . the amount of circulating mir- increased only in microvesicles, suggesting that circulating mir- was primarily present in microvesicles and that transplacental transmission may involve a microvesiclesmediated pathway . the same study also tested gavage fed synthetic sirnas ( nmol), which were found in the plasma and foetal liver. protein targeting of the sirna was dramatically reduced, suggesting that exogenous sirnas delivered from the mother to the foetus by trasnplacental transmission may regulate foetal gene expression. direct injection of microvesicles loaded with sirnas also reduced their mrna target in the foetus liver, suggesting overall that exogenous small ncrnas can be transplacentally transmitted from the mother to the foetus. although outside the scope of this review, other non-plant derived exogenous ncrnas have also been reported to exert systemic effects in animals. bovine milk-derived extracellular vesicles (bmevs) were orally administered to il- ra -/mice on the balb/c background, which spontaneously develop polyarticular arthritis, from week to week . drinking water was also supplemented and administered to a collagen-induced arthritis (cia) dba/ j mice model starting at week before immunization to day . mice receiving the higher dose of x bmevs (≈ micrograms/ml) showed a considerable delay in disease onset in the il- ra -/mice model. moreover, in the cia model the higher dose administered in drinking water . x /ml (≈ microgram/ml) reduced arthritis incidence and severity, accompanied by reduced expression of inflammatory cytokine il- and mcp- . although bmevs contained mirnas mir- a, mir- and mir- a, it is unclear if they are efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. responsible for the observed biological effects. in a similar study, oliveira et al. administered bmevs (particle concentration of . x /ml or . x /ml) in the drinking water to female dba/ j mice for weeks and observed an increase in osteocyte numbers. moreover, the highest concentration of bmevs increased the percentage of woven bone compared to the pbs group, and a reduction of the adipocyte area in the bone marrow (oliveira et al., ) . although the bmevs also contained mir- a (oliveira et al., ) , its implication in the biological effects is unknown. exogenous plant-derived ncrnas can pass the placenta and reach the foetus. by analysing small rnas components in royal jelly, honey, beebread and pollen collected during the cabbage (brassica campestris) flowering stage, zhu et al. found that honeybee srnas were present at a far higher level in royal jelly than in beebread and pollen, while the abundance of cabbage small rnas gradually increased from royal jelly to honey, beebread and pollen . several mirnas were detected in the samples, the plant mirnas occurring at far higher concentrations. moreover, beebread and polled mirnas were at a much higher concentration and their composition of individual mirnas was highly similar. the plant representative mirnas with the highest concentration were mir- a, mir- a, mir- a, mir- a, mir- a, mir- a, mir- g, mir- a, mir- a, mir- a, mir- c, mir- a, mir- a, mir- , mir- and mir- a. the same analysis in samples collected during the camellia (camellia japonica) flowering stage showed similar results and out of the enriched mirnas were similar, suggesting that plant mirnas in beebread and pollen may not be very diverse between different sources . absolute concentration of the selected plant mirnas ranged from < . fmol/µg of total rna (almost undetectable) in royal jelly and honey, to a maximum of ≈ fmol/µg total rna in pollen and beebread. feeding experiments to larvae with either rna purified from cabbage pollen (levels of mirnas similar to natural beebread mirnas levels) or a efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. synthetic mix of the plant mirnas, increased whole body accumulation of the representative plant mirnas and developed worker bee-like characteristics (reduced weight and size, extended pre-adult development time and decreased ovary size). ninety-six honeybee genes were predicted to be targeted by the plant mirnas, some of which are known to influence the developmental fate of honeybees (i.e. apis millifera tor, amtor). luciferase assays showed that mir- a directly targets aptor mrna. dietary supplementation of larvae food with synthetic mir- a decreased amtor mrna levels in honeybees and showed reduced body weight, size and ovary size . similar experiments were performed in drosophila melanogaster (dm) with total pollen rna, a synthetic mirna pool or mir- a alone, and in all cases similar phenotypes were obtained (delayed drosophila development, reduced final size, ovary size and fecundity). dmtor was also a target of mir- a. overall, this study implies that plant rnas can be transmitted between species of different kingdoms. the literature also describes some in vitro studies of plant mirnas and their possible effects on mammalian cells. for example, exposure ( h) of plant mirnas extracted from a glycyrrhiza uralensis decoction to human pbmcs produced apparent cell aggregation and increased cell number and hla-dr+ cell proportion (shao et al., ) . in silico studies have also attempted to predict possible plant mirnas with potential targets in humans . as described before (see section . ), in contrast to mammalian mirnas, plant mirnas possess a '-omethylation at the '-end that stabilizes and provides resistance to rnases and oxidation. this characteristic has been used to improve the quantification method for mature exogenous mirnas from plants in biological mammalian matrices . thus, periodate oxidation and βelimination were introduced to evaluate the origin of exogenous mirnas. male c bl/ mice were fed with corn/soy-based rodent chow diet for weeks. plant mir a, mir a and mir a were evaluated. these mirnas were detected in abundant amounts in a corn/soy-based chow diet, but after periodate oxidation and β-elimination . %, . % and . % of mir a, mir a and mir a were determined to be in methylated forms ( . , . and . fmol/g of diet, respectively). the control diet showed no differences between the diet and non-template control ct values after periodate oxidation/β-elimination. this study also suggests the use of exogenous rnas (spike-in), both methylated and un-methylated forms of mirnas, to ensure appropriate periodate oxidation/β-elimination . no detectable levels of plant mirnas were found in plasma, liver or faeces samples from the mice fed the corn/soy-based chow diet, since no differences occurred between the diet and the non-template control after periodate oxidation/β-elimination . this suggests that false positive results can be resolved using treatment with periodate oxidation/β-elimination. plant mirnas from extra virgin olive oil (evoo) and beer have also been evaluated by small rna sequencing. however, very few sequences (≤ reads) were identified, and most aligned to soybean and orange . a feeding study of a single dose ingestion of ml of evoo in healthy adult volunteers (n= ) was performed. mirnas analysis at baseline and after h ingestion was performed. no plant mirnas were detected in plasma after single ingestion of evoo . only one read count was found for other species mirna vvi-mir - p from grape, while two read counts were found from medicago trunculata, grape or brachypodium distachyon when mismatches were accepted . this suggests that beer or evoo do not contain mirnas that contribute to dietary intake, even though mirnas were reported in the pulp of olive (olea europaea) (donaire et al., ) . efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. dickinson et al. evaluated a controlled mouse feeding study with rice-containing chow diets (modified ain -g with . % of rice or rice-based with % of rice) or with a purified synthetic chow devoid of plant grain or forage for , and days (figure ) . the "synthetic chow" had negligible levels of osa-mir- a, the "balance rice chow" ( . % of rice) contained reads, while the rice grain contained reads. the concentration of osa-mir- a in each diet was . , and fmol/g of diet for synthetic chow, balance rice chow and rice chow, respectively. plasma and liver analysis of mice fed the diets did not reveal measurable uptake of any rice grain mirnas, including osa-mir- a. indeed, fewer than ten reads were detected in five out of eight samples from mice fed on rice-containing chow and four out of five samples from mice fed on synthetic chow . since the synthetic chow contained no grain or forage from plants, this low number for rice mirnas-mappable reads can only be explained by sequence errors or cross-contamination. in agreement with , the authors indicated that animals fed a chow containing high levels of uncooked rice (%) had significantly increased plasma low-density lipoproteins (ldl) levels at and days after treatment initiation. however, this increase in ldl was not observed in the balanced rice/chow group ( . % rice), which contained fmol/g osa-mir- a. the increase of ldl was attributed to nutritional imbalances between the test and control groups . ldlrap protein was also evaluated by elisa in mouse liver samples, and no changes in levels were observed in any group at any time, suggesting that dietary intake of osa-mir- a (from to fmol/g) does not produce rnai-mediated modulation of ldlrap protein levels in mouse liver . efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. a commercially available plant-based, plant mirna-rich substance containing soy and fruit materials, but not animal products, was administered by gastric gavage (≈ % of estimated blood volume for each animal) to male macaques (macaca nemestrina). relative abundance of plant mirnas mir- , mir- , mir- , mir- , mir- and mir- were analysed by qrt-pcr in the plant-based dietary substance. for instance, for mir- a consistent and efficient amplification through at least pcr cycles was detected. plant mirnas were analysed in plasma samples of macaques (n= ) before and after , and h oral gavage of plant rich mirnas. qrt-pcr results indicated late amplifications of some plant mirnas, but results were highly variable. mir- did not amplify before cycles, and mirs- , - , and - had a median cq greater than . mir- non-template controls also showed regular amplification within the same cq range. mir- exhibited a tendency to increase after ingestion, but was highly variable, and cq cycles were greater than . this was not the case for endogenous animal mirnas or the spike-in control, supporting the case that the late apparent amplification of plant mirnas in plasma was non-specific . moreover, droplet digital pcr analysis showed that counts from plasma were generally very low and of variable intensity, as occurs in non-specific amplifications. this even occurred in mir- (plant mirna with relatively high apparent count by qrt-pcr), which showed large number of counts but with variable intensity plots ("rain" appearance) (figure ) , and is consistent with non-specific amplification . overall, the study (though only subjects were included) suggests that the detection of plant mirnas in plasma does not support a general and consistent uptake of dietary plant mirnas . : substantial levels of mirna in oral diets consumed by macaque but negligible steady-state presence of diet-derived mirnas in plasma or organ tissues. snow et al. ( ) analysed the presence of exogenous plant mirnas after regular intake of a western diet containing fruits in healthy human subjects, and mir- in plasma or organ tissues of mir- -/mice . the conserved plant mirnas mir- a, mir- a and mir- a were highly the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. expressed in many fruits common in the human diet such as ripened avocado, apple, banana, orange and cantaloupe. mir- a and mir- reached up to , copy numbers/mg food. the conserved animal mirna mir- was expressed at substantial levels (≈ , copies/mg food) in meat but not in fruits. mouse vegetarian and soy-enriched diets also contained high quantities (up to , copy numbers/mg food) of mir- a and mir- a, but not mir- . in contrast, mouse diets enriched with animal products contained elevated levels of mir- , suggesting that conserved plant and animal mirnas are commonly present in oral diets for various animal organisms. although the dietary record of of the healthy adults reported the intake of fruits replete in mir- a, mir- a and mir- a the day prior to plasma harvest, these plant-derived mirnas were undetectable in plasma samples. following -week feeding with a custom animal lard diet containing mir- no detectable levels of mir- were found in plasma samples of mir- -/mice, while high quantities of this mirna in plasma were seen in control wild type mice fed the same diet . in tissues (liver, lung, kidney and stomach) mir- was either not detected or detected at exceedingly low levels corresponding to less than one copy of mir- per cell (assuming that the mammalian cell expresses roughly pg of total rna). wild type-mice showed only a non-significant trend toward increased plasma levels of mir- a after consuming either a vegetarian or soy-enriched diet when compared to mice fed a lard diet for the same period ( -week) ( figure ) ; however, levels were very low. indeed, mir- a was detected in organ tissues (liver, lung, kidney and stomach) but at levels of less than one copy of mirna per cell and regardless of diets. mir- a and mir- a were not detected at all. fresh avocado containing three specific plant mirnas was also fed to mice ( . g per mouse over h). even though the three mirnas were found in the stomach content of mice at high levels, very low levels were found in plasma and organ tissues, reaching levels lower than one copy mirna per cell in tissues. this provides no evidence of substantial steady-state presence of these plant mirnas in organ tissues after dietary intake . assuming that, on average, at least copies per cell (> copies/pg srna per cell) are necessary to achieve canonical target gene repression (brown et al., ) , at least copies of a given plant mirna must be ingested and delivered for widespread biological activity in humans (the average human body contains ≈ cells). based on the concentration in ripe cantaloupe (≈ x copies mir- a/mg fruit), an individual would need to consume , kg of cantaloupe just to release this number of copies in the gi tract . in terms of srnas, the literature describes the threshold for target gene regulation to be between and copies of mammalian mirna per cell . in an analysis of activity for hundreds of mirnas, only the most abundant mirnas in cells were found to mediate targeted repression and a large amount of mirnas had no discernible activity (mullokandov et al., ) . this suggests that only a minimum amount of mirnas need be reached to exert a biological activity, although this will also depend on the amount of target transcripts. a study mentioned above the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. analyses of other common plant mirnas reported by others to be bioavailable were not successful. this implies that although the transgenic mirnas had resistance to degradation comparable to that of mir- and high expression levels in plants, they were not readily bioavailable . huang (huang et al., ) analysed the possible bioavailability of orally-ingested corn mirnas in mice when incorporated into a rodent diet and consumed for days. in an initial experiment, male c bl/ mice (n= per group) received either water (control), random nucleotides ( µg, equivalent to the same amount of small rnas) or purified small rnas isolated from corn kernel ( µg, equivalent to the same amout of random nucleotides). in the second experiment, mice received either a control diet (ain- m), ain- + % autoclaved corn kernel powder or ain- + % fresh corn kernel powder. mirna levels in the isolated rna contained . pg mir- a, . pg mir- a, and . pg mir- a per µg corn srna isolates. the diet supplement with fresh corn powder contained pg mir- a, pg mir- a, and pg mir- a per gram of diet. autoclaving the corn powder at ºc for min reduced the amount of initial mirnas by more than %. in the first study, liver and whole blood sample mirnas (analysed by qrt-pcr), showed no differences in ct values between the three groups. after periodate oxidation followed by β-elimination, no differences were detected between the no-template control group and the experimental groups. similar observations were made in the liver and whole blood in the second study using fresh corn powder. also, after periodate oxidation followed by β-elimination, no differences were detected between the no-template control group and the experimental groups in the liver and blood samples. analysis of corn mirnas in cecal and faecal samples www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. showed similar results after periodate oxidation, although certain minor differences remained between the no-template control group and the experimental groups. in all cases, less than . % of total the mirna tested in both studies was recovered from the faecal samples. moreover, analysis of the mirnas in the content collected from different parts of the mouse gi tract from the animal studies administered via gavage or dietary intake showed that calculated recovery accounted for less than . % of the content originally ingested in the stomach, less than . % of that originally ingested in the intestine and faeces, and less than . % of that originally ingested in the colon and cecum (huang et al., ) . further experiments using in vitro digestion system suggested that after the gastric phase, over % of corn mirna mir- a from the diet or from the extract were degraded, suggesting overall that significant degradation of corn mrnas occurred during digestion, which resulted in minimal uptake of corn mirnas after oral intake (huang et al., ) . witwer re-analysed the plasma data evaluated by liu et al., and found that only one putative plant mirna mapped above a median cut-off ( reads), which was the plant mirna peu-mir- . this implies that all rnas -including previously reported exogenous mirnas (xenomirs) such as mir- a, mir- a and the plant ribosomal degradation fragment mir- -are below the level of background noise. it is not clear if mir- is a real mirna as it is found in the highly conserved and expressed subunit rrna of plants . moreover, the mir- sequence is not only a fragment of plant rrna, but also has a % coverage and % identity match with human s rrna, while others do not map to plant genomes at all . perhaps more analyses are needed when assessing possible xenomirs. after in silico xenomirs analysis using public sequencing datasets (see section . . for details), kang et al. performed controlled feeding studies to detect the transfer of exogenous plant mirnas to rat blood or from bovine milk sequences into piglet blood . adult rats (n= per group) followed a -day controlled feeding study with supplementation in three different diets: monocot plant material (rice); dicot material (potatoes); and husbandry chow containing a defined mixture of grains, cereals, vitamins, minerals and fats. small rnas were isolated from serum and sequenced. most samples showed either a lack of plant xenomirs in rat serum, or three or less read counts in certain samples . these extremely low numbers suggested a probable false positive result. in the same study, piglets were fed either cow milk or soy milk for weeks followed by weeks of feeding with maize. few cow-specific sequences were found ( reads) in the piglets fed cow milk, while piglets fed maize as well did show cow-specific sequences ( reads). this suggests likely false positive results. both studies provide no evidence of dietary transfer of xenomirs . the abovementioned studies are summarized in table . although outside the scope of this literature review, pollen plant mirnas ingested as part of a typical diet of adult honey bees were not found to be robustly transferred across the epithelial barrier under normal conditions (masood et al., ) . for example, despite the high levels of plant mir- a in pollen or its detected levels in the bee digestive tract ( . copies per pg total rna in the midgut), less than . copy per pg total rna was found in abdominal tissue (masood et al., ) . www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. large amount of plant mirnas mir- a, mir- a and mir- a detected in vegetarian, soy-enriched, or avocado diets (i.e. ≈ x copies for vegetarian soy/diets or ≈ . x copies for avocado diet of mir- a after h), but very low levels found in plasma and less than copy per/cell in tissues. after weeks mir- rich diet in mir- ko mice, no plasma levels and less than one copy of mir- per cell detected in tissues. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. after periodate oxidation, no major differences observed in mir- a, mir- a, or mir- a among treatment, control groups or no-template control in liver and blood. in cecal and fecal samples no major differences observed among control and treated groups. mirna analysis in the gi content collected from different parts of mouse showed that the recovery (calculated) account for less than . % of originally ingested in the stomach, less than . % of originally ingested in the intestine and feces, and less than . % of originally ingested in colon and cecum. in vitro digestion experiments suggested that after the gastric phase, over % of corn mirna mir- a from the diet or from the extract is degraded. (huang et al., ) the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. exogenous plant mirnas found in umbilical cord blood and amniotic fluid of healthy chinese pregnant women. plant mir- increased in plasma of the mothers, placenta and foetus liver h after oral gavage ( . nmol/mice). exogenous sirna gavage ( nmol) increased plasma levels of mother and foetus liver. protein target in foetus repressed. direct injection of microvesicles loaded with sirnas also increased its levels in foetal liver and reduced mrna expression of its target. exogenous small ncrnas can be transplacentally transmitted from mother to foetus. n.d., not determined. ko, knock out; wt, wild type; mirna, microrna; srna, small rna; dsrna, double-stranded rna; col a , collagen type i α ; α-sma, alpha smooth muscle actin; hjt, hong jing tian; www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). the available evidence presents several examples of systemic effects of plant-derived exogenous ncrnas when administered orally. however, there are also several reports that do not support these findings and contradict the hypothesis of cross-kingdom regulation. in either case, the essential question concerning the existence of this possibility is still heavily debated. important aspects such as the precise mechanism of transport of plant ncrnas from food to the circulatory system, the amount of exogenous ncrnas reaching tissues or the molecular mechanism of cellular uptake need to be understood. it remains unknown if such a transfer could be modulated in particular context (i.e. specific diet, disease status, medication). the available evidence also suggests that plant ncrnas can directly target mammalian genes through rnai effects when they are exposed in in vitro systems. without considering other physiological parameters, if a synthetic plant mirna is exposed to its mammalian target, it can effectively bind by base complementarity and exert a biological effect (i.e. gene repression). however, whether the exogenous plant ncrna can bypass all the biological barriers to reach and interact with the possible target remains to be clarified. apparently, exogenous plant-derived ncrnas are found in exosomes or macrovesicles. it is not yet known how they reach these types of structure in biological fluids, or if novel plant exosome-like nanoparticles participate in any of these processes. www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). immune cells were mainly from studies in human and mouse isolated lymphocytes. however, these studies were descriptive/not comprehensive, reporting specific effects of ncrnas in proinflammatory cytokine production using human cells, and would not allow identifying specific endpoints that can be used to evaluate changes in the regulation of immune function and homeostasis by ncrnas. retrieval of grey literature using the key words 'thesis plant rna immune' provided numerous results (i.e. google = . ), mostly regarding the function of mirnas and rna silencing in plant immunity. for instance, a document could be chosen based on title selection because it included the terms 'plant, double-stranded rna and systemic immunity' (https://uknowledge.uky.edu/plantpath_etds/ ). however, this study was not further considered since it refers to plant immunity. after analysing the documents and refining by key questions, a large number of screened publications were discarded because they refer to the role of endogenous ncrna (human and animals) as well as viral infections in relation to the immune function, which is outside the scope of this literature review. as an example, within original in vivo publications related to the key words, only studies were considered relevant for the proposed review questions. in summary, from the literature search, papers were finally selected as relevant to review of the effects of ncrnas on immunity. very few studies analyse the direct relationship of exogenous plant ncrna with immune function and homeostasis of humans or animals. albeit controversial, exogenous ncrnas derived from plants and microorganisms have also been described in human blood (detailed in section . . .). overall, the results indicate that there is scarce information on the specific impact of plant ncrnas on activation of specific immunocompetent cells. of these references, documents were used for reviewing the general features of ncrnas in immune function and immunity in humans and animals (section . . .), were used for the effect of exogenous plant ncrnas on immune system (section . . .) and for the additional section on the effects of exogenous ncrnas on gut microbial composition (section . . .) ncrnas have been shown to play important roles in immune cell development and function in normal and disease conditions (ansel, ) . human and mouse studies demonstrate that mirnas play a critical role in the regulation of immune cell development and their function. there is also evidence of the relevance of neonatal mirna-mediated immune activation (kosaka et al., ) . here, mirna molecules of maternal origin appear to be stable in the infant's gut conditions, allowing dietary intake and transfer of mirna (kosaka et al., ) . ncrna have also been identified as regulators of metabolic shifts within microbial communities (paul et al., ) . thereby, ncrnas may significantly influence the critical roles of gut microbiota on health and disease. this part of the literature review (efsa task ) presents the current knowledge on the role of exogenous ncrna molecules on the immune system of humans and animals and aims to provide information on the possible effets of dietary ncrnas on the regulation and function of the immune system. in addition, since the gut microbiota influences the immune system, a review on the possible effects of exogenous ncrnas as gut microbiota modulators is provided this chapter provides background information on the modulatory effects of ncrna on the immune system of humans and animals and serves as a general introduction to explore the possibility of dietary plant ncrnas effect on the immune system after dietary intake. sixty-four ( ) documents were finally used to review this section. when activated, the immune system tightly regulates innate and adaptive response(s), aiming at restoring homeostasis, and preventing autoimmunity. the 'amplitude' (i.e. intensity and duration) of the www.efsa.europa.eu/publications efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). interaction with the t-(tcr) and b-(bcr) cell receptors determines cell fate decisions. in the periphery, regulatory t cells (tregs) play a key role in restraining the activity of mature b and t (th , th and th ) cells, and preserving tolerance mechanism(s). these responses depend on transcriptional and epigenetic regulation including regulation by mirnas. emerging systems for measuring mirna activity during immune activation revealed the complex network of genes that may be simultaneously targeted by mirnas to tune distinct cell fate decisions (wells et al., ) . rnas exhibit an intrinsic property of base pairs that is crucial in defining their structure and, thereby, their physiological role (lu et al., ) . plant mirnas are '-o-methyl modified on their terminal nucleotide, making them more difficult to be ligated to the cloning adapter. the inhibitory effect of '-o-methyl modification at the ' end of the rna substrate for the mammalian nuclease family member nef-sp has been described (silva et al., ) . as described elsewhere, most mammalian mirnas require 'pre-processing steps' to become physiologically active. nucleotide sequence complementarity appears as a critical feature driving the predominant result exerted by mirnas. notably, it has been reported that a modification pattern involving alternating '-o-methyl rna bases improves dsirna in vitro stability and evades activation of the innate immune system (collingwood et al., ) . these data were obtained with isolated peripheral blood mononuclear cells (pbmcs) as a mixed 'lymphocyte' population of immune receptors that are encountered in vivo. however, the authors do not provide any molecular mechanism for the observed effects. sirnas have been shown to be potent stimuli of interferon- production by plasmocytoid dendritic cells (hornung et al., ) . this sirna technology induced systemic immune responses in the same range as the toll-like receptor (tlr)- ligand 'cpg', including activation of t cells and dendritic cells in spleen. notably, immunostimulation by sirna was absent in tlr -deficient mice. there is a lack of data related to the effects of plant ncrnas on pge production, implicated in immunosuppression by tlr via cox . several distinct classes of lncrnas are transcribed from different dna elements or are derived from long primary transcripts with noncanonical rna processing pathways, generating new rna species with unexpected formats. these lncrnas can be processed by several mechanisms, including ribonuclease p (rnase p) cleavage to generate mature ' ends, capped by small nucleolar rna (snorna)-protein (snornp) complexes at their ends, or the formation of circular structures. recently, it was reported that several lncrnas mediate their regulatory effects through binding to specific rna-binding proteins . in addition, expression of lncrnas has been defined as highly cell-type specific (guttman et al., ; washietl et al., ) . this high specificity has been reported to be critical for the development and activation of immune lineages. however, lncrnas-mediated regulation of innate immune activation (i.e. il- production) has not been tested for function in vivo, but only evaluated in vitro (roux et al., ) . the regulation of inflammatory mediators or cytokines by lncrnas is still poorly understood. synthetic nucleic acids, such as dsrnas, have been reported to be recognized by toll-like receptor (tlr)- and can stimulate the innate immune system and trigger a type i interferon response (marques et al., ; mian et al., ) . in plants, it has been described that dsrna with ' overhangs contributes to endogenous and antiviral rna silencing pathways (fukunaga and doudna, ) . earlier reports defined a structural basis (i.e. ' overhangs) to cause nonspecific effects on the ds-rna-activated signalling pathways through the interferon responsive factor (irf)- (marques et al., ) . similarly, immune activation by the subcutaneous route with a viral mimic dsrna (poly i:c) of c bl/ mice during neonatal early or late brain development differentially affects depression-related behaviours developed in adolescent and adult mice (majidi-zolbanin et al., ) . the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). specific molecular interactions of the exogenous rna triggering immune responses have been reported. several studies have demonstrated the role of components of innate immunity, including toll-like receptors (tlrs), the retinoic acid-inducible gene i/melanoma-differentiation factor (rig-i/mda ) and the interferon response effectors protein kinase r (pkr) and rnase l, as well as mirnas in the recognition of viral or synthetic dsrna (malathi et al., ; urcuqui-inchima et al., ) . tlrs are a family of proteins recognizing different pathogen and damage-associated molecular patterns, pamps and dmaps, respectively. rig-i/mda , pkr and rnase l constitute cytosolic rna sensing proteins. the pathways responsible for these defined signalling processes can be largely separated (figure ) , despite the existence of some degree of crosstalk between them where the engagement of interferon regulatory factors is a common feature. tlrs play an essential role in innate immune responses in mammals. in humans, ten different tlrs recognize distinct molecular patterns, where tlr- , - , - and - stimulate innate immune responses upon interaction with nucleic acids weber et al., ) . tlr is located on the plasmatic cell membrane, while tlr- , - and - display endosomal localization. it has been reported that tlr recognizes dsrna, viral or the synthetic poly i:c, and sirna has also been identified (in the presence or absence of delivery systems) as a relevant ligand . notwithstanding, rna interference is considered an unlikely mechanism to be engaged for viral sensing (cullen, ) . gurich rna from viruses or synthetic single-stranded (ss) oligoribonucleotides displays an apparent preference for stimulating human tlr / -mediated immune effects (heil et al., ; forsbach et al., ) . a minimum of four nucleotides, uugu, found in the gu-rich region are reported as necessary to stimulate cytokine responses. endogenous or exogenous rnas are internalized into the endosomal compartment. the rna sensor system is composed of i) several innate immune 'toll-like' receptors (tlrs) located at the endosomal compartment that trigger different signalling pathways engaging adaptor distinct molecules (myd and trif), and ii) a cytosolic system (rig-i/mda- ) that coordinates its signalling with tlrs. antigen processing will lead to rearrangement of mhc molecules to activate the adaptive immune response(s). type i interferons (ifn-α/β) are expressed rapidly after infection and serve as a link between the innate and adaptive immune responses. : schematic representation of the eukaryotic rna sensor (single-stranded rna and short/long double-stranded rna) and its relation to innate immunity. the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). two classes of ssrna motifs have been described that either preferentially activate tlr -mediated or tlr / -mediated immune responses. au-rich oligoribonucleotides stimulate tlr -but not tlr mediated immune activation (forsbach et al., ) . rna motifs activating tlr -associated signalling fail to induce ifn- from tlr -expressing plasmacytoid dendritic cells but induce the secretion of th like and proinflammatory cytokines from monocytes or myeloid dendritic cells. in addition, rna motifs activating the tlr / -associated signalling pathway stimulate cytokine secretion from both tlr -and tlr -positive immunocytes (forsbach et al., ) . differences in the protein sequence of tlr- between different mammals (human, monkey, chimpanzee, cattle, porcine, mouse, and rat) can be found, and species-specific recognition of ssrna via tlr / had been described (heil et al., ) . the tlr -specific rna sequences are unable to trigger cytokine responses from mouse, rat, and porcine immune cells (forsbach et al., ) . a dual function has also been suggested for the murine coreceptor cd to enhance tlr- , - and - signalling (baumann et al., ; lee et al., ) . however, further studies showed evidenced that human cd did not appear to function as a co-receptor for tlr- or tlr (weber et al., ) .the innate immune system detects rna lacking nucleoside methylation (m c, m a and m u) or otherwise (i.e. pseudouridine or '-o-methyl-u) modified as a mechanism to selectively trigger immune responses to nucleic acids from necrotic tissues or of exogenous origin. a recent systematic study on transcriptional and, especially, post-transcriptional regulation of stressresponsive lncrnas in oryza sativa showed that hundreds of lncrnas with down-regulated polyadenylation are highly conserved in stresses . in the ascidian ciona intestinalis a novel alternative polyadenylation signal activated by the prototypical tlr agonist (i.e. lps) has been described (vizzini et al., ) . moreover, alternative polyadenylation has been identified as a regulatory mechanism during the innate antiviral immune response in macrophages (jia et al., ) . these authors showed an enrichment of polyadenylated genes and mrna abundance change in tlrs as well as rig-i-like receptor, jak-stat and apoptosis-related signalling pathways. studies conducted on the crystal structure of the sensing tlr- ectodomain allowed definition of dsrna recognition of at least bp (liu et al., ; leonard et al., ) . it was concluded that tlr assembles on dsrna as stable dimers and that the minimal signalling unit is one tlr dimer. recognition by secondary structure 'alone' has been reported, independently of the sequence motif, by tlr . this dsrna length is larger than the typical helix occurring in normal mirnas and sirnas. tlr- and - have been found to recognize single-stranded rna (ssrna). the existence of a similar double horseshoe structure for tlr- , - and - upon activation has been proposed, although their binding modes to their ligands remain unclear . often, in a partial view, tlr is considered as the ssrna-sensing tlr and as complementary to tlr . however, higher structural features have been identified as important aspects in tlr rna sensing (jöckel et al., ) . in addition, small molecules that may be viewed as nucleoside analogues are able to activate tlr and/or tlr (hemmi et al., ; lee et al., ) . it has not been defined if the recognition mode for small molecules, including srnas, resembles that of tlr for rna recognition. besides tlrs, rig-i is a cytosolic multi-domain protein sensing viral rna associated to n-terminal caspase activation and recruitment domains (cards) to transmit signalling but preventing it in the absence of rna. the identification of rig-i/mda as cytoplasmic sensors for dsrna, associated to facilitators such as pkr and '- '-olygoadenylate synthetase, suggests that additional 'aspects' (i.e. dependence on atp) other than dsrna structure or short size influence innate immune responses to srnas (i.e. sirnas and mirnas). these aspects remain unclear. the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). current information on mirnas points to clear differences in cellular recognition of endogenous and exogenous dsrna. here, '-triphosphate and dsrna represent molecular patterns enabling rig-i to discriminate exogenous from endogenous rna (myong et al., ) . dsrna products as short as - bp are sensed by rig-i independently from the '-termini (marques et al., ) . similarly, the ifninducible '- '-olygoadenylate synthetase (oas) requires dsrna that must be at least nucleotides long for activity, and no modification of the ′-hydroxyl group is tolerated (sarkar et al., ) . the oas family requires synthesizing di-, tri-, and tetrameric '- '-oligoadenylates ( - a), which in turn bind and activate rnase activation of rnase l together with pkr, irf and c-jun-n-terminal kinases, constituting pro-apoptotic mediators in response to dsrna. moreover, human oas may also enhance rig-i mediated signalling by mimicking polyubiquitin . further structural and mechanistic studies identify a key role of domain duplication in the oas family, thus revealing different functions of oas- and oas- in sensing dsrna (donovan et al., ; . rna interference (rnai) as well as antisense oligonucleotides and aptamers have been postulated to exert immune stimulatory effects in mammals (agrawal and kandimalla, ; cullen, ; mustonen et al., ) . however, at present, there is still an important open debate about agents exerting physiologically relevant functions at systems level biology (cullen, ; kleinman et al., ) . moreover, clinical trials of sirna demonstrated that the only population carrying the ff coding variant for tlr was protected from sirna-induced cytotoxicity (kleinman et al., ) . this study concluded that, because multiple cells express surface tlr , the therapeutic approach with sirna might induce unanticipated vascular or immune effects. here, anti-angiogenic innate immunity triggered by sirna was found to occur without the induction of inf-α/β via trif activation being biased toward nf-ĸb rather than irf- . side effects, for example, the immunosupressive effects of t regulatory cell function are inhibited by ifn-α- β (yu et al., ) , are still only vaguely understood in association with exogenous ncrnas administration. naturally occurring mammalian rnas do not display a stimulatory potential on innate immune responses through tlr- , - , - and - in dendritic cells and tlr-expressing cells (karikó et al., ) . by contrast, it has been shown that major innate immune responses to chemically synthesized sirnas are mediated by tlr and/or tlr . notably, replacement of uridines with their '-modified counterparts in the sirnas, reduced immune activation (sioud, ; barik and lu, ) . studies conducted on synthetic sirnas have shown results directly dependent on the nucleotide sequence (judge et al., (judge et al., , hornung et al., ) , and not to silencing effects of a target gene. putative immunostimulatory motifs allowing definition of ifn-α induction by β-gal control sirna or its constituent sense (gu-rich) and antisense ssrna oligonucleotides (judge et al., ) have been identified. these studies were performed in pbmcs, and therefore lack a clear demonstration of these effects at the organism level. intravenous administration of encapsulated sirna induced substantial dose-dependent ( - g sirna) ifn-α responses in outbred institute for cancer research (icr) inbred mice. in this line, reports also exist of increased hepatotoxicity of asos containing locked nucleic acid modifications (swayze et al., ) . in addition, asos carrying '-o-methoxyethylribose modifications showed no cytotoxic effects but were able to reduce targeted mrna. by analysis of sequence variants of either ss or ds sirna, uridine-rich immunostimulatory motifs within sirna were identified in human peripheral blood cells via tlr (sioud, ; jurk et al., ; judge et al., ) . there is a report of sequence-and targetindependent angiogenesis suppression induced by sirna via tlr (kleinman et al., ) . a minimum length for dsrna of about -nucleotide or -nucleotide luc sirna, but not truncated versions the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). suppressed by choroidal neovascularization have been observed in wild-type mice (kleinman et al., ) . recently, it has been established that nuclease-resistant aptamers can play a 'dual' role in immune signalling pathways (gefen et al., ; rajagopalan et al., ) . these aptamers seem to possess a unique feature acting as both agonist and antagonist depending on their degree of oligomerization (nozari and berezovski, ) . aptamers are ss oligonucleotides ( - nucleotides) capable of recognizing, in a specific manner and with high affinity, several types of target molecules by means of a three-dimensional folding of their chain. preclinical studies in murine models have highlighted distinct regulatory points that may be influenced by oligonucleotide aptamers, including clusters of differentiation (cd) (pastor et al., ) , cell-to-cell communication such as immunoglobulins (gefen et al., ) and interleukin (il) signalling (rajagopalan et al., ) . the cd molecules can act in several ways, often acting as receptors or ligands important to the cell initiating signal cascades or serving as adhesion molecules. here, cd is one of the main co-stimulatory receptors responsible for proper activation of t lymphocytes that could be manipulated to modulate the immune response (pastor et al., ) . further studies reported enhancement of t cell functions by specific t cell immunoglobulin- aptamers through negative regulation of ifn- secretion by cd + and cd + cells (gefen et al., ) . positive effects attributed to oligonucleotide aptamer have also been associated to attenuation of il- signalling in cd + cells (rajagopalan et al., ) . while sirna and dsrna in cultured cells have been shown to trigger an interferon response (alexopoulou et al., ; sledz et al., ) , the administration of naked (dtdt) synthetic sirnas ( . mg/kg) to mice (either intraperitoneally or iv via low pressure or high pressure injection) induced no interferon response (heidel et al., ) . this suggests that sirna administration in vivo may elicit little or no immune response. also, modification of nucleotides within rna molecules was shown to reduce immune response (durbin et al., ) , which is relevant when applying nucleotide modifications to rna therapeutics. for example, n- -methyladenosine (m a) and pseudouridine seems to reduce the retinoic acid inducible gene i triggering of the innate immune signalling (durbin et al., ) . thus, rna modification seems to influence recognition of exogenous rnas, for example, ribose '-o-methylation of mrna at the '-end provides a molecular signature for the discrimination of self and non-self rna (züst et al., ) . prior to immune cells performing their immunological action on target cells, they need to receive a suitable 'maturation' signal to enhance clonal expansion and acquire the effector function. activation signals occur in peripheral lymphoid organs through a concerted interaction with mhc molecules. the major histocompatibility complex (mhc) regulates the cell-mediated immune response(s). there are recent reports on the regulatory role of mirnas on mhc class i and ii molecules (wongfieng et al., ; xie et al., ) . for instance, mhc class i chain related protein b present in natural killer (nk) cells was associated to cell-mediated antitumor immune response through engagement with the nkg d receptor (xie et al., ) . the expression level of this receptor has been shown to be regulated by exogenous mir- c mimics in nk cells . based on the temporal and spatial regulation which determines the expression patterns of mirnas, that suggestion was made for the participation of complex regulatory networks composed of several transcription factors influencing the expression level of 'each' mirna. the mhc class ii complex has been identified as a target in mammalian mirnas regulation (wongfieng et al., ) . in addition, some lncrnas have been identified as candidates to be translated into peptide fragments (guttman et al., ) suggesting, as several other 'indirect' lines of evidence, the the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). participation of ncrnas-derived peptides on mhc complexes (yewdell, ; mellman et al., ) . peptides bound to mhc molecules serve as recognizing fragments of antigen for the tcr. here, it has been shown that mirnas can influence the amplitude of tcr signalling modulating t cell fate decisions towards survival and maturation wells et al., ) . here it is worth noting the recently defined dual role of mirna in maintenance of the naive phenotype of cd t cells or clonal expansion, where modulation of let- mirnas levels is required (wells et al., ) . in a similar way to mhc class ii for t cells, bcr determines the fate of developing b cells allowing their adequate maturation or the development of self-reactive b cells leading to immunodeficiency of b cell malignancy. the mirnas set threshold for lymphocyte development targets several genes in the pi k pathway (simpson and ansel, ) . it is also important to highlight that pi k displays an inhibitory interaction on tlrs-induced production of il- , therefore limiting excessive th polarization and suggesting a potential immunoregulatory role for mirnas. a few relatively recent studies address the potential regulatory role of 'specific' mirnas in treg functions (jeker et al., ; warth et al., ) . these studies have shown that inhibition of mirna- a in c bl/ j mice led to reduced foxp expression dependent on the levels of mediators such as tgf and/or retinoic acid (ra) (jeker et al., ) . another study reported induction of treg cell differentiation regulated by a mirna 'network' influencing the promoting factor mtor (warth et al., ) . these studies revealed that the underlying molecular mediators responsible for the mirna regulation of t cell signalling pathways remain unknown. unlike animals, plant mirnas have not been associated to tgf nor ra. intersecting the rapidly emerging field of treg function, it has been discovered that ra controls both the homing and differentiation of treg. in addition, computational and systems biology approaches have identified mtor as a potential pathway to explain the cross-kingdom mirnas-mediated regulatory potential of camptotheca acuminata ). although many reports can be found regarding the immunomodulatory role of endogenous and viral ncrnas, very few evaluated the impact of plant ncrnas on immune function and homeostasis when ingested/absorbed by animals and/or humans. thus, this section was reviewed using scientific documents. however, no studies were found that directly evaluate the effect of plant exogenous ncrna and their effects on the immune system following dietary intervention trials. recent reports highlight the shortcomings of existing mirna measuring systems (i.e. qpcr) in comparison to sequencing procedures to explore mirnas in biological fluids chen et al., a) . these studies point out the considerable number of unmapped ncrnas as a major factor limiting the use of qpcr techniques. notably, the abundance level of identified exogenous rna sequences in biological fluids (i.e. human plasma and breast milk, and serum from mice) was rather low. in addition, the role of food matrix on ncrna delivery to their cellular or tissue targets has not been defined. although the identification of exogenous ncrnas in biological samples led to considerations on their role as cross-kingdom regulators of immune-related processes, the impact (i.e. influence and regulatory role) of exogenous plant ncrnas on immune function and homeostasis is inferential. exogenous rnas sequences have been found in plasma , serum (chen et al., b) and breast milk ) from humans and/or animals. these studies reported the presence of ncrnas in biological fluids, although their impact on immune function remains to be fully clarified. the recent discovery of blood circulating rnas originating from foods seems to indicate that certain dietary plant mirnas can be absorbed at significant levels . however, the potential contribution of distinct 'plant food matrices' on ncrnas the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). absorption has not been addressed as it has been described for viral rnas (seljelid et al., ; zou et al., ) . the existing studies show that adequate antigen-processing processes at the intestinal level can be avoided by ncrnas bound to coadjuvant molecules (i.e. polycations, cationic lipids, phospholipids) (seljelid et al., ; zou et al., ; vertzoni et al., ) . notably, phospholipid homeostasis is central in the response to toll like receptors (tlrs) activation through a type i interferon 'autocrine-paracrine' loop (song et al., ) . in biological samples, rna sequences have been reported from most common gm food matrices, including corn (zea mays), rice (oryza sativa japonica group), soybeans (glycine max), tomato (solanum lycopersicum) and grape (vitis vinifera) . the identification of rice mir- a at fm concentration level in various mouse tissues may indicate the possibility of a cellular active uptake system for circulating rna . these data are in agreement with the role of defined food matrices as effective delivery systems protecting ncrnas from interaction with macrophages, dendritic cells, b-lymphocytes, and t-lymphocytes found in peyer's patches and other sites of gut-associated lymphoid tissue. however, several other studies contradict these results (see section . . . for details). enrichment of mirnas with targets related to immune response has been observed in colostral milk . colostrum represents a primordial food driving the first immunization of offspring and provides the nutritional needs of their immature organs in the earliest stages of life. the facts that immune-related mirnas from host, and exogenous plant mirnas (from bamboo diet to milk exosomes of giant panda) targeting synapse organization, neuron migration or axon guidance are enriched in giant breast milk of panda (restricted to a diet primarily comprising bamboo) indicate that breast milk may facilitate dietary intake of plant mirnas by infants for possible regulation of postnatal development ). very few studies address feeding plant ncrnas to animals (i.e. weaning, adult and old age) to evaluate induction of specific adaptive immune response(s) (i.e. th , th or th ). similarly, there is a lack of data on the impact of exogenous plant ncrnas on expression of mhc-ii molecules to assess the potential allergenicity of these rnas. additionally, the different immune response(s) induced by the administration of distinct exogenous plant ncrnas and well-known weak or strong immune inducers by different routes (intraperitoneal or intragastric) have not been tested. understanding of the allergenic functions of exogenous plant ncrnas is comparatively meagre in relation to proteins. the homology of distinct ncrnas could elicit different immune response(s) as proteins showing homology values up to % on their amino acid sequences induce different igg and igg responses (adel-patient et al., ) . for example, the literature search identified one study on safety assessment of the possible effects of gm-triticale on the immune system using a murine c bl/ model (krzyżowska et al., ) . immunoblotting analysis in serum showed significantly increased il- levels, but not those of ifn-γ. it was concluded that multigenerational use of feeds for rodents containing the gm-triticale leads to expansion of the b cell compartment in the secondary lymphoid organs not associated to allergic response(s) (krzyżowska et al., ) . however, this study did not report the ncrna content in gmtriticale. efforts over the last decade have focused on elucidating the role of ncrnas in immune function. although no immunogenic potential has been found, ncrnas implications in pathways regulating the production of inflammatory mediators (witwer et al., ) , the development of hematopoietic cells (satpathy and chang, ) , regulation of tlrs signalling (carpenter et al., ) and changes in t cells needed to support clonal expansion and growth have been highlighted. papers on ncrnas-mediated regulation of the amplitude and quality of immune response(s) could not be found. the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). at the moment, knowledge on the role of ncrnas in immune responses at systems-level remains incomplete. earlier reports defined significant differences in the effects of dsrna (poly i:c) in innate immune responses as a function of 'length' ( . - . kb and . - . kb), depending on cell type (i.e. splenocytes, pbmcs, raw . and thp- ) (mian et al., ) . notably, it was reported that the 'shorter' dsrna caused higher immunomodulatory effects. poly i:c, a synthetic analogue of viral dsrna, has long been known as a strong inducer of innate immune responses by interacting with tlr in the endosome. following ligand recognition, specific ligand cascades involving interferon regulatory proteins and nuclear factor kappa b (nf-b) are activated to produce inflammatory mediators (démoulins et al., ) . similarly, signal-specific lncrnas have been reported in dendritic cells (dcs) stimulated with tlr agonists, % of which clustered with nf-b signalling components (guttman et al., ) . further approaches also identified upregulated responses in a long intergenic noncoding rna (lincrna) in response to tlr- , - , - and - , but not tlr- in macrophages (carpenter et al., ) . specifically, lincrna-cox was found to mediate both activation and repression of immune response genes dependent on interactions of lincrna-cox with heterogeneous nuclear ribonucleoprotein a/b and a /b . a further study determined that lincrna-cox plays a key role as a coactivator of nf-b for the transcription of late-primary inflammatory response genes in macrophages through modulation of epigenetic chromatin remodelling (hu et al., ) . due to the relevance of gut microbiota in immune system development and homeostasis, the possibility of modulating the gut microbiota through dietary exogenous ncrnas was reviewed. as described in section . , during the preparatory phase this novel topic was identified as relevant to this literature review. this additional section serves as a general introduction to emphasize the relevance of the possible modulation of the immune system by dietary exogenous ncrnas through the gut microbiota, and to identify gaps pointing to needs for future studies relevant to food/feed risk assessment of ncrnabased gm plants. specific details of the possible mechanisms of action, if any, are outside the scope of this review. due to the novelty of this topic and the lack of information on the scientific literature, only documents were reviewed for this section. the intestinal tract harbours a complex community (microbiota), which has an enormous impact not only on the nutritional, but also immune status of the host. a balanced gut microbiota composition confers benefits to the host, while microbial imbalances are associated with metainflammatory disorders. the dynamic processes initiated upon birth define the microbiota composition that co-evolves with the host and its environment. thus, colonization of the intestine in early life seems particularly important as it represents one of the major environmental stimuli for immune system maturation. particularly, breast feeding is crucial to influencing early intestinal colonization. as previously indicated, development of emerging 'next generation sequencing technologies' has allowed identification of a significant fraction of the circulating rna in human plasma that originated from exogenous bacterial and fungi species . this study, based on microarray profiling results, reported that plasma can contain ncrnas from common foods influencing the expression profile of a number of genes in the cells. however, several other studies suggest that the quantitative amount of ncrnas are irrelevant for influencing gene expression (see section . . . for details). though the interaction between humans and their environments, particularly microbes, raises the possibility of some sort of feedback signalling process, it is still not fully understood. the literature search showed how few publications address the role of ncrnas in shaping gut microbiota. emerging the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). evidences point out interactions between environmental factors and inter-species gene regulation facilitating host control of the gut microbiota choi et al., ) . however, the impact of exogenous plant ncrnas on gut microbiota modulation remains elusive. increasing evidence demonstrates that a significant number of ncrnas are found in the extracellular media, and it is apparent that mammalian cells release srna that might carry functions which transcend the confines of single cells (valadi et al., ) . few studies have identified changes in mirna expression in human stool. overall, they provide an inconsistent association between their level and dietary habits and lifestyle (tarallo et al., ; . in this context, the survival of exogenous (orally administered) plant mirnas in faeces from mice has been observed . although the amount of particular mirnas such as mir- that survived the gi tract reached a maximum of . %, this study does not support consistent survival rates for distinct ncrnas. a recent study identified mirnas secreted by intestinal epithelial cells in intestinal contents and supports their role in modulating gut microbiota composition . this study reports the potential of host mirnas to enter bacteria and co-localize with microbial nucleic acids. similarly, exogenous plant mirnas that were present in animal faeces were indicated as being primarily acquired orally . recently, bacterial rnas comparable in size to mirnas have been identified that could serve as signalling molecules mediating bacteria-to-human interaction (choi et al., ) . these data were obtained from outer membrane vesicles produced by periodontopathogens and were able to exert 'slight' immunosuppressive effects on t cells cytokine production (il- , il- and il- ). mirna expression levels and amount in stool appear to be modulated by diet (micro-and macronutrients, phytochemicals) without excluding the possibility that the results were affected by other lifestyle factors; however, the influence of gm plant exogenous ncrnas has not yet been exhaustively studied in animals or humans. distinct innate immune mediators participating in the recognition and discrimination between the distinct nucleic acids have been identified; thus, additional definition of the dynamic interplay between the different 'sensors' (i.e. tlrs, rig-i/mda , the interferon response effectors protein kinase r and rnase l) in response to ncrnas have to be clarified. the chemical modifications observed in plant ncrnas highlight the need to address the detection and measurement of exogenous plant ncrnas in body fluids, as well as the sensitivity and specificity of these rnas to cellular and tissue targets. although many reports focusing on synthetic ss-and dsrna mimics can be found in the literature, scientifically assessed direct hazardous impacts of food and feed on fauna and flora are scarce and conflicting. notably, there is a need to evaluate the effects of exogenous plant ncrnas at the system biology level and in human trials. current knowledge on the immune functions of exogenous ncrnas is comparatively lacking. most studies use cell models based on isolated cells that could not mimick the whole set of players that determine immune response(s), which are necessary for mechanistic studies, but need approach cell-to-cell communication in target tissues. additionally, these studies should focus on the relationship of immune responses to production of ifns since tlrs signalling converge on the production of this immune regulatory factor. to better understand the impact of exogenous ncrnas in host microbiota composition and diversity, further studies are needed to establish the microbial targets and their effect on physiological conditions. understanding the basic biological aspects of plant-derived exogenous ncrna (including their half-life and stability), gathering information on human and animal exposure by diet to these molecules, their subsequent bioavailability and possibility to exert local or systemic effects, and reviewing the experience from the development of rna-based therapeutics are relevant aspects for the risk assessment of ncrnabased gm food and feed. the literature includes very few studies evaluating the half-life of plant-derived ncrnas. while no specific studies on plant mirnas and lncrnas half-life were found in this review, the studies on plant circrnas half-life suggest that they exhibit considerable stability as compared to linear rnas (section . . . ). in mammals, the few reports related to ncrnas half-life suggest that mirnas and circrnas are generally much more stable than mrna, while lncrnas have a half-life like that of mrna. however, when assessing the stability of plant ncrnas outside the plant (section . . . .), compelling evidence exists that plant mirnas are highly stable under different conditions including food storage, processing, cooking, or simulated digestion. moreover, they seem to survive after long incubation in serum, or are detected in the gastric content of mice, suggesting that plant mirnas are more resistant to degradation than synthetic or animal mirnas. this high resistance has been attributed to the unique characteristics of plant mirnas (and sirnas), in contrast to mirnas from mammalian and other organisms; the '-omethylation at their ' end which confers plant mirnas more resistance to degradation (section . . . ). efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). the experience from development of ncrna therapeutics (section . . ) shows that unmodified nucleic acids (i.e. rnas) exhibit very low stability in biological media and are subjected to rapid nuclease mediated degradation. indeed, a plethora of rnases are encoded within most genomes, often with overlapping activities, making redundancy a general feature of rna degradation systems. consequently, different chemical modifications or conjugations have been introduced to potential rna-based therapeutics (section . . . ) to improve their rna-binding affinity, their in vivo nuclease stability and their pharmacokinetic and pharmacodynamic properties. the available literature also suggests that naked or unmodified rnas parenterally administered (e.g. iv) are rapidly cleared from circulation and their presence in the kidney and urine has been reported immediately after administration (section . . ). similarly, following oral administration naked or unmodified exogenous rnas are rapidly degraded when exposed to the gi harsh conditions, and different delivery vehicles for exogenous rnas have been developed. in general, and in contrast to chemically modified or formulated rnas, no major biological effects have been observed for naked rnas, which has probably limited further evaluation in humans (section . . . ). although certain disease conditions may influence the pharmacokinetics of exogenous rnas, there are no studies that specifically evaluate this for naked unmodified exogenous ncrnas. to exert a biological effect, an exogenous ncrna first needs to reach the intended target tissue in sufficient quantity levels, this implying the necessity to overcome many biological barriers (section . . ). considering the oral intake as the main possible entry point of exogenous plant ncrnas into both animals and humans, the first major barrier is the mammalian gi tract, which encompasses a group of extracellular and cellular barriers. extracellular barriers include the presence of different enzymes (i.e. nucleases), the harsh environment and a net negative charged mucous layer. the cellular barriers include a three-layer (epithelial cells or enterocytes, the lamina propria and the muscularis mucosa) barrier known as the intestinal mucosa. possible ncrnas passage between cells is limited by the presence of tight junctions and the pore size in the human intestine would prevent the passage of all but mirnas, which are the smallest ncrnas. although crossing the cells by transcytosis may be possible, this mechanism implies facing new intracellular barriers, such as nucleases, recycling of ncrnas back to the lumen and nuclear uptake. however, m-cells, present in the gut epithelium interspersed between enterocytes, present certain characteristics that would make them more amenable for rna uptake. a second barrier is represented by a set of obstacles during the passage from the intestine to the target tissue, which encompasses plasma and tissue nucleases. once in the circulatory system, exogenous ncrnas are subjected to distribution and elimination. for example, small rnas would be rapidly cleared by the kidney due to their small size. furthermore, ncrnas need to escape the reticuloendothelial system, the function of which is to clear foreign pathogens. finally, they would have to cross the vascular endothelial barrier to reach the target tissue, facing hurdles like those in the enterocytes. packaging or incorporation of ncrnas into extracellular transport systems (i.e. exosomes/microvesicles or inclusion in ribonucleoprotein complexes) would confer certain resistance to nucleases and other barriers. ncrnas must enter the target cells to exert their functions. the molecular mechanisms of exogenous ncrna cellular uptake have been inferred from studies performed when developing strategies for nucleic acids delivery as therapeutics and is also largely derived from invertebrates, with little data reported in mammals. descriptions exist of receptor-mediated uptake of oligonucleotides through the sidt and sidt proteins, although this is now in doubt since they have been described as cholesterol transporters. due to the charged nature of rnas, cellular uptake could be achieved by endocytosis after which they would enter the system of intracellular trafficking through multiple membrane-bound compartments. ncrnas must exit these compartments to reach their functional sublocation within the efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). cell, either the cytosol or the nucleus, while simultaneously escaping degradation in the lysosomes. specialized barriers such as the blood-brain barrier or the placental barrier represent additional obstacles to overcome. while rna content in plant-derived foods varies (≈ mg/g of tissue), it could be estimated that when consuming a daily dose of total fruit and vegetables of ≈ g/day, humans theoretically ingest mg of total rnas (dietary intake of plant rna may typically range from . - g/person/day) (section . . . ). humans following special diets, such as vegetarians and vegans, have higher intake of plant origin rnas, but estimated exposure to such rnas implies only a -fold increase on average (section . . . ). it has been estimated that small rnas make up far less than % of total rna in plants, suggesting a very low exposure to plant ncrnas under normal conditions. whether sufficient levels of exogenous plant ncrnas can be consumed in the diet to exert a biological effect is also another aspect that needs to be evaluated. some examples in the literature suggest that for certain plant mirnas (i.e. mir- a) a person would need to consume kg of cantaloupe to reach the copies of mirna per cell possibly determining an effect. these aspects should be evaluated for each plant ncrna (on a case basis), considering that some plant-derived mirnas have shown unexpectedly high resistance to degradation and greater bioavailability. the available literature indicates a widespread presence of exogenous rnas (including plant derived ncrnas, i.e. small rnas) in the biological fluids of humans and animals (section . . ). whether these rnas are derived from dietary intake is unclear. their generally low abundance and lack of enrichment in tissues mostly exposed to dietary changes (i.e. liver) suggest that some of may have be originated as technical artefacts or through contamination. very few studies address biological effects of dietary exogenous ncrnas in the gi tract and its annex glands (e.g. liver) (section . can directly target mammalian genes through rnai effects, but still clarification is needed if exogenous plant ncrnas can overcome all the biological barriers to reach and interact with their possible targets. exogenous plant-derived ncrnas have been found in exosomes or macrovesicles. how they reach these types of structures in biological fluids is unknown. in summary, supporting and contradicting evidence concerning the existence of systemic effects of dietary plant-derived exogenous ncrnas is heavily debated. important aspects such as the precise mechanism/s of transport of plant ncrnas from food into the systemic circulation, the amount of exogenous ncrnas reaching tissues or the molecular mechanisms of cellular uptake need to be determined. while several in vitro studies cover the participation of exogenous ncrnas in the dynamic network to modulate innate immunity responses, few tackle the immunomodulatory effects of plant ncrnas in adaptive immune response (section . ). none of these studies have considered the potential impact of exogenous dietary plant-derived ncrnas at the systemic level. since the first publication suggesting a possible cross-kingdom effect by dietary ncrna (through rice mir- a) the knowledge on exogenous ncrnas, and particularly plant-derived exogenous ncrnas, on efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). their fate when ingested has increased. however, the knowledge useful to support the assessment of dietary ncrna such as ncrna-based gm food and feed still faces several gaps. an important aspect needing further investigation is the stability of plant ncrnas. the meagre available literature contains a very few studies of the half-life of plant ncrnas (i.e. mirnas, lncrnas and circrnas) both inside the plant cell and outside the plant. specifically, the stability of circrnas should be assessed due to their apparently high stability. moreover, very few reports focus on assessment of plant ncrnas stability in mammalian systems, when ingested. understanding the molecular basis that confers plant ncrnas stability is of great importance and needs to be further evaluated. for instance, plant mirnas and sirnas carry modifications that do not occur in mammals. these modifications, such as the presence of a methyl group located on the ' nucleotide ribose, could hypothetically have an impact on the plant mirnas stability in mammalian systems/cells due to either the lack of the appropriate enzymes for recognition and degradation of plant mirnas by mammalian cells, or the increased stability to degradation by mammalian rnases. the circularized structure of plant circrnas would hypothetically confer them increased stability to mammalian rnases. if this is the case, plant mirnas and circrnas could exhibit a much longer half-life in mammalian systems than expected, especially when compared to mammalian mirna or circrnas. this would increase their chances of reaching the appropriate target tissue and encountering suitable target molecules within the cells. the half-life of plant ncrnas within mammalian cells is also an important aspect to consider. experiments are needed with labelled plant ncrnas, first in transfected mammalian cells (in vitro studies) and then orally administered to experimental animals (in vivo studies). radioactive probes or molecules of very low molecular weight, such as biotin or fluorescein, should be used, in order to prevent distorting the actual size of the studied plant ncrna; this is important when studying mirnas stability due to their small size. another aspect possibly relevant for risk assessment is to understand whether and how ingested gm plant-derived ncrnas can affect the expression levels of other ncrnas and other rnas, due to putative compensatory circuits and codifying rnas. whole transcriptome sequencing, comparing the rna levels of unmodified (wild type) to those of modified (transgenic) plants coul be useful to this aim. although available information suggests that many biological barriers exist in mammalian system to ingested plant exogenous ncrnas preventing these to exert a possible local or systemic effect, several gaps are still present. putative pathways of dietary exogenous ncrnas after ingestion and the many transporters potentially involved (i.e, in cellular uptake, or barrier passage transport pathways) in the transit from the gut to the target tissues should be characterized using standard molecular biology gainof-function and loss-of-function approaches. also, the many routes of mammalian intracellular trafficking to the specific intracellular target should be studied. to this aim, ncrnas isolated from human diets should be used preferentially, since most of the incomplete current knowledge comes from the rna-therapeutics field or is derived from the invertebrate world. which molecular mechanisms support the possible passage through specialized barriers (i.e. blood-brain barrier or the placental barrier), is another critical aspect requiring further studies. while there are in vitro studies describing the interaction between plant mirnas and mammalian mirna silencing complexes possibly leading to target repression, these findings need to be experimentally validated in vivo. therefore, more studies are needed to understand the possibility of processing plant ncrnas in mammalian cells. it is also unknown if ingested plant mirnas reaches or needs to reach a necessary level to exert a biological effect. for instance, studies in mammals suggest that the threshold for target gene regulation is > copies per cell for a certain study; however, another study suggests around to . copies of mammalian mirna per cell. clearly, a minimum amount of small rnas efsa supporting publication :en- the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). would be required to achieve biologically relevant effects on gene expression. on a case by case basis it would be useful to evaluate the amount of consumption of a given ncrna in a normal diet. it is also relevant to study if exposure to these plant ncrnas could change under certain pathological conditions (i.e. compromised intestinal permeability or renal function) or dietary patterns (i.e. vegetarian or vegan diets). although tissue accumulation of dietary plant exogenous ncrnas has not been reported to date, it seems clear that plant mirnas and other exogenous rnas are present in biological fluids from humans and animals. the biological significance of their presence is still unknown and needs to be addressed. methods that do not require amplification for detection should be used. more quantitative approaches to determine the levels of plant-derived exogenous ncrnas should be determined both in biological fluids and tissues to understand the magnitude of presence. for example, this could be partially obtained from studies reporting the presence of exogenous rnas in biological fluids using public small rna-seq databases. in the case of plant-derived mirnas, the use of sodium periodate oxidation of samples could be a good start to verifying the presence of genuine plant-derived mirnas from diet in human and animal biological fluids. the immunomodulatory effects of exogenous rnas have been widely described in the literature. however, there is no in vivo information regarding the immunomodulatory effects of dietary exogenous ncrnas. these types of studies should be performed both in vitro and in vivo to determine if the different types of ncrnas consumed in the diet can affect the immune system. in the case of target gene repression by plant ncrnas investigations in specific animal models could be considered. expression levels of other ncrnas and rnas, which may be modified due to putative compensatory circuits could be evaluated in these models, as well as information on the stability and half-life of the specific ncrna in specific cells and systemically following ingestion. genome-wide high-resolution mapping of exosome substrates reveals hidden features in the arabidopsis transcriptome genome-wide identification of circular rnas in arabidopsis thaliana a lariat-derived circular rna is required for plant development in arabidopsis pattern formation via small rna mobility small rnas are on the move literature review of baseline information on rnai to support the environmental risk assessment of rnai-based gm plants identification of circular rnas from the parental genes involved in multiple aspects of cellular metabolism in barley a group i plant intron accumulates as circular rna forms with extensive ' deletions in vivo toward a consensus on the binding specificity and promiscuity of prc for rna a stepwise pathway for biogenesis of -nt secundary sirnas and spreading of dna methylation site-specific phosphorylation profiling of arabidopsis proteins by mass spectrometry and peptide chip analysis hierarchical action and inhibition of plant dicer-like proteins in antiviral defense characterization of stress-responsive lncrnas in arabidopsis thaliana by integrating expression, epigenetic and structural features identification and characterization of human testis derived circular rnas and their existence in seminal plasma the rna-binding proteins hyl and se promote accurate in vitro processing of pri-mirna by dcl genome-wide discovery of circular rnas in the leaf and seedling tissues of arabidopsis thaliana intra-and intercellular rna interference in arabidopsis thaliana requires components of the microrna and heterochromatic silencing pathways dicer-like is required for rna interference and produces the -nucleotide small interfering rna component of the plant cell-to-cell silencing signal plant mobile small rnas the arabidopsis thaliana doublestranded rna binding protein drb directs guide strand selection from microrna duplexes naturally occurring variations in sequence length creates microrna isoforms that differ in argonaute effector complex specificity the xist lncrna exploits three-dimensional genome architecture to spread across the x chromosome identification of nuclear dicing bodies containing proteins for microrna biogenesis in living arabidopsis plants phased, secondary, small interfering rnas in posttranscriptional regulatory networks antisense rna polymerase ii divergent transcripts are p-tefb dependent and substrates for the rna exosome long noncoding rnas in cell-fate programming and reprogramming target mimicry provides a new mechanism for regulation of microrna activity location of a possible mirna processing site in smd /smb nuclear bodies in arabidopsis partially redundant functions of arabidopsis dicer-like enzymes and a role for dcl in producing trans-acting sirnas short integuments /suspensor /carpel factory, a dicer homolog, is a maternal effect gene required for embryo development in arabidopsis oncogenic role of fusion-circrnas derived from cancer-associated chromosomal translocations topological organization of multichromosomal regions by the long intergenic noncoding rna firre natural rna circles function as efficient microrna sponges vernalization-mediated epigenetic silencing by a long intronic noncoding rna flowering locus c's lessons: conserved chromatin switches underpinning developmental timing and adaptation transitivity-dependent andindependent cell-to-cell movement of rna silencing specific interactions between dicer-like proteins and hyl /drb-family dsrna-binding proteins in arabidopsis thaliana phosphate utilization efficiency correlates with expression of low-affinity phosphate transporters and noncoding rna, ips , in barley gene silencing by micrornas: contributions of translational repression and mrna decay cyclophilin facilitates hsp -mediated risc assembly in plants in vitro assembly of plant rna-induced silencing complexes facilitated by molecular chaperone hsp a rice cis-natural antisense rna acts as a translational enhancer for its cognate mrna and contributes to phosphate homeostasis and plant fitness the nuclear pore protein attpr is required for rna homeostasis, flowering time, and auxin signaling dicer- and r d -l catalyze microrna maturation in drosophila molecular insights into mirna processing by arabidopsis thaliana serrate covalent circularization of exogenous rna during incubation with a wheat embryo cell extract small rnas as guardians of the genome fast-forward genetics identifies plant cpl phosphatases as regulators of mirna processing factor hyl the evolution and diversification of dicers in plants passenger-strand cleavage facilitates assembly of sirna into ago -containing rnai enzyme complexes circular single-stranded rna replicon in saccharomyces cerevisiae rna-directed dna methylation: an epigenetic pathway of increasing complexity intercellular and systemic movement of rna silencing signals circular rnas are a large class of animal rnas with regulatory potency sorting of small rnas into arabidopsis argonaute complexes is directed by the ' terminal nucleotide morc family atpases required for heterochromatin condensation and gene silencing small silencing rnas in plants are mobile and direct epigenetic modification in recipient cells specificity of argonaute -mir interaction and dual functionality in tas trans-acting sirna formation evolution of animal and plant dicers: early parallel duplications and recurrent adaptation of antiviral rna binding in plants literature review of baseline information to support the risk assessment of rnai-based gm plants systemic acquired silencing: transgene-specific post-transcriptional silencing is transmitted by grafting from silenced stocks to non-silenced scions microrna is a long-distance signal for the regulation of plant phosphate homeostasis evidence for nuclear processing of plant micro rna and short interfering rna precursors nuclear processing and export of micrornas in arabidopsis carpel factory, a dicer homolog, and hen , a novel protein, act in microrna metabolism in arabidopsis thaliana rde- preferentially binds long dsrna and its dimerization is necessary for cleavage of dsrna to sirna rna exosome-regulated long non-coding rna transcription controls super-enhancer activity prediction of trans-antisense transcripts in arabidopsis thaliana genome-wide identification of long noncoding natural antisense transcripts and their responses to light in arabidopsis identification and characterization of circrnas in pyrus betulifolia bunge under drought stress analysis of noncoding transcriptome in rice and maize uncovers roles of conserved lncrnas associated with agriculture traits a long noncoding rna maintains active chromatin to coordinate homeotic gene expression arabidopsis noncoding rna mediates control of photomorphogenesis by red light identification of circular rnas and their targets in leaves of triticum aestivum l. under dehydration stress importin is a gene silencing factor that targets argonaute proteins to distinct mrnas noncoding transcription by rna polymerase pol ivb/pol v mediates transcriptional silencing of overlapping and adjacent genes expression of arabidopsis mirna genes genetic and functional diversification of small rna pathways in plants identification and characterization of wheat long non-protein coding rnas responsive to powdery mildew infection and heat stress by using microarray analysis and sbs sequencing bidirectional promoters generate pervasive transcription in yeast functions of long intergenic non-coding (linc) rnas in plants widespread noncoding circular rnas in plants a systemic small rna signaling system in plants wavy leaf , an ortholog of arabidopsis hen , regulates shoot development by maintaining microrna and trans-acting small interfering rna accumulation in rice vigs, higs and figs: small rna silencing in the interactions of viruses or filamentous organisms with their plant hosts control of coleopteran insect pests through rna interference the expanding world of small rnas in plants genome-wide analysis of long noncoding rna stability structural variations and stabilising modifications of synthetic sirnas in mammalian cells circular rnas are long-lived and display only minimal early alterations in response to a growth factor host-delivered rnai: an effective strategy to silence genes in plant parasitic nematodes analysis of microrna turnover in mammalian cells following dicer ablation polysome arrest restricts mirna turnover by preventing exosomal export of mirna in growth-retarded mammalian cells a versatile monosaccharide transporter that operates in the arbuscular mycorrhizal fungus glomus sp is crucial for the symbiotic relationship with plants the drosophila rna methyltransferase, dmhen , modifies germline pirnas and single-stranded sirnas in risc uridylation of mature mirnas and sirnas by the mut nucleotidyltransferase promotes their degradation in chlamydomonas circular rnas are abundant, conserved, and associated with alu repeats regulation of small rna stability: methylation and beyond methylation protects mirnas and sirnas from a ' end uridylation activity in arabidopsis assessing the survival of exogenous plant microrna in mice effective detection and quantification of dietetically absorbed plant micrornas in human plasma adenylation of plant mirnas covalent circularization of exogenous rna during incubation with a wheat embryo cell extract silencing a cotton bollworm p monooxygenase gene by plant-mediated rnai impairs larval tolerance of gossypol rna-specific ribonucleotidyl transferases oral delivery of sirna and antisense oligonucleotides toxicogenomics of non-viral drug delivery systems for rnai: potential impact on sirna-mediated gene silencing activity and specificity nonviral delivery of synthetic sirnas in vivo a theoretical basis for a biopharmaceutic drug classification: the correlation of in vitro drug product dissolution and in vivo bioavailability oral nucleic acid therapy using multicompartmental delivery systems the adherent gastrointestinal mucus gel layer: thickness and physical state in vivo polymeric nano-and microparticle technologies for oral gene delivery isolation and properties of an extracellular nucleases of serratia marcescens guidance for industry on waiver of in vivo bioavailability and bioequivalence studies for immediate-release solid oral dosage forms based on a biopharmaceutics classification system issues in oral nanoparticle drug carrier uptake and targeting in vitro and in vivo models for the study of oral delivery of nanoparticles alginate/poly-l-lysine microparticles for the intestinal delivery of antisense oligonucleotides multi-compartmental oral delivery systems for nucleic acid therapy in the gastrointestinal tract extracellular ribonucleases from bacillus subtilis. i. crystallization and specificity intestinal delivery of non-viral gene therapeutics: physiological barriers and preclinical models first-pass elimination. basic concepts and clinical consequences key issues in non-viral gene delivery polymeric nanoparticle drug delivery technologies for oral delivery applications effect of sodium caprate on the intestinal absorption of two modified antisense oligonucleotides in pigs drug delivery of sirna therapeutics: potentials and limits of nanosystems oral delivery of antisense oligonucleotides in man the human microbiome project consortium. structure, function and diversity of the healthy human microbiome milk-derived exosomes for oral delivery of paclitaxel delivery of sirna to the mouse brain by systemic injection of targeted exosomes orally delivered sirna targeting macrophage map k suppresses systemic inflammation oral administration of bovine milk derived extracellular vesicles attenuates arthritis in two mouse models oral nucleic acid therapy using multicompartmental delivery systems oral delivery of rnase p ribozymes by salmonella inhibits viral infection in mice protection and systemic translocation of sirna following oral administration of chitosan/sirna nanoparticles interleukin gene transfer prevents experimental colitis in rats inhibition of trkb limits development of the zebra finch song system commercial dairy cow milk micrornas resist digestion under simulated gastrointestinal tract conditions oral il- gene delivery in a microsphere-based formulation for local transfection and therapeutic efficacy in inflammatory bowel disease the role of nucleotides in human nutrition retroviral delivery of rna interference against marek's disease virus in vivo biogenesis, secretion, and intercellular interactions of exosomes and other extracellular vesicles systemic exosomal sirna delivery reduced alpha-synuclein aggregates in brains of transgenic mice pancreatic cancer exosomes initiate pre-metastatic niche formation in the liver proteomics of extracellular vesicles: exosomes and ectosomes dietary yeast rna supplementation reduces mortality by aeromonas hydrophila in rohu (labeo rohita l.) juveniles effects of mannose density on in vitro and in vivo cellular uptake and rnai efficiency of polymeric nanoparticles peg-pla nanoparticles facilitate sirna knockdown in adult zebrafish heart exosome-mediated delivery of hydrophobically modified sirna for huntingtin mrna silencing oral delivery of small rna and dna colonic gene silencing using sirna-loaded calcium phosphate/plga nanoparticles ameliorates intestinal inflammation in vivo a randomized controlled trial of preoperative oral supplementation with a specialized diet in patients with gastrointestinal cancer a novel class of small rnas: trna-derived rna fragments (trfs) small non-coding rnas transfer through mammalian placenta and directly regulate fetal gene expression assessing the survival of exogenous plant microrna in mice effective detection and quantification of dietetically absorbed plant micrornas in human plasma human milk exosomes and their micrornas survive digestion in vitro and are taken up by human intestinal cells detection of plant mirnas abundance in human breast milk in silico identification of plant mirnas in mammalian breast milk exosomes--a small step forward? detection of dietetically absorbed maizederived micrornas in pigs exosomal micrornas in giant panda (ailuropoda melanoleuca) breast milk: potential maternal regulators for the development of newborn cubs unsuccessful detection of plant micrornas in beer, extra virgin olive oil and human plasma after an acute ingestion of extra virgin olive oil a novel chemopreventive strategy based on therapeutic micrornas produced in plants annotation of the goat genome using next generation sequencing of microrna expressed by the lactating mammary gland: comparison of three approaches expressional analysis of immune-related mirnas in breast milk identification of stable, high copy number, medium-sized rna degradation intermediates that accumulate in plants under non-stress conditions determination of the potential bioavailability of plant micrornas using a simulated human digestion process human breast milk mirna, maternal probiotic supplementation and atopic dermatitis in offspring the effect of dietary ginseng polysaccharide supplementation on the immune responses involved in porcine milk-derived esrnas immune modulatory function of abundant immunerelated micrornas in microvesicles from bovine colostrum uptake and function studies of maternal milk-derived micrornas mining of public sequencing databases supports a nondietary origin for putative foreign mirnas: underestimated effects of contamination in ngs literature review of information supporting food/feed risk assessment of rnai-based gm plants www.efsa.europa this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document alternative mirnas: human sequences misidentified as plant mirnas in plant studies and in human plasma transfer and functional consequences of dietary micrornas in vertebrates: concepts in search of corroboration: negative results challenge the hypothesis that dietary xenomirs cross the gut and regulate genes in ingesting vertebrates, but important questions persist real-time quantitative pcr and droplet digital pcr for plant mirnas in mammalian blood provide little evidence for general uptake of dietary mirnas: limited evidence for general uptake of dietary plant xenomirs the intestinal transport of bovine milk exosomes is mediated by endocytosis in human colon carcinoma caco- cells and rat small intestinal iec- cells the levels of human milk micrornas and their association with maternal weight characteristics detection of dietary plant-based small rnas in animals anomalous uptake and circulatory characteristics of the plant-based small rna mir the atypical genesis and bioavailability of the plant-based small rna mir : bulking up while breaking down bioavailability of transgenic micrornas in genetically modified plants exogenous plant mir a specifically targets mammalian ldlrap : evidence of cross-kingdom regulation by microrna analysis of plant-derived mirnas in animal small rna datasets immune-related micrornas are abundant in breast milk exosomes honeysuckle-encoded atypical microrna directly targets influenza a viruses one step forward, two steps back; xeno-micrornas reported in breast milk are artifacts small rnas from plants, bacteria and fungi within the order hypocreales are ubiquitous in human plasma dietary microrna database (dmd): an archive database and analytic tool for food-borne micrornas lack of detectable oral bioavailability of plant micrornas after feeding in mice survey of + data sets from human tissue and body fluid reveals xenomirs are likely artifacts plant mirnas found in human circulating system provide evidences of cross kingdom rnai detection of plant mirnas abundance in human breast milk in silico identification of plant mirnas in mammalian breast milk exosomes--a small step forward? optimization of enzymatic reaction conditions for generating representative pools of cdna from small rna computational characterization of exogenous micrornas that can be transferred into human circulation mining of public sequencing databases supports a nondietary origin for putative foreign mirnas: underestimated effects of contamination in ngs the complex exogenous rna spectra in human plasma: an interface with human gut biota? the spectrum of circulating rna: a window into systems toxicology detection of an abundant plant-based small rna in healthy consumers oral administration of bovine milk derived extracellular vesicles attenuates arthritis in two mouse models exosomes as a nanodelivery system: a key to the future of neuromedicine? endogenous microrna can be broadly exploited to regulate transgene expression according to tissue, lineage and differentiation state identification of dietetically absorbed rapeseed (brassica campestris l.) bee pollen micrornas in serum of mice lack of detectable oral bioavailability of plant micrornas after feeding in mice high-throughput sequencing of rna silencingassociated small rnas in olive (olea europaea l.) plant-derived phosphocholine facilitates cellular uptake of anti-pulmonary fibrotic hjt-srna-m autosomal recessive hypercholesterolemia caused by mutations in a putative ldl receptor adaptor protein exosomes as therapeutic drug carriers and delivery vehicles across biological membranes: current perspectives and future challenges extensive degradation and low bioavailability of orally consumed corn mirnas in mice an improved method to quantitate mature plant microrna in biological matrices using modified periodate treatment and inclusion of internal controls this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document cross talk between adipose tissue and placenta in obese and gestational diabetes mellitus pregnancies via exosomes grape exosome-like nanoparticles induce intestinal stem cells and protect mice from dss-induced colitis survey of + data sets from human tissue and body fluid reveals xenomirs are likely artifacts cross-kingdom regulation of putative mirnas derived from happy tree in cancer pathway: a systems biology approach small non-coding rnas transfer through mammalian placenta and directly regulate fetal gene expression assessing the survival of exogenous plant microrna in mice reply to dr. witwer's letter to the editor effective detection and quantification of dietetically absorbed plant micrornas in human plasma plant mirnas found in human circulating system provide evidences of cross kingdom rnai detection of dietetically absorbed maizederived micrornas in pigs negligible uptake and transfer of diet-derived pollen micrornas in adult honey bees the transport mechanism of extracellular vesicles at the blood-brain barrier unsuccessful detection of plant micrornas in beer, extra virgin olive oil and human plasma after an acute ingestion of extra virgin olive oil a novel chemopreventive strategy based on therapeutic micrornas produced in plants interspecies communication between plant and mouse gut host cells through edible plant derived exosome-like nanoparticles high-throughput assessment of microrna activity and function using microrna sensor and decoy libraries milk extracellular vesicles accelerate osteoblastogenesis but impair bone matrix formation corn rootwormactive rna dvsnf : repeat dose oral toxicology assessment in support of human and mammalian safety a -day oral toxicity evaluation of small interfering rnas and a long double-stranded rna targeting vacuolar atpase in mice citrus limon-derived nanovesicles inhibit cancer cell proliferation and suppress cml xenograft growth by inducing trail-mediated cell death this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document ineffective delivery of diet-derived micrornas to recipient animal organisms adipose-derived circulating mirnas regulate gene expression in other tissues uptake and function studies of maternal milk-derived micrornas contamination or artifacts may explain reports of plant mirnas in humans alternative mirnas: human sequences misidentified as plant mirnas in plant studies and in human plasma real-time quantitative pcr and droplet digital pcr for plant mirnas in mammalian blood provide little evidence for general uptake of dietary mirnas: limited evidence for general uptake of dietary plant xenomirs detection of an abundant plant-based small rna in healthy consumers anomalous uptake and circulatory characteristics of the plant-based small rna mir bioavailability of trasngenic micrornas in genetically modified plants exosome delivered anticancer drugs across the blood-brain barrier for brain cancer therapy in danio rerio molecular basis of mammalian transmissibility of avian h n influenza viruses and their pandemic potential exogenous plant mir a specifically targets mammalian ldlrap : evidence of cross-kingdom regulation by microrna honeysuckle-encoded atypical microrna directly targets influenza a viruses plant micrornas in larval food regulate honeybee caste development to evaluate oral toxicity of exogenous ncrnas in mice, petrick et al. administered for days a repeated oral dose of sirnas and dsrna and evaluated several parameters including body weight, food consumption, clinical observations, clinical chemistry, haematology, gross pathology, or histopathology endpoints it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). signs (mortality, abnormalities, and sign of pain and distress), body weight and food consumption, haematology, organ weight, or pathology results. the minor differences observed in certain parameters were limited to single intervals, were not dose-related, and were attributed to interanimal variability torula yeast rna (rna negative control) at mg/kg/day or a control vehicle were used. ten animals per sex and per group were used. no treatment-related effects were observed on clinical signs (mortality, or sign of pain and distress), body weight and food consumption, haematology, organ weight, or pathology results. minor changes in selected parameters in selected groups were observed, but these were attributed by the authors to normal variability and not treatment-related effects this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document ecological risk assessment for dvsnf rna: a plant-incorporated protectant with targeted activity against western corn rootworm literature review of baseline information on rnai to support the environmental risk assessment of rnai-based gm plants a comparative evaluation of the regulation of gm crops or products containing dsrna and suggested improvements to risk assessments endogenous small rnas in grain: semi-quantification and sequence homology to human and animal genes literature review of baseline information to support the risk assessment of rnai-based gm plants corn rootwormactive rna dvsnf : repeat dose oral toxicology assessment in support of human and mammalian safety a -day oral toxicity evaluation of small interfering rnas and a long double-stranded rna targeting vacuolar atpase in mice rnai technologies in agricultural biotechnology: the toxicology forum th annual summer meeting no impact of dvsnf rna on honey bee (apis mellifera l.) adults and larvae in dietary feeding tests microstructure and ultrastructure of highamylose rice resistant starch granules modified by antisense rna inhibition of starch branching enzyme a -day toxicology study of high-amylose transgenic rice grain in sprague-dawley rats a three generation reproduction study with sprague-dawley rats consuming high-amylose transgenic rice high-amylose rice improves indices of animal health in normal and diabetic rats studies performed with human cells were mainly carried out ex vivo using lymphocytes and, to a significant extent, macrophages. from the studies retrieved during the screening phase on immune cells in relation to exogenous ncrna %) from rat and ( . %) from mouse. and within the studies on immune cells other than lymphocytes (i.e. macrophages), the studies were allocated into ( . %) from human, ( . %) from rat, and ( . %) from mouse. all these data clearly show that studies retrieved in relation to ncrnas and www this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document immunological and metabolomic impacts of administration of cry ab protein and mon maize in mouse role of toll-like receptors in antisense and sirna recognition of double-stranded rna and activation of nf-kappab by toll-like receptor rna regulation of the immune system therapy of respiratory viral infections with intranasal sirnas cd is a coreceptor of toll-like receptors and a long noncoding rna mediates both activation and repression of immune response genes identification of dietetically absorbed rapeseed (brassica campestris l.) bee pollen micrornas in serum of mice identification of dietetically absorbed rapeseed (brassica campestris l.) bee pollen mirnas in serum of mice chemical modification patterns compatible with high potency dicer-substrate small interfering rnas is rna interference involved in intrinsic antiviral immunity in mammals? poly (i:c) induced immune response in lymphoid tissues involves three sequential waves of type i ifn expression structural basis for cytosolic double-stranded rna surveillance by human oligoadenylate synthetase structural mechanism of sensing long dsrna via a noncatalytic domain in human oligoadenylate synthetase rnas containing modified nucleotides fail to trigger rig-i conformational changes for innate immune signaling this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document identification of rna sequence motifs stimulating sequence-specific tlr -dependent immune responses dsrna with ′ overhangs contributes to endogenous and antiviral rna silencing pathways in plants a tim- oligonucleotide aptamer enhances t cell functions and potentiates tumor immunity in mice ab initio reconstruction of cell type-specific transcriptomes in mouse reveals the conserved multi-exonic structure of lincrnas ribosome profiling provides evidence that large noncoding rnas do not encode proteins chromatin signature reveals over a thousand highly conserved large noncoding rnas in mammals lack of interferon response in animals to naked sirnas species-specific recognition of single-stranded rna via toll-like receptor and small anti-viral compounds activate immune cells via the tlr myd -dependent signaling pathway sequencespecific potent induction of ifn-alpha by short interfering rna in plasmacytoid dendritic cells through tlr lincrna-cox promotes late inflammatory gene transcription in macrophages through modulating swi/snf-mediated chromatin remodeling microrna a marks regulatory t cells the role of alternative polyadenylation in the antiviral innate immune response the '-o-methylation status of a single guanosine controls transfer rna-mediated toll-like receptor activation or inhibition design of noninflammatory synthetic sirna mediating potent gene silencing in vivo sequence-dependent stimulation of the mammalian innate immune response by synthetic sirna immunostimulatory potential of silencing rnas can be mediated by a non-uridine-rich toll-like receptor motif suppression of rna recognition by toll-like receptors: the impact of nucleoside modification and the evolutionary origin of rna sequence-and target-independent angiogenesis suppression by sirna via tlr microrna as a new immune-regulatory agent in breast milk this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document the effect of multigenerational diet containing genetically modified triticale on immune system in mice cross-kingdom regulation of putative mirnas derived from happy tree in cancer pathway: a systems biology approach double-stranded rna-mediated tlr activation is enhanced by cd molecular basis for the immunostimulatory activity of guanine nucleoside analogs: activation of toll-like receptor the tlr signaling complex forms by cooperative receptor dimerization mir- a is an intrinsic modulator of t cell sensitivity and selection assessing the survival of exogenous plant microrna in mice structural basis of toll-like receptor signaling with double-stranded rna the host shapes the gut microbiota via fecal microrna in silico identification of plant mirnas in mammalian breast milk exosomes--a small step forward? exosomal micrornas in giant panda (ailuropoda melanoleuca) breast milk: potential maternal regulators for the development of newborn cubs microrna- c promotes natural killer cell cytotoxicity via up-regulating the expression level of nkg d neonatal immune activation during early and late postnatal brain development differently influences depressionrelated behaviors in adolescent and adult c bl/ mice small self-rna generated by rnase l amplifies antiviral innate immunity a structural basis for discriminating between self and nonself double-stranded rnas in mammalian cells cancer immunotherapy comes of age length of dsrna (poly i:c) drives distinct innate immune responses, depending on the cell type oligonucleotide-based pharmaceuticals: non-clinical and clinical safety signals and non-clinical testing strategies cytosolic viral sensor rig-i is a '-triphosphate-dependent translocase on double-stranded rna aptamers for cd antigens: from cell profiling to activity modulation cd aptamers as powerful immune response modulators influences of diet and the gut microbiome on epigenetic modulation in cancer and other diseases this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document aptamer-targeted attenuation of il- signaling in cd + t cells enhances antitumor immunity catalog of differentially expressed long non-coding rna following activation of human and mouse innate immune response. front immunol , long noncoding rna in hematopoiesis and immunity the effect of poly-l-lysine on the uptake of reovirus doublestranded rna in macrophages in vitro characterization of the mammalian rna exonuclease /nef-sp as a testis-specific nuclear ' → ' exoribonuclease microrna regulation of lymphocyte tolerance and autoimmunity advances in rna sensing by the immune system: separation of sirna unwanted effects from rna interference activation of the interferon system by short-interfering rnas type i interferons function as autocrine and paracrine factors to induce autotaxin in response to tlr activation antisense oligonucleotides containing locked nucleic acid improve potency but cause significant hepatotoxicity in animals microrna expression in relation to different dietary habits: a comparison in stool and plasma samples interplay between dengue virus and toll-like receptors, rig-i/mda and micrornas: implications for pathogenesis exosome-mediated transfer of mrnas and micrornas is a novel mechanism of genetic exchange between cells dissolution media simulating the intralumenal composition of the small intestine: physiological issues and practical aspects lps injection reprograms the expression and the ' utr of a cap gene by alternative polyadenylation and the formation of a gait element in ciona intestinalis the complex exogenous rna spectra in human plasma: an interface with human gut biota? induced mir- a expression represses mtor cooperatively with mir- to promote regulatory t-cell differentiation evolutionary dynamics and tissue specificity of human long noncoding rnas in six mammals toll-like receptor (tlr) immune modulation by unformulated small interfering rna or dna and the role of cd (in tlrmediated effects) homology modeling of human tolllike receptors tlr , , and ligand-binding domains modulation of let- mirnas controls the differentiation of effector cd t cells this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document microrna regulation of ifn-beta protein expression: rapid and sensitive modulation of the innate immune response . '-utr and '-utr regulation of micb expression in human cancer cells by novel micrornas microrna- is downregulated by histone deacetylase inhibitors and confers resistance to natural killer cytotoxicity in hepatocellular carcinoma cells ncrna-and pc methylation-dependent gene relocation between nuclear structures mediates gene activation programs plumbing the sources of endogenous mhc class i peptide ligands infalpha- b inhibitory effects on cd (+)cd (+)foxp (+) regulatory t cells in the tumor microenvironment of c bl/ j mice with melanoma xenografts stress-responsive regulation of long noncoding rnas' polyadenylation in oryza sativa exogenous plant mir a specifically targets mammalian ldlrap : evidence of cross-kingdom regulation by microrna antiviral activity of human oligoadenylate synthetases-like (oasl) is mediated by enhancing retinoic acid-inducible gene i (rig-i) signaling lipid-mediated delivery of rna is more efficient than delivery of dna in non-dividing cells ribose '-omethylation provides a molecular signature for the distinction of self and non-self mrna dependent on the rna sensor mda the present document has been produced and adopted by the bodies identified above as authors. this task has been carried out exclusively by the authors in the context of a contract between the european food safety authority and the authors, awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the authors. the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). (zhou et al., ) . twenty female and ten male rats per group in the first generation (f ) were fed the different diets. their pups, weanling/female/group and weanling/male/group were randomly selected as an f generation. the f generation was acquired by using the same procedure described above. at weaning, f generation rats were chosen, and rats/sex/group provided the corresponding diets and observed for weeks. no major differences in animal survival, health status, behaviour, body weight, food consumption, or reproductive capacity were observed with the different diets. some statistically significant differences were observed in animals given the transgenic rice diets compared to those receiving the standard diet for certain clinical chemistry parameters, in certain generations (alt, alkp, ldh, cholesterol, hdlc, ldlc). these changes were not considered adverse or biologically significant (zhou et al., ) . moreover, no evidence of altered incidence or altered severity of background changes was observed in any organ or tissues of the rats fed the three diets (zhou et al., ) . although minor changes in the mean relative weight of epididimides in male f adults was observed in rats fed the rice diets compared to the control group, no difference was observed between the transgenic rice fed rats and their near-isogenic line controls, suggesting overall that the transgenic line was unlikely to cause any risk to rat health in reproduction or development, even when consumed for up to three generations (zhou et al., ) . however, these studies did not evaluate any aspect related to rnai.heinemann et al. compared the history of risk assessment of gmos producing dsrnas in australia, new zealand and brazil, with a focus on regulatory context (heinemann et al., ) . the authors suggested some processes to properly assess the safety of dsrna-producing gm plants before their release or marketing. these include i) bioinformatics analysis to identify any likely, unintended targets of the dsrna in human and animals; ii) experimental procedures that would identify all new intended and unintended dsrna molecules in the gm product; iii) testing animal and human cells in tissue cultures for a response to intended and unintended effects of dsrnas from the product; iv) long-term testing on animals; and possibly v) clinical trials on human volunteers (heinemann et al., ) . in response to this article, the regulatory agency food standards australia new zealand (fsanz) published a document addressing regulation of gm crops and foods developed using gene silencing (http://www.foodstandards.gov.au/consumer/gmfood/pages/response-to-heinemann-et-al-on-theregulation-of-gm-crops-and-foods-developed-using-gene-silencing.aspx). this document criticised some of the comments of the above risk assessment review. some key points included the lack of weight of scientific evidence (published up to ) to support the view that small dsrnas in foods are likely to have adverse consequences in humans; the lack of a scientific basis for suggesting that small dsrnas present in gm foods have different properties than naturally-occurring ones; or the adequate acknowledge of many barriers (in the uptake, distribution and targeting) during oral development of small dsrna therapies, among others. the overall suggestion was that there was no need to consider additional studies as proposed by heinemann et al. ( ) .commentary documents reviewing scientific meetings in the context of rnai technologies in agricultural biotechnology discuss some of the aspects reviewed in this document .additional relevant documents of baseline information to support the risk assessment and environmental risk assessment of rnai-based gm plants can be obtained from efsa external scientific reports the present document has been produced and adopted by the bodies identified above as author(s). this task has been carried out exclusively by the author(s) in the context of a contract between the european food safety authority and the author(s), awarded following a tender procedure. the present document is published complying with the transparency principle to which the authority is subject. it may not be considered as an output adopted by the authority. the european food safety authority reserves its rights, view and position as regards the issues addressed and the conclusions reached in the present document, without prejudice to the rights of the author(s). key: cord- -mqrx q authors: seabra, claudia; reis, pedro; abrantes, josé luís title: the influence of terrorism in tourism arrivals: a longitudinal approach in a mediterranean country date: - - journal: annals of tourism research doi: . /j.annals. . sha: doc_id: cord_uid: mqrx q abstract this longitudinal study examines the impact that terrorist attacks within a representative group of european countries can have on the tourism demand of a south european country with no record of terrorism attacks. in order to analyze the connections between terrorist attacks and tourists' arrivals, occurred between and the end of , an unrestricted vector autoregressive model was used for multivariate time series analysis. the main results show that terrorist attacks have a strong impact on tourist arrivals and confirm the existence of terrorism spillover, namely the substitution and generalization effects phenomena. terrorism has become an important and recurring topic in the public discourse over the last decades due to the increased frequency of terrorist attacks witnessed (national consortium for the study of terrorism and responses to terrorism, ). since the beginning of the century, and particularly over the last years, several developed and stable countries have witnessed high levels of terrorist events (lanouar & goaied, ) . new york, united states ( ) , madrid and barcelona, spain ( , london and manchester, united kingdom ( , ) , tuusula, finland ( ) , apeldoorn, netherlands ( ), utoya, norway ( ), paris and nice, france ( , brussels, belgium ( ) , sousse, tunisia ( ) , berlin, germany ( ) , istanbul, turkey ( , christchurch, new zealand ( ), colombo, sri lanka ( ) were some of the places that have experienced terrorist attacks, but the list goes on (national consortium for the study of terrorism and responses to terrorism, ). revolutionary and destructive groups carry out attacks all over the world every single day, transforming terrorism into a constant in modern life (coca-stefaniak & morrison, ) . according to the "prospect theory" (tversky & kahneman, ) , tourists are rational consumers who, when confronted with two different options, tend to choose the option that will bring them more potential gains and lower risk (seabra, kastenholz, abrantes, & reis, ) . when tourists perceive the existence of any risk of terrorism, they become more careful as they plan their travel and tend to adopt risk-reducing strategies (fuchs & reichel, ) . tourists are sensitive to terrorism threats and can be flexible in their destination choices (neumayer & plümper, ) , so they will avoid destinations they believe to be connected with that sort of threat (rittichainuwat & chakraborty, turning into a single market, which leads to the standardization of consumer lifestyles and the rising of "global consumers" (hollensen, ) . tourism is one of the most global economic activities and is, therefore, vulnerable to risks that affect any other global business (fennell, ) . the world has become interdependent and tourism crises in one part of the world can have strong repercussions on other locations (lanouar & goaied, ; ritchie, ) . when tourists realize that a destination is unsafe, they replace it with others they consider safer and, in doing so, may damage entire regions that are affected by violence or that tourists consider threatened by terrorism (beirman, ; gu & martin, ; lutz & lutz, ; mansfeld, ; sönmez, ) . this fact draws attention to the spatial patterns of terrorism impacts caused by spillover effect on destinations/regions (Öcal & yildirim, ) . the topic of terrorism spillover effects has produced a considerable amount of literature (veréb, nobre, & farhangmehr, ) . the decline in tourists' arrivals and receipts caused by terrorism is well documented in several countries and regions since the s and has affected countries like spain (enders & sandler, ) , european countries ( (enders, sandler, & parise, ; radić, dragičević, & sotošek, ) , the mediterranean region (drakos & kutan, ) , non-democratic countries and africa (blomberg, hess, & orphanides, ) , the usa (bonham, edmonds, & mak, ; goodrich, ) , israel (eckstein & tsiddon, ; fleisher & buccola, ; morag, ; pizam & fleischer, ) , italy (greenbaum & hultquist, ) , nepal (baral, baral, & nigel, ) , ireland (o'connor, stafford, & gallagher, ) , fiji and kenya (fletcher & morakabati, ) , nigeria (adora, ) ; turkey (feridun, ; ozsoy & sahin, ) , pakistan (raza & jawaid, ) , the middle east (bassil, ) , the caribbean (lutz & lutz, ) , tunisia (lanouar & goaied, ) , and worlwide (liu & pratt, ; llorca-vivero, ; neumayer & plümper, ) . despite all past research conducted on the impacts of terrorism on tourism industry, and bearing in mind recent disturbing events, many sectors are calling for further in-depth analysis of this issue (almuhrzi, scott, & alroiyami, ; saha & yap, ) . people have to learn how to deal with the changes that this new global terrorism context is bringing to tourism (veréb et al., ) and it is crucial to find ways to cope with the disruption in tourist flows in the wake of terrorism events (cohen & cohen, ) . more investigation is required on the impact that terrorism may have on tourist arrivals and it will have to take into account the changes that are affecting the spatial patterns of tourism flows and, specifically, the spillover effects of terrorism (neumayer, ; prideaux, ) . while most empirical studies report the negative spillover effects on tourism demand and receipts caused by the substitution effect affecting countries and regions suffering from terrorism activity, there is no study, to our knowledge, dealing with the effects those events have on a safe country with no record of terrorism attacks. portugal is currently one of the most popular european tourist destinations (world tourism organization, ) and has no history of terrorist attacks ever (national consortium for the study of terrorism and responses to terrorism, ). the country is considered the third most peaceful country in the world by global peace index (institute for economics & peace, ) and clearly benefits from this safe tourist destination image. the main goal of this study is to analyze the influence that terrorist attacks carried out in a representative group of european countries can have on a safe country. this particular study will focus on portuguese tourism demand and will take into account the possible consequences of the so-called spillover effect, particularly the substitution and generalization effects. this paper begins with an outline of the current literature available on tourism consumer behavior regarding terrorism risk and the spillover effects caused by terror. the literature review was used to develop the study's conceptual framework. discussion on the research methodology was the next logical step. using a longitudinal study, this work uses an unrestricted vector autoregressive model to test the conceptual framework. arguments about theoretical and management implications are discussed in the last part of the article. the results of this study demonstrate the existence of the so-called substitution effect caused by terrorism events and add information to the existing literature about the spatial spillover effects of terrorism on tourism activity. it was also possible to create a tour-terror index allowing managers to observe the potential impacts of attacks on tourists' arrivals and receipts in different regions. the definition of terrorism is troublesome. according to previous research (lutz & lutz, ) terrorism is conceptualized, in the global terrorism database, as "the threatened or actual use of illegal force and violence by a non-state actor to attain a political, economic, religious, or social goal through fear, coercion, or intimidation" (national consortium for the study of terrorism and responses to terrorism, ). terrorism attacks generally "appear to be random and dispersed in order to create tension in the widest possible audience" (Öcal & yildirim, , p. ) . tourism demand, as previously established, is particularly sensitive to terrorist attacks, since tourists' choices value safety, tranquility and peace (araña & león, ) . past research has repeatedly proved the strong exposure of tourism industry to terrorism and violence and showed that tourism is one of the economic sectors that are most vulnerable to this threat (araña & leon, ; avraham, ; lanouar & goaied, ; saha & yap, ) . several studies have proved its negative impacts on tourism activity as a whole in the short and long term (araña & león, ; baker, ; liu & pratt, ; pizam & smith, ; sönmez, ) . this is mainly because of the effects that terrorist threats have on tourists' decision-making process (neumayer, ; pizam & fleischer, ; ritchie, ; seabra, abrantes, & kastenholz, ; seabra et al., ; sönmez, apostolopoulos, & tarlow, ; sönmez & graefe, ) . safety is clearly one of tourists' main concerns. it is a basic human need. as such, it affects human behavior in general and consumer behavior in particular (isaac & velden, ) . when people travel, they do not want to feel exposed to situations that will threaten their integrity. tourists are concerned with travelling to a destination where they will be able to fulfill their desires with as little complications and threats to their safety as possible (seabra, dolnicar, abrantes, & kastenholz, ) . safety on vacation and leisure is an expected requirement for any visitor in any destination (baker, ) . therefore, stability is one of the key factors for the development of tourism industry (almuhrzi, scott, & alroiyami, ) . if some event disrupts this balance and causes a risk perception among tourists, it can have both a powerful and negative impact on demand. risk perception relates to the uncertainty of consequences and potential loss (dholakia, ) and is associated with a large number of factors that may influence tourists' decision making. since the first studies on leisure and travelling, seven types of risk perceptions have been considered (rohel & fesenmaier, ) : i) psychological: how travel will affect one's own self-image; ii) social: impacts on social image; iii) financial: the value of money; iv) time: the cost of the time wasted while planning and travelling; v) physical: impacts on physical and psychological wellbeing; vi) satisfaction: probability of experiencing some kind of dissatisfaction with the trip or one of its components; and vii) functional: associated with bad trip organization and malfunction of tourist equipment or transportation. those risks are associated with three travel dimensions: vacation risk, physical-equipment risk and destination risk. focusing specifically on the risk factors connected with tourism destinations, subsequent research drew attention to more specific issues such as: i) health: factors that may cause physical danger, injury or sickness (baker, ; jonas, mansfeld, paz, & potasman, ) ; ii) communication: language and communication barriers that may hinder the connections with locals or service providers (reisinger & mavondo, ) ; iii) natural disasters: the exposure to natural hazards (becken & hughey, ; pearlman & melnik, ) ; iv) crime: several forms of criminality and physical violence (brunt, mawby, & hambly, ; chesney-lind & lind, ; dimanche & leptic, ; michalko, ) ; v) political instability and violence (fletcher & morakabati, ; ioannides & apostolopoulos, ; saha & yap, ; sönmez, ; yap & saha, ) ; and vi) terrorism: probability of being involved in a terrorist attack (baker, ; lanouar & goaied, ; saha & yap, ; seabra et al., ) . terrorism and political instability are the threats that tourists fear the most (sönmez, ) . in fact, the younger generations rank "war, terrorism and political tension" as their top concerns (coca-stefaniak & morrison, ) . previous research reports that terrorism and political instability are "particularly intimidating risks due to the uncontrollable, involuntary and random nature of the potential harm involved in visiting destinations struck by such incidents" (kapuściński & richards, , p. ; see also cavleck, ; heng, ) . that way, the perceived risk of terrorism is more effective and will influence tourists' behaviors and decision-making, regardless of their country of origin or of their cultural background (seabra et al., ) . feridun ( ) suggests the existence of a negative causal effect of terrorism on tourism. raza and jawaid ( ) also proved the unidirectional causal relationship between terrorism and tourism. in their opinion, there is a unique direction of causality that goes from terrorism to tourism. the changes in tourists' decision making caused by perceived risks of terrorist attacks are mainly due to the so-called memory effect. memory effect means any feeling, apprehension, or panic that lead individuals to change their usual behaviors (shin, ) . people create a memory effect after experiencing or after hearing about devastating events that have occurred in a certain place. terrorist events can give rise to a great sensitivity between the tourists' initial memory of a destination andthe posterior image of such destination. this leaves a persistent recollection or memory effect (baggio & sainaghi, ) . past attacks directly influence tourist arrivals in the affected countries due to those negative memories. consequently, tourists will replace their usual leisure destinations with other places they consider safer (lutz & lutz, ) . eventually they will return to their favorite destinations and this is the reason why this memory effect is not permanent, although its effects may last for quite a long time as stated in previous research (baggio & sainaghi, ) . tourists show a high preference for more stable and peaceful geographic areas (araña & león, ; neumayer, ; reisinger & mavondo, ) since their main goal is to relax in an insouciant holiday environment (neumayer, ) . they have a big range of destinations available and, normally, they do not even consider travelling to places located near sites where risky incidents have occurred. they choose alternative destinations with similar characteristics but with a more stable environment (neumayer, ) . this has some negative economic impacts not only on the affected destinations (lanouar & goaied, ; sönmez et al., ) , but also on other nearby places. this is due to the spatial spillover effects (isaac & velden, ; walters & beirman, ) that will affect several destinations and entire regions (radić et al., ) . according to the institute for peace economics (institute for economics & peace, ), the contribution of the tourism sector to gross domestic product in was twice larger in countries where there had been no occurrence of terrorist attacks targeting tourists. between and , tourism average contribution to gross domestic product growth reached . % in countries that had experienced no terrorist attacks against tourists. in countries where attacks deliberately targeted tourists, it amounted to . % only. nowadays, safety is one of tourists' major concerns when they have to choose their travel destination (world tourism organization, ). spatial spillover effects of terrorism on tourism activity: generalization and substitution effects spillover effect, a term which comes from economics, refers to the positive or negative externalities that an economic activity can have on any element not directly associated with that given activity. in tourism, spillover effects are closely related to a spatial perspective. it refers to the inadvertent effects that the tourism industry of a certain region has on tourism flows to other regions. positive spillover effects occur when a region benefits from their neighbors' tourism growth, while the opposite can occur when a destination is affected by negative factors occurred in neighboring regions (yang & wong, ) . previous research on the spillover effects found the existence of two major impacts of terrorism on tourism industry: the generalization and the substitution effects. the "generalization effect" refers to the cognitive biases of tourists who tend to generalize the perceived image of a given destination to a whole region (saha & yap, ) . the generalization effect may cause completely safe countries to witness strong drops in their tourism arrivals and revenues as a result of insecurity episodes in nearby countries (enders et al., ) . empirical evidence showed that perceptions of terrorist threats, war, political instability, and violence in one country tend to make tourists assume that entire regions are risky (sönmez, ; walters & beirman, ) . although some studies have shown that neighboring countries can sometimes benefit from being considered immediate substitutes, it has also been proved that there is always a negative impact on tourist demand in the region where the incident has occurred (gu & martin, ) . mansfeld ( ) proved that tourists who had experienced terrorist events during the arab-israeli conflicts transferred their fear of terrorism to the middle east as a whole. this spillover effect resulted in a negative contagion effect in that region but, on the other hand, had a positive impact on the tourist activity of the mediterranean area. neighboring countries, or countries that share identical characteristics, gain tourists in the same proportion as the less safe countries lose visitors. the different patterns of substitution that affect destinations are commonly represented by concentric rings. the "inner ring" that includes more unstable destinations like egypt, israel, jordan, lebanon, and syria was replaced by an "outer ring" composed of more peacefull and terrorism-free countries, such as cyprus, greece and turkey (mansfeld, ) . wahab ( ) drew attention to the fact that intraregional tourists may be more sensitive to this effect since they seek more peaceful destinations located near places considered to be of higher risk. in his study, he claimed the existence of a positive spillover effect in the case of egypt. however, an interregional tourist may also generalize conflict to an entire region, which may explain the decline in tourist arrivals in iraq's neighboring countries during the gulf war (sönmez, ) . during that period, middle east and eastern mediterranean countries experienced a big decline in tourist arrivals. cyprus, for example, experienced a drop of . million visitors in and of . million in as a result of the perceived risk of terrorism and political instability (goodrich, ) . drakos and kutan ( ) stated that any terrorist or violence event taking place in a particular middle eastern and north african destination will naturally affect tourist's perceptions of another country in that region (drakos & kutan, ) . some years later, kozak and his colleagues also concluded that tourists form overall negative images of entire regions due to terrorism acts (north america and the middle east), health issues (severe acute respiratory syndrome in asia) and political conflicts (africa). tourists change their travel plans when they realize the existence of any potential threat or risk. they assume that incidents that take place in a specific country will affect its neighboring countries as well. this assumption results in a global devastating impact on the region (kozak, crotts, & law, ) . recently, more empirical evidence supported findings that random terrorism acts perpetrated in some countries have an impact on nearby areas not directly subject to such attacks. this was true for kenya, africa (masinde, buigut, & mung'atu, ) and jordan because the violent uprisings in neighboring countries have deeply affected the image of the whole geographic area (liu, schroeder, pennington-gray, & farajat, ) . the "substitution effect" in tourism refers to customers' tendency to replace one destination with another, due to a number of factors that include changes in the elasticity of demand, shifts in price, a desire to try new products or experiences (tribe, ) or because of risk perception (prideaux, ) . using an unrestricted vector autoregressive model to analyze monthly data for spain covering a period of time between and , enders and sandler ( ) identified negative unidirectional causality between terrorism attacks perpetrated by the separatist basque group eta and tourism activity in the country. the authors concluded that the biggest drop in tourists' arrivals occurred three months after the attacks and that a typical transnational terrorist incident can prevent about tourists from visiting a certain country (enders & sandler, ) . enders et al. ( ) , a year later, studied a large sample of european countries between and and determined that tourism reactions occurred mostly within a period of six to nine months after the incident and that there was clear evidence of the replacement of those destinations with others located in different regions (enders et al., ) . gu and martin ( ) analyzed the substitution effect based on tourist arrivals at orlando airport between and . the authors concluded that there was a direct correlation between the increase in terrorist attacks in the middle east and in europe and the number of arrivals recorded at that airport (gu & martin, ) . during their analysis conducted on three mediterranean countries -greece, israel and turkey -, using italy as a control variable, drakos and kutan ( ) tested the cross-country effects of terrorism on tourism in the mediterranean region between and . they concluded that the intensity of terrorism events, measured according to the location (urban vs rural) and the number of fatal casualties, led to shifts in tourist visits that caused positive and negative spillover. the country where the attack took place registered drops in tourist arrivals, while other nearby countries experienced an increase in their visitors. their empirical results revealed that israeli and turkish tourism industries are more sensitive to terrorism than their greek counterpart (drakos & kutan, ) . a study conducted on tourism flows after / in the united states and in hawaii reinforced the findings of earlier studies that showed the existence of that substitution effect. tourists change their travel plans instead of canceling them, giving priority to safer domestic destinations they perceive as less risky. the study results indicate that united states citizens replaced international destinations with hawaii balancing the significant drop of foreign arrivals that followed the / events (bonham et al., ) . focusing on the same period, araña and león ( ) used a discrete choice approach to assess the impact those worldwide events had on the way tourists evaluate the qualities of alternative and competing destinations located in the mediterranean area. the authors concluded that countries with higher proportion of islamic population, such as tunisia and turkey, suffered a more severe negative impact on their competitive value than other destinations, such as canary islands and the balearics, where this proportion was lower (araña & león, ) . using two estimation techniques, neumayer ( ) tested the impact of various forms of political violence on tourism. the cross-country analysis provides empirical evidence for intraregional, negative spillover and cross-regional substitution effects for countries where human rights are violated or where there is evidence of conflicts and other politically motivated violent events. the author concluded that tourists choose alternative destinations with similar characteristics but that are more stable (neumayer, ) . while examining the effects of terrorism on tourism activity in turkey, yaya ( ) found strong evidence that terrorism events in the country have affected the industry and that the madrid bombings had a positive effect on tourist arrivals in turkey. this happened because tourists perceive those two countries as close substitutes for one another (yaya, ) . other studies concluded that while some countries experience a negative indirect effect, terrorist attacks increase tourism demand in other neighboring countries that are known for their low or moderate risk level (saha & yap, ) . this is the case of dubai, for instance, that represents a safe regional alternative to lebanon or syria, two former popular tourism destinations destroyed by war (clancy, ) . a study dealing with the reactions of german tourists to unanticipated shocks and the respective impacts on risk perception and tourism destination selection analyzed several terrorism episodes: / ( ), egypt ( ), tunisia ( ), morocco ( ) and indonesia ( ) . the findings proved that those shocks heavily affected islamic countries and provided a temporary substitution effect in favor of (southern) european countries (ahlfeldt, franke, & maennig, ) . neumayer and plümper ( ) apply a model to explain tourists' arrivals using a lagged dependent variable and other lagged independent variables representing terrorist attacks proxies. using a spatial dyadic approach, the authors concluded that terrorist attacks on tourist destinations in a given country reduce tourist flows due to a substitution effect that is visible at several levels: i) from the countries in which the attacks took place and other similar source markets to a different destination or country; and ii) from the same tourism source market to other similar destinations (neumayer & plümper, ) . as stated, to this day there has been no record of terrorist events in portugal. other european countries such as luxembourg, monaco, lithuania, andorra, malta, montenegro, san marino, and slovenia are safe countries as well. however, none of these countries kept a record of tourism arrivals or receipts. with a large international recognition and several tourism awards granted to its many regions, cities and destinations, portugal has been one of the countries with the best performances in tourism. the country has experienced an average annual growth of % in tourists' arrivals since (world tourism organization, ) . in , portugal received > . million international tourists and . million domestic tourists. the arrivals had a massive impact on tourism receipts that have been growing in a sustained way over these last few years and amounted to > billion american dollars in (instituto nacional de estatística, ; world tourism organization, ). this growth is directly connected to the strategic positioning of the "destination portugal" brand that, in recent years, has brought portugal and other specific portuguese destinations several international awards. over the last years, portugal has won several international awards at the world travel awards: i) portugal "world's best tourist destination" (for the rd year in a row); (ii) lisbon "world's best citybreak destination"; iii) madeira "the best insular destination" (for the th year in a row); iv) sintra-monte da lua parks "the best example of heritage recovery"; v) portugal "the best european and world golf destination" (for the th year in a row); vi) "turismo de portugal" considered the world's best tourism organization, among many others. the image of portugal as a safe tourism destination has led to an exponential increase in tourist arrivals. in fact, portugal was the third safest country in the world in (institute for economics & peace, ) and is currently a huge european tourist attraction. portugal was the country chosen for this study because of its importance and potential and especially because it is regarded as a safe country with no record of terrorist events since ever. in the next subsections, sample and data will be presented. data includes tourists' arrivals in portugal, sorted by region of origin, and the number of terrorist attacks that have occurred in the selected european countries, from to the end of . the main database that supports this study was built upon two different time series sub-databases: i) tourism arrivals in portugal between and and ii) the terrorism events occurred worldwide during the same time period. the first sub-database was obtained from the portuguese public statistical entity (instituto nacional de estatística, ). data provided information about the number of tourists who stayed in any accommodation facility in portugal, between and the end of , sorted by region of origin. a reasonable medium-term time series is required if the analysis applies lagged variables (bell & jones, ) to achieve significant results. the number of guests has significantly increased, surpassing million tourists in (see fig. ). the compound annual growth rate from to was . %, and the number of tourists doubled during that period. over the last years, tourists coming from europe have exceeded the number of domestic tourists. these results reveal the increasing dynamics of the portuguese tourist sector and reflect the country's good and stable political, economic, and social environment. data collected regarding tourists' arrivals included all contributing countries and showed that the most representative emission markets in portugal are: europethe united kingdom, spain, france, and germany; africa -angola; america -brazil, the united states, and canada; asia -china and japan (instituto nacional de estatística, ). the second sub-database, obtained from global terrorism database (national consortium for the study of terrorism and responses to terrorism, ), gathers information about terrorist attacks per country and other aggregate geographic areas between and (see table ). terrorist attacks, according to global terrorism database, are violent incidents triggered by bombing or explosion, arm attacks, assassination, and facility/infrastructure attack caused by terrorist organizations or individuals in a domestic c. seabra, et al. annals of tourism research ( ) as well as in a transnational and international context. for each global terrorism database incident, information is provided on the date and location of the incident, the weapons used and the nature of the target, the number of casualties, and, when identifiable, the group or individual responsible for the attack (national consortium for the study of terrorism and responses to terrorism, ). table provides information about the number of terrorist attacks sorted by country, considering also the injuries and fatalities caused by the events. the main criterion to select the considered countries was the number of terrorist attacks they have experienced between and . the selected countries represent the biggest terrorist targets for each european region, as seen in table (highlighted in light gray). however, some representative countries were selected to avoid multicollinearity. those were countries with the highest rate of terrorist attacks and that had some potential connection with portugal (social, economic or tourist connections). the importance of those countries as emissive markets was also taken into consideration. keeping in mind the tourism regions considered by world tourism organization (world tourism organization, ), the sample includes the united kingdom, for northern europe; the western europe sample includes france and germany; the central/eastern europe sample incorporates russia -russia and ukraine attacks are highly correlated, so this tourism market was an obvious choiceand lastly israel, greece, and spain were part of the southern/mediterranean europe sample. israel was chosen over turkey considering that turkey is strongly correlated with germany that represents an important tourist market for portugal. in addition to being the countries with a higher incidence of terrorist attacks, france, england, germany, greece, and spain are countries that, along with portugal, belong to the european union. they also have ancient and strong foreign commercial relations with portugal (base de dados portugal contemporâneo, a). the first three countries considered are destinations chosen by many portuguese emigrants. therefore, strong communities have been created over the years and strong social relationships were established between different generations of families living both in their home countries and in the different host countries (base de dados portugal contemporâneo, b). those are, furthermore, the most relevant emission markets to portugal. as for central/eastern europe, russia was the country that registered the higher number of attacks. portuguese relationships with russia date back to (russian federation, ) and those relationships are strengthened by commercial links (oil and gas imports) (belyi, ) . currently, a large number of russian tourists choose portugal as their tourism destination. they represent an important asset for tourism due to their number and to their high purchasing power. as far as the south /mediterranean region is concerned israel is, along with spain and greece, the country that has experienced more terrorist attacks. portugal established diplomatic relations with israel in and those relations were reinforced in with the signing of cultural, economic, industrial, technical, and scientific cooperation agreements (república portuguesa, ). the main goal of this study is to analyze the potential connections between terrorist attacks in specific european countries and tourist arrivals in portugal. the countries' political and economic stability, safety and security can be advantages that may attract tourists who have proven to be loyal to destinations that have recently been targeted by terrorist attacks. this study follows the procedure applied by neumayer and plümper ( ) when they used lagged dependent variables. the information about the number of terrorist attacks occurred in a certain country was also taken into account to provide an explanation on tourist arrivals per country of origin. the conceptual autoregressive model has two distinctive independent variables' groups. ( ) one is related with the one-lag period and the two-lag period of the first differences (t -t − period) of the dependent variable values tourist arrivals from any continent or an autoregressive variable. the second, ( ) is related with the first differences (t -t − period) of the one-lag period values of attacks perpetrated against the selected countries, as described in fig. . then, evidence was collected to prove that all the series were non-stationary at levels (time periods) using kendall's tau statistic indicator, also known as dickey-fuller test (gujarati, ). ordinary least squares cannot be trusted because they may cause sweden united kingdom western europe greece israel italy turkey spurious (non-sense) regressions. therefore, differencing the series to make them stationary can be a solution, but it could cause the loss of important long run relationships between levels (gujarati, ). co-integration tests were also performed and led to negative results. thus, and in accordance with previous works, it became clear that an unrestricted vector autoregressive model, using first difference variables would be the best solution (esso, ; gospodinov & pesavento, ; toda & yamamoto, ) to resolve potential non-stationarity series problems (shin, ) . the unrestricted vector autoregressive model method resembles simultaneous-equation modeling as it considers several endogenous variables together. nevertheless, each endogenous variable is explained by its lagged values or past lagged values. the first difference between the variables and their respective lagged values were used towards non-co-integration and non-stationarity at levels (statacorp, ) . the unrestricted vector autoregressive model assumed that all the attacks against the respective countries were exogenous variables in the model. it also considered that there were no or few relationships among them and that the aim would be to study one-way causality relationship between attacks and tourists' arrivals in portugal and not the opposite. since there may be a time lag (one or two-year delay effect) in the impact caused by the terrorist attacks on tourists' arrivals, this study opts to use a one period and a two-period time lag. the study also includes post estimation robustness tests for autocorrelation (lagrange-multiplier test) (statacorp, ) and residual distribution tests, like the jarque-bera test (lutkepohl, ) . fig. shows the specification models used in the estimation. a granger causality test is used (asteriou & hall, ; zivot & wang, ) , suggesting that there is short run causality between the significant independent variables and the dependent variable. tourists' arrivals are an endogenous variable because its value is determined within the model. the terrorists' attacks against countries are exogenous variables, since their value is determined outside the model. in other words, they do not depend on tourist arrivals. there is no correlation between terrorist attacks and the error term whose data represents relevant input to the model. the objective of the analysis is to use terrorists' attacks as determinant variables. this is in line with the methodology used by neumayer and plümper ( ) in which terrorist attacks are exogenous variables that explain the arrivals of western tourists to islamic countries. table demonstrates the descriptive statistics of all variables and table shows the results of the model. data clearly suggests that tourists' arrivals from any country to portugal reflect the impact of a certain memory effect. hence, the first difference of the lagged autoregressive variable is statistically significant with a . % alpha for every equation. the granger causality test applied to the model, and to its respective sub models, one for each dependent variable, demonstrates the existence of unidirectional causal relationship between terrorist attacks and tourist arrivals in this country. the second autoregressive lagged dependent variable has an important impact on tourist arrivals from portugal, europe, africa, and america, but, in contrast, exerts negative impact on the addition of more tourists. tourist arrivals from asia are positively related with the attacks perpetrated in greece, france and in the united kingdom; however, attacks perpetrated in spain move those specific tourists away from portugal and force them to look for other destinations. portuguese domestic tourists appear to be more sensitive to attacks committed in france, spain, and greece. in this case, there is a positive correlation, even if in the case of spain, the level of significance is slightly over %. terrorist attacks perpetrated in israel, the united kingdom, greece, germany, and france directly influence tourist arrivals from europe. in contrast, terrorist attacks perpetrated in spain and russia contribute to dislocate tourists from portugal to other regions. all variables are significant at the . % level. even though there is evidence that all variables concerning tourists from africa are significant at the . % level, the coefficient direction changes in the case of israel, russia, spain, and greece. in fact, only terrorist attacks that took place in israel and greece affect negatively tourist arrivals from africa in portugal, while attacks perpetrated in other countries positively influence arrivals in the country. as far as tourists from america are concerned, it seems that attacks in russia, israel and spain move them away from portugal and force them to choose other destinations, while attacks in france, germany and greece bring those tourists to portugal. tourists from oceania are negatively sensitive to attacks committed in russia and spain. a phenomenon that involves extremely sensitive reaction to terrorist attacks makes tourists replace a destination with other safer countries. post estimation tests to the models revealed the absence of auto-correlation after using the lagrange multiplier test, as null hypothesis of zero autocorrelation could not be rejected and as the null hypothesis under jarque-bera statistic of residuals' normality (statacorp, ) could not be rejected either. despite the importance granted to the effects of terrorism on tourism and in spite of a considerable amount of research conducted on such issue, there have been recent calls for new research on the topic, especially on the consequences that this phenomenon can exert on economy and tourism activity (almuhrzi, scott, & alroiyami, ; saha & yap, ) . this paper represents an effort to close this gap by establishing the connections between terrorist events in some regions and tourist arrivals in other regions. the study takes into account previous literature on spillover effects, specifically those that trigger the substitution and generalization effects that make tourists reject and replace destinations considered unsafe with others they consider safer (gu & martin, ; neumayer, ) . whereas past research analyzes terrorism negative spillover effects on tourism demand, the present study focuses on the effects that terrorism events have on the tourists' arrivals in a safe country that has no record of terrorism attacks. portugal is one of the european tourism destinations that have experienced a higher growth in tourism demand and receipts over the last decades. according to the institute for economics and peace, portugal is one of the world's most peaceful countries (institute for economics & peace, ) and has no record of terrorism events (national consortium for the study of terrorism and responses to terrorism, note: + p < , ; * p < . ; ** p < . ; *** p < . ; d first difference; l, l lags of respective variable; first number in the table is the variable coefficient and the second one in the same column is the z statistics. these are the significant values c. seabra, et al. annals of tourism research ( ) ). the present study demonstrates that terrorist attacks in other countries have an impact on the demand for portuguese tourist accommodations. in turn, the results also show that instability in other countries can have consequences for portuguese tourism arrivals confirming the existence of spillover effect. as far as the effects of terrorist attacks on tourists' arrivals in portugal are concerned, this study allowed the uncovering of interesting results. terrorist events perpetrated in the united kingdom, greece, and france positively affect tourist arrivals from asia. attacks perpetrated in spain, on the contrary, have a negative contribution to tourists' arrivals from asia. in the cases of france and greece, this occurs probably because they are both mediterranean destinations similar to portugal. this result confirms the existence of the substitution effect as well as the presence of the generalization effect (mansfeld, ) . asian countries have a strong historic, economic and even social connection with the united kingdom. natives of those countries are well aware of the attacks committed in the united kingdom through media coverage or through personal information provided by emigrants residing in the country. on the other hand, for asian tourists, spain and portugal belong to the same iberian territory since they are located in the same geographical and risk area. this belief confirms the existence of the generalization effect (enders & sandler, ) . terrorist attacks in france, greece and spain strongly affect portuguese domestic tourism. these are important tourism destinations to portuguese tourists. these results confirm earlier research on unsafety effects and prove that during unsafety times tourists do not stop travelling; alternatively, they choose domestic destinations they consider much safer (bonham et al., ) . terrorist events perpetrated in the united kingdom, france, germany, greece, and israel positively affect tourist arrivals in portugal from european countries. on the other hand, terrorist attacks occurred in russia and spain exert a negative impact on tourists' arrivals from europe. this happens possibly because those countries are geographically located in the center of europe. accordingly, european tourists will choose peripheral countries. this assumption is in accordance with the results obtained by mansfeld ( ) . the united kingdom, france and germany are also important emissive markets to portugal. that means that tourists who come from those parts of the world can feel safer in portugal than in their own countries. since they are intraregional tourists, they have enough information to know which countries are the safest (wahab, ) . in the cases of tourist destination such as greece or israel, the substitution effect involves portugal, probably because the three markets share the same emissive tourist markets. spain and portugal are neighboring countries. terrorist attacks in spain have severe impacts on tourists who come to portugal from asia, oceania, america, and from other european countries. however, tourists coming from africa, namely from angola, a portuguese-speaking african country (palop) and member of the community of portuguese speaking countries (cplp), or from portugal are unbiased and, thus, do not feel that effect. for those tourists, spain and portugal are two different and unrelated nations, even though they are both iberian countries. the overall findings show the existence of the so-called generalization effect (drakos & kutan, ; enders et al., ; kozak et al., ; liu et al., ; masinde et al., ) : a safe country like portugal with no terrorist events can suffer from the impact of episodes of insecurity that took place in spain, a neighboring country. such impact will affect tourism demand. moreover, this work confirms that the generalization effect that over-assumes the similarities of neighboring countries enhances the causal relationship between terrorist attacks and tourist arrivals (feridun, ; raza & jawaid, ) . the attacks perpetrated in russia do not produce changes in the destination choices of tourists coming from european traditional markets and tourists remain faithful to traditional destinations like spain, france, italy, greece, and tunisia, among others. in consequence, there is a decrease in tourists' arrivals in portugal. the attacks committed in the united kingdom, russia, spain, france, and in germany have a positive impact on the increase in tourists from africa who choose portugal as their destination. on the other hand, terrorist incidents in greece and israel seem to have a negative impact on their choice. these results may be due to the cultural and economic proximity of most african countries and specifically of angola, which is portugal's most important african market. terrorist attacks in greece and israel are responsible for the decrease in african tourists' arrivals, probably because instead of coming to portugal they rather choose high level shopping tourism destinations such as italy, france, the united kingdom, and germany. terrorism in greece, germany, and france positively affects tourist arrivals from america, while terrorist events occurred in israel, russia and spain will have a negative effect on the number of american tourists who choose portugal as their destination. the substitution effect is, once again, clear as far as greece is concerned since that destination is quite similar to portugal. germany and france are important markets for brazilian, north american and canadian tourists. however, due to the attacks in germany and france, those tourists decide to come to portugal instead. tourists from america are well aware of the repeated and regular instability and unsafety felt in russia and israel, so any negative event in those countries will force them to choose other traditional or domestic destinations. these findings are in close accordance with previous research conducted by pizam and fleischer ( ) . the influence of the memory effect is clear when past tourist arrivals influence current tourist arrivals. this occurs in all the models except in the case of tourist arrivals from oceania. this means people want to relive pleasant past experiences and return to destinations where they had spent enjoyable moments. this positive feeling reinforces the role played by memory effect when time comes to choose a destination. therefore, the present study reinforces the work carried out by baggio and sainaghi ( ) . asian, european, african, american, and portuguese tourists' experiences in portugal will have a strong and positive impact on the flow of tourists who will choose to visit the country in the coming years. this paper provides a number of theoretical contributions to tourism literature and helps explain the factors that may influence tourist flows. evidence collected show that tourist arrivals and the demand for tourist accommodation depend on international markets and, more specifically, on the existence of terrorist attacks in other countries. these assumptions confirm the existence of c. seabra, et al. annals of tourism research ( ) "global consumers" (hollensen, ) . this study clearly demonstrates the existence of the so-called substitution effect caused by terrorism events. tourists choose destinations perceived as safer instead of places they see as potential terrorist targets, as stated in previous studies (e.g. ahlfeldt et al., ; araña & león, ; bonham et al., ; drakos & kutan, ; enders et al., ; enders & sandler, ; gu & martin, ; neumayer, ; neumayer & plümper, ; yaya, ). secondly, it was possible to confirm the existence of a one-year consumer short memory effect. the increase in tourist arrivals recorded during a certain year-long period has a direct impact on the increase of tourist arrivals that will occur in the following years. this result may be related with the positive image created by tourists' word of mouth or through the repetition effect. however, a longer, two-year memory effect shows different results. tourist arrivals from portugal, europe, africa, and america in a two-year lag reveal an opposite impact on the arrival of tourists in portuguese destinations, confirming the findings of the study conducted by enders and sandler ( ) . this study also contributes to add information to the existing literature that analyzes the effect of terrorist attacks on tourist arrivals according to their geographical origin. the results unveiled by this study allow public and private tourism organizations to create new predictive models or to update those already in motion, by adding new variables to improve their efficiency and increase the knowledge needed to understand and anticipate international tourist behaviors, specifically those that are related with the existence of terrorist attacks in other european regions. moreover, using the theoretical background provided by this paper, it was possible to create an index -tour-terror -that allows managers to observe the potential impacts of attacks perpetrated in a given region on the economy of recipient countries and that will lead to an increase or to a decrease in tourist arrivals. those fluctuations are related with the tourists' characteristics such as their cultural, social, and economic origin. these features may differ from the residents' usual standards but also from those of the average tourist who usually visits the country. accordingly, the tour-terror index determinants of a country may include independent variables like: i) the level of change in tourism demand and supply due to terrorist attacks; ii) the tourist price level structure; iii) tourists' purchasing power parity; iv) the duration of the stay; v) tourist accommodation capacity; vi) the capacity of other related facilities; and vii) human labor capacity. managers should face the fact that terrorist attacks provoke a substitution effect on destination choice behaviors. tourists will replace destinations considered unsafe due to terrorist threats with others considered safer. the physical distance of the tourists' home countries and their cultural and socio-economic traits influence this replacement behavior. in addition, managers must take into account that the substitution effect occurs between the european countries located in central areas and those located in peripheral areas. when terrorist attacks occur in the center of europe, tourists avoid those regions and choose peripheral destinations like portugal. evidence also made it clear that the opposite effect also happens. given the randomness of terrorist attacks, tourism managers should be prepared to alter quickly their marketing strategy, namely their market targeting strategies and promotion campaigns to prevent substitution effect. these specific practical implications are particularly important to tourist companies located in portugal. tourists from angola, brazil, the united states, and canada have proven to be very profitable since they stay longer and purchase high level and luxury products. for that reason, portuguese companies should make them their main target. the first limitation refers to the research settings, namely the fact that the study only considers one country and a -year time lag. more countries or bigger regions and a longer time lag could improve the analysis. a broader period of analysis could prove the relevance of additional factors like price level, crime rates, accommodation profit, price effect and quantity effect registered in portugal, among others. a more detailed analysis could use sub-regions or even countries instead of continents for tourist arrivals. further analysis should study the countries' macro-economic and social-cultural fundamentals to improve the results. managing tourism in nigeria: the security option terrorism and international tourism: the case of germany tourism in the arab world: an industry perspective the impact of terrorism on tourism demand vector autoregressive (var) models and 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of uncertainty the fear of terrorism and shift in cosmopolitan values tourism and terrorism: sinthesis of the problem with emphasis on egypt marketing the middle east in times of political instability-the case of jordan a spatial econometric approach to model spillover effects in tourism flows do political instability, terrorism, and corruption have deterring effects on tourism development even in the presence of unesco heritage? a crosscountry panel estimate terrorism and tourism: the case of turkey vector autoregressive models for multivariate time series she is doing her post-phd on "terrorism and the eu : impact on citizens and organizations" in novasbe. her research interests are: ) safety and terrorism, and ) tourism. email: cseabra@uc pedro reis is a professor at polytechnic institute of viseu. pedro develops his research in: ) finance, ) accounting josé luís abrantes develops his research in: ) marketing, ) tourism, and ) pedagogy key: cord- -lgdsi m authors: lin, feng; huang, yi; zhang, huifang; he, xu; yin, yonghua; liu, jiaxin title: evaluating the different control policies for covid- between mainland china and european countries by a mathematical model in the confirmed cases date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: lgdsi m this study focuses on evaluating the different policies of controlling the outbreak of covid- in mainland china and in some european countries. the study is based on mathematical model which is a modified susceptible-infected-recovered (sir) model. the model takes death and recovery into consideration which in convenience is called the susceptible-infected-recovered-death (sird) model. the criterion for the recovered patients is assumed by covid- nucleic acid testing negative. the mathematical model is constructed by retrospective study. determination of the parameters in the model is based on the epidemic bulletin supplied by the chinese center for disease control and prevention (cdc) and national health commission of the people's republic of china (nhc) from jan to mar . the data cover the date when the epidemic situation is reported and the data showed that the epidemic situation is almost under control in china. the mathematical model mainly simulates the active cases and the deaths during the outbreak of covid- . then apply the mathematical model to simulate the epidemic situations in italy and spain, which are suffering the outbreak of covid- in europe. the determination of the parameters for the european countries is based on the data supplied by worldometers. by comparing the difference of the parameters based on the same mathematical model, it is possible to evaluate the different policies in different countries. it turns out that the relatively easing control policies might lead to rapid spread of the disease. the current outbreak of coronavirus disease was first reported from wuhan, china, on december [ ] . when the confirmed cases in more than countries worldwide raised concern on an international scale, it is closely monitored by researchers and public alike [ ] [ ] [ ] [ ] . according to the infection, the data of covid- supplied by the chinese center for disease control and prevention (cdc) and national health commission of the people's republic of china (nhc) in mainland china, the outbreak of covid- in mainland china is almost under control, while in european countries the epidemic situation is severe as the number of deaths increases rapidly [ ] . in january the disease spread very fast in china. according to the information supplied by nhc and the chinese cdc, the number of confirmed cases was on jan , which increased to more than on mar . as the increasing rate of the active cases is close to in march, it could be deduced that the epidemic situation is almost under control. however the epidemic situation in european countries was severe in march. as there was a relatively complete process from epidemic outbreak to control in china, it could supply a model by retrospective study. a mathematical model is proposed by modified susceptibleinfected-recovered (sir) model, which for convenience is named as susceptible-infected-recovered-death (sird) model. the determination of parameters relies on case number data provided by the chinese cdc which updates the number of active cases both in china and the global per day [ ] and nhc which mainly updates the number of the active cases, the close contacts and the recovered cases in china [ ] . using the mathematical model proposed by the epidemic situation in china, the epidemic situation in european countries could also be estimated in the same way. the determination of the parameters in european countries relies on the data from worldometer [ ] . in this study european countries italy and spain where the epidemic situation is quite severe are chosen . the sir model is an effective model to predict the growth of the confirmed cases [ ] . the parameters in the sir model are usually constant [ ] [ ] [ ] [ ] . and the mortality is not taken into consideration. so far, the researchers have already taken banning of traffic into consideration. by retrospective study, it is found that the parameters are actually time-dependent. and the mortality may be an important variable during the outbreak of the disease. even though the death is much less than the infected population, it needs to be considered. in the traditional sir model, this kind of information is not taken into consideration. with these modifications, a new model proposed in this study is called the susceptible-infected-recovered-death (sird) model. the mathematical model in this study only contains kinds of variables: susceptible group s(t), infected group i(t) (the confirmed cases), recovered group r(t) (the nucleic testing negative after infection) and the mortality group d(t). the transportation control is not involved in this model. the mechanism only reflects the spread of disease in the healthy population. the result in this model could be compared with the transportation control which is the actual result. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. before introducing the mathematical model, the observed results are shown in figure . the sub-figures are (a)- (d). the observation of the confirmed cases, the death, the close contacts and the recovered cases lasts for days. it began from jan to mar . figure (a) and figure (b) imply that the growth of the close contacts and the confirmed cases slowed down on the th day. and figure (d) implies that the growth of the deaths slowed down on the th day. according to the data, rapid increase of the confirmed cases implies that the growth of it might be in the exponential form [ ] . comparing figure (a) with figure (b) , it implies that the close contacts are actually the susceptible population rather than the total population in china or in hubei province. the transportation policies in china [ ] [ ] [ ] [ ] [ ] [ ] is proposed for controlling the outbreak of the disease. if some one was traveling by train as was diagnosed, all the people in that train should be confined in a isolated area. this means that all the people in that train were assumed to be susceptible one, while people who are not in that trip are not regarded as the close contacts. other policies should also been taken into consideration, such as active cases were either quarantined or put under a form of self-quarantine at home [ ] . the suspicious cases were also either quarantined or put under a form of self-quarantine [ ] . the policies imply that the number of the susceptible population only depends on the cumulated number of the close contacts. fig. (a) ), the active cases ( fig. (b) ), the recovered cases ( fig. (b) ) and the death ( fig. (d) ) in this work, the variable s(t) which denotes the number of the susceptible cases in sir model is related to the number of the close contacts. by retrospective study, the total number of the close contacts is . × . however, as the epidemic situation is changeable, it is difficult to give the exact number of close contacts, which are the susceptible population. moreover the number of the close contacts is increasing day by day, while the infected number of population is growing. so it could be deduced that in fact the exact number of the close contacts is unknown. for these reasons, the mechanism of infection-recovering-death is given by ( )-( ): formula ( ) means that the number of the susceptible population consists of parts: the number of confirmed cases would increase the number of the susceptible population by the rate of r , and the susceptible population is converted into the confirmed cases when they are in contact with the confirmed cases with the rate r . when the confirmed cases were in contact with those healthy ones, the healthy ones might be transmitted into the infected cases. the confirmed cases are described as variable i(t), by formula ( ). according to the nucleic testing, the recovered cases could be defined by the infected cases whose nucleic testing negative. in this model, it is assumed that the recovered population with nucleic acid testing positive again might change into the active cases, as it is reported that some recovered covid- patients appeared nucleic acid testing positive [ ] . the increase of i(t) depends on changing the susceptible cases into the active cases and the recovered population with nucleic acid testing positive. the recovery rate µ and the death rate δ eliminate the number of the active cases. the number of the recovered patients depends on the variable i(t) in equation ( ). the number of the death is described in equation ( ). the transmission coefficients would be estimated by the observed results. the determination of the transmission coefficients mainly reported in appendix. in this section we only give an introduction of determining the transmission rate. the transmission rate r calculated by the observed results is shown in figure (a), and the death transmission rate δ calculated by observed results all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted april , . according to the statistical character of r , its mean value is . . the shape of its distribution function skew-right (skewness> ), and the distribution function is "thinner" than normal distribution (kurtosis> ). so the most probable number of r is larger than . . in this work we choose r = . . the detailed approach of determining the formula of the transmission rates would be shown in appendix. similarly, the recovery parameter is also time-dependent. the details of determination of the time-dependent function of δ(t) and µ(t) are shown in appendix. there only show the results. δ(t) is chosen in the formula as ( ) . the parameter ρ = . , as the incidence of the recovered patients with nucleic acid testing positive is very low. r is to be set according to the observed number. r must let the peak number of the confirmed cases similar to the observed ones. as the epidemic situations in european countries is changeable, the model ( )-( ) is also used for prediction. the curing rate is assumed to be the same, as there is no reason to assume the medical resources in european countries are better or worse than it in china. the difference stems from the medical resources management, which are concerned with δ, r and r . the data have been in collection since feb [ ] . according to the observed data, the parameters r and r should be adjusted. the death rate needs to be re-estimated as it reflects the management of medical resources in european countries. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted april , . . https://doi.org/ . / . . . doi: medrxiv preprint lack of medical resources is also reflected in european countries. comparing with the cases in china, it might be more severe in european countries. the death rate of italy and spain is estimated as in the formula ( ): the formula ( ) is estimated in the same way as that was done for china. it is obvious that the death rate is larger in european countries than that in china. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted april , . (a) and fig. (b) ) is the number of the population that infected by the disease. the brown line ( fig. (a) ) is the population that are recovered. and the dashed red line is the number of the death ( fig. (a) and fig. (c) ). the parameters are r = . , r = . , ρ = . . µ(t) and δ(t) are referred to formular ( ) and formula ( ) . the initial conditions are s(t ) = , i(t ) = , r(t ) = , d(t ) = all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. comparing the simulated results with the actual results would reflect the infection under kinds of cases: one is the ad libitum transportation, the other is the transportation under control. it has already been mentioned in the previous introduction, the mechanism of this model did not take the transportation control into consideration. so this result could reflect how the infection would evolve without transportation control. figure shows the comparison of the simulated number with the actual number supplied by the chinese cdc. figure (a) gives the comparison of the number of the infected population every day in mainland china with the simulated result. and figure (b) shows the comparison of the number of the death in mainland china (blue dots), hubei province china (red dots) and the simulated result. in fig. (a) , the data of the active cases in hubei province china is limited, so the number of active cases is not available. generally, the increase of the active cases and the death simulated from the mathematical model is not as large as that in the actual data, and the key performances such as the peak number of the active cases, the number of the death are consistent with the actual information. in european countries, the mechanism is the same as that in china. the time-dependent parameter the recovery rate is the same as that in china. the difference stems from the parameters like r , ρ in the model. the death rate might be high at the beginning, because of lack of medical resources and curing experience. later the death rate is slowed down, while the recovery rate would increase slowly. the policies in european country is different from that in china. the prediction results of italy are shown in figure (a)- (b). all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted april , . ( ) and ( ) . the initial conditions are s(t ) = , i(t ) = , r(t ) = , d(t ) = similar to the simulation in china, the increase rate in simulation is not as large as that in the actual data. as it has been reported that the susceptible cases might be very inexplicit [ ] [ ] [ ] , the initial number of the susceptible cases s(t ) is not accurate. it would be shown in the section of discussion. the model simulation for china and european countries are with the same mechanism. the only difference are the parameters r and ρ. and it results in the difference of δ. according to equations ( )- all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted april , . . https://doi.org/ . / . . . doi: medrxiv preprint ( ), r is related to the increasing of the susceptible cases and ρ is related to the number of the recovered patients with nucleic acid testing positive. both of the parameters are related to the transportation policies. the parameters are determined by the actual data, so the simulation in figures - are close to the actual data. the parameter r and ρ are adjusted to fit the observed data. though the simulation result is different from the observed data, mainly in the increase rate, the relative quantity of r and ρ in china and european countries implies whether the epidemic control policies is rigorous. in figures - , it is clear that the simulation result is delayed comparing with the actual result. there might be reasons: one is that the mechanism does not explicitly contain banning of the transportation [ , , ] , the other is that the initial number of susceptible cases is not accurate due to lack of access to covid- testing in the early weeks [ ] [ ] [ ] . for the second reason, we change the initial number of s(t ) to check whether this deduction is right. if in fact the initial number of the susceptible cases was rather than in china, and was rather than in italy and spain, then the active cases and the death is shown in figures - . the number of the active cases were the same as those in the previous simulation, the only change is the initial number of the susceptible population and the coefficient r . it turns out that r for european countries is still larger than that for china. it implies that the banning of transportation for china is more rigorous. moreover, the initial number of the susceptible population is larger than that was reported regardless of the country due to lack of access to covid- test kits in the early weeks. in view of the simulation by mathematical model, the initial number of the susceptible cases might be difficult to be determined. this might be demonstrated by the report that % of active cases might have no symptoms but could infect other healthy people [ ] [ ] [ ] . and it is reported that the infected cases are of strong infection when they appeared to have no symptoms [ ] . so when the first infected person appears in some area, the infected ones with no symptoms might convert the healthy people into the susceptible ones without records. in the hypothesised simulation, the actual data seemed to be closer to the simulated curves. but the increasing rate of the simulation is not as large as that in the observed data. the reason might be that the transportation control was not considered in this mechanism, which has already been mentioned in the previous section. this implies that banning of the traffic is an effective way to prevent the spreading of disease. the authors have declared no competing interest. the infection coefficient is calculated in this way: all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted april , . . the reason why we wrote before the transition rate of infection is that it is assumed that the contact in fact occurs one by one quite often. the dimension of is per person i.e. i(t) is dimensionless. for calculation convenience, we just figure out the variable part. according to the data given by chinese cdc and nhc, the bound of infection coefficient is r ∈ ( . , . ). it is hard to give a formula to describe r . the best way is to choose an exact number in its % confidence interval to set r . the statistical character of infection coefficient is given by table . the confidence interval is figured by the population mean estimated from the actual data: the death coefficient is calculated in this way: the death coefficient is like some exponential function of t. by fitting the time to the ln δ(t). it changes from time to time. however, the tendency of the death coefficient is like some exponential function of t. so by changing the bases of exponential function, it could turn out a series of fitted function. some fitted function is shown in figure china. the data of the death and the patients could be checked on the website of chinese cdc and nhc. in figure , the black line is: the brown line is all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted april , . in this study we choose the formula fitted as black line: the reason is that when t goes to infinity, δ → . . the recovery coefficient is calculated in this way: µ(t) is like some power function of t. by changing the power of t, the result gives: figure : the recovery coefficient per day is shown in dots, and the simulated curing coefficients is shown in the line all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted april , . . https://doi.org/ . / . . . doi: medrxiv preprint the fitted formula of black line is µ(t) = . t . ( ) the brown line is the fitted formula µ(t) = . t ( ) however, the blue line is the best: the fitted formulas are given below: in this study we use the formula: effective containment explains sub-exponential growth in confirmed cases of recent covid- outbreak in mainland china how many infections of covid- there will be in the" diamond princess modelling-based evaluation of the effect of quarantine control by the chinese government in the coronavirus disease cmmid ncov working group, et al. interventions targeting air travellers early in the pandemic may delay local outbreaks of sars-cov- . medrxiv coronavirus cases in the world chinese center for disease control and prevention. covid- distribution in the world national health commission of the people's republic of china modeling infectious diseases in humans and animals the world needs to follow china methods to combat epidemic hubei raises health emergency response to highest level coronavirus battle in china: process and prospect yunnan adopts more measures to control virus outbreak lockdowns resolute move that needs national support clinical characteristics of the recovered covid- patients with re-detectable positive rna test. medrxiv covid- transmission through asymptomatic carriers is a challenge to containment. influenza and other respiratory viruses asymptomatic and human-to-human transmission of sars-cov- in a -family cluster, xuzhou, china. emerging infectious diseases asymptomatic and presymptomatic sars-cov- infections in residents of a long-term care skilled nursing facility)king county, washington association of public health interventions with the epidemiology of the covid- outbreak in wuhan, china hopkins experts present latest coronavirus information on capitol hill coronavirus death toll in u.s. almost certainly higher than official count universal screening for sars-cov- in women admitted for delivery temporal dynamics in viral shedding and transmissibility of covid- . medrxiv key: cord- -zisujjsx authors: sabat, iryna; neuman-böhme, sebastian; varghese, nirosha elsem; barros, pedro pita; brouwer, werner; van exel, job; schreyögg, jonas; stargardt, tom title: united but divided: policy responses and people's perceptions in the eu during the covid- outbreak date: - - journal: health policy doi: . /j.healthpol. . . sha: doc_id: cord_uid: zisujjsx to understand the public sentiment toward the measures used by policymakers for covid- containment, a survey among representative samples of the population in seven european countries was carried out in the first two weeks of april . the study addressed people's support for containment policies, worries about covid- consequences, and trust in sources of information. citizens were overall satisfied with their government's response to the pandemic; however, the extent of approval differed across countries and policy measures. a north-south divide in public opinion was noticeable across the european states. it was particularly pronounced for intrusive policy measures, such as mobile data use for movement tracking, economic concerns, and trust in the information from the national government. considerable differences in people's attitudes were noticed within countries, especially across individual regions and age groups. the findings suggest that the epidemic acts as a stressor, causing health and economic anxieties even in households that were not directly affected by the virus. at the same time, the burden of stress was unequally distributed across regions and age groups. based on the data collected, we draw lessons from the containment stage and identify several insights that can facilitate the design of lockdown exit strategies and future containment policies so that a high level of compliance can be expected. the outbreak of covid- triggered a wide range of responses from governments in the european union. given that the disease was new and effective medical countermeasures did not exist in early , governments had to adopt non-medical measures aiming at the containment and mitigation of covid- . with the aim of "flattening the curve," these policies included bans on public gatherings, closures of academic institutions and public places, national and international mobility restrictions, confinement, and several others [ ] . italy was the first country in europe to apply intervention measures from the beginning of march in response to the severity of the covid- outbreak. other eu countries followed soon afterward, using similar countermeasures around mid-march [ ] . the adoption of these policies varied in their scale, stringency, and pace across countries. while most european states implemented confinement measures, the extent of limitations of people's freedoms differed across individual countries. lockdowns were usually strictest where the pandemic was deadliest (italy, spain, and france), imposing severe limitations on population movements. some governments chose less stringent versions of confinement or no lockdown at all, for instance, "an intelligent lockdown" in the netherlands or "freedom under responsibility" in sweden [ ] . forced to react swiftly to the unfolding epidemic situation, policymakers in every country tried to balance the implementation of containment policies against numerous important factors with the priority mostly given to the protection of the population's health. consequently, there has been a lot of debate in every society about whether measures taken by the government were appropriate or not. some parts of the population have been voicing support for more severe containment policies to minimize the spread of the virus. such attitudes were likely fueled by people's worries about their health and the potential of their national healthcare system to withstand the epidemic. meanwhile, others expressed their concerns about the social and economic consequences of such policies, thereby advocating for less severe containment measures [ ] . as the pandemic began to abate, governments started designing the lockdown exit strategies and restarting their economies. however, the risk that the new wave of the epidemic may happen did not disappear, especially given that the vaccine development takes a long time, and herd immunity was not achieved [ ] . in this light, the issue of lifting lockdowns has become a new subject of public debate across and within european countries raising discussions about the appropriateness of timing, risks, and potential consequences of ending the confinement [ ] . lifting lockdown restrictions creates acute dilemmas to the policymakers since the economic and human costs of any exit strategy seem to be closely linked together. taking a utilitarian approach in this situation could backfire if the society's understanding is not preliminarily secured or expectations are not fulfilled. policymakers and public health experts have to persuade their citizens to make behavior changes and respect future containment interventions while facing the difficulty of enforcing such regulations. therefore, it becomes crucial to understand people's worries about the pandemic and their perceptions of the effects of containment policies, so that the design of further policies and contingency measures is well-informed, and a high level of compliance can be expected from the population. moreover, trust in the government and social institutions may become central to achieving a successful implementation of future measures, whereas lack of it may turn detrimental to the fight against the pandemic. hence it is of paramount importance to understand who people trust most so that public health messages can be amplified using correct means of communication. we provide a timely description of the current situation and draw lessons from the containment stage to inform the design and implementation of the lockdown exit policies. in order to understand the public sentiment towards the covid- containment measures and to inform future policy development, we collected information on people's support for these policies, their worries in relation to the unfolding epidemic, and their trust in different sources of information. we surveyed over , people representative of the adult population in seven european countries: denmark, france, germany, italy, portugal, the netherlands, and the uk. the fieldwork was conducted online during april - , , using multi-sourced online panels provided by the market research company dynata. to ensure that the sampling frame was representative given the online nature of the study, the company applied diverse recruiting procedures to reach the general population (through open recruitment, loyalty programs, affiliate networks, mobile apps). it then used quotas to match the national census shares in each country. the questionnaire was designed by the authors of the study except for the worry items that were adopted from the world health organization (who) covid- snapshot monitoring project [ ] . the questionnaire was carefully translated into six other languages by native speakers and then implemented using the qualtrics platform first as a pilot ( % of the sample in every country) and next as a large-scale survey. the data from the pilot study were included in the total sample. in each country, we collected data from a sample of , respondents representative of the national population in terms of region, age, gender, and education. given that the italian region lombardy was the most severely hit by the covid- outbreak, we collected additional responses in this region representative in terms of age and gender. learning about perceptions and attitudes of people who reside there could provide essential insights to researchers and policymakers. the extra data collected from lombardy were not included in the representative sample of italy. thus, no weighting was used as the additional lombardy sample was analyzed separately and denoted as lombardy in the results section. we assessed people's approval of policy measures that were taken (or were likely to be taken) by their national government in response to the covid- outbreak. in particular, we covered such issues as school closures, bans on public gatherings, border closures, bans imposed on the export of medical equipment, fines for quarantine violations, random temperature checks, curfews, public transport suspensions and utilization of mobile phone data for tracking covid- cases and their contacts. on average, % of people in the seven european countries approved of the policies taken in their country in response to the pandemic, implying considerable public support. nevertheless, the extent of approval differed by country and by policy measure. the most approved measures were fining -day quarantine violations, ban of public gatherings, and border closures (each supported by % of respondents). by the time of the survey's fieldwork, restrictions on public gatherings had been adopted in all countries covered by the study, whereas international travel controls had been imposed to a certain extent everywhere, except the uk [ ] . prior to complete border closures in mid-march , some countries (for example, italy, france, germany, denmark) had been requiring screening and -day quarantine for arrivals from high-risk regions already since february. in contrast, other countries, such as portugal and the netherlands, started later and turned directly to strict measures, such as banning arrivals from high-risk areas and imposing partial border closures. the latter typically implied either limitation on entries of nonresidents or closure of only certain types of borders (land, sea, air), while ensuring "green lanes" for freight vehicles transporting goods. however, complete border closures occurred haphazardly and led to disrupted commerce and stranding citizens. among countries covered in our study, denmark was the first to close all borders in mid-march, whereas the uk did so only in the second half of may . moreover, at the time of fieldwork, the uk did not have routine screenings at its airports or quarantine requirement for travelers [ , ] . thus, the results for the uk showed the extent of public support that these measures would have received, had they been implemented earlier. meanwhile, the most opposed containment policies were public transport suspension ( % of respondents against it), ban of medical export, use of mobile phone data for tracking, and the imposition of a curfew (each disapproved by approximately % of respondents). these trends might reflect within-country regional and age structure of the population. for example, older individuals and those living in remote areas tended to be the most strongly opposed to public transport suspension. in fact, among countries covered by the survey, public transport suspension was implemented only in italy, whereas its volume was reduced in all other states except for germany [ ] . the stay-at-home orders were most significantly opposed by the youngest respondents aged below . this measure was enforced in all countries covered by the survey except for denmark, where it was introduced as a recommendation [ ] . overall, a north-south gradient could often be noticed in the eu regarding policy support: people living in the southern states (portugal, italy, and france) tended to approve of the containment policies more than residents in the northern countries (denmark, germany and the netherlands). noteworthy, the largest share of supporters for every containment measure was noticed among the residents of italy and particularly in lombardy. here, on average, % of the population approved of the government's response to the pandemic. interestingly, the most significant share of the population who explicitly opposed each of the containment policies taken by their government was identified in denmark. here, for example, % of respondents disapproved of school closures and % disapproved of the imposition of a curfew. in comparison, the average disapproval of these measures in other countries was around % for schools and % for curfews. the most polarizing opinions were observed concerning the use of mobile data for tracking covid- cases and their contacts. the most significant share of people explicitly opposing such policy was identified in denmark ( %), the netherlands ( %), and germany ( %). it was particularly disfavored by the youngest age group ( % of respondents aged below against it). this policy received significant media attention as some countries and the european commission started the collaboration with telecom providers to access individual geolocation data for prediction and surveillance of covid- spread [ , ] . as of march , deutsche telekom provided german authorities with the anonymized data on the movement of its users. in italy, vodafone, windtre and telecom italia offered aggregated user data provision to the government for the same purpose. authorities in the lombardy region used mobile phone data to check compliance with the lockdown restrictions [ , , ] . other countries either initiated the development of their own mobile phone tracking apps or cooperated on the creation of common software, such as the pan-european privacy-preserving proximity tracing (pepp-pt) project led by germany. however, the launch of the pepp-pt was delayed at the end of april due to the data protection concerns voiced by experts and even some of the project participants [ ] . while proponents of the contact-tracing measures claim that using mobile data is of paramount importance in response to the covid- pandemic, many people worry about the government's use of technology due to possible privacy violations, thereby raising debates about the appropriateness of such social control measures [ , , ] . according to our data, people in some european countries expressed considerable reluctance about supporting such policy, which therefore makes future compliance questionable. moreover, such privacy disputes, as in the case of the pepp-pt project launch, might trigger higher reluctance among the potential users to use any contact-tracing app in the future, which could be detrimental for the implementation of a viable tracing technology [ ] . to better understand public opinion on certain policies, it is essential to look at the big picture and place obtained results into the national contexts. people's attitudes were likely based on their perceptions of the general state of affairs in their country, particularly in terms of the epidemic situation and restrictions they were subject to at that moment. in view of that, table summarizes the scale of the pandemic and the stringency of government's response in seven european countries at four points of time spaced around april (when the survey's fieldwork was % complete in every country). the public health situation in each state is described using total confirmed cases of covid- and total deaths attributed to covid- , both measured per million people and reported by the european centre for disease prevention and control [ ] . the stringency of government's response is measured with the covid- government response stringency index, a composite measure of containment policies ranging from to , where a higher value denotes a stricter response [ ] . at the time of the survey's fieldwork, the epidemic situation was worst, and the stringency index was highest in italy and france [ , ] . clearly, there was a north-south gradient in the stringency of government response: italy, france and portugal imposed more demanding policies than denmark, germany, the netherlands and the uk. nevertheless, although people in southern countries were exposed to more severe containment measures, they approved of them more than people residing in northern states, who experienced less stringent restrictions. turning now to within-country variations, we observed considerable heterogeneity of attitudes towards many policy responses within individual countries with particularly marked differences between regions and age groups in italy, france, and the netherlands. hereinafter, we grouped regions based on the severity of the covid- outbreak distinguishing between the most and the least affected areas. noteworthy, lombardy denotes the extra sample collected in italy and was analyzed separately from the representative italian sample. overall, we did not find significant differences in policy support between lombardy and the rest of italy. to illustrate within-country differences, fig. . reflects regional and age-related heterogeneity of public opinions in france and italy toward banning the export of medical equipment, such as masks. in fact, this measure was briefly undertaken by germany and france at the onset of the pandemic in early march , leading to political tensions between the eu member states. germany declared that the reason was to avoid shortages of masks, gloves and safety glasses within the country, whereas france argued that the ban was needed for the assessment of inventory and storage capacity [ ] . following the call for solidarity, both countries lifted the within-eu export ban on equipment in mid-march [ ] . while support for this policy tended to be similar in the most and the least severely affected parts of italy and france, the approval of the export ban conspicuously differed across age groups. older individuals approved more of this policy than younger people, which, besides other factors, may be related to the levels of worry people in these age categories have about the risks that covid- poses to their health. we found that % of french and % of italian respondents aged above perceived risks to their health from covid- as high or very high, while the corresponding share among people aged below equaled % in france and % in italy. to address the mental health implications of the covid- outbreak and subsequent containment measures, we assessed levels of worry prevailing in european societies over several domains (health, economic, emotional, work, and future). more specifically, we addressed concerns about losing a close person, becoming unemployed, health system getting overloaded, school closures, small companies running out of business, recession, restricted access to food supplies, blackouts, and society getting more egoistic. these items were adopted from the who covid- snapshot monitoring project, which will allow future comparisons with similar data collected for other countries and at different points in time [ ] . we found that the mean trend was similar in all countries: people worried most of all about the health system getting overloaded so that the capacities could become insufficient to cope with the surge in covid- cases. we observed that even in case of households that had not been directly hit by the novel coronavirus (above % of respondents in the total sample), the pandemic might have acted as a stressor causing health and economic anxieties. fig. presents people's worry about selected issues across seven eu countries (measured on a likert scale from -not worry at all to -worry a lot), where the higher intensity of color reflects a larger share of the population who worry "quite a bit" or "a lot". cross-country differences look substantial, and a north-south divide in the worry caused by the covid- outbreak is conspicuous. fig. . the proportion of respondents who worry "quite a bit" or "a lot" for instance, % of respondents in portugal and % in italy mentioned that they worried "quite a bit" or "a lot" about the national health system becoming overloaded, while the corresponding shares in denmark and germany were % and %, respectively. these health concerns might have reflected the development of the pandemic. as showed in table , the progress of the epidemic had a north-south pattern with more covid- cases and deaths per million of the population in southern states than in northern. the exception was the uk, where the epidemic was third deadliest after italy and france, but government response was less strict than in countries with a better epidemiological situation [ , ] . similarly, more people in portugal and italy were concerned with the economic consequences of the pandemic than in other european countries. for example, % of portuguese and % of italians were worried about losing their jobs, while respective shares in the netherlands and denmark were % and %, correspondingly. these cross-country differences in economic anxieties may be related to people's perceptions of the economic and financial countermeasures taken by their national government and the eu. during the pandemic, european countries implemented several fiscal and monetary measures to mitigate the economic impact of the covid- outbreak. these policies typically included support of wages under the reduced-hour scheme, postponement of tax payments for companies, direct financial supports and grants to small enterprises and self-employed, the extension of unemployment benefits, provision of capital buffers to banks, etc. [ ] . nevertheless, there were substantial variations in the timing and specific content of these countermeasures across the states. to briefly overview the scale of economic support provided by the government in each of the seven countries, table summarizes values of the economic support index, a composite measure reflecting income support and debt/contract relief provided by the national government to households [ ] . it is measured on a to scale, where a higher value refers to a more substantial economic assistance. at the time of the survey's fieldwork, all countries provided some type of economic relief to their residents. nevertheless, the extent of such support was conspicuously different: france and the uk ranked highest, while denmark, germany, and italy ranked lowest [ ] . hence, it may be possible that higher levels of economic concerns in some countries indicated people's beliefs in the insufficiency of the government's response, which will be subject to the analysis in the next waves of the survey. moreover, the composition of employment varies across the eu, especially in terms of informal and temporary employment. temporary contracts provide lower levels of social protection and job security to employees, but their prevalence has increased over the last years, particularly in the netherlands, italy, and france. as of , the share of temporary employees in the total number of employed was highest in southern european countries: portugal ( . %), france ( . %), and italy ( . %). in contrast, it was significantly lower in northern states: the uk ( . %), denmark ( . %), and germany ( . %). the only exception was the netherlands, where temporary workers constituted . % of all employees [ ] . thus, such differences in the employment composition may be in part responsible for the cross-country dissimilarities in economic concerns. we also observed differences in the levels of concern within individual countries. fig. shows the extent of worry about the health system and a recession in italy. we grouped regions based on the severity of the covid- outbreak and distinguished the levels of anxiety across age categories. higher intensity of the color reflects a greater extent of worry. overall, the level of worry in the highly affected regions of the country was not higher than elsewhere in italy, except for the youngest age group. however, economic concerns tended to be unequally distributed across the age groups. for instance, worries about the recession and small companies running out of business were higher among older individuals than younger age cohort. this pattern was similar in all countries covered by the survey. we asked people about the main sources of information from where they received news about covid- . the data show that overall % of respondents closely followed the news on the situation with covid- , implying a high level of public awareness. regarding the sources of information, % of respondents mentioned receiving updates from the tv and % additionally searched for information on the internet. presumably, reliable information presented through the television emerged as the best channel to reach the population at large. next, we assessed the extent of people's trust in the information received from various sources in the context of the covid- situation. the trust in the following information sources was addressed: national government, the eu, the who, hospitals and gps, national news channels and newspapers, social media, relatives and friends. fig. shows mean values of trust in information from six selected sources across seven european states (measured on a likert scale from -no trust at all to -trust very much). higher intensity of the color reflects a higher level of trust in the information from a specific source. the data show that overall people had the highest levels of trust in information from hospitals, family doctors, and the who, followed by information from the national government and main national news channels. this ranking of sources by trust was similar in all countries covered by the survey, except for france, where citizens had a high level of confidence only in healthcare providers and placed relatively little trust in all other sources. moreover, a north-south divide could be noticed in the level of trust in information from the national government. trust was highest in denmark and the netherlands (more than % of respondents trusted "much" or "very much"), whereas it was lowest in france ( % of respondents had a high level of trust). furthermore, a similar north-south gradient was observed concerning the trust in the eu: trust was highest in denmark ( %), germany ( %), the netherlands ( %) and the uk ( %), whereas it was lowest in italy ( %) and france ( %). portugal was an exception to this case since the corresponding value here constituted %. finally, we also observed considerable regional heterogeneities in levels of trust within countries with particularly noticeable differences across individual regions in italy, france, and germany. fig. shows people's trust in information from the national government in the context of covid- in germany and france as an example, where the higher intensity of the color indicates a greater extent of trust. while trust did not differ significantly between regions grouped with respect to the covid- severity, it was heterogenous across the age groups. although the survey asked about the level of trust in information from different sources in the context of the covid- situation and not about the overall trust in institutions, these two are likely to be related. generally, trust reflects people's perceptions of whether institutions are doing what is right. thus, trust in the information they provide can be considered an indicator of the confidence that citizens have in these institutions [ ] . the covid- pandemic raised new challenges for policymakers across the eu. the imminent threat to public health at the onset of the pandemic led most governments to impose a lockdown on society. however, as the peak of the pandemic abated, the focus of attention turned to the social and economic consequences of the containment measures. given that without acquired herd immunity the risk of a new wave of the epidemic remains high, and the production and distribution of vaccines may take to months [ ] , governments must try to strike the right balance between effects on public health, social life and the economy when considering possible exit-strategies from the current lockdown situation. in the absence of medical intervention, policymakers and public health officials must resort to non-medical behavioral interventions. lifting the lockdown requires that citizens support and adhere to the policy measures that aim to contain the spread of the virus as social and economic activity gradually restarts. given the difficulty of enforcing such regulations, future measures need to be both well-designed and well-communicated to the public. the more people are willing to comply voluntarily with the new measures, the less enforcement and supervision will be needed to achieve high compliance. for this, people's perceptions and attitudes need to be factored in at the policy-design and implementation stages. our survey sought to capture the public sentiment toward measures previously taken by policymakers to contain covid- and addressed people's support for policies, worries about the consequences of covid- , and trust in different sources of information. the first insights obtained from the data showed that containment and mitigating policies undertaken by national governments in response to the initial stages of the covid- pandemic were generally wellreceived by the population in all countries covered by the survey. nevertheless, the extent of approval varied across states and specific policy measures. several lessons can be drawn for the design and implementation of policies for the prolongation or gradual removal of lockdown restrictions. first, we observed a north-south divide in people's perceptions, worries and trust across the european countries. this finding suggests that further containment measures and lockdown exit strategies need to be balanced against the factors that worry people in each specific country. one noteworthy example is the level of importance that people in european countries attribute to the concepts of individual freedom and privacy. using mobile data for tracking covid- cases and their contacts may be a controversial decision to take even though it is believed by many experts to be a useful tool to manage the covid- outbreak. the effectiveness of this policy critically depends on a sufficient level of adoption of the technology by the population [ ] . our data suggest that this may not be achieved easily in some european countries. a clear takeaway is that an open dialogue with society on this matter is needed. explaining the need for and the advantages of such intrusive policies through trusted means of communication, while addressing people's concerns explicitly and being open about the risks of using such policy measures may help raise the support and compliance in society to a sufficient degree. another critical issue is the balance between saving lives and saving livelihoods. according to the survey, people in southern european countries are substantially more concerned about the economic aspects of the covid- outbreak than people in northern european countries. economic anxieties, if left unaddressed, may have adverse effects on the mental health and wellbeing of the population, as well as cause downward adjustments in consumption behavior, thereby exacerbating the economic situation in a country if the recession indeed happens. second, we found considerable heterogeneities in people's approval of policies within individual countries. this tendency was particularly noticeable in france and italy. one possible determinant of regional differences in public support could be the extent of the devolution of decision-making in the country. on the one hand, devolution could enable regional or local authorities to make better decisions due to their better awareness of region-specific circumstances. on the other hand, it could harm the coordination of policy responses between the central and regional authorities within individual countries. thus, it is crucial to understand the determinants of such differences and address them to secure public support of future policies and ensure high compliance with government measures. furthermore, our results showed that the burden of stress tended to be unequally distributed across and within countries. even in case of households that were not directly hit by covid- , the pandemic may have acted as a stressor causing health and economic anxieties. such worries may be detrimental to individual mental health and wellbeing, and they may become further exacerbated by the imposition of self-isolation policies. thus, it may be reasonable to consider an asymmetric approach to the design of exit strategies taking region-specific levels of support and worry into account. this includes the identification of vulnerable categories of the population not only in terms of health risks but also with respect to social and economic activities, and addressing their concerns satisfactorily. third, during a pandemic, public trust in the government and the information it provides is of paramount importance. to expect high compliance over extended periods of time, policymakers need to adopt effective strategies and means of communication whereby securing a sufficient level of trust and confidence from the society. as our results suggest, some countries were more successful in this respect than others. society needs to be well-informed about the dilemmas faced by policymakers, and for this, the communication between the government and the citizens must be clear and transparent. the data showed that % of respondents closely followed the news on the situation with covid- mainly using television to keep themselves updated. thus, television emerged as the best channel to reach the population at large, suggesting that presenting reliable information through this means is an effective strategy to follow. nevertheless, given that the data show regional and age-related heterogeneities in trust and policy support, it may be worth tailoring messages and means of communication to specific groups of the society. for example, cooperation with public figures and well-known experts can be used to deliver government and public health messages in a simple language, or local voices could be used to amplify such messages in individual regions of the country. overall, information provision, public education and effective communication strategies should be among the key guidelines for policymakers when implementing exit strategies and designing future containment measures so that these policies have public support and high compliance. additional waves of the survey are scheduled in june and august . this will allow us to investigate in more detail how the population copes with the health, social and economic consequences of the covid- pandemic as the situation evolves. declarations of interest: none estimating the number of infections and the impact of non-pharmaceutical interventions on covid- in european do low-trust societies do better in a pandemic? lockdown fatigue hits as europe enforces coronavirus restrictions answering the right questions for policymakers on covid- . the lancet global health the new york times covid- snapshot monitoring (cosmo): monitoring knowledge, risk perceptions, preventive behaviours, and public trust in the current coronavirus outbreak oxford covid- government response tracker. blavatnik school of government temporary reintroduction of border control tracking and tracing covid: protecting privacy and data while using apps and biometrics how will governments know when to lift restrictions? european mobile operators share data for coronavirus fight deutsche welle on the responsible use of digital data to tackle the covid- pandemic complete our world in data covid- dataset seeks solidarity as nations after intense discussions, i welcome that de and fr now allow for export of #covid protective equipment. i will continue to follow supply to it closely. no single eu country can win this battle alone. #cooperation #solidarity temporary employees as percentage of the total number of employees trust in government, policy effectiveness and the governance agenda the race against covid- this project has received funding from the european union's horizon research and innovation programme under the marie skłodowska-curie grant agreement no , the work was supported by funding under the excellence strategy by the german federal and state governments, as well as by the university of hamburg, erasmus university rotterdam, and nova school of business & economics lisbon -chair bpi | "fundação la caixa" on health economics. we thank our colleagues for their feedback and work on the adoption of the survey to national contexts: helen banks, joana pestana, maarten husen, laurie rachet jacquet, nicolai fink simonsen.