key: cord-0284805-4f17asfe authors: Cardenas, M.; Patinoa, L.; Pernia, B.; Erazo, R.; Munoz, C.; Valencia-Avellan, M.; Lozada, M.; Regato-Arrata, M.; Barrera, M.; Aquino, S.; Moyanoc, S.; Fuentes, S.; Duque, J.; Velazquez-Araque, L.; Carpio, B.; Mendez-Roman, C.; Calle, C.; Cardenas, G.; Guizado-Herrera, D.; Lucia Tello, C.; Bravo-Basantes, V.; Zambranod, J.; Francis, J. D.; Uyaguari, M. title: Detection of fecal coliforms and SARS-CoV-2 RNA in sewage and recreational waters in the Ecuadorian Coast: a call for improving water quality regulation date: 2022-01-05 journal: nan DOI: 10.1101/2022.01.04.22268771 sha: 0618a72f2d73f07caa97d7ee7c2642897bfe5020 doc_id: 284805 cord_uid: 4f17asfe Wastewater surveillance represents an alternative approach for the diagnosis and early detection of infectious agents of public health importance. This study aimed to evaluate SARS-CoV-2 and other quality markers in oxidation lagoons, estuarine areas and seawater at Guayas and Santa Elena in Ecuador. Sample collections were conducted twice at 42 coastal sites and 2 oxidation lagoons during dry and rainy seasons (2020-2021). Physico-chemical and microbiological parameters were evaluated to determine organic pollution. Quantitative reverse transcription PCR was conducted to detect SARS-CoV-2. Results showed high levels of Escherichia coli and low dissolved oxygen concentrations. SARS-CoV-2 was detected in sea-waters and estuaries with salinity levels between 34.2-36.4 PSU and 28.8 degrees celsius -31.3 degrees celsius. High amounts of fecal coliforms were detected and correlated with the SARS-CoV-2 shedding. We recommend to decentralized autonomous governments in developing countries such as Ecuador to implement corrective actions and establish medium-term mechanisms to minimize a potential contamination route. Monitoring of fecal contamination in recreational waters is not usually enforced in developing 120 countries. Despite this, recreational waters are widely used and leisure activities represent one of 121 the main economic incomes for the local population. In tropical countries such as Ecuador, 122 beaches are visited by tourists from all over the country and abroad throughout the year. The 123 beaches are crowded especially on holidays, four of them (Christmas, New Year´s Eve, Carnival 124 and Easter) occurring during the rainy season of the Ecuadorian Coast (December-April) making 125 these coastal areas susceptible to runoff events. Ecuador has a total of 421 wastewater treatment 126 plants; of these, 129 are located in the coastal region (INEC, 2016) , however, the effectiveness of 127 their treatments and capacity for the growing population is poorly known. 128 129 Ecuador has a total of 421 wastewater treatment plants, 129 The main economic activities in both cantons are fishing, and sun and beach tourism that take 134 place throughout the year. These places receive national tourists in the months of December to 135 April (coastal region holiday season); and the month from August to October (highland region 136 holiday season). Santa Elena has a sanitary system that covers 60% of the urban area. This 137 sanitary network collects wastewater to a pumping station and then towards oxidation lagoons, 138 where they also receive influents from Salinas, Santa Rosa, Muey and La Libertad. Wastewater 139 receives a secondary treatment with oxidation and subsequent disinfection with chlorine to be 140 discharged into the sea while Playas has a facultative lagoon (Córdova, 2013). 141 142 The use of recreational waters promotes one of the main economic activities in Ecuador, 143 therefore, the survey of its quality to guarantee users health should become a priority. The 144 purpose of this study was to assess water quality and the presence of SARS-CoV-2 at the most 145 visited beaches in Ecuador and at two oxidation lagoons at Guayas and Santa Elena during 146 restrictions (during the dry season in 2020) and opening of beaches (during the rainy season in 147 2021). 148 149 Materials and Methods 150 151 Study sites 152 153 The study area comprised marine waters located between latitude 1º 56' 5,81" S and 2º 43' 154 24,368" S. and longitude 80º 43' 24,6" W and 80º 18' 18,493" W along the Ecuadorian coastal 155 area. Fifteen beaches and two oxidation lagoons were selected along the Guayas and Santa Elena 156 provinces. They included artisanal fishing ports, estuaries, recreational beaches and bays, and a diving area located 10 km from the coast. Description of the sampling sites is shown in Table 1 . 158 All areas were sampled during the Ecuadorian confinement established for reducing the impact 159 of the COVID-19 pandemic during the dry season in July 2020 (A1-O2) and after the 160 confinement in the rainy season in January 2021 (A12-O22). 161 162 163 164 Water sampling and in situ analyses 165 166 A total of 84 surface seawater samples were collected at 15 beaches (A1-O22) and two oxidation 167 lagoons, in Playas (OL-A) and Punta Carnero (OL-B) ( Figure 1 , (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. recreational purposes through primary contact ( Table 6 ) and Limits of discharges to a body of 292 seawater (Table 10) . 293 294 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 January 5, 2022. ; Water physicochemical parameters 297 298 Maximum, minimum and mean values obtained from physical and chemical parameters at each 299 sampling site for dry season (2020) and rainy season (2021) are shown in Table S1 . The 300 temperature ranged from 23.6 °C to 34.4 °C with higher values in the rainy season (January) and 301 lowest in dry season (July), due to differences between the season period and time of survey 302 (early in the morning or past midday). A higher temperature was observed in Estero Valdivia and 303 Esterillo. The higher TSS were observed at Ayangue in the Estero Valdivia (698 mg/L) in an 304 area close to a pig farm, Fisherman area (114 mg/L) and Esterillo (86 mg/L) located near 305 restaurants in Playas, mainly in the rainy season (Table S1 ). Dissolved oxygen concentration 306 fluctuated between 2 to 9 mg/L, with higher concentration in the rainy season (July) and lowest 307 concentration associated with the estuarine area called Estero Valdivia and Area de Pescadores. 308 Concentrations lower than 5 mg/L (below the recommended concentration for the preservation of 309 the flora and fauna) were found at fishing facilities (Anconcito), estuaries (Esterillo and 310 Valdivia) and fishing ports (Area de Pescadores). COD ranged from 27mg/L to 820 mg/L. The pH ranged from 6.88 to 8.75, the lowest values were in the July of 2020 at Estero Data 320 (6.88) and in the rain season (7.72) at Esterillo. Salinity ranged from 0.5 to 60.2 PSU; an 321 alteration in salinity was observed in intertidal zones due the influence of wastewater from 322 Playas county, and in landing and commercialization of fishing in Area de Pescadores; estuarine 323 area (Esterillo). The highest salinity values were found in the external area from Estero Valdivia 324 where high temperatures increase evaporation rate, increasing salinity in the area near the mouth 325 of the sea during the rainy season. Tensoactives showed values between 0.004-0.085 mg/L, the 326 highest concentrations were observed in Punta Carnero 1 during February 2021 (Table S1 ). In 327 addition, ammonium ranged from 0.12 to 8 mg/L with highest levels at intertidal zones where 328 domestic wastewater is discharged (Area de Pescadores), and estuarine zones such as Esterillo 329 and Estero Valdivia (Table S1 ). The concentrations of residual free chlorine and surfactants were 330 less than the detection limit <10 mg/L and <0.05 mg/L, respectively. There were only significant 331 (p <0.05) differences between sampling sites for Temp, Sal and OF. In this context, OF ranged 332 from 0.07 to 2.6 mg/L, exhibiting the highest values in rainy season (January 2021 higher levels and number of sites were observed in the rainy season ( Figure 3 ). 347 348 Both FC and EC were consistently higher in seawater and wastewater discharged in estuarine 349 zones compared to shoreline sand. Values were high at a fish landing and management facility 350 located at Anconcito Artesanal Fishing Port, at a channel of domestic wastewater (Esterillo 1 and 351 2), in areas next to restaurants and public toilets, on the coastline near to urbanizations (Portete 352 Grande Casa del Sol), and an estuarine area influenced by urban settlement (Palmar 1) ( Figure 353 3 Relation between seawater water quality and environmental factors 370 371 The non-metric multidimensional scaling ordination showed that there was differentiation of the 372 water quality generated by seasonality between the dry and rainy seasons ( Figure 4 ). The 373 similarity of studied sites was influenced by the levels of TENS, Temp, pH, BOD, Sal, Cl, EC, 374 an DO. The SIMPER routine showed the variables with most contribution were TENS (10.54%); 375 Temp (9.94%); pH (8.87%), BOD (7.46%) and salinity (7.15%). 376 377 378 379 The main microbiological, physical and chemical parameters in OLs are shown in Table 2. The 380 concentration of FC in the oxidation lagoons fluctuated between 1.8 to 9´200,000 MPN/100ml. 381 The highest levels were showed in Santa Elena during 2020, with maximum levels registered in 382 the entrance of Punta Carnero (OL-B, 9´200,000 MPN/100 mL) while lower concentrations were 383 recorded in the Playas´s lagoons (120,000 MPN/100 mL). The levels observed in the output of 384 both oxidation lagoons were high, with values above the allowed limit (2,000 MPN/100 mL) for 385 discharge from wastewaters to seawater in Ecuador. The high levels of FC recorded at the output 386 of the oxidation ponds analyzed show low performance and low efficiency in the removal of 387 organic matter and microorganisms in these systems. 388 389 SARS-CoV-2 Analysis 390 391 The water and food. Our results emphasize the need for secondary or biological treatment such as 505 activated sludge or moving bed bioreactors, rather than primary/physical treatment as conducted 506 in these areas with oxidation ponds. Moreover, there is a need to develop an education program 507 for owners of hotels and restaurants at tourist beaches, regarding good practices in food 508 manufacturing and prevention of food-borne diseases (ETA). 509 510 Conclusions 511 In this work, water quality of seawater and wastewater discharges in two regions of Ecuador 512 were analyzed during two seasons during the COVID pandemic. It was found that 81% of 513 beaches were not suitable for recreation, as they exceeding the maximum limit allowed in water 514 for recreational activities, according to Ecuadorian environmental legislation. The virus SARS-515 CoV-2 was present in seawater and outfalls of estuaries, highlighting the need to reinforce 516 prevention actions that guarantee the health of the users of these ecosystems. Fecal 517 contamination could lead to the introduction of other human enteric viruses into aquatic 518 ecosystems. Therefore, it is recommended to implement a water quality monitoring program 519 including physical, chemical, and microbiological parameters (including the presence of SARS-520 CoV-2), especially in wastewater to avoid possible bacterial or viral outbreaks to residents and 521 tourists. 522 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 January 5, 2022. ; The authors thank authorities of Prefectura (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 January 5, 2022. ; (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (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 January 5, 2022. ; Tables 895 Table 1 (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. 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 January 5, 2022. ; beaches are yellow in the dry season and orange in the rainy season. 927 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. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (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 January 5, 2022. (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 January 5, 2022. (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 January 5, 2022. (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 January 5, 2022. ; Dahiya 662 COD and BOD removal from domestic wastewater generated in 663 decentralised sectors Durowoju 668 Impact of wastewater on surface water quality in developing countries: A 669 case of study in South Africa 18 Publisher: In Tech Editors: Hlanganani Tutu SARS-CoV-2 in river water: Implications in low sanitation countries Merck food microbiology applications Kitajima 685 First environmental surveillance for the presence of SARS-CoV-2 RNA in 686 wastewater and river water in Japan 691 Emerging contaminants in municipal wastewaters and their relevance for the 692 surface water contamination in the tropical coastal city Instituto Nacional de Estadística y Censos (INEC) 697 Población demográfica Estadística de información ambiental económica en gobiernos autónomos 703 descentralizados municipales 2015 (agua y alcantarillado) Octubre 710 SARS-CoV-2 in the environment: Modes of transmission, early detection and 711 potential role of pollutions The burden and etiology of diarrheal illness in developing countries Bather Shedding as a Source of Human Fecal Markers to a Recreational 722 Beach 727 Potential secondary transmission of SARS-CoV-2 via wastewater SARS-CoV-2 in wastewater: State of the knowledge and research needs Coronaviruses in the Sea 744 Prevalence of Antibiotic-Resistant Bacteria ESKAPE among Healthy People 745 Estimated by Monitoring of Municipal Wastewater