key: cord-0332668-zum9l60t authors: de Moura Manoel Bento, Flavia; Cecília Darolt, Josiane; Laís Merlin, Bruna; Penã, Leandro; Wulff, Nelson Arno; Cônsoli, Fernando Luis title: The molecular interplay of the establishment of an infection – gene expression of Diaphorina citri gut and Candidatus Liberibacter asiaticus date: 2021-04-06 journal: bioRxiv DOI: 10.1101/2021.04.06.438581 sha: e3ab52fa63eecc0ddbbd4fed945b9ef9dfc8d32c doc_id: 332668 cord_uid: zum9l60t Candidatus Liberibacter asiaticus (CLas) is one the causative agents of greening disease in citrus, an unccurable, devastating disease of citrus worldwide. CLas is vectored by Diaphorina citri, and the understanding of the molecular interplay between vector and pathogen will provide additional basis for the development and implementation of successful management strategies. We focused in the molecular interplay occurring in the gut of the vector, a major barrier for CLas invasion and colonization. We investigated the differential expression of vector and CLas genes by analyzing a de novo reference metatranscriptome of the gut of adult psyllids fed of CLas-infected and healthy citrus plants for 1-2, 3-4 and 5-6 days. CLas regulates the immune response of the vector affecting the production of reactive species of oxygen and nitrogen, and the production of antimicrobial peptides. Moreover, CLas overexpressed peroxiredoxin in a protective manner. The major transcript involved in immune expression was related to melanization, a CLIP-domain serine protease we believe participates in the wounding of epithelial cells damaged during infection, which is supported by the down-regulation of pangolin. We also detected that CLas modulates the gut peristalsis of psyllids through the down-regulation of titin, reducing the elimination of CLas with faeces. The up-regulation of the neuromodulator arylalkylamine N-acetyltransferase implies CLas also interferes with the double brain-gut communication circuitry of the vector. CLas colonizes the gut by expressing two Type IVb pilin flp genes and several chaperones that can also function as adhesins. We hypothesized biofil formation occurs by the expression of the cold shock protein of CLas. We also describe the interplay during cell invasion and modification, and propose mechanisms CLas uses to invade the host hemocel. We identified several specific targets for the development of strategies directed to interfere with the successful utilization of the psyllid vector by this pathogen. Author Summary Huanglongbing (HLB) or greening is an incurable disease causing severe damage to citrus production, making citrus industrial activity unsustainable in several countries around the world. HLB is caused by three species of Candidatus Liberibacter. Ca. L. asiaticus (CLas), vectored by the psyllid Diaphorina citri, is the prevalent species. Attempts to apply new technologies in the development of strategies for disease and pest management are been made. However, we still miss basic information on this system to efficiently apply the current technologies and envisage the implementation of new approaches for pest control, despite the relevant scientific contribution available. One major gap is regarded to the molecular interplay between CLas and its vector. We focused our attention in the molecular interplay occurring at the first relevant interaction of CLas and D. citri, represented by the gut barrier. We report the transcriptional activity of CLas during the invasion and establishment of the infection in the gut of the vector, as well as the transcriptional activity of the vector in response to the infection. We identified several host genes that are targeted and regulated by CLas as well as several CLas genes that are promising targets for the application of new management strategies. mechanisms that were activated in the gut epithelium of psyllids when exposed to Ca. 177 Since we could not detect relevant read countings against transcripts belonging 178 to Ca. L. asiaticus when using samples obtained from nymphs, only samples collected 179 at the adult stage were were subjected to differential expression analysis. Gene were expressed in A1CLas + , 766 in A2CLas + and 804 in A3CLas + (Fig 1) . One 187 transcript was expressed only in A1CLas + and A2CLas + , 22 in A1CLas + and A3CLas + , 188 and 63 in A2CLas + and A3CLas + . We also identified CLas genes that were exclusively adults (S1 Table) . One-hundred transcripts out of the over 700 transcripts detected in 206 the three sampling times differed in their abundance (S1 Table) . Differences in the level 207 of expression were detected for 80 transcripts when comparing A1CLas + and A3CLas + 208 and 20 transcripts in comparisons of A2CLas + and A3CLas + (S1 Table) . CLas 209 expression was always higher in A3CLas + when compared to the others. No differences 210 in gene expression between A1CLas + and A2CLas + were detected (S1 Table) . CLas-infected plants differed from controls specifically at each particular stage (Fig 2) . interesting, the majority of DE transcripts detected were specific to each one of the 249 periods of feeding psyllids remained exposed to CLas-infected plants. Gene expression of CLas in the gut of adult psyllids after different times of 251 feeding on CLas-infected citrus plants were quite different. CLas transcription in the gut 252 of adult psyllids was highly active soon after adult feeding started, but expression of a 253 large set of genes was significantly increased at later stages of feeding (A3CLas + ). 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