| Congresso Brasileiro de Microbiologia 2023 | Resumo: 1023-1 | ||||
Resumo:Bacterium from intestinal microbiota interacts with each other and with pathogens, sharing metabolites that could determine the establishment of disease allowing the pathogen to recognize its preferred niche and modulate its behavior. Bacteroides thetaiotaomicron is a symbiotic bacterium commonly found in the intestinal microbiota of humans. New-found evidence revealed its ability to positively modulate virulence phenotype and genotype of enteric pathogens such as Escherichia coli. Enteroaggregative E. coli (EAEC) is an emerging pathogen associated with acute diarrheic infections, persistent diarrhea, and chronic intestinal inflammation. EAEC is an extremely versatile pathogen, capable of infecting children, infants, travelers and susceptible adults, such AIDS patients. In a previous study we showed that EAEC virulence phenotypes were modified in the presence of B. thetaiotaomicron metabolites, in anaerobic conditions. Adhesion of EAEC 042 to HEp-2 cells and laminin, and biofilm production was significantly increased when bacteria is grown with metabolites of B. thetaiotaomicron. Motility of EAEC 042 was reduced 6-fold in a petri dish dispersion assay. Based on these initial results, we hypothesize that metabolites of B. thetaiotaomicron may alter EAEC 042 major virulence encoding gene expression. First, EAEC 042 strain was grown in the presence and absence of B. thetaiotaomicron supernatant in anaerobic and aerobic environment. Further, RNA of samples were extracted, followed by cDNA synthesis. Primer for major virulence encoding genes were designed based on EAEC 042 complete genome sequence at GeneBank (FN554766 and FN554767). Target selected were major virulence regulator aggR; adhesins aap, aafA and aafB; toxins shf, Pet, Pic and astA; biofilm associated shf and fis; motility associated fliC and lafA fliC; and chemotaxis cheY encoding genes. Gene expressions were accessed by RT-qPCR and calculated by 2-ΔΔCt method. Statistical significance was set at p<0,05. We found out that 12 of the 13 gene targets were upregulated in the presence of B. thetaiotaomicron metabolites when cultured in anaerobiosis. In anaerobic conditions, metabolites increased in 3,5-fold the expression of the major virulence regulator aggR, when compared to not treated strain. Its product regulates aafA, aafB, pet and shf, which also showed significant increases of 2,5-fold, 3,3-fold, 13-fold and 3,7-fold, respectively, when strain was cultured with metabolites. Genes that are not regulated by aggR were also upregulated: aap (3,5-fold), pic and astA (5-fold), fis (4-fold), fliC (23,5-fold). While the supernatant did not show any effect on expression of most genes in aerobiosis, the genes encoding for dispersin (aap), for the toxin EAST-1 (astA), for flagellin (fliC) and for the chemotaxis protein CheY (cheY) were significantly up-regulated. lafA fliC was the only target that was not modulated in any condition. None of the genes tested were downregulated in any condition. These results showed that expression of EAEC 042 genes involved in colonization, chemotaxis, regulation, and virulence such as adhesion, biofilm and toxins production, is positively modulated in response to metabolites released from B. thetaiotaomicron, in anaerobic conditions. Therefore, interactions among these bacteria may increase EAEC efficiency of colonization of the gastrointestinal mucosa, facilitate its persistence, and increase its virulence potential.
Palavras-chave: Bacteroides thetaiotaomicron, Enteroaggregative Escherichia coli, Gene expression, Virulence, Microbiota Agência de fomento:CNPq, CAPES, FAPERJ | |||||