The role of the human gut microbiota in modulating virulence of Enterohemorrhagic E. coli

人类的胃肠道是数以万亿计的微生物的家园，统称为肠道微生物区系，它们在维持人类健康方面发挥着关键作用，为宿主提供营养，促进免疫系统的成熟，并防止病原体的定居。然而，最近人们认识到，微生物区系的某些成员实际上可以加剧与肠道病原体相关的疾病。因此，迫切需要研究共生物种对肠道病原菌毒力的影响。肠出血性大肠杆菌(EHEC)是一种重要的人类食源性病原体，每年在美国导致265,000例出血性腹泻，严重时可发展为溶血性尿毒症综合征，甚至死亡。EHEC编码两个导致疾病的主要毒力因子：编码在被称为肠细胞消失点(Lee)的致病岛上的3型分泌系统(T3SS)和志贺毒素。以前的研究已经证明，微生物区系衍生的产物可以调节这些重要毒力因子的表达，并影响疾病的严重性。这项建议的目的是确定人类肠道共生对EHEC相关疾病的影响。我们的假设是，微生物群的成员通过调节毒力因子的表达和活性并直接处理毒力因子来影响EHEC疾病。我们之前已经证明，类杆菌产生的琥珀酸能增强Lee的表达，导致更严重的EHEC疾病。我们将调查其他常见肠道共生体对EHEC疾病的影响，并使用代谢组学来识别可能引起这些影响的代谢物。我们以前也证明过，在EHEC感染的小鼠模型中，共同的肠道共生杆菌thetaiotaomicron(B.theta)的存在会导致宿主粘液层的破坏增强。我们将研究这种增强粘液降解的机制，并确定共生粘液多糖降解在这一过程中的作用。最后，我们证明了B.theta产生的蛋白酶能够将EHEC T3SS的关键成分EspB切割成两个更小的产物。我们将鉴定负责这种切割的B.theta蛋白水解酶，并从功能上表征所产生的切割产物。这项拟议工作的成功完成将揭示人类肠道微生物区系调节EHEC毒力的新机制。更多地了解影响疾病进展的因素将使我们能够为这种重要的人类病原体设计更有效的治疗方法。

The human gastrointestinal tract is home to trillions of microbial organisms, collectively known as the gut microbiota, that play a critical function in maintaining human health by providing the host with nutrients, promoting maturation of the immune system and preventing colonization by pathogenic organisms. However, more recently it has been appreciated that certain members of the microbiota can actually exacerbate disease associated with enteric pathogens. Therefore, there is a critical need to investigate the effect of commensal species on the virulence of intestinal pathogens. Enterohemmorhagic E. coli (EHEC) is an important human food borne pathogen that causes 265,000 cases of bloody diarrhea year in the United States, which in severe cases can progress to hemolytic uremic syndrome or even death. EHEC encodes two principal virulence factors that contribute to disease: a type three secretion system (T3SS) encoded on a pathogenicity island known as the locus of enterocyte effacement (LEE), and Shiga Toxin. Previous studies have demonstrated that microbiota-derived products can modulate expression of these important virulence factors and affect the severity of disease. The objective of this proposal is to determine the effect of human gut commensals on EHEC associated disease. Our hypothesis is that members of the microbiota affect EHEC disease by modulating expression and activity of, and directly processing virulence factors. We have previously demonstrated that Bacteroides- produced succinate enhances LEE expression, leading to more severe EHEC disease. We will investigate the effect of other common gut commensals on EHEC disease and use metabolomics to identify the metabolites that may be eliciting these effects. We have also previously demonstrated that the presence of a common gut commensal, Bacteroides thetaiotaomicron (B. theta) leads to enhanced destruction of the host mucus layer during a mouse model of EHEC infection. We will investigate the mechanism of this enhanced mucus degradation and define the role of commensal mucosal glycan degradation in this process. Finally, we have demonstrated that B. theta-produced proteases are capable of cleaving EspB, a critical component of the EHEC T3SS, into two smaller products. We will identify the B. theta proteases responsible for this cleavage and functionally characterize the resulting cleavage products. Successful completion of the proposed work will reveal novel mechanisms of EHEC virulence modulation by the human gut microbiota. Understanding more about the factors that affect disease progression will allow us to design more effective therapies for this important human pathogen.