NOVEL E. COLI 0157:H7 INTESTINAL COLONIZATION FACTORS

描述（改编自应用程序） 典型E.大肠杆菌O157：H7感染为定植 的肠道。迄今为止，只有一种细菌因子被证明是 通过O157：H7介导肠粘附，O157：H7是外膜蛋白， 最初是在首席研究员的实验室里发现的 然而，各种观察表明，额外的肠道 定殖因子可能在E.杆菌O157：H7和 产滋贺毒素E. O157：H7以外血清型的大肠杆菌 本项目的主要目的是研究E.杆菌O157：H7和 非O157：H7肠出血性E.大肠杆菌在肠内定植。我们的方法将 要聚焦四个具体目标。其中三个具体目标将侧重于 三个新发现的潜在的殖民因素的表征 肠出血性大肠杆菌这些因子中的两个与先前描述的粘附素具有同源性， 第三因子与其他肠道致病菌无明显同源性 先前描述的粘附素。对于每一个因素，我们将构建 结构基因中的同基因非极性突变，并测试突变体的 在不同的肠上皮细胞系中粘附性降低，新鲜 收获人肠上皮细胞，并在特定菌仔猪模型中 疾病。三个新殖民地中的任何一个 肠粘附因子将为疫苗提供新的靶点 发展第四个目标将集中在一个新的监管机制， EHEC定植因子，intimin。我们最近发现， 内膜粘附素和III型分泌系统的体外表达 LEE致病岛上编码的基因受自诱导因子的正调控 正常植物群E.大肠杆菌菌株，即，群体感应我们 我建议通过构建一个 O157：H7衍生物，不再响应群体感应和测试它的 在猪模型中的定殖。如果群体感应在体内是活跃的， 为基于抑制的潜在治疗干预开辟了一个新领域， 群体感应机制，从而减少肠 殖民因素

DESCRIPTION (adapted from the application) The essential first stage of typical E. coli O157:H7 infection is colonization of the intestinal tract. To date, only one bacterial factor has been shown to mediate intestinal adherence by O157:H7, the outer membrane protein intimin which was originally discovered in the Principal Investigator's laboratory. However, a variety of observations indicate that additional intestinal colonization factors may play a role in disease due to E. coli O157:H7 and Shiga toxin-producing E. coli of serotypes other than O157:H7 The overall goal of this project is to investigate the mechanisms by which E. coli O157:H7 and non-O157:H7 enterohemorrhagic E. coli colonize the intestine. Our approach will be focused in four specific aims. Three of the specific aims will focus on characterization of three newly discovered potential colonization factors of EHEC. Two of these factors have homology to previously described adhesins in other enteric pathogens and the third factor has no obvious homology to previously described adhesins. For each of these factors we will construct isogenic non-polar mutations in the structural gene and test the mutants for decreased adherence in differential intestinal epithelial cell lines, freshly harvested human intestinal epithelial cells, and in a gnotobiotic piglet model of disease. The demonstrated involvement of any of the three novel colonization factors in intestinal adherence would provide new targets for vaccine development. The fourth aim will focus on a novel regulatory mechanism of the best-characterized EHEC colonization factor, intimin. We recently showed that in vitro expression of the intimin adhesin and the type III secretion system encoded on the LEE pathogenicity island is positively regulated by autoinducer molecules expressed by normal flora E. coli strains, i.e., quorum sensing. We propose to test whether quorum sensing is active in vivo by constructing an O157:H7 derivative that no longer responds to quorum sensing and testing it for colonization in a pig model. If quorum sensing is active in vivo, this would open up a new area for potential therapeutic intervention based on inhibiting the quorum sensing mechanism, thereby decreasing expression of intestinal colonization factors.