Shiga Toxin Production and Role in Pathogenesis

描述（由申请人提供）：大肠杆菌O157:H7是一种重要的新兴病原体。由大肠杆菌O157:H7引起的疾病以腹泻、出血性结肠炎和可能致命的并发症溶血性尿毒症综合征（HUS）为特征。志贺毒素（Stx）是大肠杆菌O157:H7的主要毒力因子。志贺毒素的两种主要抗原变体Stx1和Stx2具有55%的氨基酸同源性，但Stx2的产生与包括溶血性尿毒综合征在内的严重疾病的进展有关。我们拟从两个方面探讨志贺毒素在大肠杆菌O157致病机制中的作用：H7 -肠内志贺毒素产生的调控以及Stx1和Stx2效价差异的分子基础。志贺毒素的基因被编码在细菌病毒上。毒素产生的调节与影响病毒生命周期的因素密切相关，特别是诱发病毒分解周期的因素，例如环丙沙星等抗生素。在特异性目标1中，我们将描述影响Stx2在体外和体内表达的因素。除了产生毒素的数量外，感染期间产生的志贺毒素的类型也会影响疾病的严重程度，其中Stx2是迄今为止更有效的毒素。虽然Stx2可以诱导细胞死亡是明确的，但尚不清楚溶血性尿毒综合征的发生是否需要细胞死亡，毒性可能发生在机体水平。在特异性目标2中，我们将描述Stx2/Stx1杂交突变体，以确定为什么Stx2比Stx1使用体外细胞培养模型更有效。在特异性目标3中，我们将使用小鼠疾病模型确定杂交突变体在体内的效力。了解影响志贺毒素产生和效力的病毒、细菌和宿主因素可能会导致减少致命疾病发病率的治疗方法。

DESCRIPTION (provided by applicant): E. coli O157:H7 is an emerging pathogen of major importance. Disease caused by E. coli O157:H7 is characterized by diarrhea, hemorrhagic colitis, and the potentially fatal complication, hemolytic uremic syndrome (HUS). Shiga toxin (Stx) is a major virulence factor of E. coli O157:H7. Two major antigenic variants of Shiga toxin, Stx1 and Stx2, share 55% amino acid homology, however Stx2 production has been associated with progression to severe disease, including HUS. We propose to examine two aspects of Shiga toxin in the pathogenesis of disease caused by E. coli O157:H7 - regulation of Shiga toxin production in the intestine and the molecular basis for the difference in potency between Stx1 and Stx2. The genes for Shiga toxin are encoded on bacterial viruses. Regulation of toxin production is strongly linked to factors that affect the virus life cycle, specifically factors that induce the viral lytic cycle, for example antibiotics such as ciprofloxicin. In Specific Aim 1 we will characterize factors that influence Stx2 expression in vitro and in vivo. In addition to amount of toxin production, the type of Shiga toxin produced during infection can influence the severity of disease, with Stx2 being by far the more potent toxin. While it is clear that Stx2 can induce cellular death, it is not clear that cellular death is required for development of HUS, and toxicity may occur at the organismal level. In Specific Aim 2 we will characterize hybrid Stx2/Stx1 mutants to determine why Stx2 is more potent than Stx1 using in vitro cell culture models. In Specific Aim 3 we will determine the potency of the hybrid mutants in vivo using a mouse model of disease. Understanding the viral, bacterial and host factors that influence Shiga toxin production and potency could lead to treatments that reduce the incidence of fatal disease.