Long Polar Fimbriae of Attaching and Effacing Escherichia coli

描述（由申请人提供）：附着和去除大肠杆菌（AEEC）毒力因子的表达是一个严格调控的过程，在某些情况下，这些因子的鉴定一直很困难，因为它们要么在体外被抑制，要么表达条件未知。虽然很明显，某些毒力因子的表达与人类疾病密切相关，但AEEC菌株中存在的与其致病过程相关的其他因素仍不清楚。缺乏对编码这些额外毒力因子的基因如何被控制的充分了解是很重要的，因为如果没有这些知识，我们就不可能了解AEEC菌株的总体致病特性。因此，我们的目标是确定AEEC菌株的长极性（LP）菌毛如何促进发病机制，并使用这些菌毛编码基因作为检测毒力菌株的标记。核心假设是，除了已经表征的定植因子（例如，免疫球蛋白介导的粘附）外，AEEC菌株还具有高度调控的LP菌毛，该菌毛在定植过程中起作用，尽管编码这些菌毛的基因广泛分布于致病性大肠杆菌菌株中，但某些LP菌毛类型仅存在于特定的AEEC菌株中。我们将通过三个具体目标来验证这一假设，即：1)确定Ler和H-NS是否作为控制AEEC菌株LP菌毛表达的选择性沉默/抗沉默防御系统；2)鉴定非典型EPEC中控制LP毛表达的调节蛋白，并在兔模型中测定LP毛在定植过程中的功能；3)对AEEC菌株中LP纤毛基因簇的分布特征进行表征，确定某些LP纤毛亚基类型是否可作为不同致病性AEEC菌株的可靠标记。为了实现我们的目标，我们将在体外和体内（幼兔定植模型）条件下充分表征Ler、H-NS和非典型肠致病性大肠杆菌编码调节因子的功能，并对致病性AEEC菌株特定亚群中lpf基因的流行率进行详细研究。我们的研究工作是创新的，因为它利用了我们关于AEEC菌株的新定植因子及其在治疗和诊断中的潜在应用的发现。研究控制LP菌毛表达的调控网络的结果具有重要意义，因为我们将能够发现根本的差异，以解释不同AEEC菌株的组织趋向性，并确定LP菌毛的沉默是否是AEEC菌株对水平获得性基因的防御系统的一个例子。此外，家兔模型的使用将使我们对AEEC菌株的发病机制和定植特性有新的认识。了解AEEC在胃肠道定植的机制不仅将进一步加深我们对这些生物发病机制的了解，而且还将为降低感染率和改进针对这些生物制剂的治疗方案提供机会，这些生物制剂因其潜在的食品安全威胁而被分类为B类病原体。公共卫生相关性：附着和清除大肠杆菌（AEEC）代表了几个国家中涉及人类腹泻的不同分离群。大多数AEEC菌株具有高度调控的长极性（LP）菌毛，这有助于一些AEEC菌株的肠道定植过程。它们的完整表征将为AEEC定植过程提供新的和更深入的见解，并更好地了解其控制表达的调控机制，这将为开发新的治疗方法和简化诊断测试奠定基础

DESCRIPTION (provided by applicant): The expression of Attaching and Effacing Escherichia coli (AEEC) virulence factors is a tightly regulated process, and, in some cases, the identification of these factors has been difficult because they are either repressed in vitro or the conditions of expression are unknown. While it is evident that expression of certain virulence factors is strictly associated with human disease, the additional factors present in AEEC strains that are linked to their pathogenic process remain unclear. Lack of a full understanding of how the genes encoding these additional virulence factors are controlled is important, because, without this knowledge, we are unlikely to understand the overall pathogenic properties of AEEC strains. Thus, our objective is to determine how the Long Polar (LP) fimbriae in AEEC strains contribute to pathogenesis and to use these fimbrial-encoding genes as markers to detect virulent strains. The central hypothesis is that, in addition to the already characterized colonization factors (e.g., intimin-mediated adhesion), AEEC strains possess a highly regulated LP fimbriae, that plays a role in the colonization process, and although the genes encoding these fimbriae are widely distributed in pathogenic E. coli strains, some LP fimbriae types are found exclusively in specific AEEC strains. We will test this hypothesis through three specific aims, which are to: 1) Define whether Ler and H-NS act as a selective silencing/anti-silencing defense system that controls LP fimbriae expression in AEEC strains; 2) Identify the regulatory protein(s) controlling LP fimbriae expression in atypical EPEC and determine in a rabbit model the function of LP fimbriae during colonization; and 3) Characterize the distribution of the LP fimbrial gene clusters among AEEC strains and determine whether certain LP fimbrial subunit types are reliable markers of different pathogenic AEEC strains. To accomplish our aims, we will fully characterize the functions of Ler, H-NS, and atypical enteropathogenic E. coli-encoded regulators under in vitro and in vivo (infant rabbit colonization model) conditions and perform a detailed study of prevalence of the lpf genes in specific subsets of pathogenic AEEC strains. Our research work is innovative because it capitalizes on our findings regarding novel colonization factors in AEEC strains and their potential application in therapeutics and diagnostics. The results from studies of the regulatory networks controlling LP fimbriae expression have significance, because we will be able to identify fundamental differences to explain the tissue tropism of different AEEC strains and to determine whether silencing of LP fimbriae is an example of a defense system that AEEC strains have against horizontally acquired genes. In addition, the use of the rabbit model will give us new insight into the pathogenesis and colonization properties of AEEC strains. An understanding of the mechanisms underlying AEEC colonization to the gastrointestinal tract will not only further our knowledge of the pathogenesis of these organisms but also provide opportunities for reducing infection rates and improving treatment options against these biological agents classified as category B pathogens due t their potential use as a food safety threat. PUBLIC HEALTH RELEVANCE: Attaching and effacing Escherichia coli (AEEC) represent a diverse group of isolates implicated in diarrhea in humans in several countries. Most AEEC strains possess highly regulated Long Polar (LP) fimbriae, which contribute to the intestinal colonization process of some of the AEEC isolates. Their full characterization will provide new and deeper insights into the process of AEEC colonization, and a better understanding of their regulatory mechanisms controlling expression, which will be a basis for developing novel therapeutics and simplified diagnostic tests