Mechanisms of Dysbiosis by Attaching and Effacing Pathogens

附着和消灭病原体造成生态失调的机制

• 批准号: 8717328
• 负责人: Christopher Lopez
• 金额: $ 3.63万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-02-01 至 2017-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8717328
• 关键词: Address; Adherence; Animal Feed; Area; Bacteria; Biological Assay; Cell Culture Techniques; Cell Respiration; Cells; Child; Chronic Disease; Citrobacter rodentium; Complex; Confocal Microscopy; Country; Crohn' s disease; Data; Developed Countries; Developing Countries; Development; Disease; Disease Outbreaks; Environment; Epithelial; Epithelial Cells; Escherichia coli; Food; Formate dehydrogenase; Formates; Foundations; Future; Gastrointestinal tract structure; Generations; Genes; Germ-Free; Goals; Growth; Health; Human; Immune response; Immune system; In Vitro; Indigenous; Individual; Infantile Diarrhea; Infection; Inflammation; Ingestion; Intestines; Invaded; Lead; Life; Location; Metabolic; Methods; Microbe; Microfilaments; Microscopy; Modeling; Molecular; Mus; Nutrient; Oral; Oxidases; Oxygen; Pathogenesis; Pathology; Physiology; Population; Predisposition; Production; Proteins; Public Health; Resources; Respiration; Role; Salmonella; Scientist; Serotyping; Structure; Surface; Systems Development; Testing; Therapeutic; Training; Virulence; Work; Writing; base; commensal microbes; cytokine; design; disease transmission; electron donor; enteric pathogen; enteropathogenic Escherichia coli; fitness; foodborne; foodborne illness; gastrointestinal; improved; in vivo; member; pathogen; pathogenic Escherichia coli; pathogenic bacteria; prevent; public health relevance; research study; respiratory enzyme; skills; stem; therapy development; tissue culture; transmission process

DESCRIPTION (provided by applicant): Enteropathogenic E. coli (EPEC) are common causes of gastrointestinal illness in very young children in developing nations. These and the related murine pathogen, Citrobacter rodentium, are classified as attaching and effacing (A/E) bacteria because upon ingestion, they intimately attach to and cause effacement of host intestinal cells. A/E bacteria induce shifts in the bacteria groups typically present naturally in the gastrointestinl tract, resulting in dysbiosis and disease. While the molecular basis of A/E bacteria-induced pathology is well studied, the mechanisms that enable these pathogens to successfully compete for resources and drive dysbiosis are not well understood. Additionally, the fitness benefit of intimate adherence to host cells through formation of pedestal-like structures, a feature unique to A/E pathogens, is currently unknown. The long-term objectives of this study are to define the mechanisms that result in pathogen-induced intestinal dysbiosis. Specifically, we aim to 1) illustrate the role of respiration to outgrowth of A/E pathogens during infection and 2) define the mechanisms that govern the fitness benefit provided by C. rodentium intimate attachment to host cells. Our hypothesis states that close adherence to epithelial cells provides A/E pathogens with access to oxygen for respiration, therefore allowing for more efficient energy generation over fermenting commensal microbes. To address our hypothesis, C. rodentium strains will be constructed with disruptions in key respiratory enzymes (bd oxidase and formate dehydrogenase) and attachment genes (pedestal-inducing effector protein EspH) and characterized in vitro with a combination of growth assays, tissue culture assays, and confocal microscopy. Mice will then be orally gavaged with single or mixed inocula for competitive infections to test the benefit derived from respiration and attachment in vivo. Germ-free mouse infections will then allow us to isolate interactions between pathogen and specific members of the microbiota. Quantitative PCR on host and pathogen genes will provide information on the levels of cytokine expression as part of the host response as well as a gauge to judge the metabolic state of C. rodentium under different experimental conditions. Completion of this study will provide valuable information on the mechanisms that result in pathogen-induced gut dysbiosis and a strong foundation on which to develop future strategies to prevent A/E pathogen transmission and disease.

性状（由申请方提供）：肠致病性大肠杆菌。大肠杆菌（EPEC）是发展中国家幼儿胃肠道疾病的常见原因。这些和相关的鼠病原体，啮齿柠檬酸杆菌，被归类为附着和消除（A/E）细菌，因为在摄入后，它们紧密附着并导致宿主肠细胞的消除。A/E细菌诱导通常天然存在于胃肠道中的细菌群的变化，导致生态失调和疾病。虽然A/E细菌诱导的病理学的分子基础得到了很好的研究，但使这些病原体能够成功竞争资源并驱动生态失调的机制还没有得到很好的理解。此外，目前还不清楚通过形成类尿道结构（A/E病原体特有的特征）与宿主细胞紧密粘附的健身益处。本研究的长期目标是确定导致病原体诱导的肠道生态失调的机制。具体来说，我们的目标是1）说明呼吸作用在感染期间A/E病原体的生长中的作用，2）定义呼吸作用的作用。 控制C.啮齿动物与宿主细胞的紧密附着。我们的假设指出，与上皮细胞的紧密粘附为A/E病原体提供了呼吸氧气的途径，因此可以比发酵的微生物更有效地产生能量。为了验证我们的假设，C.将构建具有关键呼吸酶（bd氧化酶和甲酸脱氢酶）和附着基因（呼吸诱导效应蛋白EspH）的破坏的啮齿动物菌株，并在体外用生长测定、组织培养测定和共聚焦显微镜的组合进行表征。然后用单一或混合接种物经口管饲小鼠以进行竞争性感染，以测试体内呼吸和附着的益处。无菌小鼠感染将使我们能够隔离病原体和微生物群特定成员之间的相互作用。对宿主和病原体基因的定量PCR将提供作为宿主反应的一部分的细胞因子表达水平的信息，以及判断C.在不同的实验条件下。这项研究的完成将提供有关导致病原体诱导的肠道生态失调的机制的有价值的信息，并为制定未来预防A/E病原体传播和疾病的战略奠定坚实的基础。