Host-pathogen interactions in experimental cholera

项目摘要 霍乱是由霍乱弧菌引起的一种严重的脱水腹泻疾病。这种革兰氏阴性杆菌有 非同寻常的定植小肠和引起爆炸性流行病的能力。在这里，我们将解决 霍乱弧菌与宿主相互作用中的基本问题，利用我们的许多方法和工具 在过去的几十年里创造了。引起霍乱大流行的O1型霍乱弧菌分为小川型 和稻叶血清型，它们的不同之处仅在于末端O-抗原甲基化的存在与否 分别是糖。霍乱流行期间小川和稻叶血清型的转换 已经被认识了一个多世纪，但血清型转换对病原体适合性的影响并不是 安全。我们发现Ogawa血清型比稻叶血清型在体内有更大的适合性，并且 这两种血清型在体内的生长依赖于不同的代谢过程。因此，有意想不到的直接或 O抗原甲基化对霍乱弧菌体内生长的间接表型和生理影响。 霍乱弧菌血清型在体内适应性差异的后果和机制 将在AIM 1中确定。霍乱毒素(CT)触发肠液分泌，这在很大程度上占了 胆汁性腹泻。我们发现，CT还导致数百种宿主蛋白的分泌，这些蛋白被鉴定为 腹泻液，并重新模拟霍乱弧菌对肠上皮细胞的转录反应。其中许多 蛋白质和转录本与先天免疫反应有关，我们发现其中一种蛋白质， 表面活性蛋白D(SP-D)，限制霍乱弧菌在肠道内的生长。在目标2中，我们将分析 SP-D介导的保护机制及额外霍乱弧菌诱导的作用 分泌宿主蛋白以阻止病原体的定植，以揭示保护宿主的固有轴心 抗霍乱弧菌感染。霍乱流行经常以极快的速度传播，并通过 增加霍乱弧菌的传染性，但霍乱弧菌和控制霍乱传播的宿主基因 很大程度上是未知的。在目标3中，我们将利用我们在病原体条形码和新的计算方法方面的经验 框架，以扩展在AIMS 1和2中开发的分析，检查血清型和CT在 调节宿主对感染性的启动。控制弧菌的其他病原体途径和寄主过程。 还将阐明霍乱的传染性，以加深对霍乱传播的了解。总体而言， 拟议中的研究将对控制霍乱弧菌的相互关联的过程产生新的理解 肠道定植和传染性以及限制定植和感染的宿主因素和机制 控制传输。这项工作将为霍乱弧菌两个血清型的生物学提供新的视角， 以及霍乱毒素在刺激宿主对病原体的天然防御反应中的作用。我们的 这一发现也将对设计新的霍乱疗法和 疫苗。

Project Summary Cholera is a severe dehydrating diarrheal disease caused by Vibrio cholerae. This Gram-negative rod has the unusual capacity to colonize the small intestine and to cause explosive epidemics. Here we will address fundamental questions in V. cholerae-host interactions, leveraging many of the approaches and tools we have created in the past decades. V. cholerae O1, the cause of pandemic cholera, is divided into Ogawa and Inaba serotypes, which differ only by the presence or absence of methylation of the terminal O-antigen sugar respectively. Switching of the Ogawa and Inaba serotypes during cholera epidemics has been recognized for over a century, but the consequences of serotype conversion on pathogen fitness are not clear. We discovered that the Ogawa serotype has greater in vivo fitness than the Inaba serotype and that the two serotypes rely on distinct metabolic process for growth in vivo. Thus, there are unexpected direct or indirect phenotypic and physiological consequences of O-antigen methylation on V. cholerae growth in vivo. The consequences and mechanisms that underlie the in vivo fitness differences of the V. cholerae serotypes will be determined in Aim 1. Cholera toxin (CT) triggers the intestinal fluid secretion that largely accounts for choleric diarrhea. We found that CT also leads to the secretion of hundreds of host proteins identified in diarrheal fluid and re-models the intestinal epithelial transcriptional response to V. cholerae. Many of these proteins and transcripts are linked to innate immune responses and we found that one of these proteins, surfactant protein D (SP-D), restricts V. cholerae growth in the intestine. In Aim 2, we will analyze the protective mechanisms mediated by SP-D and investigate the roles of additional V. cholerae-induced secreted host proteins in impeding the pathogen’s colonization, to uncover host innate axes that protect against V. cholerae infection. Cholera epidemics often spread extremely rapidly, and host passaging increases V. cholerae infectivity, but the V. cholerae and host genes that govern cholera transmission are largely unknown. In Aim 3, we will leverage our experience with pathogen barcoding and a new computational framework to extend analyses developed in Aims 1 and 2, examining the roles of serotype and CT in modulating host-priming of infectivity. Additional pathogen pathways and host processes that control V. cholerae infectivity will also be elucidated, to deepen understanding of cholera transmission. Collectively, the proposed research will yield new understanding of the interconnected processes that govern V. cholerae intestinal colonization and infectivity as well as the host factors and mechanisms that limit colonization and control transmission. This work will provide new perspectives on the biology of the two V. cholerae serotypes, and on the actions of cholera toxin in stimulating innate host defense responses against the pathogen. Our findings will also have important translational implications for design of new cholera therapeutics and vaccines.