SE REGIONAL CENTER FOR EXCELLENCE FOR EMERGING INFECTIONS & BIODEFENSE

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 图拉氏方济各氏菌是一种高度传染性的A类细菌病原体，会导致图拉热症，这是一种潜在威胁人类生命的疾病。方济各氏菌致病的关键是其在巨噬细胞内复制和颠覆宿主免疫反应的能力。我们最近在小鼠身上采用了一种强大的全球体内负选择筛选，以确定新城疫杆菌致病所需的基因，这是方济氏菌的一个亚种，在小鼠身上引起疾病，而不是在人类身上。这一方法导致鉴定出164个毒力所需的基因，其中44个似乎编码新的毒力因子。我们发现164个基因中有65个是在体外巨噬细胞中复制所必需的。巨噬细胞复制所必需的表型最强的基因之一编码一种未知功能的新蛋白质。缺失该蛋白的缺失突变体证实该基因(我们称之为fpeA-Francisella噬菌体逃逸A)是新城疫霉菌在巨噬细胞中复制和体内毒力所必需的。电子显微镜显示突变株有缺陷，不能逃脱巨噬细胞吞噬小体，解释了其复制表型减弱的原因。这种无法快速逃脱吞噬小体和复制的能力与弗朗西塞氏菌致病岛(FPI)基因表达减少有关，而这些基因是这些过程所需的。我们目前正在确定FpeA控制FPI表达的分子机制。我们还在阐明在我们的屏幕上发现的其他几个重要毒力基因的功能。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Francisella tularensis is a highly infectious Category A bacterial pathogen that causes tularemia, a potentially life-threatening disease in humans. Critical to Francisella's pathogenesis are its ability to replicate within macrophages and to subvert the host immune response. We recently employed a powerful global in vivo negative selection screen in mice to identify genes required for the pathogenesis of F. novicida, a subspecies of Francisella that causes disease in mice but not humans. This approach resulted in the identification of 164 genes that are required for virulence, 44 of which appear to encode novel virulence factors. We found that 65 of the 164 genes are required for replication in macrophages in vitro. One of the genes with the strongest phenotypes that was essential for replication in macrophages encodes a novel protein of unknown function. A deletion mutant lacking the protein confirmed that this gene (which we refer to as fpeA - Francisella phagosome escape A) is required for F. novicida replication in macrophages as well as virulence in vivo. Electron microscopy revealed that the mutant strain is defective for escape from the macrophage phagosome, explaining its attenuated replication phenotype. This inability to rapidly escape the phagosome and replicate correlated with decreased expression of Francisella pathogenicity island (FPI) genes that are required for these processes. We are currently determining the molecular mechanism by which FpeA controls FPI expression. We are also elucidating the function of several other important virulence genes identified in our screens.