Typhoid Pathogenesis and Immunity

 描述(申请人提供)：人类病原体伤寒沙门氏菌(STY)每年导致2000多万人感染和20万人死亡。我们实验室开发了一种新的伤寒模型，使用从脐带血中获取的人造血干细胞移植到人源化小鼠。这是第一个概括的小动物模型。 急性链球菌感染的特征性病理和致死性。我们还获得了证据表明，STY通过促进另一种激活途径在人类巨噬细胞中持续存在，并表明人源化小鼠可以用减毒活疫苗株免疫，以产生保护性适应性免疫反应。这一应用建立在我们的初步观察基础上，以检验中心假设，即伤寒沙门氏菌通过促进替代巨噬细胞激活来逃避人类的先天免疫，并需要适应性免疫来清除。该应用的具体目的是：1.确定使STM在人巨噬细胞中持续存在的机制。我们发现STM在培养的人巨噬细胞中持续存在，而STM会导致细胞迅速死亡。这是理解伤寒和非伤寒沙门氏菌感染发病机制差异的重要线索。我们将对巨噬细胞中与巨噬细胞极化和凋亡相关的细胞表面标志物、信号通路和细胞因子/趋化因子进行检测，以了解沙门氏菌与人巨噬细胞相互作用中不同血清型的差异。2.分析人源化小鼠毒力所需的STY遗传位点。一个高度复杂的转座子突变文库已经确定了人源化小鼠中STY增殖所需的细菌基因。筛选出了已知和新的毒力决定因素，包括参与Vi衣壳多糖的产生和输出、转录调控、中枢代谢、O-抗原修饰和IV型菌毛生物合成的基因。深入的分析将确定抗沉默转录调节因子、新的渗透酶复合体、铁摄取系统和IV型菌毛在伤寒发病中的作用。3.人源化小鼠对STY疫苗的保护性适应性免疫反应特征。我们已经证明，人源化小鼠在接种减毒活疫苗后会产生保护性反应。详细的免疫学特征将分析人源化小鼠接种疫苗后的适应性免疫反应，并确定疫苗诱导保护的机制。相关性：这些研究将促进我们对伤寒发病机制的理解，并为伤寒候选疫苗的评估建立一个新的研究平台。

 DESCRIPTION (provided by applicant): The human pathogen Salmonella Typhi (STy) causes more than 20 million infections and 200,000 deaths each year. Our laboratory has developed a novel model of typhoid fever that uses humanized mice engrafted with human hematopoietic stem cells obtained from umbilical cord blood. This is the first small animal model to recapitulate the characteristic pathology and lethality of acute STy infection. We have also obtained evidence to suggest that STy persists in human macrophages by promoting an alternative pathway of activation and shown that humanized mice can be immunized with a live attenuated STy vaccine strain to engender protective adaptive immune responses. This application builds upon our preliminary observations to test the central hypothesis that Salmonella Typhi evades human innate immunity by promoting alternative macrophage activation and requires adaptive immunity for clearance. The specific aims of the application are to: 1. Determine the mechanisms that allow STy to persist in human macrophages. We have found that STy persists in cultured human macrophages whereas STm causes rapid cell death. This is an important clue to understanding differences in the pathogenesis of typhoidal and non-typhoidal Salmonella infections. Cell surface markers, signaling pathways, and cytokines/chemokines associated with macrophage polarization and apoptosis will be assayed in human macrophages to understand serovar-specific differences in Salmonella interaction with human macrophages. 2. Analyze STy genetic loci required for virulence in humanized mice. A high-complexity transposon mutant library has identified bacterial genes required for STy proliferation in humanized mice. The screen has identified both known and novel virulence determinants including genes involved in Vi capsular polysaccharide production and export, transcriptional regulation, central metabolism, O-antigen modification, and type IV pilus biosynthesis. In-depth analysis will determine the roles of a counter-silencing transcriptional regulator, a novel permease complex, iron uptake systems and type IV pili in typhoid pathogenesis. 3. Characterize protective adaptive immune responses to STy vaccines in humanized mice. We have shown that humanized mice develop protective responses following vaccination with live attenuated STy. Detailed immunological characterization will analyze adaptive immune responses to vaccination of humanized mice and determine the mechanism of vaccine-elicited protection. Relevance: These studies will advance our understanding of typhoid pathogenesis and establish a new research platform for the evaluation of candidate typhoid vaccines.