MYCOBACTERIAL ENVIRONMENT WITHIN MACROPHAGES

鸟分枝杆菌和结核分枝杆菌是感染单核吞噬细胞并在组织巨噬细胞内存活的细胞内病原体。 肺结核每年造成数百万人死亡，M.禽流感是引起艾滋病患者全身性疾病的最常见病原体。 这两种细菌都在巨噬细胞的膜结合空泡内存活。这些生物体的细胞内定位表明它们很好地适应了在宿主中的生存，并且它们可能可以在宿主细胞内获得足够量的营养。 已知环境中元素（如铁）的存在或缺乏会影响细菌中的基因表达。 据推测，巨噬细胞空泡缺乏铁，作为宿主防御感染的机制。提高我们对分枝杆菌空泡内环境以及分枝杆菌如何对空泡中存在的条件作出反应的理解将为致病机制提供新的见解。我们建议：（1）测定M中铁、钾、磷、硫、钙、镁、锰、铜、锌等重要元素的存在、总浓度和有效浓度;巨噬细胞和树突状细胞中的鸟空泡与M.使用两种方法：（a）硬X射线显微镜，和（B）细菌涂片系统。(2a)确定分枝杆菌空泡内环境对分枝杆菌推定毒力决定因子表达的作用;（B）确定这些决定因子在毒力中的重要性。这是我们第一次能够获得分枝杆菌吞噬体中单一元素浓度的数据。我们相信，拟议的研究将产生重要的信息吞噬体环境和分枝杆菌对它的反应。

Mycobacterium avium and Mycobacterium tuberculosis are intracellular pathogens that infect mononuclear phagocytes and survive within tissue macrophages. Tuberculosis is responsible for millions of deaths annually, and M. avium is the most common pathogen causing systemic disease in AIDS patients. Both bacteria survive inside membrane-bound vacuoles in macrophages. The intracellular localization of these organisms suggest that they are well adapted for survival in the host and that they probably can obtain sufficient amounts of nutrients inside the host cells. The presence or absence of elements, such as iron, in the environment is known to influence gene expression in bacteria. It has been assumed that macrophage vacuoles are deficient in iron, as a host defense mechanism against infection. Improving our understanding about the environment within the mycobacterial vacuole and how mycobacteria respond to the conditions present in the vacuoles will provide new insights into pathogenic mechanisms. We propose to: (1) Determine the presence and total concentration as well as the available concentration of important elements such as iron, potassium, phosphorus, sulphur, calcium, magnesium, manganese, copper and zinc in M. avium vacuole in macrophages and dentritic cells compared to M. smegmatis vacuoles by using two methods: (a) hard X-ray microscopy, and (b) bacterial reporte systems. (2a) Establishing the role of the environment within mycobacterial vacuoles on the expression of mycobacterial putative virulence determinants; (b) determine the importance of these determinants in virulence. This is the first time that we will be able to obtain data on the concentrations of single elements in mycobacterial phagosomes. We believe that the proposed studies will generate important information about the phagosome environment and the mycobacterial response to it.