EARLY ESTABLISHMENT STAGES OF ANTHRAX INFECTION

这项建议调查了炭疽感染的最初几个小时；炭疽芽孢杆菌在体内的萌发、巨噬细胞的存活、营养杆菌的生长和逃逸。在寄主之外，内孢子保持代谢休眠，即使暴露在恶劣的环境条件下也保持毒力。内孢子是炭疽病的传染病，进入体内后被区域巨噬细胞吞噬。内生孢子“感觉”到新的环境，萌发并生长到营养状态。我们的初步数据定义了在这些步骤中的每一步都受阻的离散突变体。逃跑后，大量的菌血症、毒血症和死亡接踵而至。我们的数据还表明，炭疽菌内孢子在体内具有独特的感官和信号机制来触发萌发。在培养的巨噬细胞中迅速萌发。非致病性芽孢杆菌。内生孢子在巨噬细胞中没有增加萌发。炭疽杆菌转座子介导的诱变系统允许选择不能在巨噬细胞中萌发但完全能在细菌介质中萌发和长出的单个内孢子突变体。对几类独特的突变体进行了鉴定。其中一个这样的基因座名为GERP(萌发质粒)，位于毒力(毒素)质粒pXO1上。因此，GERP的突变消除了寄主特有的萌发，但不能消除一般的萌发反应。这项建议的目的是：A.定义和表征炭疽杆菌的萌发基因和宿主化学信号，以确定它们在宿主特异性萌发反应中的作用；B.确定营养细菌用来生存和逃脱巨噬细胞的明确的细胞内事件和细菌基因；以及C.了解炭疽杆菌宿主特异性萌发系统和早期细胞内事件在小鼠模型中致病机制方面的相关性。对炭疽病这些关键的“建立”阶段的了解可能为接触炭疽菌内孢子后的早期干预提供目标。了解这种快速而戏剧性的转变，从内孢子的绝对新陈代谢休眠到不断生长的毒力杆菌，可以利用炭疽作为一种有效的模型来研究细菌感染周期的早期阶段。

This proposal investigates the first hours of anthrax infections; in vivo germination of the Bacillus anthracis endospore, macrophage survival, growth and escape of the vegetative bacilli. Outside the host, endospores remain metabolically-dormant, preserving virulence even when exposed to harsh environmental conditions. Endospores are the anthrax contagion, entering the body where they are phagocytosed by regional macrophages. Endospores "sense" the new locale, germinate and outgrow to a vegetative state. Our preliminary data defined discrete mutants blocked at each of these steps. After escape, massive bactermia, toxemia and death ensues. Our data also indicate that anthrax endospores have unique in vivo sensory and signaling mechanisms for triggering germination. Germination occurs rapidly in cultured macrophages. Non-pathogenic Bacillus sp. endospores show no increased germination in macrophages. A B. anthracis transposon-mediated mutagenesis system allowed selection of individual endospore mutants incapable of germination in macrophages but fully capable of germination and outgrowth in bacterial media. Several unique classes of mutants were characterized. One such loci, named gerP (germination Plasmid), is located on the virulence (toxin) plasmid pXO1. Thus mutations in gerP eliminate host-specific germination but not general germination responses. The aims of this proposal are to: a. define and characterize the germination genes of B. anthracis and host chemical signals to determine their roles in the host- specific germination response; b. determine defined intracellular events and bacterial genes used by the vegetative bacilli allowing for survival and escape from the macrophage, and; c. understand the relevance of B. anthracis host-specific germination systems and early intracellular events in terms of pathogenesis in the murine model. Knowledge of these critical "establishment" stages of anthrax may provide targets for early intervention after exposure to anthrax endospores. Understanding this rapid and dramatic switch, from absolute metabolic dormancy of the endospore to growing virulent bacilli allows anthrax to be exploited as an effectual model for examining the earliest stages of bacterial infectious cycles.