B. burgdorferi Hematogenous Dissemination

描述（由申请人提供）：莱姆病是美国最流行的节肢动物传播疾病，由螺旋体伯氏疏螺旋体感染引起。尽管近年来研究越来越深入，但在分子水平上对感染的发病机制知之甚少。我们已经确定了伯氏疏螺旋体临床分离株的遗传多样性，并表明螺旋体在不同基因型感染的患者和小鼠中的传播差异显著。这些结果表明，不同基因型的伯氏疏螺旋体在感染宿主中具有不同的传播潜力。我们假设这些差异是不同基因型之间基因含量和/或表达差异的结果。我们建议使用功能基因组学和蛋白质组学来深入了解具有不同血液传播能力的伯氏疏螺旋体菌株之间基因/蛋白表达的可能差异。该项目的长期目标是阐明介导伯氏疏螺旋体毒力的基因和/或蛋白质。提出以下具体目标：1)利用基因阵列技术监测不同基因型的伯氏疏螺旋体临床分离株在不同环境下的全球基因表达；2)利用蛋白质组学分析不同基因型的蛋白表达差异；3)通过实时实时RT-PCR检测早期莱姆病患者和感染小鼠中一些最有希望的候选基因的表达，评估候选毒力基因在莱姆病发病机制中的作用；4)选择候选毒力基因，然后靶向插入失活和互补。这种基因操作的效果应该直接证明这些基因在螺旋体发病机制中可能发挥的关键作用。

DESCRIPTION (provided by applicant): Lyme disease, the most prevalent arthropod-borne disease in the United States, is caused by infection with the spirochete, Borrelia burgdorferi. Despite intensive study in recent years little is known regarding the pathogenesis of infection at the molecular level. We have determined the genetic diversity among clinical isolates of B. burgdorferi and shown that spirochete dissemination varies significantly in patients and mice infected with distinct genotypes. These results demonstrate that different genotypes of B. burgdorferi possess varying potential for dissemination in an infected host. We hypothesize that these differences are the result of variations in gene content and/or expression among different genotypes. We propose to employ functional genomics and proteomics to gain insight into possible differences in gene/protein expression between B. burgdorferi strains with differing capacities for hematogenous dissemination. The long-term objective of this project is elucidation of genes and/or proteins that mediate B. burgdorferi virulence. The following specific aims are proposed: 1) global gene expression of B. burgdorferi clinical isolates of differing genotype in varying environments will be monitored by gene array; 2) differences in protein expression among the various genotypes will be assessed by proteomics; 3) the roles of candidate virulence genes in pathogenesis of Lyme disease will be evaluated by monitoring expression for a number of the most promising candidate genes will be monitored in early Lyme disease patients and infected mice by real-time RT-PCR; 4) selected candidate virulence genes will then be targeted for insertional inactivation and complementation. The effects of such genetic manipulation should provide direct demonstration of the critical role such genes may play in spirochete pathogenesis. \The combination of functional genomics, in vivo and genetic approaches proposed here should provide for the comprehensive assessment of the molecular basis of pathogenesis for B. burgdorferi isolates with varying potential for bloodstream dissemination and result in identification of determinants required for spirochete dissemination. This will provide new insights into the natural history of B. burgdorferi infection in humans, which, in turn, may have implications for prevention, treatment, and diagnosis of Lyme disease.