Genetic determinants of mammalian host adaptation by the Lyme disease spirochete.

描述(申请人提供)：伯氏疏螺旋体是一种专性寄生虫，在自然界中通过一个复杂的循环维持，涉及到扁虱和哺乳动物宿主。这两种非常不同的寄主类型之间的转换需要通过基因表达的变化来快速适应的能力。人工感染研究已经提供了一些关于这些基因的信息，这些基因可能在类似于脊椎动物宿主的条件下差异表达。尽管取得了这些进展，但我们对一些基因在哺乳动物宿主适应中的作用和功能的理解仍然存在着根本的差距。我们的长期目标是鉴定和表征伯氏杆菌适应壁虱和哺乳动物宿主环境所需的蛋白质。这项应用的总体目标是利用小鼠模型识别特定适应哺乳动物宿主所需的伯氏杆菌候选基因。中心假说是大量未知功能的伯氏杆菌基因具有适应性表达。此外，我们假设一些对宿主适应重要的基因要么是瞬时表达的，要么是只在小鼠感染后期才被激活的。这项拟议研究的基本原理是，一旦确定了与宿主环境适应有关的基因，就有可能确定它们在确定哺乳动物感染方面的各自作用。因此，拟议的研究与NIH使命中与发展基础知识相关的部分相关，这可能有助于减轻人类疾病和残疾的负担。在已发表的伯氏杆菌基因组序列的指导下，并引用其他小组的工作，我们的假设将通过追求两个特定目标来验证：1)识别参与伯氏杆菌适应哺乳动物宿主环境的基因；2)识别对哺乳动物感染至关重要的伯氏杆菌晚期宿主适应因子。在第一个目标下，我们将利用In Vivo Expression Technology(IVET)。该系统将利用伯氏杆菌基于启动子的基因组DNA文库，该文库被设计成只允许选择那些在小鼠宿主中唯一活跃的启动子。第二个目标将利用IVET系统的改进版本，称为基于重组酶的IVET(RIVET)。该系统还将利用伯氏杆菌基于启动子的基因组DNA文库，但被设计为只允许选择那些在小鼠宿主中瞬时或在晚期感染期间活跃的启动子。这项拟议的工作具有创新性，因为它利用了特定伯氏杆菌蛋白的基本活性，作为识别重要毒力因子的新手段。当应用时，拟议研究的结果有望允许靶向该系统，以便显著降低该病原体在哺乳动物宿主中建立感染的能力。 公共卫生相关性：拟议的研究是莱姆病研究的一个重要领域，对了解伯氏疏螺旋体的传染性和发病机制具有适用性。这项拟议的研究与公共卫生相关，因为由此产生的发现有可能从根本上推进伯氏杆菌适应性基因表达领域，并可能对其他动物和人类病原体的宿主适应系统具有广泛的影响。因此，这些发现最终有望适用于人类的健康。

DESCRIPTION (provided by applicant): Borrelia burgdorferi is an obligate parasite, and is maintained in nature through a complex cycle involving both a tick and mammalian host. The transition between these two very different host types requires the ability to rapidly adapt through changes in gene expression. Artificial infection studies have provided information on a number of these genes that may be differentially expressed under conditions similar to that of the vertebrate host. Despite this advance, there remains a fundamental gap in our understanding of the role and function of a number of genes for mammalian host adaptation. Our long-term goal is to identify and characterize the proteins necessary for adaptation of B. burgdorferi to both the tick and mammalian host environments. The overall objective of this application is to identify B. burgdorferi candidate genes required for adapting specifically to the mammalian host using a mouse model. The central hypothesis is that a large number of B. burgdorferi genes of unknown function are adaptively expressed. Furthermore, we hypothesize that a number of genes important for host adaptation are either transiently expressed, or are turned on only during late-stage infection of mice. The rationale for the proposed research is that, once the genes involved in adapting to the host environment have been identified, it will be possible to determine their respective roles in establishing mammalian infection. Thus, the proposed research is relevant to that part of NIH's mission that pertains to developing fundamental knowledge that will potentially help to reduce the burdens of human illness and disability. Guided by the published B. burgdorferi genome sequence and cited work by other groups, our hypothesis will be tested by pursuing two specific aims: 1) Identify genes involved in the adaptation of B. burgdorferi to the mammalian host environment; and 2) Identify late-stage host adaptation factors of B. burgdorferi important for mammalian infection. Under the first aim, we will make use of In Vivo Expression Technology (IVET). This system will utilize a promoter-based genomic DNA library of B. burgdorferi that is designed to allow for the selection of only those promoters that are active exclusively within the mouse host. The second aim will utilize a modified version of the IVET system, termed Recombinase-based IVET (RIVET). This system will also make use of a promoter-based genomic DNA library of B. burgdorferi, but is designed to allow for the selection of only those promoters that are active transiently or during late-stage infection in the mouse host. The proposed work is innovative because it capitalizes on the essential activities of specific B. burgdorferi proteins as a new means of identifying important virulence factors. When applied, the results from the proposed studies are expected to allow the targeting of this system in order to significantly reduce the ability of this pathogen to establish infection in the mammalian host. PUBLIC HEALTH RELEVANCE: The proposed studies are of an important area of Lyme disease research that has applicability to understanding infectivity and pathogenesis by Borrelia burgdorferi. The proposed research has relevance to public health because the resulting discoveries have the potential to fundamentally advance the field of B. burgdorferi adaptive gene expression, and may have broad implications for host adaptation systems in other animal and human pathogens. Thus, the findings are ultimately expected to be applicable to the health of human beings.