Molecular Genetic Basis of the Infectious Cycle of Borrelia burgdorferi

莱姆病是美国和欧洲最常见的扁虱传播疾病。它是由伯氏疏螺旋体引起的，这是一种细菌性病原体，在自然界中存在于各种小型哺乳动物和硬蜱媒介之间的人畜共患病循环中。莱姆病螺旋体的一个特征是其不同寻常的节段性基因组，由一个线性染色体和大约20个线性和环状质粒组成。越来越多的数据表明，质粒编码的功能对于伯氏杆菌在其自然感染周期中成功适应不同的环境至关重要。我们已经开发了基因工具来研究伯氏杆菌不寻常的基因组组织、细胞结构和新陈代谢的基本方面。我们已经将这项研究扩展到活体环境中，使用了一个实验系统，该系统非常接近自然节肢动物媒介/啮齿动物宿主的感染循环。通过对生物体的基本分子生物学的了解，我们希望深入了解这种重要的媒介传播病原体所采用的感染策略，从而促进预防、诊断和治疗莱姆病的努力。 替代的西格玛因子rpos在莱姆病螺旋体的关键适应反应中起着核心作用，该反应是在蜱类摄食过程中诱导的，并为螺旋体感染脊椎动物宿主做好准备。我们实验室以前的工作证实，质粒编码的BBD18蛋白是rpos的负调控蛋白，但在野生型螺旋体中bbd18基因的失活从未实现。我们利用一个条件bbd18突变体，全基因组转录，代谢和蛋白质谱，以及第一次工程置换所有cp32原噬菌体质粒，来研究感染性伯氏杆菌中伴随着BBD18耗尽和未调节的rpos产生的致死表型。此外，我们进行了体内研究，以确定在小鼠-壁虱感染周期中螺旋体需要BBD18生存的确切阶段和位置。作为这些实验的结果，我们提供了第一个关于转导噬菌体和莱姆病螺旋体在扁虱摄食过程中经历的依赖rpos的适应性反应之间的联系的报告。我们推测，疏螺旋体基因组的多片段结构促进了噬菌体介导的遗传交换和重新分配，从而促进了螺旋体在自然地方病循环中维持所需的多样性水平。 注：首席调查员于2023年4月退休，该项目将终止/停用。

Lyme disease is the most common tick-borne illness in the United States and Europe. It is caused by Borrelia burgdorferi, a bacterial pathogen that is maintained in nature in a zoonotic cycle between various species of small mammals and an ixodid tick vector. A hallmark of the Lyme disease spirochete is its unusual, segmented genome, comprising a linear chromosome and approximately 20 linear and circular plasmids. An increasing body of data demonstrates that plasmid-encoded functions are critical for successful adaptation of B. burgdorferi to the different environments that the spirochete encounters during its natural infectious cycle. We have developed genetic tools to investigate basic aspects of the unusual genomic organization, cellular structure and metabolism of B. burgdorferi. We have extended this investigation to an in vivo setting with an experimental system that closely mimics the natural arthropod vector/rodent host infectious cycle. Through an understanding of the basic molecular biology of the organism, we hope to gain insight into the infectious strategy utilized by this significant vector-borne pathogen and thereby facilitate efforts to prevent, diagnose and treat Lyme disease. The alternative sigma factor RpoS plays a central role in a critical adaptive response of the Lyme disease spirochete that is induced during tick feeding and prepares the spirochete for infection of the vertebrate host. Previous work from our lab identified the plasmid-encoded BBD18 protein as a negative regulator of RpoS, but inactivation of the bbd18 gene in wild-type spirochetes was never achieved. We utilized a conditional bbd18 mutant, genome-wide transcriptomic, metabolomic, and protein profiling, and the first engineered displacement of all cp32 prophage plasmids, to investigate the lethal phenotype that accompanies BBD18 depletion and unmodulated production of RpoS in infectious B. burgdorferi. Additionally, we conducted in vivo studies to identify the precise stage and location during the mouse-tick infectious cycle at which spirochetes require BBD18 for survival. As a result of these experiments, we provide the first report of a link between transducing phage and the RpoS-dependent adaptive response that Lyme disease spirochete undergo during tick feeding. We infer that the multipartite structure of the Borrelia genome facilitates phage-mediated genetic exchange and reassortment, and thereby fosters the level of diversity that is required for spirochete maintenance in the natural enzootic cycle. NOTE: The Principal Investigator retired in April 2023 and this project will be terminated/inactivated.