Virulence Determinants of Borrelia burgdorferi

伯氏疏螺旋体的毒力决定因素

• 批准号: 8889600
• 负责人: STEVEN J. NORRIS
• 金额: $ 43.08万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-01 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8889600
• 关键词: Adherence; Affect; Arthritis; Arthropods; Bacteria; Biological Assay; Black-legged Tick; Blood capillaries; Borrelia; Borrelia burgdorferi; C3H/HeN Mouse; Cardiovascular system; Chemotaxis; Communities; Data; Dermal; Development; Diagnosis; Disease; Evaluation; Feeding Methods; Gene Expression Regulation; Genes; Genetic study; Genome; Goals; Grant; Health; Heart; Human; Immersion Investigative Technique; Infection; Inflammatory Response; Invaded; Ixodes; Joints; Libraries; Lyme Disease; Mammals; Membrane Proteins; Methodology; Methods; Mus; Mutagenesis; Mutate; Mutation; Nervous system structure; Neurologic; North America; Nutrient; Order Spirochaetales; Organism; Parents; Pathogenesis; Pathway interactions; Phase; Plasmids; Prevention; Process; Proteins; Protocols documentation; Replicon; Research; Research Personnel; Resource Sharing; Resources; Role; Sampling; Screening procedure; Shapes; Site; Site-Directed Mutagenesis; Skin; Specimen; Symptoms; Techniques; Testing; Tick Infestations; Ticks; Time; Tissues; Toxin; United States; Virulence; base; capillary; cell motility; design; feeding; gene complementation; improved; interest; mutant; novel; pathogen; prevent; screening; transmission process

DESCRIPTION (provided by applicant): Borrelia burgdorferi is the causative agent of Lyme disease in North America and is transmitted by ticks of the genus Ixodes. It is a highly invasive spirochete that can cause infection and manifestations in humans and other mammals that persist for months to years. The infection has localized, disseminated, and persistent phases, and B. burgdorferi appears to cause dermal, neurologic, cardiovascular, and arthritic symptoms primarily though the ability to invade almost any tissue, establish long-term infection, and induce inflammatory responses. The bacterium produces no known toxins, and its mechanisms of pathogenesis are largely unknown. Genetic studies using low-passage, infectious B. burgdorferi have been challenging due to exceedingly low transformation rates and plasmid loss; as a result, fewer than 50 genes have been investigated by allelic exchange or other site-directed mutagenesis methods for their importance in the mammal-tick infectious cycle. During the prior grant period, a sequence-defined library of 4,479 signature-tagged mutagenesis (STM) transposon mutants was generated in a transformable, infectious clone of B. burgdorferi B31. The plasmid content of the clones in this library was also determined using a novel Luminex-based high throughput strategy. Using the library, we have already performed STM screening of mouse infectivity of 484 transposon mutant clones in 434 different genes. In Aim 1, we will complete the systematic analysis of the roles of the 790 mutated protein-encoding genes in the infection of C3H/HeN mice using the STM mutant library. For this analysis, we will employ the STM Luminex Assay protocol developed during the prior grant period to analyze each mutant clone in 3 mice per group, 5 different tissues, and two time points in a high throughput manner. The goal of Aim 2 is to determine the effects of the transposon mutations on infectivity, persistence and transmission of B. burgdorferi in Ixodes scapularis ticks. Groups of the STM mutants will be transmitted to mice by larval ticks infected by immersion with a mixture of organisms with different mutations and signature tags. The survival of the STM clones will be analyzed before and after feeding on mice, and transmission of the clones to the mice will also be determined by the STM Luminex Assay. In Aim 3, the findings in Aims 1 and 2 will be used to select classes of mutants for detailed infectivity, complementation, and functional analysis. These analyses will include studies of genes involved in chemotaxis, motility, nutrient transport, surface proteins involved in adherence and other roles, novel gene regulation pathways, and additional gene sets identified during the STM screening procedure. The STM library will also be made available to any interested investigator through BEI-Resources, providing maximal resource sharing and stimulation of Borrelia research. We anticipate that this project will continue to fuel new discoveries useful in improving the diagnosis, treatment, and prevention of Lyme borreliosis.

描述（由申请人提供）：Borrelia burgdorferi是北美莱姆病的病因，并由IXODES属的tick虫传播。这是一种高度侵入性的螺旋体，可能会导致人类和其他哺乳动物的感染和表现，这些螺旋体持续了数月至数年。该感染具有局部，传播和持续的阶段，而B. burgdorferi似乎会引​​起皮肤，神经系统，心血管和关节炎症状，主要是尽管能够侵入任何组织，建立长期感染并引起长期感染并引起。 炎症反应。细菌没有产生已知的毒素，其发病机理的机理在很大程度上未知。由于转化率极低和质粒损失，使用低通道，传染性B. burgdorferi进行了遗传研究。结果，在等位基因交换或其他位置定向的诱变方法中，已经研究了少于50个基因，因为它们在哺乳动物的传染周期中的重要性。在上一批授予期间，在B. burgdorferi B31的可转化的，传染性的克隆中生成了一个4,479个标记诱变（STM）转座子突变体的序列定义的文库。该库中克隆的质粒含量也使用基于新型的基于Luminex的高吞吐量策略确定。使用该库，我们已经对434个不同基因中的484个转座子突变克隆的小鼠感染性进行了STM筛选。在AIM 1中，我们将使用STM突变库中790个突变蛋白编码基因在C3H/HEN小鼠感染中的作用进行系统分析。为了进行此分析，我们将采用先前赠款期间开发的STM Luminex分析方案来分析每组3只小鼠中的每个突变克隆，5个不同的组织和两个时间点以高通量方式分析。 AIM 2的目的是确定转座子突变对ixodes capapularis tick虫中的感染性，持续性和传播的影响。 STM突变体组将通过浸入浸入具有不同突变和签名标签的生物的混合物感染的幼虫壁虱传播给小鼠。在小鼠进食之前和之后，将分析STM克隆的存活，将克隆传播到小鼠之前，也将由STM Luminex测定法确定。在AIM 3中，目标1和2中的发现将用于选择突变体的类别，以详细的感染，补充和功能分析。这些分析将包括对参与趋化性，运动性，营养转运，参与依从性和其他角色的表面蛋白的基因的研究，新型基因调节途径以及在STM筛选过程中确定的其他基因集。 STM库还将通过Bei-Resources提供给任何感兴趣的研究人员，从而提供最大的资源共享和刺激Borrelia研究。我们预计该项目将继续推动新发现，可用于改善莱姆毛毛病的诊断，治疗和预防。