An analysis of Borrelia burgdorferi motility

描述(申请人提供)：莱姆病是美国最常见的节肢动物传播的传染病，自发现以来发病率一直在稳步上升。伯氏疏螺旋体是病原体。关于给宿主带来疾病的伯氏杆菌毒力属性，人们知之甚少。目前的建议集中在伯氏杆菌的趋化性和运动性，以及它们在疾病过程中的作用。趋化性和移动性很可能是重要的毒力属性，因为这些微生物渗透到其他细菌入侵的组织和器官中。在分子水平上理解趋化性的基础上，在目标1中，我们将识别那些用作引诱剂的化合物。从伯氏杆菌基因组序列和已知新陈代谢中提取的信息将确定可能的趋化物质。这些化合物的趋化能力将使用已建立的方法进行测试。高传代(无毒)和两代(强毒)菌株都将在适当的环境条件下进行测试。在目标2中，我们建议构建特定运动性和趋化性基因的插入-缺失和框内缺失突变。高、低传代菌株均可作为亲本菌株。我们假设，这些突变体中的每一个都将具有不同的表型。这些表型将从运动性、趋化性、结构、蛋白质印迹、蛋白质组学和使用DNA微阵列技术的转录分析方面进行分析。为了确保给定的表型是由于正在研究的目标突变所致，我们将使用基因互补分析。在目标3中，我们将检验运动性和趋化性是伯氏杆菌重要毒力属性的假设。为了验证这一假设，我们将使用标准的细胞渗透试验和莱姆病的小鼠模型。我们预测，在这两种检测中，移动性和趋化性突变体将比亲本菌株侵袭性和毒力更小。所提出的实验将提供有关伯氏杆菌趋化性和运动性在莱姆病中的作用的信息，并可能为新的预防和治疗方法奠定基础。

DESCRIPTION (provided by applicant): Lyme disease is the most prevalent arthropod borne infection in the United States, and has shown a steady increase in incidence since its discovery. The spirochete Borrelia burgdorferi is the causative agent. Very little is known about the B. burgdorferi virulence attributes that bring disease to the host. The present proposal focuses on the chemotaxis and motility of B. burgdorferi, and their rote in the disease process. Chemotaxis and motility are likely to be important virulence attributes, as these organisms penetrate into tissues and organs where other bacteria fall to invade. Basic to an understanding of chemotaxis on a molecular level, in Aim 1 we will identify those compounds that serve as attractants. Putative chemoattractants will be identified from information drawn from the genome sequence and known metabolism of B. burgdorferi The chemotactic ability of these compounds will be tested using established methodology. Both high passage (avirulent) and tow passage (virulent) strains will be tested under appropriate environmental conditions. in Aim 2, we propose to construct both insertion-deletion and in-frame deletions mutations in specific motility and chemotaxis genes. Both high and low passage strains will serve as parental strains. We hypothesize that each of these mutants will have a distinct phenotype. These phenotypes will be analyzed with respect to motility, chemotaxis, structure, western blot, proteomics and transcriptional analysis using DNA-microarray technology. To insure that a given phenotype is due to the targeted mutation being studied, we will use gene-complementation analysis. in Aim 3, we will test the hypothesis that motility and chemotaxis are important virulence attributes of B. burgdorferi. To test this hypothesis, we will use standard cell penetration assays, and the mouse model of Lyme disease. We predict that the motility and chemotaxis mutants will be less invasive and virulent than the parental strains in both of these assays. The experiments proposed should yield information with respect to the roles of chemotaxis and motility of B. burgdorferi in Lyme disease, and perhaps lay the foundation for new methods for prevention and treatment.