Biology of Rickettsia

立克次体生物学

• 批准号: 8745434
• 负责人: David (Ted) Hackstadt
• 金额: $ 48.84万
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8745434
• 关键词: Actins; Animal Model; Awareness; Biological Models; Biology; Bite; Cells; Characteristics; Clinical; Collection; Complement; DNA; Diagnosis; Disease; Disease Outbreaks; Disease Progression; Epidemic; Exanthema; Extravasation; Fever; Genes; Genetic; Genome; Genomics; Goals; Hereditary Disease; Human; Infection; Knock-out; Laboratory Animals; Libraries; Life; Life Style; Liquid substance; Methods; Molecular; Montana; Mutagenesis; Nature; Organism; Outcome; Pathogenesis; Pathogenicity; Plasmids; Reporter; Rickettsia; Rickettsia Infections; Rickettsia rickettsii; Rocky Mountain Spotted Fever; Severities; Site; Spottings; System; Ticks; Typhus; Variant; Vascular Endothelial Cell; Vascular Permeabilities; Virulence; Virulence Factors; Virulent; base; biodefense; comparative genomics; design; human disease; improved; interest; member; mutant; pathogen; transmission process

Rickettsia rickettsii is the tick-borne etiologic agent of Rocky Mountain spotted fever. R. rickettsii is the prototypic spotted fever group rickettsia. Several other species, R. conorii, R. siberica, R. japonica, R. akari, and others cause diseases of lesser severity. Still other species in the spotted fever group, R. montana, R. peacockii, R. belli, and R. rhipicephali, are considered avirulent as they have never been associated with human disease nor do they cause overt disease in standard laboratory animals. The typhus group of rickettsia, typified by R. prowazeki, the agent of epidemic typhus, include some of the historically most devastating disease agents known to mankind. The typhus group also includes species of lesser or no virulence potential to humans. R. prowazeki and R. rickettsii are classified as Biodefense Catagory B and C agents, respectively. Rickettsia rickettsii is a member of the spotted fever group rickettsiae and the etiologic agent of Rocky Mountain spotted fever (RMSF). R. rickettsii is a small obligate intracellular Gram-negative organism maintained in its tick host through transovarial transmission. Infection with R. rickettsii occurs through the bite of an infected tick 33. Once the organism gains access to the host it is able to replicate within the host vascular endothelial cells and spread from cell to cell by polymerizing host cell actin. Damage to vascular endothelial cells by R. rickettsii leads to increased vascular permeability and leakage of fluid into the interstices causing the characteristic rash observed in RMSF. Infection with R. rickettsii results in a severe and potentially life threatening disease if not diagnosed and treated properly. While much is known about the progression of disease, the molecular mechanisms involved in the pathogenesis of RMSF are poorly understood. Strains of Rickettsia rickettsii vary dramatically in their virulence in animal model systems and severity of human disease. The obligate intracellular lifestyle of rickettsiae and the lack of tractable genetic systems make it difficult to identify genes involved in virulence. With the completed sequences of multiple rickettsial species, it has become possible to investigate differences between virulent and avirulent strains of rickettsiae through comparative genomics. In efforts to generate isogenic mutant pairs to definitively identify rickettsial virulence determinants, we used the mariner-based transposon mutagenesis system to generate a collection of R. rickettsii mutants. Direct sequencing of purified genomic DNA from each of the clones using primers from within the transposon delineated the precise insertion sites. Transposition sites were randomly distributed throughout the rickettsial genome. Random transposon mutagenesis is being applied to generate a library of mutants for analysis. In addition, plasmid transformation to introduce and complement genes is being developed. Fusions of various reporters to rickettsial genes of interest is ongoing. In addition, methods to directly knock out specific rickettsial genes are being explored.

立克次体是落基山斑疹热的蜱传病原体。立克次体是典型的斑点热族立克次体。其他几种物种，如康氏悬钩子（R. conorii）、西伯利亚悬钩子（R. siberica）、粳稻悬钩子（R. japonica）、赤角悬钩子（R. akari）等，会引起严重程度较轻的疾病。斑疹热组中的其他物种，如蒙大拿立克次体、孔雀立克次体、贝利立克次体和扇头立克次体，被认为是无毒的，因为它们从未与人类疾病相关，也不会在标准实验动物中引起明显的疾病。以流行性斑疹伤寒病原体普罗瓦泽基氏菌为代表的斑疹伤寒立克次体群包括一些人类已知的历史上最具破坏性的疾病病原体。斑疹伤寒组还包括对人类具有较小毒力或无毒力的物种。 R. prowazeki 和 R. rickettsii 分别被归类为生物防御类别 B 和 C 类病原体。 立克次体是斑疹热立克次体的成员，也是落基山斑疹热 (RMSF) 的病原体。 立克次体是一种小型专性细胞内革兰氏阴性生物，通过跨卵巢传播维持在其蜱宿主体内。 立氏立克次氏体感染是通过受感染蜱虫的叮咬而发生的 33。一旦该生物体进入宿主，它就能够在宿主血管内皮细胞内复制，并通过聚合宿主细胞肌动蛋白在细胞之间传播。 立克次体对血管内皮细胞的损伤导致血管通透性增加和液体渗漏到间隙中，从而导致 RMSF 中观察到的特征性皮疹。 如果诊断和治疗不当，立克次体感染会导致严重且可能危及生命的疾病。 虽然人们对疾病的进展了解很多，但对 RMSF 发病机制所涉及的分子机制知之甚少。 立克次体菌株在动物模型系统中的毒力和人类疾病的严重程度差异很大。 立克次体的专性细胞内生活方式和缺乏易处理的遗传系统使得鉴定与毒力相关的基因变得困难。 随着多个立克次体物种的完整序列，通过比较基因组学研究立克次体强毒株和无毒株之间的差异成为可能。 为了生成等基因突变体对以明确鉴定立克次体毒力决定子，我们使用基于水手的转座子诱变系统来生成立克次体突变体的集合。 使用转座子内的引物对每个克隆的纯化基因组 DNA 进行直接测序，描绘出精确的插入位点。 转座位点随机分布在整个立克次体基因组中。 随机转座子诱变被用来生成突变体文库以供分析。 此外，正在开发用于引入和补充基因的质粒转化。 各种报告基因与感兴趣的立克次体基因的融合正在进行中。 此外，正在探索直接敲除特定立克次体基因的方法。