The Role of Plasmids in Rickettsia Biology

质粒在立克次体生物学中的作用

基本信息

批准号：
8586290
负责人：
Ulrike Gertrud Munderloh
金额：
$ 37.17万
依托单位：
UNIVERSITY OF MINNESOTA
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-01-15 至 2015-12-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): The genus Rickettsia contains important pathogens and consists of small gram-negative bacteria that replicate within eukaryotic host cells. Many Rickettsia species are associated with blood-feeding arthropods (ticks, lice, fleas and mites) that serve as vectors for their transmission to vertebrate hosts. Various Rickettsia species are the etiologic agents of typhus and spotted fevers in humans but others are apparently confined to arthropod hosts as endosymbionts. Members of the Rickettsia are the closest known microbial relatives of mitochondria with which they share the characteristics of highly reduced genomes and intimate metabolic interactions with their host cells. Historically, the slow and obligate intracellular growth of rickettsiae and their resistance to genetic manipulation presented substantial technical challenges to their study. Our laboratory has been at the forefront of both the recently developed capability to genetically manipulate rickettsiae and the discovery of rickettsial plasmids, which are extra-chromosomal autonomously replicating genetic elements. Plasmids in pathogenic bacteria are often associated with virulence functions involving particular metabolic capabilities or antibiotic resistance and are an important source of genetic diversity because they may be mobile, or genetically transmissible, via conjugation between bacteria of the same or different species. We hypothesize that plasmids are involved in rickettsial host adaptation/virulence and genetic plasticity. We propose a comprehensive analysis of the distribution, phylogeny and role of plasmids in the biology of rickettsiae associated with blood-feeding arthropods. We will use pulsed field gel electrophoresis and Southern blot techniques to define the distribution of plasmids within the genus. We will physically isolate and clone plasmids from Rickettsia species that range from non-pathogenic arthropod endosymbionts to human pathogens. The cloned plasmids will be sequenced with high throughput pyro-sequencing technology (454 Life Science Corp.) and subjected to phylogenetic analysis to determine the probable origin(s) and evolutionary history of plasmids within the genus, and obtain clues to their potential host adaptive functions. We will test our hypotheses that plasmids confer host adaptive functions and genetic diversity within rickettsiae through plasmid curing and phenotype analyses and plasmid conjugation experiments. The proposed research will delineate the distribution of plasmids and test their role in host adaptation/virulence in a group of medically important obligate intracellular bacteria that, until recently, were not believed to harbor plasmids.

描述（由申请人提供）：立克次体属含有重要的病原体，由在真核宿主细胞内复制的小型革兰氏阴性细菌组成。许多立克次体物种与吸血节肢动物（蜱、虱、跳蚤和螨）有关，这些节肢动物是立克次体向脊椎动物宿主传播的载体。多种立克次体是人类斑疹伤寒和斑疹热的病原体，但其他立克次体显然仅限于作为内共生体的节肢动物宿主。立克次体成员是已知与线粒体最接近的微生物亲戚，它们具有基因组高度减少以及与宿主细胞密切代谢相互作用的特征。从历史上看，立克次体缓慢且专性的细胞内生长及其对基因操纵的抵抗力给他们的研究带来了巨大的技术挑战。我们的实验室在最近开发的立克次体基因操纵能力和立克次体质粒（染色体外自主复制遗传元件）的发现方面一直处于领先地位。病原细菌中的质粒通常与涉及特定代谢能力或抗生素抗性的毒力功能相关，并且是遗传多样性的重要来源，因为它们可以通过相同或不同物种的细菌之间的接合而移动或遗传传播。我们假设质粒参与立克次体宿主适应/毒力和遗传可塑性。我们建议对质粒在与吸血节肢动物相关的立克次体生物学中的分布、系统发育和作用进行全面分析。我们将使用脉冲场凝胶电泳和 Southern 印迹技术来定义该属内质粒的分布。我们将从立克次体物种中物理分离和克隆质粒，范围从非致病性节肢动物内共生体到人类病原体。克隆的质粒将采用高通量焦磷酸测序技术（454 Life Science Corp.）进行测序，并进行系统发育分析，以确定属内质粒的可能起源和进化历史，并获得其潜在宿主适应性的线索功能。我们将通过质粒固化和表型分析以及质粒接合实验来检验我们的假设，即质粒赋予立克次体宿主适应性功能和遗传多样性。拟议的研究将描绘质粒的分布，并测试它们在一组医学上重要的专性细胞内细菌的宿主适应/毒力中的作用，直到最近，人们还不认为这些细菌含有质粒。