Recombination and Genetics in Chlamydia spp.

描述(由申请人提供)：本申请涉及广泛的挑战领域(15)转译科学，以及具体的挑战主题，15-AI-106：转译研究侧重于高优先级病原体，基础研究侧重于抗性机制。沙眼衣原体和肺炎衣原体的感染会导致人类生殖道、眼睛和呼吸道的疾病，这些感染的并发症会导致慢性问题，包括沙眼、盆腔炎、宫外孕，或许还有其他情况。全世界有数百万人受到这些疾病的影响，目前还没有有效的疫苗。由于缺乏实用的遗传系统，有效的疫苗和可能是针对感染的新疗法的发现变得复杂。这个项目的总体目标是通过对衣原体水平基因转移机制的研究，开发一个实用的遗传系统。最近在我们和其他实验室的实验表明，衣原体在体外和很可能在体内都能活跃地重组。我们已经确定，这些生物能够在不同的菌株和物种之间调动三种抗生素耐药性基因中的任何一种，并在没有选择的情况下稳定地保留这种DNA。这包括可测量的和频繁的四环素耐药性标记物从猪链球菌的体外转移到沙眼衣原体。这种转移的机制尚不清楚。以下目的是利用我们正在开发的技术和积累的初步数据来阐明衣原体自然遗传交换的机制，并利用这些发现来开发这些生物的实用遗传系统。在目标1中，我们将使用实验室中已有的不同抗药性的衣原体菌株来探索衣原体的重组机制。目标2将使用重组策略作为工具来定位与沙眼衣原体已知表型相关的MAP基因。最终目标将探索开发将基因导入沙眼衣原体的实用技术的可能途径。每个目标都将使用一套通用的研究工具，这些工具基本上已经在我们的研究小组中开发出来了。这些相互作用目标的完成将显著影响人们对衣原体如何繁殖的看法。在活体内相互作用，并可能提供一种实用的转化技术，我们可以向研究界提供。他们还将研究可能允许沙眼衣原体在患者群体中获得或转移耐药基因的机制。 公共卫生意义：数百万人患有由沙眼衣原体引起的严重眼部或生殖道疾病。由于缺乏实用的遗传系统，对可能的候选疫苗或合理的治疗靶点的研究受到限制。我们的实验室已经开发出重组技术，可以研究衣原体重组所涉及的机制。阐明衣原体中的重组机制将有助于采取合理的方法进行衣原体转化，这也在本提案中有所描述。随着研究人员探索对抗这些细菌的疫苗策略，开发一个可行的遗传系统将是重要的。

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (15) Translational Science, and specific Challenge Topic, 15-AI-106: Translational research focused on high priority pathogens and basic research focused on resistance mechanisms. Infections by Chlamydia trachomatis and C. pneumoniae cause diseases of the genital tract, eye, and respiratory tract in humans, and complications from these infections lead to chronic problems including trachoma, pelvic inflammatory disease, ectopic pregnancy, and perhaps other conditions. Millions of people worldwide are affected by these conditions, for which there are no effective vaccines. Discovery of useful vaccines and perhaps novel therapies against infection is complicated by the lack of a practical genetic system. The overall goals of this project work toward the development of a practical genetic system, through an examination of the mechanism of horizontal gene transfer by chlamydiae. Recent experimentation in our and other laboratories demonstrates that chlamydiae are actively recombinogenic in vitro and, very likely, in vivo. We have determined that these organisms are capable of mobilizing any of three antibiotic resistance genes between different strains and species, and stably retaining this DNA in the absence of selection. This includes measurable and frequent in vitro transfer of a tetracycline resistance marker to C. trachomatis from the pig pathogen C. suis. The mechanism of this transfer is not yet known. The following aims are proposed to utilize our developing technologies and accumulating preliminary data to elucidate the mechanism of natural genetic exchange in chlamydiae, and to exploit these findings to develop a practical genetic system for these organisms. In Aim 1 we will explore the mechanisms of recombination in Chlamydia spp., using differently resistant chlamydial strains already available in the laboratory. Aim 2 will use a recombination strategy as a tool for the mapping of map genes involved with known phenotypes in C. trachomatis. The final Aim will explore possible avenues for developing a practical technique for introducing genes into C. trachomatis. A common set of research tools will be used for each Aim, and these are largely already developed in our research group. Completion of these interactive Aims will significantly affect perceptions of how Chlamydia spp. interact in vivo, and may provide a practical transformation techniques that we can make available to the research community. They will also examine possible mechanisms that may allow C. trachomatis to acquire or transfer resistance genes in patient populations. PUBLIC HEALTH RELEVANCE: Millions of humans suffer serious diseases of the eye or genital tract caused by Chlamydia trachomatis. The study of possible vaccine candidates or logical therapeutic targets is limited by the lack of a practical genetic system. Our laboratories have developed recombination technologies that allow a study of the mechanisms involved in chlamydial recombination. Elucidating the mechanism of recombination in the chlamydiae will facilitate a rational approach to chlamydial transformation, which is also described in this proposal. Development of a workable genetic system will be important as researchers explore vaccine strategies to fight these bacteria.