Molecular Epidemiology, Virulence, and Genomic Characterization of Ureaplasmas

描述（申请人提供）：脲原体在许多健康人体内定植，但也可能引起侵袭性疾病。它们在某些情况下是共生的，而在其他情况下产生全身感染的原因尚不清楚。越来越多的证据表明，一些脲原体血清型可能比其他的具有更大的致病潜力。然而，这个概念的证据是不完整的。由于分型方法不精确、交叉反应、缺乏商用试剂以及多种血清型可能同时存在，先前的发病机制研究受到了阻碍。关于两种脲原体和单个血清型的不同致病性的相互矛盾的发现也表明，可能存在使用较旧的、歧视性较低的技术未检测到的毒力因素。我们推测，通过分析临床分离株和14个血清型脲原体的基因型，可以解释脲原体的不同致病性，以确定遗传差异和可能不同的毒力因子表达。本研究将检查侵袭性感染患者的脲原体，并将其与非侵袭性感染患者的脲原体进行比较。我们的具体目标是：(1)利用PCR技术确定临床分离的各种不同条件和共生生物中脲原体种类和血清型的发生率；(2)完善并进一步发展脉冲场凝胶电泳（PFGE）和限制性片段长度多态性，用于确定脲原体血清型之间以及致病与共生分离株血清型内的遗传相关性；(3)比较致病性和共生分离株的多条带抗原中串联氨基酸重复序列的大小和数量，以评估这一特征作为疾病预测因子的方法；(4)利用全局转座子诱变技术鉴定编码潜在毒力因子lgA1蛋白酶和磷脂酶A1、A2和C活性的基因，并对所有14种血清型和选定的临床分离株进行比较基因组分析，寻找这些基因的可能候选基因，然后克隆和表达这些基因，以确定它们的存在或表达是否与致病结果相关。识别入侵菌株的独特特征可以指导诊断工具的开发，并指导未来的研究，以提高对影响包括孕妇及其婴儿在内的弱势群体的疾病的理解，预防策略和管理。

DESCRIPTION (provided by applicant): Ureaplasma spp. colonize many healthy persons, yet they may also cause invasive diseases. Reasons they are commensals in some instances and produce systemic infections in others are unknown. There is increasing evidence that some Ureaplasma serovars may have a greater pathogenic potential than others. However, proof of this concept is incomplete. Prior attempts to study pathogenesis were hampered by imprecise typing methods, cross-reactions, lack of commercial reagents, and the fact that multiple serovars may be present simultaneously. Contradictory findings regarding differential pathogenicity of the 2 Ureaplasma species and individual serovars also suggests the possibility there may be virulence factors that were not detected using older, less discriminatory techniques. We hypothesize that differential pathogenicity of Ureaplasma spp. may be explained by analyzing clinical isolates and 14 serovars genotypically to identify genetic differences and possibly dissimilar expression of virulence factors. This research will examine Ureaplasma spp. from persons with invasive infections and compare them with others from persons without these conditions. Our Specific Aims are to: (1) Determine occurrence of Ureaplasma species and serovars in clinical isolates from a variety of different conditions and in commensal organisms using PCR; (2) Refine and further develop pulsed field gel electrophoresis (PFGE) and restriction fragment length polymorphism for use in determining genetic relatedness between Ureaplasma serovars and within serovars of pathogenic versus commensal isolates; (3) Compare the size and number of tandem amino acid repeats in multiple banded antigens of pathogenic versus commensal isolates a means to assess this characteristic as a predictor of disease; (4) Identify genes that encode potential virulence factors lgA1 protease and phospholipase A1, A2, and C activities using global transposon mutagenesis and comparative genomic analysis of all 14 serovars and selected clinical isolates to search for likely candidates for these genes, followed by cloning and expressing the genes to determine if their presence or expression correlates with pathogenic outcome. Identification of distinctive features of invasive strains may guide development of diagnostic tools and guide future studies to improve understanding of diseases affecting vulnerable populations including pregnant women and their infants, preventive strategies, and management.