Contribution of Novel Frimbriae to Proteus Mirabilis Virulence

描述（由申请人提供）：尿路感染是美国最常见的细菌感染之一。奇异变形杆菌（肠杆菌科成员）是复杂尿路患者感染的主要原因。虽然已经在奇异变形杆菌中鉴定出了几种毒力因子，但最近完成的该基因组测序有可能大大增加我们对奇异变形杆菌发病机制的理解。菌毛是从细菌细胞表面延伸的结构，通常参与定植和疾病进展。值得注意的是，我们在奇异变形杆菌基因组中注释了17个潜在的伴侣-引导菌毛，其中只有5个以前被鉴定过。本研究的第一个具体目的是研究这些菌毛的一般表达特征，并选择一种新的菌毛进行进一步的详细研究。RT-PCR，在小鼠模型中实验性尿路感染期间表达的基因的微阵列分析，以及表面表达蛋白质的质谱分析将用于磨练与疾病过程相关的功能性菌毛。将采用敲除和过表达研究来阐明菌毛的功能，使用的测定包括粘附细胞培养物或细胞外基质组分和小鼠泌尿道的实验感染。奇异变形杆菌的17个潜在菌毛操纵子中有10个含有编码转录调节因子的基因。大多数这些监管机构，确定其同源性的mrp菌毛操纵子中的mrpJ，抑制鞭毛运动。初步结果表明，mrpJ旁系同源物可能也调节其他功能，包括其他菌毛的表达。该提案的第二个具体目标关注与具体目标1中概述的菌毛相关的mrpJ parasites。将mrpJ paradox的过表达和敲除菌株与已知菌毛的微阵列研究和分析相结合。从这些研究中获得的数据不仅将增加我们对奇异变形杆菌发病机制的了解，并可能导致对UTI的新措施，而且还将与粘膜表面的其他病原体相关。

DESCRIPTION (provided by applicant): Urinary tract infections are one of the most common bacterial infections in the U.S. Proteus mirabilis, a member of the Enterobacteriaceae, is a major cause of infection in individuals with complicated urinary tracts. Although several virulence factors have been identified in P. mirabilis, the recent completion of the sequencing of this genome has the potential to greatly increase our understanding of P. mirabilis pathogenesis. Fimbriae, structures that extend from the bacterial cell surface, are often involved in colonization and disease progression. Remarkably, 17 potential chaperone-usher fimbriae were annotated by us in the P. mirabilis genome, only five of which had been previously identified. The first Specific Aim of this proposal is to study general expression characteristics of these fimbriae, and to choose one of the novel fimbriae to study in further detail. RT-PCR, microarray analysis of genes expressed during experimental urinary tract infection in a mouse model, and mass spectrometry of surface-expressed proteins will be used to hone in on a functional fimbria relevant to the disease process. Knockout and overexpression studies will be employed to elucidate the function of the fimbria using assays including adherence to cell culture or extracellular matrix components and experimental infections of the murine urinary tract. Ten of the 17 potential fimbrial operons in P. mirabilis contain a gene encoding a transcriptional regulator. The majority of these regulators, identified by their homology to mrpJ in the mrp fimbrial operon, inhibit flagellar motility. Preliminary results suggest that mrpJ paralogs likely also regulate other functions, including expression of other fimbriae. The second Specific Aim of this proposal focuses on the mrpJ paralog associated with the fimbria outlined in Specific Aim 1. Overexpression and knockout strains of the mrpJ paralog will be coupled with microarray studies and analysis of known fimbriae. Data gained from these studies will not only increase our knowledge about P. mirabilis pathogenesis and potentially lead to new measures against UTIs, but also will be relevant to other pathogens of mucosal surfaces.