The long term objective of this proposal is a comprehensive analysis of virulence factors in pathogenic yeast required for colonization and persistence in the urinary tract. Candida glabrata and Candida albicans are the two major organisms responsible for funguria, as well as vaginal, oropharyngeal and systemic candidiasis. Little is known about the factors that allow Candida to colonize or persist in the urinary tract. Here, we will take a genetic approach to understanding Candida infections of the urinary tract. Unlike C. albicans, C. glabrata is a genetic approach to understanding Candida infections of the urinary tract. Unlike C. albicans, C. glabrata is haploid making it an excellent model for genetic analysis of virulence using already established genetic tools. We propose first to identify genes specifically involved in colonization and persistence in the urinary tract. We will use two different complementary global approaches. First, a variation of signature-tagged mutagenesis that we have established in C. glabrata will be used to screen a bank of . glabrata mutants for those that are unable to colonize and replicate in the bladder. Second, a library of genomic fragments fused to GFP will be constructed and screened for promotes specifically induced in the bladder or kidney but not under laboratory conditions These two global approaches will yield overlapping sets of genes relevant to the host interaction... Lastly, we will analyze in detail the role of any identified genes in urinary tract infections. Identified genes will first be prioritized for further study based on sequence homology. Mutants will be characterized with a set of in vitro assays for different aspects of the host interaction, including adherence to host tissue, interaction with phagocytes and growth under in vitro conditions mimicking the bladder environment. Different genes are likely to have roles in different aspects of infection which can be assayed only if the dynamics of the infection are known. We will therefore also use our signature-tagged strains to analyze the dynamics of UTI infection for wild type and mutant strains of C. glabrata, distinguishing between initial colonization and outgrowth. These approaches will provide insight into diverse aspects of the yeast- host interaction and an improved understanding of the processes contributing to fungal UTI, ultimately permitting enhanced therapeutic intervention.
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