Identification and characterization of potential virulence factors expressed by Candida albicans during oropharyngeal candidiasis in HIV-infected patients.

摘要

Candida albicans is an important human fungal pathogen able to cause a wide variety of infections. We applied an antibody-based screening strategy, in vivo induced antigen technology (IVIAT), in order to identify potential C. albicans virulence factors expressed during oropharyngeal candidiasis (OPC) in HIV-infected patients. Pooled sera from 24 HIV-infected patients with OPC was depleted for antibodies against C. albicans proteins expressed in culture. This adsorbed sera was used to screen a genomic expression library consisting of 24 C. albicans clinical isolates. I identified five C. albicans antigenic proteins by screening the genomic expression library with the adsorbed sera. These proteins (RBF1, CDC24, IPF11959, ALG5, and IPF15632) were analyzed in this thesis. Genes encoding the antigenic proteins were further shown to be expressed within pseudomembranes from thrush samples. Both RBF1 and CDC24 are involved in the yeast-to-hyphae transition and have been previously identified as virulence factors. PTH1 encodes a hypothetical protein with homology to S. cerevisiae YGR046w, an essential gene. ALG5 encodes a putative dolichol-phosphate glucosyltransferase involved in N-glycosylation of cell wall proteins. IPF15632 has no homology with any protein in GenBank or the S. cerevisiae database.;I further characterized ALG5 and IPF15632 role in C. albicans pathogenesis. I found that ALG5 does not contribute to adherence to primary human buccal epithelial cells nor does it play a role in a murine model of DC, in spite of minor cell wall changes due to under glycosylation of cell wall proteins. However, mutants with disruption of IPF15632 produced small colonies on YPD solid media compared to the wild type and their virulence was attenuated in the murine model of DC. IPF15632 mutants were able to colonize mouse kidneys, spleens, and liver as well as the wild type 1 day post-infection. However, the mutant cells were cleared from the kidneys and spleens faster than the wild type 4 days post-infection, suggesting their elimination by the immune system. Furthermore, I created a triploid C. albicans strain that contains a duplication of all or part of chromosome 6. This strain has cell wall defects that interfere with virulence in a murine model of OPC but not of DC.