Localized and systemic mycoses caused by pathogenic fungi (e.g. the yeasts Candida albicans and Cryptococcus neoformans) represent a serious health threat, especially to immunocompromised patients, among whom invasive candidiasis has high morbidity and mortality. Currently, identification of fungal pathogens requires isolation of pure cultures, followed by testing for substrate assimilation and fermentation patterns. Our long-term objective is development of a series of innovative diagnostic tests for specific human fungal pathogens in clinical samples using rapid nucleic acid hybridization methodology. Our immediate goal is to focus on DEVELOPMENT OF A DIAGNOSTIC TEST FOR CANDIDA ALBICANS BASED ON HYBRIDIZATION OF A SPECIFIC DNA PROBE WITH HIGHLY AMPLIFIED RIBOSOMAL RNA (rRNA) SEQUENCES IN DISRUPTED YEAST CELLS. In Phase I, Candida albicans rRNA gene fragments will be cloned using purified in vitro radiolabeled 18S rRNA as a probe. Subclones found to map in a putative unique region corresponding to nucleotides 642 to 702 of previously described eukaryotic 18S rRNAs will be DNA sequenced. DNA probes homologous to this and other regions will be isolated (or synthesized) and tested for specific hybridization to Candida albicans total RNA in pure and mixed cultures. In Phase II, test format procedures will be optimized for use with clinical samples.
