The object of the research is to devise a method of genetic analysis for Candida albicans. Previous work has shown that the cells are diploid, that many natural isolates contain recessive auxotrophic mutations, and that these mutations can be exposed by UV- and nitrous acid-induced mitotic crossing over. Spheroplast fusion can also be used to construct novel phenotypes and identify alleles. We will continue our mapping and complementation studies using mitotic crossing-over and spheroplast fusion. We will also use a C. albicans clone bank to devise a transformation system, based on the techniques used in Saccharomyces. The clone bank has been constructed in the chimeric E. coli-S. cerevisiae vector YEp13 and consists of 4463 colonies, each containing about 7.5 kb of Canadida DNA. We will use this bank to look for C. albicans sequences which complement S. cerevisiae nutritional deficiencies and for C. albicans sequences which complement C. albicans auxotrophies. We will also try to use known S. cerevisiae genes( ARG4, TRP1, TRP5, H1S3) to complement C. albicans deficiencies. We will isolate mutants in properties of C. albicans thought to be associated with virulence and test the mutants for their virulence in mice. These studies will further our knowledge of the genetic basis of virulence in this important human pathogen and may serve as a model system for parasexual analysis of other pathogens which are presently recalcitrant to genetic investigations.
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