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The taxonomy of the genusSaccharomyces has undergone significant changes recently with the use of genotypic rather than phenotypic methods for the identification of strains to the species level. The sequence of rRNA genes has been utilized for the identification of a variety of fungi to the species level. This methodology, applied to species ofSaccharomyces , allows unknownSaccharomyces isolates to be assigned to the type strains. It was the aim of the present study to assess whether typing of the intergenic spacer region by using restriction fragment length polymorphisms of PCR products (intergenic transcribed spacer PCR [ITS-PCR] ribotyping) could distinguish among type strains of the 10 accepted species ofSaccharomyces and further to assess if this method could distinguish strains that were interspecific hybrids. Cellular DNA, isolated after the lysis of protoplasts, was amplified by PCR using ITS1 and ITS4 primers, purified by liquid chromatography, and digested with restriction endonucleases. Ribotyping patterns using the restriction enzymesMae I andHae III could distinguish all species ofSaccharomyces from each other, as well as fromCandida glabrata ,Candida albicans , andBlastomyces dermatitidis . The only exception to this was the inability to distinguish betweenSaccharomyces bayanus andS. pastorianus (S. carlsbergensis ). Furthermore, interspecific hybrids resulting from the mating of sibling species ofSaccharomyces were shown to share the ITS-PCR ribotyping patterns of both parental species. It should now be possible, by this simple PCR-based technique, to accurately identify these strains to the species level, thereby allowing an increase in our understanding of the characteristics required by these interspecific hybrids for their particular ecological niches.

Intergenic Transcribed Spacer PCR Ribotyping for Differentiation of Saccharomyces Species and Interspecific Hybrids