Parasexual genetics of Torulopsis glabrata

Abstract

Prototrophic hybrids were generated in the asexual yeast Torulopsis glabrata by the fusion of spheroplasts derived from parent strains which bore complementing auxotrophic markers. The DNA content (per cell) of two hybrids was essentially that predicted by summing the corresponding parental values. UV irradiation of these two hybrids resulted fi the formation of sectored colonies with genetic properties consistent with their origin by either mitotic recombination or chromosomal nondisjunction.The asexual yeast Torulopsis glabrata is an opportunistic pathogen (1). In a study of fungemia in immunosuppressed patients, T. glabrata was isolated in a significant number of episodes of fungemia (26 of 136) (9). Recently, Hickey et al.(7) described a lethal infection in an immunosuppressed patient and provided a review of the literature.Previous basic studies with T. glabrata were concerned mainly with the structure of its mitochondrial genome and with the genetic basis of its killer activity against other yeasts (3,16,17). In a study of hybridization, Galeotti et al.(5) obtained T. glabrata hybrids by inducing the fusion of spheroplasts derived from parents (presumed haploid); however, those hybrids contained less DNA per cell than was predicted by summing the observed parental values of DNA per cell. Those researchers suggested, on the basis of their finding that the dose-response curves for X-ray killing of hybrids were indistinguishable from the corresponding doseresponse curves for parents, that T. glabrata lacked a recombinational pathway for repair of DNA damage.Development of a parasexual system based upon euploid T. glabrata hybrids was the object of the present study. We report here that euploid hybrids may be obtained by fusion of spheroplasts, and we provide evidence that these hybrids give rise to variants which express parental markers.MATERIALS AND METHODS Strains. T. glabrata G4 was described previously (21). The haploid Saccharomyces cerevisiae reference strain XP660-14D was provided by T. R. Manney. Candida albicans reference strain 208R1 was a derivative of clinical isolate MS24, which was described previously (18).Culture methods. The undefined complete medium YEPD agar and the defined minimal medium MIN agar were described previously (4). MIN agar was supplemented (with one or more of the compounds L-tryptophan, L-histidine, and L-methionine, to a final concentration of 40 mg/liter) as appropriate (see below). Cultures were grown at 300C, except when otherwise indicated.Induction of mutation. T. glabrata G4 was grown on YEPD agar (2 days). A sample of this confluent culture was suspended in water, spread on YEPD agar to yield approximately 600 CFU per petri dish, and exposed for 15 s to UV light delivered by a Sylvania G8T5 germicidal lamp. The dose rate at the agar surface, 32 ergs/mm2 per s, was estimated with a Blak-Ray UV meter (Ultra-Violet Products, San Gabriel, Calif.). Irradiated cultures and unirradiated * Corresponding author. controls were incubated for 2 days, and colonies were replica pl...

Genetic Structure ofCandida glabrataPopulations in AIDS and Non-AIDS Patients

“…Among the 33 loci used, no heterozygous profile could be found. Considering the polymorphism observed for these strains, this result supports the haploid chromosomal structure for this yeast, as suggested by other studies (43,44,46).…”

Electrophoretic karyotypes of Torulopsis glabrata

“…The results could be consistent-with T. glabrata having a haploid number of 14 if each band represented one chromosome. The conclusion that T. glabrata was haploid would agree with conclusions drawn from protoplast fusion hybrid analysis and irradiation studies of this species (5,22,23). However, if one assumes that intense bands represent comigration of homologs and narrower bands represent a single homolog of an individual chromosome, the data would be consistent with T. glabrata being diploid with a minimum haploid number of 7.…”

“…There was significant variation of the electrophoretic karyotype patterns among the population of these clinical isolates, although all isolates studied had identical phenotypic characteristics. Within single isolates recovered from 33 patients, 22 distinct electrophoretic karyotype patterns were discernible. These patterns were distinguished by both differences in the number of chromosomal bands observed and differences in the migration of the chromosomal bands assumed to be chromosome length polymorphism due to genetic translocations and deletions.…”

Genetic differences between type I and type II Candida stellatoidea

“…The auxotrophic strains obtained were CA329pro, B-4252met, and B-4365ade. Crosses were performed according to established methodology (18). Hybrids were obtained by selecting for growth on minimal medium (yeast nitrogen base without amino acids) with 2.0% glucose.…”