Polyphasic taxonomy of the basidiomycetous yeast genus Rhodosporidium: Rhodosporidium kratochvilovae and related anamorphic species.

Abstract

The phenotypic and genetic heterogeneity of the basidiomycetous yeast species Rhodosporidium kratochvilovae was investigated in a group of recent isolates and collection strains. A polyphasic taxonomic approach was followed which included micromorphological studies, nuclear staining, determination of sexual compatibility, physiological characterization, comparison of electrophoretic isoenzyme patterns, PCR fingerprinting, determination of mol % GMC, DNA-DNA reassociation experiments and 26S and ITS rDNA sequence analysis. The results allowed a more natural circumscription of the species, both from the genetic and phenotypic perspectives. The relationships with anamorphic species of the genus Rhodotorula were studied and isolates previously identified as Rhodotorula glutinis were found to belong to Rhodosporidium kratochvilovae. Other isolates included in the study were found to represent members of Rhodotorula glutinis var. dairenensis. Rhodosporidium kratochvilovae was found to include heterothallic strains, besides those already known to be self-sporulating. A total of 17 isolates, which were found to belong to this species, were heterothallic, selfsporulating and anamorphic strains. It is anticipated that integrated polyphasic studies of basidiomycetous yeasts will provide a more coherent classification system and the basis for accurate identification schemes, which in turn are essential for detailed ecological studies.

Cryptococcus terrestris sp. nov., a tremellaceous, anamorphic yeast phylogenetically related to Cryptococcus flavescens

“…Phylogenetic trees were constructed by using the neighbourjoining method and bootstrapping was done based on 1000 random samplings. MS-PCR fingerprinting followed the protocols described in Sampaio et al (2001), using the primer (GTG) 5 . Gel electrophoresis images were acquired with the GelDoc XR System software (Bio-Rad).…”

Reappraisal of the Sporobolomyces roseus species complex and description of Sporidiobolus metaroseus sp. nov.

“…In fact, we could observe the sexual stage in only four of more than 20 strains that were studied. Initially, we reasoned that, besides the self-fertile strains, heterothallic (selfsterile) strains would be found after mating experiments were carried out, as happened before for Rhodosporidium kratochvilovae and Sporidiobolus ruineniae (Sampaio et al, 2001;Valério et al, 2002). We performed crosses on cornmeal agar on several occasions but failed to observe the development of mycelium with teliospores.…”

“…For determination of the extent of DNA relatedness, total genomic DNA was extracted and purified using the procedures described by Sampaio et al (2001). For DNA-DNA reassociation experiments, a Gilford Response UV-VIS spectrophotometer and its thermal programming software were used and the methods of Kurtzman et al (1980) were followed.…”

“…For rDNA sequence analysis, total DNA was extracted using the protocol of Sampaio et al (2001) and amplified using primers ITS5 (59-GGAAGTAAAAGTCGTAACAAGG) and LR6 (59-CGCCAGTT-CTGCTTACC). Cycle sequencing of the 600-650 bp region at the 59 end of the 26S rDNA D1/D2 domains employed forward primer NL1 (59-GCATATCAATAAGCGGAGGAAAAG) and reverse primer NL4 (59-GGTCCGTGTTTCAAGACGG).…”

Cryptococcus bromeliarum sp. nov., an orange-coloured basidiomycetous yeast isolated from bromeliads in Brazil

“…PCR fingerprinting followed the protocols described in Sampaio et al (2001), using the primers M13 (59-GAGGGTGGCGGTTCT) and (GTG) 5 . Gel electrophoresis images were acquired with the GelDoc XR System software (Bio-Rad).…”