Molecular Characterization and Application of Random Amplified Polymorphic DNA Analysis of Mrakia and Sterigmatomyces Species

Abstract

The qualitative and quantitative monosaccharide spectra of purified yeast cell walls revealed that there are three phylogenetically distinct lineages of sterigma-forming basidiomycetous yeasts: (i) Kurtzmanomyces and Sterigmatomyces species, which contain high levels of mannose; (ii) Tilletiopsis species, which contain glucose, galactose, and small amounts of mannose; and (iii) Fellomyces, Kockovaella, Sterigmatosporidium, and Tsuchiyaea species, which appear to be closely related on the basis of their high levels of glucose and the presence of xylose. The yeast cell wall neutral sugars of Sporobolomyces antarcticus and Sterigmatomyces aphidis were similar to those of members of the genus Tilletiopsis. However, the possibility that these taxa are conspecific was eliminated by the results of a random amplified polymorphic DNA (RAPD) analysis. The conspecificity of Mrakia jiigida and Mrakia nivalis, the conspecificity of Mrakia gelida and Mrakia stokesii, and the conspecificity of Sterigmatomyces halophilus and Sterigmatomyces indicus were confirmed by RAPD analysis results. RAPD analysis was found to be a simple and highly sensitive method which can be used to differentiate species at the DNA level; it can replace nuclear DNA-nuclear DNA hybridization experiments for species identification, characterization, and delimitation.The genus Mrakia was proposed for Leucosporidium fngidum as a new genus of the Filobasidiaceae by Yamada and Komagata (51). The following four species that formerly had been placed in the genus Leucosporidium were transferred to the genus Mrakia: Mrakia Jizgida, Mrakia gelida, Mrakia nivalis, and Mrakia stokesii (7, 8, 51). A homothallic, apomictic life cycle, self-sporulating simple holobasidia bearing terminal, sessile basidiospores, fermentation of sugars, the presence of ubiquinone Q-8, production of extracellular amyloid compounds, and yeast cell walls containing xylose are the most distinctive features of Mrakia species (14,26,32,51). The fact that the genus Mrakia should be separated from phragmobasidial, smut-like Leucosporidium species (31) has been corroborated by the results of determinations of parts of the 18s and 26s rRNA sequences (6, 9,48) and ultrastructural septa1 pore data (37).The Mrakia species originally defined on the basis of their ability to ferment glucose and their ability to utilize nitrate or various carbon compounds (cellobiose, maltose, maltose, melibiose, raffinose) for growth (7, 8) turned out to be questionable taxa after investigations of the electrophoretic enzyme patterns (39, 52) and parts of the 18s and 26s rRNA sequences (6, 9, 48). The electrophoretic enzyme patterns (52) suggested that M. nivalis and M. stokesii are synonyms of M. fngida and M. gelida, respectively.The genus Sterigmatomyces was introduced by Fell (5) for nonfilamentous, yeast-like fungi that are characterized by a unique method of cell division. The yeast cells produce one or more sterigmata, each of which gives rise to a single conidium.

Enrichment of a continuous culture of Saccharomyces cerevisiae with the yeast Issatchenkia orientalis in the production of ethanol at increasing temperatures

“…DNA isolation for RAPD-PCR DNA was extracted using hexadecyltrimethylammonium bromide or CTAB and puriWed following procedures as described by Messner et al [25]. The puriWed DNA preparation was diluted with TE buVer to a concentration of approximately 5 ng l ¡1 and stored at ¡20°C.…”

“…Conditions and procedures for the separation of the RAPD-PCR fragments were carried out as described by Messner et al [25]. The primers were selected based on the procedure used by Prillinger et al [31] for the identiWcation of yeasts in cheese.…”

Heterogeneity of the genus Myrothecium as revealed by cell wall polysaccharides

“…Recently, molecular techniques have been used to classify fungi and to study their evolution. Monosaccharides from the cell wall are useful markers in yeast and fungal taxonomy (Weijman and Golubev 1987;Prillinger et al 1990Prillinger et al , 1991Prillinger et al , 1993Messner et al 1994). This topic has been recently reviewed by Pfyffer et al (1998).…”

Hanseniaspora meyeri sp. nov., Hanseniaspora clermontiae sp. nov., Hanseniaspora lachancei sp. nov. and Hanseniaspora opuntiae sp. nov., novel apiculate yeast species

“…The 14 strains segregated into five distinct groups, which clustered with the other Hanseniaspora and Kloeckera species at a similarity level below 40 %. On the basis of previous correlation between similarity values derived from RAPD-PCR analysis and DNA-DNA reassociations (Messner et al, 1994;Molnár et al, 1995), we predicted that the five groups were genetically divergent from currently accepted species of Hanseniaspora.…”