Rapid identification of medically important yeasts by electrophoretic protein patterns

Abstract

A simple electrophoretic method for yeast identification was evaluated. Whole cells were extracted by SDS and the protein profiles obtained in SDS-PAGE after Coomassie blue staining were compared for 52 strains from 9 species of yeast or yeast-like fungi commonly isolated from man (Candida albicans, C. glabrata, C. guilliermondii, C. krusei, C. parapsilosis, C. pseudotropicalis, C. tropicalis, Geotrichum candidum, Saccharomyces cerevisiae). The corresponding patterns showed 30 to 45 polypeptides in the range 95-20 kDa and were clearly different for the 9 species. No differences could be detected between strains from the same species. The characteristic patterns were obtained within 24 h allowing rapid identification of the most commonly encountered clinical yeast isolates.

Analysis of parity between protein-based electrophoretic methods for the characterization of oral Candida species

“…The reproducibility of electrophoretic profiles on different slab SDS-PAGE gels was evaluated by the inclusion of molecular mass markers, besides protein extract of a organism from a non-correlated genus (Costas et al 1989, Bruneau & Guinet 1989 and gave similarity correlation values S SM = 0.853 for three repetitions of S. cerevisiae and S SM = 1.000 for three repetitions of molecular mass markers. These values are in agreement with the minimum acceptable proposed by Sneath and Johnson (1972) that was 0.800.…”

“…After cell disruption, the micro-centrifuge tubes were centrifuged at 10,000 g for 2 min, and the supernatant's protein concentration were determined according to Bradford (1976) and adjusted to 80 µg/ml (Ames 1974). The MLEE supernatants were applied on Whatman 3 filter paper wicks of 5x12 mm (Selander et al 1986), and for SDS-PAGE technique equal volumes of supernatant and loading buffer of Bruneau and Guinet (1989) (5mM Tris, 2.5% 2-mercaptoethanol, 1.5% SDS, 0.025% bromophenol blue) were combined and heated in a boiling water bath for 10 min.…”

Evaluation of biochemical and serological methods to identify and clustering yeast cells of oral Candida species by CHROMagar test, SDS-PAGE and ELISA

“…The preparation of the proteins for each yeast strain was made according to the Bruneau & Guinet (1989) method. Equal volumes of supernatant and loading buffer (5 mM Tris-HCl, 2.5% 2-β-mercaptoetanol 1.5% SDS, 0.025% bromophenol blue, pH 6.8, were combined and heated for 5 minutes at 95-100 o C, and SDS-PAGE was performed according to Laemmli (1970), using 12% (w/v) polyacrylamide gels.…”

SDS-Page and numerical analysis of Candida albicans from human oral cavity and other anatomical sites

“…Dendrograms, represented by non-rooted trees, based on SSM values were generated by the unweighted pairgroup arithmetic average (UPGMA) clustering method (17,51,52). The type-strain of C. albicans CBS562 and molecular weight markers (Bovine Serum Albumin 66,000Da, Bovine Pancreas Trypsinogen 24,000Da, Bovine Milk b-Lactoglobulin 18,400Da -Sigma-Aldrich Co.) were included in this experiment in order to establish the degree of similarity among the C. albicans samples and to determinate reproducibility (8,13,14,58).…”

“…Commensal Candida species inhabiting the oral cavity, vaginal canal, and gastrointestinal tract of host may begin the infectious process (15,20,29,32). Different types of electrophoretic techniques have been used for the characterization or typing of Candida species including separation of chromosomes, DNA fragments, isoenzymes, cell-wall glycoproteins and whole-cell proteins (5,8,9,28,30,34,45,47,50). In regard to whole-cell proteins, their separation has been employed satisfactorily in the characterization of bacteria and yeast (13,22,26,55,56,57,58).…”