RESUMEN
The incidence of superficial or deep-seated infections due to Candida glabrata has increased markedly, probably because of the low intrinsic susceptibility of this microorganism to azole antifungals and its relatively high propensity to acquire azole resistance. To determine changes in the C. glabrata proteome associated with petite mutations, cytosolic extracts from an azole-resistant petite mutant of C. glabrata induced by exposure to ethidium bromide, and from its azole-susceptible parent isolate were compared by two-dimensional polyacrylamide gel electrophoresis. Proteins of interest were identified by peptide mass fingerprinting or sequence tagging using a matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometer. Tryptic peptides from a total of 160 Coomassie-positive spots were analyzed for each strain. Sixty-five different proteins were identified in the cytosolic extracts of the parent strain and 58 in the petite mutant. Among the proteins identified, 10 were higher in the mutant strain, whereas 23 were lower compared to the parent strain. The results revealed a significant decrease in the enzymes associated with the metabolic rate of mutant cells such as aconitase, transaldolase, and pyruvate kinase, and changes in the levels of specific heat shock proteins. Moreover, transketolase, aconitase and catalase activity measurements decreased significantly in the ethidium bromide-induced petite mutant. These data may be useful for designing experiments to obtain a better understanding of the nuclear response to impairment of mitochondrial function associated with this mutation in C. glabrata.
Asunto(s)
Candida glabrata/química , Proteínas Fúngicas/análisis , Mutación/genética , Proteoma/análisis , Antifúngicos/farmacología , Azoles/farmacología , Candida glabrata/efectos de los fármacos , Candida glabrata/genética , Farmacorresistencia Fúngica/genética , Electroforesis en Gel Bidimensional , Proteínas Fúngicas/genética , Regulación Fúngica de la Expresión Génica , Proteoma/genética , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
The incidence of superficial or deep-seated infections due to Candida glabrata has increased markedly, probably because of the low intrinsic susceptibility of this microorganism to azole antifungals and its relatively high propensity to acquire azole resistance. To determine changes in the C. glabrata proteome associated with petite mutations, cytosolic extracts from an azole-resistant petite mutant of C. glabrata induced by exposure to ethidium bromide, and from its azole-susceptible parent isolate were compared by two-dimensional polyacrylamide gel electrophoresis. Proteins of interest were identified by peptide mass fingerprinting or sequence tagging using a matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometer. Tryptic peptides from a total of 160 Coomassie-positive spots were analyzed for each strain. Sixty-five different proteins were identified in the cytosolic extracts of the parent strain and 58 in the petite mutant. Among the proteins identified, 10 were higher in the mutant strain, whereas 23 were lower compared to the parent strain. The results revealed a significant decrease in the enzymes associated with the metabolic rate of mutant cells such as aconitase, transaldolase, and pyruvate kinase, and changes in the levels of specific heat shock proteins. Moreover, transketolase, aconitase and catalase activity measurements decreased significantly in the ethidium bromide-induced petite mutant. These data may be useful for designing experiments to obtain a better understanding of the nuclear response to impairment of mitochondrial function associated with this mutation in C. glabrata.
Asunto(s)
Candida glabrata/química , Proteínas Fúngicas/análisis , Mutación/genética , Proteoma/análisis , Antifúngicos/farmacología , Azoles/farmacología , Candida glabrata/efectos de los fármacos , Candida glabrata/genética , Farmacorresistencia Fúngica/genética , Electroforesis en Gel Bidimensional , Proteínas Fúngicas/genética , Regulación Fúngica de la Expresión Génica , Proteoma/genética , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
BACKGROUND: In order to identify intraspecific variations in Trichophyton rubrum and to correlate them to the immunological status of the host, sixty strains isolated from AIDS, HIV-positive and HIV-negative patients were compared for the production of extracellular enzymes and for their susceptibility to several antifungal drugs. METHODS: The isolates were tested for their ability to secrete keratinases, proteinases, phospholipases, lipases and DNases. Likewise, we investigated their susceptibility to amphotericin B, ketoconazole, ciclopiroxolamine, griseofulvin, miconazole and tolnaftate. RESULTS: Variations in the Minimal Inhibitory Concentration (MIC80)) values were observed for all antifungals tested, but they were similarly distributed among the three clinical groups. Griseofulvin showed the most prominent differences among the three groups of isolates. Regarding enzyme secretion, all samples secreted keratinases and DNases, while none secreted phospholipases. Proteinases and lipases were secreted by some of them. CONCLUSIONS: The differences among isolates of the three groups were not statistically significant and therefore could not be ascribed to a given clinical status. Intraspecific variations similarly occurred in each group, irrespective of the immunological status of the patients.