Abstract:

Objective To investigate the mechanism of azole resistance of clinical Candida glabrata isolates. Methods Thirty-eight clinical Candida glabrata isolates were collected from 28 patients of five hospitals between 2009 and 2010. The minimum acteriostatic concentrations of five commonly used antifungal agents (amphotericin B, 5-flucytosine, fluconazole, itraconazole, and voriconazole) were determined by the broth microdilution antifungal susceptibility test. The expressions of efflux-related genes (CDR1, CDR2, and SNQ2), ERG11, and transcription factor PDR1 were detected by the Real-Time PCR. The efflux ability of rhodamine 6G was analyzed by the flow cytometry. The function of the efflux pump of cell membrane was investigated. The whole sequence of ERG11 gene and the major functional domains of PDR1 were amplified by the PCR, sequenced, and compared to standard sequences. Results All 38 Candida glabrata isolates were sensitive to amphotericin B and 5-flucytosine and 17 of which were azole resistant. The expression of CDR1 and the efflux ability of rhodamine 6G of azole resistant isolates were significantly higher than those of sensitive isolates (P=0.001). The difference of the expression of ERG11 of sensitive isolates and azole resistant isolates was not statistically significant (P=0.283). No mutations of ERG11 were found in azole resistant isolates. The difference of the expression of PDR1 of 17 azole-resistant isolates and sensitive isolates was not statistically significant (P=0.086). But one sense mutation of PDR1 was found in each azole resistant isolate. Conclusion Mutations of transcription factor PDR1, up-regulation of efflux pump related gene CDR1, and the increase of the function of efflux pumps are the main mechanisms of clinical zole resistance of Candida glabrata.

Key words: azole resistance, Candida glabrata, efflux pump, ERG11 gene, transcription factor PDR1