Changes in karyotype and azole susceptibility of sequential bloodstream isolates from patients with Candida glabrata candidemia

Abstract

We examined the changes in genotypes and azole susceptibilities among sequential bloodstream isolates of Candida glabrata during the course of fungemia and the relationship of these changes to antifungal therapy. Forty-one isolates were obtained from 15 patients (9 patients who received antifungal therapy and 6 patients who did not) over periods of up to 36 days. The isolates were analyzed using pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) and tested for antifungal susceptibility to fluconazole, itraconazole, and voriconazole. PFGE typing consisted of electrophoretic karyotyping and restriction endonuclease analysis of genomic DNA by use of NotI (REAG-N). The 41 isolates yielded 23 different karyotypes and 11 different REAG-N patterns but only 3 MLST types. The sequential strains from each patient had identical or similar REAG-N patterns. However, they had two or three different karyotypes in 6 (40%) of 15 patients. The isolates from these six patients exhibited the same or similar azole susceptibilities, and five patients did not receive antifungal therapy. Development of acquired azole resistance in sequential isolates was detected for only one patient. For this patient, an isolate of the same genotype obtained after azole therapy showed three- or fourfold increases in the MICs of all three azole antifungals and exhibited increased expression of the CgCDR1 efflux pump. This study shows that karyotypic changes can develop rapidly among sequential bloodstream strains of C. glabrata from the same patient without antifungal therapy. In addition, we confirmed that C. glabrata could acquire azole resistance during the course of fungemia in association with azole therapy.

FIG. 1.

Representative genotyping patterns of C. glabrata obtained by EK and REAG-N. Sequential BSI isolates of C. glabrata were obtained from five patients who did not receive antifungal therapy (patients 1 to 5) and from four patients who received antifungal therapy (patients 7 to 10). Table 2 shows detailed information on each isolate. The sequential isolates from each of patients 1 to 4 showed two or three different karyotypes, which demonstrates that karyotype change can occur without antifungal therapy. The sequential isolates from each of patients 3 to 5 and 8 to 10 showed one- or two-band differences in the REAG-N patterns, suggesting that microevolution occurs frequently among bloodstream isolates. Stars indicate the positions of added or deleted bands for clonal strains from the same patient. Two DNA types were shared by isolates from two patients (patients 9 and 10), which suggests nosocomial transmission. A, C. glabrata ATCC 90030. M, Saccharomyces cerevisiae DNA concatemers as a molecular size marker.