Phenotypic and genotypic characterization of two closely related subgroups of Candida rugosa in clinical specimens

In this study, six clinical isolates (two from blood, two from urine and one each from a bronchoalveolar lavage and a vaginal swab) were identified as Candida rugosa based on carbohydrate assimilation profiles using API 20C AUX and ID32 C kits (bioMérieux). Sequence analysis of the D1/D2 domain of the yeasts differentiated the isolates into two subgroups, A and B (three isolates per subgroup), which were closely related (99.1-99.6 % nucleotide similarity) to C. rugosa strain ATCC 10571. Compared with the C. rugosa type strain, the intergenic transcribed spacer (ITS) nucleotide similarity for subgroup A was only 89.2 % (29 mismatches and one deletion) and for subgroup B was 93.7 % (20 mismatches). All isolates grew green colonies on Oxoid Chromogenic Candida Agar, with darker pigmentation observed for subgroup A. All isolates were able to grow at 25-42 °C but not at 45 °C. The isolates had identical enzymic profiles, as determined by API ZYM (bioMérieux) analysis, and produced proteinase. High amphotericin MICs (≥1 µg ml(-1)) were noted for two isolates from each subgroup. Dose-dependent susceptibility to fluconazole (MIC 32 µg ml(-1)) was noted in a blood isolate. The biofilms of the isolates demonstrated increased resistance to amphotericin and fluconazole. The greater ITS sequence variability of subgroup A isolates is in support of this yeast being recognized as a distinct species; however, further verification using more sophisticated molecular approaches is required. A sequence comparison study suggested the association of subgroup A with environmental sources and subgroup B with clinical sources. Accurate identification and antifungal susceptibility testing of C. rugosa are important in view of its decreased susceptibility to amphotericin and fluconazole. The ITS region has been shown to be a valuable region for differentiation of closely related subgroups of C. rugosa.