Fluconazole susceptibility testing of Candida inconspicua clinical isolates: comparison of four methods.

Sir, During the past 4 years, isolation of Candida inconspicua from clinical samples has increased in our laboratory. Fluconazole MIC values for C. inconspicua isolates are relatively high, as determined by standard broth microdilution (BMD), Etest or Fungitest, – 4 and according to the standard BMD method they fall mainly into the susceptible-dose dependent (S-DD) or resistant (R) categories, with few susceptible (S) isolates. – 3 Since fluconazole is the most widely used drug in the clinical setting, and in the case of rare Candida species susceptibility test results may precede definitive identification (24 or 48 h versus 48 or 72 h), the method used for susceptibility testing should be suitable to correctly determine the fluconazole susceptibility not only of the four most frequent species, but also of relatively frequently isolated rare species. The aim of our study was to evaluate Etest, Fungitest and modified BMD methods as alternatives for determination of fluconazole susceptibility of our C. inconspicua clinical isolates and to compare their agreement with the standard method. The isolates studied included 57 C. inconspicua strains isolated from 48 patients during a 3 year period (2001–2003). Twenty-two of the 42 hospitalized patients were immunocompromised and 15 were hospitalized in seven different intensive care units. Eleven patients received fluconazole previously. The majority of specimens were from the upper and lower respiratory tract (16 and 24, respectively), but wound, blood and genital isolates were also obtained. Inpatients were treated at 10 separate clinics at different times. Identification of the isolates was carried out as described earlier. The reference BMD method was carried out according to the guidelines of the NCCLS document M27-A2. The pure fluconazole powder (Pfizer) was dissolved in sterile distilled water, and the final concentration range was 0.25–128 mg/L. Yeast suspensions were prepared in RPMI 1640 medium and were adjusted to a final concentration of 10 cells/mL (standard inoculum size), or in the modified BMD method 10 cells/mL (large inoculum size). Drug-free purity controls and growth controls were included in each plate. The plates were incubated at 358C and read visually after 24 and 48 h. Fluconazole MIC was defined as the lowest concentration that produced a prominent decrease in turbidity compared with that of the drug-free growth control. Quality control strains of Candida parapsilosis (ATCC 22019) and Candida krusei (ATCC 6258) were included in each test. Etest (AB Biodisk, Solna, Sweden) and Fungitest (Bio-Rad SDP, formerly Sanofi Diagnostics Pasteur, France) methods were carried out according to the manufacturers’ instructions. Etest MICs were recorded at 24 and 48 h. Fungitest was read