Comparison of EUCAST and CLSI Reference Microdilution MICs of Eight Antifungal Compounds for Candida auris and Associated Tentative Epidemiological Cutoff Values

Abstract

Candida auris is an emerging multidrug-resistant yeast. So far, all but two susceptibility testing studies have examined ≤50 isolates, mostly with the CLSI method. We investigated CLSI and EUCAST MICs for 123 C. auris isolates and eight antifungals and evaluated various methods for epidemiological cutoff (ECOFF) determinations. MICs (in milligrams per liter) were determined using CLSI method M27-A3, and the EUCAST E.Def 7.3. ANOVA analysis of variance with Bonferroni's multiple-comparison test and Pearson analysis were used on log₂ MICs (significance at P values of <0.05). The percent agreement (within ±0 to ±2 2-fold dilutions) between the methods was calculated. ECOFFs were determined visually, statistically (using the ECOFF Finder program and MicDat1.23 software with 95% to 99% endpoints), and via the derivatization method (dECOFFs). The CLSI and EUCAST MIC distributions were wide, with several peaks for all compounds except amphotericin B, suggesting possible acquired resistance. Modal MIC, geometric MIC, MIC₅₀, and MIC₉₀ values were ≤1 2-fold dilutions apart, and no significant differences were found. The quantitative agreement was best for amphotericin B (80%/97% within ±1/±2 dilutions) and lowest for isavuconazole and anidulafungin (58%/76% to 75% within ±1/±2 dilutions). We found that 90.2%/100% of the isolates were amphotericin B susceptible based on CLSI/EUCAST methods, respectively (i.e., with MICs of ≤1 mg/liter), and 100%/97.6% were fluconazole nonsusceptible by CLSI/EUCAST (MICs > 2). The ECOFFs (in milligrams per liter) were similar across the three different methods for itraconazole (ranges for CLSI/EUCAST, 0.25 to 0.5/0.5 to 1), posaconazole (0.125/0.125 to 0.25), amphotericin B (0.25 to 0.5/1 to 2), micafungin (0.25 to 0.5), and anidulafungin (0.25 to 0.5/0.25 to 1). In contrast, the estimated ECOFFs were dependent on the method applied for voriconazole (1 to 32) and isavuconazole (0.125 to 4). CLSI and EUCAST MICs were remarkably similar and confirmed uniform fluconazole resistance and variable acquired resistance to the other agents.

Keywords: Candida auris; ECOFF; MIC; amphotericin B; antifungal susceptibility testing; azoles; echinocandins; in vitro.

FIG 1

EUCAST spectrophotometer concentration versus OD curves for a C. auris isolate with significant partial growth inhibition over several dilutions (resulting in less steep growth curves), in particular for anidulafungin (a) and isavuconazole (b). (c) For comparison, the amphotericin B inhibition curve is shown. Partial growth inhibition ranges are encircled. Ani, anidulafungin; Isa, isavuconazole; Amb, amphotericin B; mean pos control, the mean positive control line, which corresponds to the mean optical density calculated from 8 replicate drug-free growth control wells (upper dotted line), whereas the 50% cutoff line corresponds to the 50% growth inhibition endpoint after subtracting the background OD (lower dotted line).