Echinocandin Susceptibility Testing ofCandidaSpecies: Comparison of EUCAST EDef 7.1, CLSI M27-A3, Etest, Disk Diffusion, and Agar Dilution Methods with RPMI and IsoSensitest Media

Abstract

This study compared nine susceptibility testing methods and 12 endpoints for anidulafungin, caspofungin, and micafungin with the same collection of blinded FKS hot spot mutant (n ‫؍‬ 29) and wild-type isolates (n ‫؍‬ 94). The susceptibility tests included EUCAST Edef 7.1, agar dilution, Etest, and disk diffusion with RPMI-1640 plus 2% glucose (2G) and IsoSensitest-2G media and CLSI M27A-3. Microdilution plates were read after 24 and 48 h. The following test parameters were evaluated: fks hot spot mutants overlapping the wild-type distribution, distance between the two populations, number of very major errors (VMEs; fks mutants misclassified as susceptible), and major errors (MEs; wild-type isolates classified as resistant) using a wild-type-upper-limit value (WT-UL) (two twofold-dilutions higher than the MIC 50 ) as the susceptibility breakpoint. The methods with the lowest number of errors (given as VMEs/ MEs) across the three echinocandins were CLSI (12%/1%), agar dilution with RPMI-2G medium (14%/ 0%), and Etest with RPMI-2G medium (8%/3%). The fewest errors overall were observed for anidulafungin (4%/1% for EUCAST, 4%/3% for CLSI, and 3%/9% for Etest with RPMI-2G). For micafungin, VME rates of 10 to 71% were observed. For caspofungin, agar dilution with either medium was superior (VMEs/MEs of 0%/1%), while CLSI, EUCAST with IsoSensitest-2G medium, and Etest were less optimal (VMEs of 7%, 10%, and 10%, respectively). Applying the CLSI breakpoint (S < 2 g/ml) for CLSI results, 89.2% fks hot spot mutants were classified as anidulafungin susceptible, 60.7% as caspofungin susceptible, and 92.9% as micafungin susceptible. In conclusion, no test was perfect, but anidulafungin susceptibility testing using the WT-UL to define susceptibility reliably identified fks hot spot mutants.

Candida palmioleophila: Characterization of a Previously Overlooked Pathogen and Its Unique Susceptibility Profile in Comparison with Five Related Species

“…This finding contrasts with the diverse susceptibility patterns suggested by the anidulafungin and micafungin EUCAST MIC results and the caspofungin Etest end points. We previously reported on variability in caspofungin microdilution MIC values across microdilution methods, time, and country (3,4). So far, no data suggest a variable susceptibility to the three echinocandins, and we believe that the elevated caspofungin EUCAST MIC ranges for the most anidulafungin-and micafungin-susceptible species in this study reflect an in vitro phenomenon rather than true differences in susceptibility.…”

A Novel Flucytosine-Resistant Yeast Species, Candida pseudoaaseri, Causes Disease in a Cancer Patient

“…The results for the single antifungal agents were comparable, independent of the system that was used (CLSI reference method, Etest, Vitek 2, or Micronaut). It is known that the Etest correlates well with the CLSI reference method for determining the susceptibility of Candida isolates (3,4,5,11,34) and that the results from the YST card of Vitek 2 have a good correlation with the results from the standard method (9,11,36,37). In contrast to both C. aaseri strains, VK065094…”

Rapid Detection of FKS -Associated Echinocandin Resistance in Candida glabrata

“…Breakpoints were specifically developed to identify C. glabrata harboring FKS mutations by considering multiple factors, such as ␤-1,3-D-glucan synthase enzyme kinetics and echinocandin pharmacokinetic and pharmacodynamic data (6,9). However, despite standardized methodologies, important limitations with the in vitro susceptibility testing cannot be ignored: first, the assay has a time-consuming setup and intrinsically slow turnaround time, requiring 24 to 48 h after isolate recovery; second, interlaboratory variability of caspofungin MICs has limited direct testing of this drug (10); and third, susceptible and resistant populations overlap (11).…”