Caspofungin Etest Susceptibility Testing of Candida Species: Risk of Misclassification of Susceptible Isolates of C. glabrata and C. krusei when Adopting the Revised CLSI Caspofungin Breakpoints

Abstract

bThe purpose of this study was to evaluate the performance of caspofungin Etest and the recently revised CLSI breakpoints. A total of 497 blood isolates, of which 496 were wild-type isolates, were included. A total of 65/496 susceptible isolates (13.1%) were misclassified as intermediate (I) or resistant (R). Such misclassifications were most commonly observed for Candida krusei (73.1%) and Candida glabrata (33.1%). The revised breakpoints cannot be safely adopted for these two species. The CLSI breakpoints for the three echinocandins have been revised (16). The motivation behind the change was emerging data suggesting that the initial breakpoint defining susceptibility (S) for all Candida species and echinocandins (Յ2 mg/liter) failed to identify isolates with FKS hot spot gene mutations as resistant (1, 2, 4, 6, 9-11, 14, 16). Such isolates have recently been associated with a decreased response rate in animal models (1,19, 20, 22) and with a higher risk of failure in clinical practice (1,8,12,17). EUCAST has established breakpoints for anidulafungin and recommends anidulafungin MIC testing as a marker for the echinocandin class of drugs (3). EUCAST has not set caspofungin breakpoints and does not currently recommend caspofungin MIC testing for clinical decision making. This is due to unacceptably high variation among caspofungin MIC values from different centers which, at least in part, has been linked to high variability in the performance of different lots of pure caspofungin substance (1, 3, 4). Whereas the CLSI and EUCAST reference methodologies, including the associated breakpoints, have been carefully validated using wild-type and mutant isolates, it is less well understood how these breakpoints perform when applied on commercial susceptibility tests. We have recently shown that by using caspofungin Etest and the revised CLSI breakpoints, an FKS1 mutant isolate was successfully identified in a Swedish collection of blood isolates (5). However, it was noted that the caspofungin MICs for Candida glabrata and Candida krusei, as detected by Etest, were very close to the susceptibility breakpoint, suggesting a risk of misclassification of susceptible isolates among these two species (5). We therefore investigated the performance of caspofungin Etest using a collection of Danish blood isolates, including a higher number of C. glabrata and C. krusei isolates. The isolates were, with one exception, all classified as echinocandin susceptible using the EUCAST method and anidulafungin testing as a marker for echinocandin susceptibility (3, 18).A total of 497 blood isolates were included (278 Candida albicans, 9 Candida dubliniensis, 136 C. glabrata, 26 C. krusei,

Use of Micafungin as a Surrogate Marker To Predict Susceptibility and Resistance to Caspofungin among 3,764 Clinical Isolates of Candida by Use of CLSI Methods and Interpretive Criteria

“…In addition to providing a strategy for predicting caspofungin susceptibility and resistance among Candida spp., these results provide strong support for concerns regarding the issue of crossresistance between micafungin and caspofungin (5,15,16,18,22,26). By using an extensive global collection of clinically important isolates, including fks mutant strains, we validate concerns originating from single-center case series and rightly focus attention on C. glabrata as the species most likely to demonstrate cross-resistance between these two studied echinocandins.…”

“…Whereas the CLSI has developed clinical breakpoints (CBPs) for anidulafungin, caspofungin, and micafungin against the six most common species of Candida (9), the European Committee on Antimicrobial Susceptibility Testing (EUCAST) has elected to establish CBPs for anidulafungin and micafungin but not for caspofungin (10). Furthermore, the EUCAST does not currently recommend caspofungin MIC testing for clinical decision making due to unacceptably high variation among the caspofungin MIC values obtained from different centers (4,5,11,12). This variation is evident not only using the EUCAST BMD method (4,11) but also using the CLSI method (13).…”

“…The in vitro activities of caspofungin and micafungin against Candida spp. have been documented using broth dilution, agar disk diffusion, and Etest methods (4)(5)(6)(7)(8), and most recently, clinically relevant interpretive breakpoints for broth microdilution (BMD) MIC testing of Candida spp. were established by the Clinical and Laboratory Standards Institute (CLSI) (9).…”

Multidrug-Resistant Candida auris Misidentified as Candida haemulonii: Characterization by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry and DNA Sequencing and Its Antifungal Susceptibility Profile Variability by Vitek 2, CLSI Broth Microdilution, and Etest Method

“…The lower Etest MIC values than CLSI-BMD MIC values for CAS have also been reported earlier for other Candida species (27). Recently, the performance of the CAS Etest based on the recently revised CLSI breakpoints for Candida isolates showed that 13.1% were misclassified as intermediate or resistant (28). Also, marked interlaboratory variation has been observed with both CLSI-BMD and the EUCAST method for CAS susceptibility (29).…”

Evaluation of the “Dip Effect” Phenomenon in Antifungal Susceptibility Testing of Candida spp. against Echinocandins by Use of Gradient Concentration Strips

“…Different degrees of dip effect may affect the MICs of gradient concentration strips, particularly for isolates with CLSI MICs of Յ0.25 mg/liter, as found in the present study in a greater degree with the MTS strips and a lesser degree with the Etest strips. Because the dip effect occurred for isolates with MICs close to the CLSI susceptibility breakpoints for caspofungin, namely, 0.25 mg/ liter for C. albicans, C. tropicalis, and C. krusei and 0.125 mg/liter for C. glabrata, correct MIC determination is very important in order to avoid classification errors in the susceptibility of isolates as previously reported for caspofungin (12).…”