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We investigated the activities of fluconazole, caspofungin, anidulafungin, and amphotericin B againstCandida species in planktonic form and biofilms using a highly sensitive assay measuring growth-related heat production (microcalorimetry).C. albicans ,C. glabrata ,C. krusei , andC. parapsilosis were tested, and MICs were determined by the broth microdilution method. The antifungal activities were determined by isothermal microcalorimetry at 37°C in RPMI 1640. For planktonicCandida , heat flow was measured in the presence of antifungal dilutions for 24 h.Candida biofilm was formed on porous glass beads for 24 h and exposed to serial dilutions of antifungals for 24 h, and heat flow was measured for 48 h. The minimum heat inhibitory concentration (MHIC) was defined as the lowest antifungal concentration reducing the heat flow peak by ≥50% (≥90% for amphotericin B) at 24 h for planktonicCandida and at 48 h forCandida biofilms (measured also at 24 h). Fluconazole (planktonic MHICs, 0.25 to &gt;512 μg/ml) and amphotericin B (planktonic MHICs, 0.25 to 1 μg/ml) showed higher MHICs than anidulafungin (planktonic MHICs, 0.015 to 0.5 μg/ml) and caspofungin (planktonic MHICs, 0.125 to 0.5 μg/ml). AgainstCandida species in biofilms, fluconazole's activity was reduced by &gt;1,000-fold compared to its activity against the planktonic counterparts, whereas echinocandins and amphotericin B mainly preserved their activities. Fluconazole induced growth of planktonicC. krusei at sub-MICs. At high concentrations of caspofungin (&gt;4 μg/ml), paradoxical growth of planktonicC. albicans andC. glabrata was observed. Microcalorimetry enabled real-time evaluation of antifungal activities against planktonic and biofilmCandida organisms. It can be used in the future to evaluate new antifungals and antifungal combinations and to study resistant strains.

Activities of Fluconazole, Caspofungin, Anidulafungin, and Amphotericin B on Planktonic and Biofilm Candida Species Determined by Microcalorimetry