In vitro interactions among echinocandins against Aspergillus fumigatus: lack of concordance among methods.

Drug combinations are increasingly being explored as a means of improving outcomes in cases of invasive aspergillosis. However, the optimal methods for assessing in vitro drug combinations are unclear. We investigated whether echinocandin drug combinations have potentially useful interactions against Aspergillus fumigatus. We explored the in vitro interactions of three echinocandins (caspofungin [CSP], anidulafingin [ANF], and micafungin [MCF]) by three methods, i.e., (i) checkerboard assay, (ii) disk diffusion assay, and (iii) E-test/agar dilution. The checkerboard experiments revealed different interactions between echinocandin pairs depending on the drug combination tested. Specifically, the combination of CSF and either ANF or MCF yielded a fractional inhibitory concentration (FIC) index range of 0.15-2.0. Whereas the combination of MCF and ANF yielded an FIC index range of 0.19-0.31, consistent with synergistic interaction. In contrast, all echinocandin pairs appeared indifferent by the disk diffusion method. Moreover, the combination of MCF and ANF appeared antagonistic when the two drugs were tested using the E-test/agar dilution method with both the FKS1 wild-type and echinocandin-resistant fks1 mutant strains. Our results highlight methodological problems inherent in in vitro antifungal combination testing. We did not find compelling evidence of inter-echinocandin synergy that could serve as a basis for further in vivo experimentation.