Abstract
In this thesis two parameters of in vitro antifungal susceptibility testing of filamentous fungi, the medium and the quantification of fungal growth, were investigated and different drug interaction models were used for analyzing the in vitro combination of various antifungal drugs. A microbroth kinetic system was developed in order to analyze the growth curves of three species of filamentous fungi in five different nutrient media. Two colorimetric methods based on the reduction of the tetrazolium salts, MTT and XTT, was evaluated for the MIC determination of different antifungal drugs against different species of filamentous fungi. High levels of agreement were found between the colorimetric medthods and the NCCLS method for all drugs and species tested. The in vitro combination of terbinafine with three azoles, namely itraconazole, voriconazole and miconazole was tested against clinical S. prolificans isolates using the spectrophotometric method, the colorimetric MTT method and a modified MTT method. In addition, the results were analyzed with parametric and non-parametric approaches of the Loewe additivity and Bliss independence zero interaction theories. Strong synergy was found between terbinafine and the three azoles with miconazole and voriconazole showing the strongest synergy based on Loewe additivity and Bliss independece theory, respectively. Less variable results were obtained with the modified MTT method. Despite its simplicity, results of the FIC index model depended on the choice of MIC endpoints and interpretation endpoints. Furthermore it lacked of good summary. Although the model described by Prichard et al. resulted in a concentration dependent mosaic of interactions, replicates were required for statistical evaluation of the results, good summary was absent and the results were dependent on the concentration range used. The fully parametric model described by Greco et al., although it did not describe precisely the response surface of antifungal combinations, was able to distinguish synergistic and antagonistic interactions and summarized the interaction with a non-unit concentration independent interaction parameter including statistically significance levels without requiring replicates
