Susceptibility profiles of Malassezia species against antifungal drugs of clinical use.

Objective: Malassezia yeasts have been recognized as members of the normal skin biota of humans and other warm-blooded animals. These yeasts are the aetiological agents of pityriasis versicolor and Malassezia folliculitis and can be associated as agent or contributory factor in other skin disorders. In addition, Malassezia sp. has been associated with systemic infections in neonates and immunocompromised patients. Due to a defect in their ability to synthesize fatty acids, all these yeasts are obligatory lipophilic, and most of them lipid-dependent. Broth microdilution reference method is not applicable to Malassezia species, due to their strict nutritional requirements. Therefore modifications to allow Malassezia growth are required, such as use of supplemented medium, different inoculum size, temperature and reading time. The aim of this study was to evaluate the antifungal susceptibility of three Malassezia species against fluconazole (FZ), voriconazole (VZ), ketoconazole (KZ), itraconazole (IZ) and amphotericin B (AB). Methods: 130 Malassezia isolates, including 64 M. furfur, 38 M. sympodialis and 28 M. globosa were studied. Identification was performed using PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) method. Minimal inhibitory concentration (MIC) was determined according to CLSIM27”A3 reference document, with some modifications to support optimal growth of these yeasts. Results were reported as MIC ranges, MIC50 and MIC90 (MIC values that inhibited 50% and 90% of the isolates growth) and geometric mean (GM). Results: MIC (μg/mL) ranges against FZ, AB and VZ were very variable, M. furfur (FZ: ≤0.125→64, AB: 0.25→16, VZ:0.03-2), M. sympodialis (FZ: ≤0.125-16, AB: 0.125-16, VZ:0.03-1) and M. globosa (FZ: ≤0.125-64, AB: 0.06-8, VZ: 0.03- 0.5). In contrast, all isolates were inhibited at low concentrations of IZ and KZ: M. furfur (MIC50 :0.03, MIC90 :0.06 for both KZ and IZ), M. sympodialis (MIC50 and MIC90: 0.03 for KZ and MIC50 : 0.03 and MIC90 : 0.06 for IZ) and M. globosa (MIC50 and MIC90 : 0.03 for both KZ and IZ). M. furfur showed the highest MIC values against all drugs tested (GM FZ: 4.90, KZ: 0.04, AB: 2.38, VZ: 0.09 and IZ: 0.04) and M. sympodialis (GM FZ: 1.44, KZ: 0.03, AB: 1.72, VZ: 0.08 and IZ: 0.04) and M. globosa (GM FZ: 1.39, KZ: 0.03, AB: 0.85, VZ: 0.06 and IZ: 0.03) were lower. Conclusion: IZ and KZ were highly effective drugs against Malassezia species tested. These yeasts were inhibited at low concentrations of these drugs and presented slight interspecies variability. Although VZ showed an active drug for most of the isolates studies, some high MIC values were obtained for the three species. In contrast, susceptibility profiles of the yeasts against FZ, AB were very variable, displaying high values and wide ranges. This is important considering that these are more commonly used drugs. M. furfur showed the highest MIC values against all drugs tested, while M. globosa and M. sympodialis were more susceptible. Differences observed emphasize the needing to perform species identification and antifungal susceptibility testing of Malassezia yeasts. Further investigations and collaborative studies are needed for correlation in vitro results with clinical outcomes.