Microdiluição em caldo para teste de susceptibilidade de Malassezia furfur, Malassezia obtusa e Malassezia sympodialis

Abstract

In the Amazon, the tropical climate is the main factor that favors the development of superficial fungal infections and opportunistic, especially the high temperature and relative humidity, providing ideal conditions for continuity in the life cycle of pathogenic and opportunistic fungi. Among the most prevalent mycosis in this region, is pityriasis versicolor (PV), a major skin diseases treated at health care in dermatology, can affect people of both genders, age, race and social class. Different species of Malassezia spp., Some of which recently described by means of genetic studies, are responsible for PV and, despite its high prevalence in several regions of the world and in Brazil, there are few studies with more detailed approach on epidemiological, clinical and laboratory features of this mycosis in our region. Research on the susceptibility of these species to antifungal drugs is fundamental to the advancement of scientific knowledge in order to better clinical management and treatment of PV. The testing for evaluation of in vitro susceptibility to antifungal agents for yeasts and molds (vitro antifungal susceptibility), constitutes an important tool for monitoring the resistance of fungal strains and also to assist in choosing the best therapeutic regimen. Given this, the Clinical and Laboratory Standards Institute (CLSI), formerly National Committee for Clinical Laboratory Standards (NCCLS) have developed methods for the assessment of fungal susceptibility, however, due to different nutritional requirements of these species, there is still no specific protocol for testing with Malassezia spp. that has been recommended by this or any other body accredited for the regulation of clinical and laboratory procedures. The present study with the main objective to contribute to the development of a simplified protocol for assessing the susceptibility of three species of Malassezia, being isolated from 01 M. furfur, 01 and 01 M.obtusa M. sympodialis front to drugs ketoconazole, itraconazole and fluconazole. For this, bioassay were performed using the method in broth at different experimental conditions, which were defined from some methodological variants found in publications in the last 10 years, for the optimum conditions for growth, which were tested three different concentrations of inoculum, four types of culture media, incubation temperatures and two different criteria for interpretation of results. According to the results, the conditions that led to better performance during the experiments and the growth curves in bioassays with the drugs were: inoculum concentration of 0.5 to 2.5 x104 cells / mL and incubation temperature of 32 ° C; medium modified Leeming-Notman with some changes in its composition, and spectrophotometric reading at 620 nm after 3 days of incubation. Accordingly, the MIC values obtained were: MIC ≤ 0.03 mg / mL for ketoconazole and itraconazole to inhibit 50% and 80% of the growth of M. obtusa and M. furfur. For the fluconazole MICs were found for 2 and 4 mg / mL to inhibit 50% of the growth of M. sympodialis and M. obtusa respectively, and MIC of 4:08 g / mL for inhibition of 80%, while for M. furfur found an MIC ≤ 0.03 mg / mL of this drug to inhibit 50% and 80% of fungal growth. Therefore, although preliminary, considering the small number of samples analyzed, the results of this study provided important information in view of developing a protocol for susceptibility testing of Malassezia species.