Miconazole-tolerant strains of Malassezia pachydermatis generated by culture in medium containing miconazole.

Abstract

BACKGROUND: Tolerance of Malassezia pachydermatis to azole drugs has been reported worldwide, from strains isolated from dogs. Canine Malassezia dermatitis often is treated with shampoos containing 2% miconazole (MCZ) or other topical MCZ products. OBJECTIVES: In the in vitro study herein, it was investigated whether MCZ-induced amino acid substitutions in the lanosterol 14-alpha-demethylase (ERG11) gene 1 lead to azole tolerance in M. pachydermatis. METHODS AND MATERIALS: Toleranced to MCZ was induced in an azole-susceptible strain of M. pachydermatis (CBS1879) by culture in medium containing MCZ. Antifungal susceptibility to MCZ, clotrimazole (CTZ) and itraconazole (ITZ) was assessed using the modified broth microdilution (BM) method. To assess the potential mechanism of tolerance in the three MCZ-resistant strains, ERG11 was sequenced. The interaction between the calcineurin inhibitor tacrolimus and MCZ in the azole-tolerant isolates also was examined. RESULTS: Three strains (NUBS19001 to NUBS19003) from CBS1879cultured in medium containing MCZ exhibited minimum inhibitory concentrations (MICs) of 40 mg/L to MCZ, 5 mg/L to ITZ and >32 mg/L to CTZ, meaning that the isolates were tolerant. The combination of MCZ and tacrolimus exerted an indifferent effect against the MCZ-tolerant strain. BLAST analysis using the NCBI database showed mutations in the cytochrome p450 encoded by ERG11 in the MCZ-tolerant strains. CONCLUSIONS: In the present in vitro study, it was shown that MCZ exposure can induce amino acid substitutions in ERG11 and subsequent tolerance of M. pachydermatis to several azoles. Whether topical therapy with azole-containing products can exert a similar effect in vivo is a question that requires further research.