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 (CBS1879T) 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 CBS1879T cultured 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.

Malassezia Yeasts in Veterinary Dermatology: An Updated Overview

“…Kano et al showed that an isolate of M. pachydermatis with MICs of itraconazole and ketoconazole of >32 mg/L by Etest had mis-sense mutations in the ERG11 gene that encodes lanosterol 14 -alpha-demethylase, the target site for antifungal azoles (Kano et al, 2019b). Mutations in the same gene were described in field isolates with tolerance to ravuconazole (Kano et al, 2019a) and in miconazole-resistant clones of CBS1879 (the neotype culture of M. pachydermatis) selected by serial passage on miconazole supplemented media (Kano and Kamata, 2019). Azole resistance in M. pachydermatis has also been associated with quadruplication of the ERG11 gene (Kim et al, 2018).…”

Antifungal Resistance Regarding Malassezia pachydermatis: Where Are We Now?

“…Other studies generated mutants resistant/tolerant to NYS [74], KTZ [72], and MCZ. [112] to investigate possible mechanisms of resistance (see Section 12) using the following methods: exposure to N-methyl-N'-nitrosoguanidine and UV radiation [74] and serial subcultures of a yeast colony on a solid medium containing increasing concentrations of KTZ [72] and MCZ [112].…”

“…Kano et al (2019) [83] showed that an isolate with proved clinical resistance (see Section 4) had missense mutations in the ERG11 gene that encodes lanosterol 14 α demethylase, the target site for antifungal azoles. Mutations in the same gene were described in two field isolates with multi-azole in vitro resistance [85] and MCZ-resistant clones of CBS 1879 selected by serial passages on MCZ-supplemented media [112].…”

Use of a modified Burow's solution to treat canine otitis externa: A randomised comparative clinical study

“…This may have helped disperse it more widely within the inflamed ear canal and contributed to its lower failure rate in general, and better clinical efficacy in dogs over 20 kg compared with Surolan. Tolerance of M. pachydermatis to azole agents has been reported in strains isolated from dogs worldwide 12 . This may be one of the reasons why the cure rate for OE caused by yeast in this study was higher for the modified Burow's solution than for Surolan, which contains miconazole as an antifungal agent.…”

“…Tolerance of M. pachydermatis to azole agents has been reported in strains isolated from dogs worldwide. 12 This may be one of the reasons why the cure rate for OE caused by yeast in this study was higher for the modified Burow's solution than for Surolan, which contains miconazole as an antifungal agent.…”