Non-synonymous ERG11 mutations in M. restricta and M. arunalokei: impact on azole susceptibility

Malassezia are commensal lipid-dependent yeasts and opportunistic pathogens that cause superficial mycoses and systemic infection. Azole antifungals target cell wall ergosterol synthesis and are the first line of antifungal treatment. ERG11 gene mutations and overexpression are major mechanisms conferring azole resistance and resulting in antifungal therapy failure. Malassezia restricta is found ubiquitously on healthy and diseased skin, with azole-resistant isolates described. Malassezia arunalokei is a relatively new, closely related common skin species. Ketoconazole and itraconazole were the most effective at inhibiting both species. Isolates of M. restricta and M. arunalokei from healthy skin of Singapore subjects were cultured, evaluated, and generally susceptible to common over-the-counter azoles, including clotrimazole, except for select less-susceptible strains. Some less-susceptible strains have novel or reported non-synonymous mutations in the ERG11 gene, such as R88C. The QK178RQ ERG11 sequence variation was observed to be associated with differences in M. restricta and M. arunalokei as independent species. In the absence of identified ERG11 mutations, strains with elevated MICs were observed to have elevated ERG11 expression and drug efflux pump expression/activity. We conclude that antifungal susceptibility is determined by a combination of intrinsic (e.g., mutations, gene expression, efflux pump activity) and extrinsic (e.g., skin condition, prior antifungal exposure) factors and that the skin microbiome serves as a reference for the emergence of new mutations and strain phenotypes.

Malassezia over colonization is associated with conditions such as dandruff and seborrheic dermatitis, which give rise to unpleasant itching and swelling on the skin. Azole antifungals such as ketoconazole, clotrimazole, and miconazole are the primary treatments of choice available as over-the-counter creams or shampoos. However, the emergence of antifungal resistance leads to a loss of treatment efficacy and persistent fungal infection. To understand the mechanisms underlying antifungal resistance, we profiled the susceptibility profiles of commensal Malassezia isolates from the skin and identified novel ERG11 mutations. Our results indicate that antifungal susceptibility is determined by a combination of factors (mutations, efflux pump activity, gene expression, copy number) and suggest that the healthy skin microbiome serves as a reference for the emergence of new mutations and strain phenotypes.