Phenotypic Characterization and Comparative Genomics of the Melanin-Producing Yeast Exophiala lecanii-corni Reveals a Distinct Stress Tolerance Profile and Reduced Ribosomal Genetic Content

The black yeast Exophiala lecanii-corni of the order Chaetothyriales is notable for its ability to produce abundant quantities of DHN-melanin. While many other Exophiala species are frequent causal agents of human infection, E. lecanii-corni CBS 102400 lacks the thermotolerance requirements that enable pathogenicity, making it appealing for use in targeted functional studies and biotechnological applications. Here, we report the stress tolerance characteristics of E. lecanii-corni, with an emphasis on the influence of melanin on its resistance to various forms of stress. We find that E. lecanii-corni has a distinct stress tolerance profile that includes variation in resistance to temperature, osmotic, and oxidative stress relative to the extremophilic and pathogenic black yeast Exophiala dermatitidis. Notably, the presence of melanin substantially impacts stress resistance in E. lecanii-corni, while this was not found to be the case in E. dermatitidis. The cellular context, therefore, influences the role of melanin in stress protection. In addition, we present a detailed analysis of the E. lecanii-corni genome, revealing key differences in functional genetic content relative to other ascomycetous species, including a significant decrease in abundance of genes encoding ribosomal proteins. In all, this study provides insight into how genetics and physiology may underlie stress tolerance and enhances understanding of the genetic diversity of black yeasts.

Identification of opportunistic and nonopportunistic Exophiala species using high resolution melting analysis

Exophiala is a genus comprising several species of opportunistic black yeasts. Exophiala species identification by morphological, physiological, and biochemical characteristics is challenging because of the low degree of phenotypic differences between species and its polyphyletic nature. We aimed to develop a high-resolution melting (HRM) assay based on the internal transcribed spacer (ITS) region to differentiate between pairs of clinical and environmental Exophiala species. HRM primers were designed based on the conserved ITS region of five Exophiala species (E. dermatitidis, E. phaeomuriformis, E. heteromorpha, E. xenobiotica, and E. crusticola). Environmental and clinical Exophiala isolates representing these five species (n = 109) were analyzed. The HRM assay was optimized using clinical and environmental reference isolates (n = 22), and then the results were compared with those obtained with nonreference isolates of Exophiala (n = 87) using two designed primer sets. The designed HRM assay was based on the normalized melting peak approach and two primer sets, and successfully distinguished between the five Exophiala species. The HRM1 primer set provided sufficient resolution, with a melting temperature (Tm) difference of approximately 2.5°C among the analyzed species and of approximately 1°C between E. dermatitidis and E. phaeomuriformis. HRM typing results were in agreement with those of ITS-sequence typing (100% sensitivity and specificity). The developed HRM assay can be used to ascertain the identity of Exophiala species, which may differ in clinical significance, with high accuracy. Its application to identify species directly in clinical samples and/or environmental niches may be possible in the future.