Culturable Diversity of Lichen-Associated Yeasts through Enrichment Strategies

Lichens are symbiotic partnerships between a filamentous fungus and a photosymbiotic “alga”. Studies show that lichens harbor endothallic fungi, but that some taxa have been difficult to isolate from the main filamentous thallus-forming fungus and other faster growing lichenicolous/endothallic fungi. Therefore, we aimed to develop and evaluate liquid yeast-enrichment strategies to (1) isolate lichen-associated yeasts in pure culture, and (2) determine the taxonomic placement and breadth of the diversity of culturable yeasts. Eighty-two lichen samples were collected and washed with distilled water, and healthy thalli were ground up and added to seven different yeast-enrichment broths. Yeast colonies were isolated in pure culture and identified using molecular techniques. Initial isolates were identified using BLASTn analysis, and a taxonomic refinement was completed using PhyML analysis. In total, 215 isolates were obtained. The most prevalently isolated ascomycetous yeasts were within the Dothideomycetes (Aureobasidium, Plowrightia, and Dothiora), while the most frequently isolated basidiomycetous yeasts belonged to the genera Curvibasidium, Sporobolomyces, and Tremella. The generic placements could not be determined for 17 isolates, and in total 25 novel species were recovered. The results of this research indicate that (1) lichen-associated yeasts are diverse, (2) employing liquid enrichment strategies is effective for isolating many of these, and (3) lichen thalli represent a valuable untapped reservoir of diverse and novel yeast species.