A catalogue of cultivable yeasts from the microbiota of grape berries cv. Vinhão and Loureiro

The relevance of non-Saccharomyces native yeasts for improving the complexity of wine aroma is increasingly recognised, besides their use in a range of biotechnological applications. Previously, we used a metagenomic approach to assess the effects of vineyard calcium sprays on the microbiota of grape berries cv. Vinhão and Loureiro. In this study, we aimed at assembling a catalogue of the cultivable yeasts from grape berries of these cultivars and to investigate the direct effect of calcium and other elicitors on the isolated taxa. Seventeen unique colony morphologies were identified, and the sequencing of the region spanning the rDNA internal transcribed spacers or the rDNA D1/D2 domain revealed that they are distributed into 13 different genera. Although the cultivar and vintage influenced the taxa abundance and diversity, Aureobasidium pullulans and Hanseniaspora uvarum were the most abundant species, ubiquitously populating the fruits together with Pichia terricola, Rhodotorula glutinis, Starmerella bacillaris and Vishniacozyma heimaeyensis. The rarest genera comprised Candida, Trichosporon and Zygoascus. Analysis of yeast growth in the presence of specific elicitors revealed that Pichia spp. are the most sensitive to exogenous calcium, while Sporobolomyces roseus is the most sensitive to copper exposure. Moreover, A. pullulans, H. uvarum and S. bacillaris displayed the highest sensitivity to osmotic stress. In turn, Wickerhamomyces anomalus, Metschnikowia pulcherrima, P. fermentans, P. kluyveri and P. kudriavzevii showed mild resistance to ethanol, unlike the remaining non-Saccharomyces yeasts. Correlation networks expressing the overall yeast performance showed that H. uvarum and S. bacillaris share the strongest positive interspecies correlation, followed by the pairs P. fermentans–P. kluyveri, B. albus–R. glutinis and A. pullulans–V. heimaeyensis. The strongest negative correlations were found for the pairs P. kluyveri–S. roseus, P. terricola–W. anomalus, P. kudriavzevii–V. heimayensis, P. kluyveri–V. heimayensis, and A. pullulans–P. kluyveri. This study exposed microbial niches with a predictably common response to abiotic stress.