The conversion of grape juice to wine starts with complex yeast communities consisting of strains that have colonised the harvested grape and/or reside in the winery environment. As the conditions in the fermenting juice gradually become inhibitory for most species, they are rapidly overgrown by the more adaptable Saccharomyces strains, which then complete the fermentation. However, there are environmental factors that even Saccharomyces cannot cope with. We show that when the sugar content is extremely high, osmotolerant yeasts, usually considered as “spoilage yeasts“, ferment the must. The examination of the yeast biota of 22 botrytised Tokaj Essence wines of sugar concentrations ranging from 365 to 752 g∙L⁻¹ identified the osmotolerant Zygosaccharomyces rouxii, Candida (Starmerella) lactis-condensi and Candida zemplinina (Starmerella bacillaris) as the dominating species. Ten additional species, mostly known as osmotolerant spoilage yeasts or biofilm-producing yeasts, were detected as minor components of the populations. The high phenotypical and molecular (karyotype, mtDNA restriction fragment length polymorphism (RFLP) and microsatellite-primed PCR (MSP-PCR)) diversity of the conspecific strains indicated that diverse clones of the species coexisted in the wines. Genetic segregation of certain clones and interactions (antagonism and crossfeeding) of the species also appeared to shape the fermenting yeast biota.

The interaction between plant defensive metabolites and different plant-associated fungal species is of high interest to many disciplines. Volatile organic compounds (VOCs) are natural products that are easily evaporated under ambient conditions. They play a very important role in inter-species communication of microbes and their hosts. In this study, the VOCs produced by 43 different fungal isolates of endophytic and soil fungi during growth on horseradish root (Armoracia rusticana) extract or malt extract agar were examined, by using headspace-gas chromatography-mass spectrometry (headspace-GC-MS) and a high relative surface agar film as a medium. The proposed technique enabled sensitive detection of several typical VOCs (acetone, methyl acetate, methyl formate, ethyl acetate, methyl butanol isomers, styrene, beta-phellandrene), along with glucosinolate decomposition products, including allyl cyanide and allyl isothiocyanate and other sulfur-containing compounds—carbon disulfide, dimethyl sulfide. The VOC patterns of fungi belonging to Setophoma, Paraphoma, Plectosphaerella, Pyrenochaeta, Volutella, Cadophora, Notophoma, and Curvularia genera were described for the first time. The VOC pattern was significantly different among the isolates. The pattern was indicative of putative myrosinase activity for many tested isolates. On the other hand, endophytes and soil fungi as groups could not be separated by VOC pattern or intensity.

Pathogenic fungi often cause serious infections mainly in immunocompromised persons. The number of infections caused by the non-albicans Candida or other species has significantly increased over the last years. These infections present a major challenge in the health sector because these pathogenic fungi have strong virulence and often show resistance to the commonly used antifungal treatments. To solve the problems caused by the drug resistant pathogenic fungi, it is necessary to find new antifungal agents and their sources. The aim of this study was to give evidence that yeasts can effectively fight against strains which belong to pathogenic fungi and reveal those yeasts which are able to inhibit growth of Kodamaea ohmeri, Pichia kudriavzevii, Naganishia albida or Candida tropicalis. Furthermore, we wanted to determine the effects of certain culturing factors on the growth inhibition. Our screening revealed that although the strains belonging to pathogenic species were much more tolerant to the yeast-produced bioactive agents than the non-disease-associated yeasts, growth of Kodamaea ohmeri and Candida tropicalis could be inhibited by Metschnikowia andauensis, while Naganishia albida could be controlled by Pichia anomala or Candida tropicalis. Our data proved that the experimental circumstances could have a serious impact on the inhibitory capacity of the yeasts. Appearance of inhibition strongly depended on media, pH and temperature. Our data also shed some light on the fact that Pichia kudriavzevii must have high natural resistance to the yeast-produced agents, while other species, such as Saccharomycopsis crataegensis belonged to the easily inhibitable species. Our study suggests that yeast-produced bioactive agents could be potential growth inhibitory agents against the disease-associated fungi and yeasts can also contribute to alternative approaches to combat against pathogenic fungi. Our data revealed an important role of the culturing factors in inhibition and pointed to the complex nature of antagonism.

The yeast Starmerella (Candida) lactis-condensi is considered a food contaminant microorganism. The aim of our research was to determine why St. lactis-condensi could become the dominant species of Essences, the top sweet wine speciality of Tokaj wine region in Hungary. We investigated the physiological properties of these yeasts based on parameters that may influence their ability to selectively proliferate and persist during maturation in wines with very high sugar content. These include glucose and fructose, alcohol, and sulphur tolerance. Our studies have shown that St. lactis-condensi is a fructophilic yeast that is able to adapt quickly to very high sugar concentrations (up to 500 g/L) in the Essences. The high glucose concentration inhibits its growth, as well as that of the St. bacillaris (Candida zemplinina) strains tested. The type and amount of sugars in the Essences, together with the sulphur and alcohol content, influence the composition of the dominant yeast biota. Analysis of (GTG)₅ microsatellite in the nuclear genome and mtDNA-RFLP studies demonstrate that a diverse population of St. lactis-condensi occurs in the Tokaj wine region, in the Essences. This yeast species is characterised by both physiological and genetic biodiversity. GC-MS analysis of Essences colonised exclusively with these yeasts showed no deterioration in quality.

The glucosinolates of Brassicaceae plants are converted into bioactive isothiocyanates and other volatiles during a challenge by pathogens and other biotic stressors. However, the role of alternative downstream products with weaker potency (e.g., nitriles) is far from being fully understood. This study tested the possible synergistic antifungal interaction between various glucosinolate-derived nitriles and 2-phenylethyl isothiocyanate (PEITC) on 45 fungal strains, including endophytes from horseradish roots (Brassicaceae) and soil fungi, using an airtight system enabling the accurate study of extremely volatile antifungal agents. The median minimal inhibitory concentrations (MICs) were 1.28, 6.10, 27.00 and 49.72 mM for 1H-indole-3-acetonitrile (IAN), 3-phenylpropanenitrile (PPN), 4-(methylsulfanyl)-butanenitrile (MSBN) and 3-butenenitrile (BN, = allyl cyanide), respectively. Thus, nitriles were considerably weaker antifungal agents compared to PEITC with a median MIC of 0.04 mM. For the same nitriles, the median fractional inhibitory concentration indices (FICIs) of the combinations were 0.562, 0.531, 0.562 and 0.625, respectively. Altogether, 47.7%, 56.8%, 50.0% and 27.3% of tested fungal strains showed a synergistic antifungal activity (FICI ≤ 0.5) for the nitrile–isothiocyanate combinations, respectively. Hypocreales strains showed the least sensitivity towards the GSL decomposition products and their combinations. The mean MIC values for PEITC showed 0.0679 ± 0.0358, 0.0400 ± 0.0214, 0.0319 ± 0.0087 and 0.0178 ± 0.0171 mM for Hypocreales, Eurotiales, Glomerellales and Pleosporales, respectively. In addition, nitriles, especially IAN, also showed significant differences. For the same fungi, the median FICI values fell in the ranges of 0.61–0.67, 0.52–0.61, 0.40–0.50 and 0.48–0.67, respectively, depending on the nitrile. Our results suggest that glucosinolate-derived nitriles may enhance isothiocyanate antifungal activity and that they may play an active role in shaping the plant microbiome and contribute to the filtering of microbes by plants.

Starmerella bacillaris (Candida zemplinina) is a genetically heterogeneous species. In this work, the diversity of 41 strains of various origins is examined and compared by the analysis of the length polymorphism of nuclear microsatellites and the RFLP of mitochondrial genomes. The band patterns are analysed with UPGMA, neighbor joining, neighbor net, minimum spanning tree and non-metric MDS algorithms. The results and their comparison to previous analyses demonstrate that different markers and different clustering methods can result in very different groupings of the same strains. The observed differences between the topologies of the dendrograms also indicate that the positions of the strains do not necessarily reflect their real genetic relationships and origins. The possibilities that the differences might be partially due to different sensitivity of the markers to environmental factors (selection pressure) and partially to the different grouping criteria of the algorithms are also discussed.