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Hyphopichia pseudoburtonii, is emerging as a potential biocontrol agent against various phytopathogens. These traits have been attributed to the production of various antifungal compounds in the presence of target pathogens. However, the broad molecular mechanisms involved in the antifungal activity are not yet understood. This study employed RNA sequencing to assess the temporal changes in H. pseudoburtonii Y963 gene expression patterns when co-cultivated with Botrytis cinerea. Genes differentially expressed in H. pseudoburtonii in co-culture with B. cinerea, compared to the monoculture were evaluated after 24, 48, and 120 h of growth. Up-regulation of genes encoding major core histones (H2A, H3, H4) and ribosomes in the first 24 h suggested an abundance of cells in the S phase of the cell cycle. At 48 h, the genes up-regulated highlight mitotic cell cycle activity and induction of filamentous growth, while in later stages, up-regulation of genes encoding high affinity transporters of sugars, copper and iron, as well as those involved in the retention and utilization of siderophore-iron was evident. Altogether, the data allude to competition for space and nutrients as key mechanisms activated in H. pseudoburtonii in the presence of B. cinerea. This research offers new insights into H. pseudoburtonii transcriptomic response to B. cinerea and illuminates the adaptive strategies and molecular mechanisms behind its antifungal activity., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2025 Maluleke et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Grapes harbour a plethora of non-conventional yeast species. Over the past two decades, several of the species have been extensively characterised and their contribution to wine quality is better understood. Beyond fermentation, some of the species have been investigated for their potential as alternative biological tools to reduce grape and wine spoilage. However, such studies remain limited to a few genera. This work aimed to evaluate the antagonistic activity of grape must-derived non-conventional yeasts against Botrytis cinerea and non-Botrytis bunch-rotting moulds and to further elucidate mechanisms conferring antifungal activity. A total of 31 yeast strains representing 21 species were screened on different agar media using a dual culture technique and liquid mixed cultures, respectively. Pichia kudriavzevii was the most potent with a minimum inhibitory concentration of 10 2 cells/mL against B. cinerea but it had a narrow activity spectrum. Twelve of the yeast strains displayed broad antagonistic activity, inhibiting three strains of B. cinerea (B05. 10, IWBT FF1 and IWBT FF2), a strain of Aspergillus niger and Alternaria alternata. Production of chitinases and glucanases in the presence of B. cinerea was a common feature in most of the antagonists. Volatile and non-volatile compounds produced by antagonistic yeast strains in the presence of B. cinerea were analysed and identified using gas and liquid chromatography mass spectrometry, respectively. The volatile compounds identified belonged mainly to higher alcohols, esters, organosulfur compounds and monoterpenes while the non-volatile compounds were cyclic peptides and diketopiperazine. To our knowledge, this is the first report to demonstrate inhibitory effect of the non-volatile compounds produced by various yeast species., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Maluleke, Jolly, Patterton and Setati.)