Antagonistic yeasts suppress plant pathogenic fungi by various mechanisms, but their biocontrol efficacy also depends on the ability to compete and persist in the environment. The goal of the work presented here was to quantify the composition of synthetic yeast communities in order to determine the competitiveness of different species and identify promising candidates for plant protection. For this purpose, colony counting of distinct species and matrix‐assisted laser desorption ionization time‐offlight mass spectrometry (MALDI‐TOF MS; MALDI biotyping) were used to distinguish different yeast species and to quantify the composition of a synthetic community of six yeasts (Aureobasidium pullulans, Candida subhashii, Cyberlindnera sargentensis, Hanseniaspora sp., Metschnikowia pulcherrima and Pichia kluyveri) over time, on apples and in soil, and in different growth media. These studies revealed important characteristics that predispose the different species for particular applications. For example, the competitiveness and antagonistic activity of C. subhashii was strongly increased in the presence of N‐acetylglucosamin as the sole carbon source, M. pulcherrima and A. pullulans were the strongest competitors on apple, and C. sargentensis competed the best in soil microcosms. Based on these laboratory studies, M. pulcherrima and A. pullulans are promising candidates for biocontrol applications against fungal phyllosphere diseases, while C. sargentensis may hold potential for use against soilborne fungal pathogens. These results document the potential of MALDI‐TOF MS for the quantitative analysis of synthetic yeast communities and highlight the value of studying microorganisms with relevant functions in moderately complex, synthetic communities and natural substrates rather than as individual isolates.
