Unveiling the culturable and non-culturable yeast biodiversity in chaste honey produced by the sympatric Apis cerana and Apis mellifera from eastern China.

Honey's unique physicochemical properties create a restrictive environment for most microorganisms, yet support the specialized osmotolerant yeasts with significant ecological roles and biotechnological potential. In this study, we employed an integrated culture-dependent and culture-independent approach to systematically characterize yeast communities in sympatric monofloral chaste (Vitex agnus-castus) honeys from Apis cerana and Apis mellifera colonies in Qingdao, East China. Results consistently showed that A. mellifera honey harbored significantly higher yeast diversity than A. cerana honey across both methods. Culture-independent profiling identified Zygosaccharomyces as the predominant genus in both honey types, while A. mellifera honey contained a notably higher proportion of unclassified fungal taxa. Through cultivation, 13 distinct yeast species and one filamentous fungus were isolated from the honey samples, with several species exclusively associated with either A. cerana or A. mellifera honey. Importantly, despite the shared geographic and floral source, both approaches confirmed a clear bee species-specific yeast community structure, highlighting that host identity drives microbial differentiation. Physiological assays further demonstrated that all isolated yeasts exhibited exceptional tolerance to high osmotic stress and acidic pH-key traits adaptive to the extreme environment of honey. These adaptive characteristics, coupled with diverse metabolic capabilities, highlight the substantial biotechnological potential of these honey-associated yeasts, with promising applications in fermentation, bioactive metabolite synthesis, and probiotics. This study enhances our understanding of host-specific microbial associations in honey ecosystems and positions honey as a valuable reservoir of functionally diverse yeasts for biotechnological exploration.