Investigating the Dynamic Variation of Skin Microbiota and Metabolites in Bats During Hibernation

<i>Pseudogymnoascus destructans</i> (<i>Pd</i>) invades the skin tissue of bats, leading to severe population declines. The skin microbiome plays a crucial role in protecting hosts from fungal infection and exhibits pronounced spatiotemporal dynamics in its structure and function. Meanwhile, metabolites derived from microbial communities reflect the host physiological state and participate in microbe-pathogen interactions. In this study, we investigated the spatiotemporal dynamics of skin bacterial communities and metabolites during hibernation in <i>Rhinolophus ferrumequinum</i> by integrating 16S rRNA sequencing with untargeted metabolomics and experimentally verified the antifungal effects of microbially derived potential metabolites against <i>Pd</i>. Our results revealed that the structure of the skin bacterial community varied significantly across sampling contexts, with its assembly primarily governed by stochastic processes. Bacterial diversity reached its lowest level during middle hibernation, accompanied by a simplified co-occurrence network dominated by cooperative or mutualistic interactions. Additionally, metabolomic analyses demonstrated systematic metabolic remodeling of bat skin across hibernation stages, marked by significant enrichment of multiple pathways closely involved in host antimicrobial defense. Furthermore, metabolite profiles differed across locations, and the abundance patterns of several metabolites were strongly correlated with <i>Pd</i> infection levels. Integrated analyses identified multiple metabolites that showed significant correlations with bacterial genera capable of synthesizing the corresponding compounds. In vitro validation confirmed that nine metabolites effectively inhibited the growth of <i>Pd</i>, among which melatonin exhibited the strongest antifungal activity. Collectively, this study reveals the dynamics of the skin microbiome and metabolites of <i>R. ferrumequinum</i> during hibernation, providing novel insights into the defensive role of skin-associated microbes and metabolites in maintaining population health and resilience against fungal pathogens.