<b>Dietary Supplementation Changes Gut Microbiome and Urinary VOCs in Male Asiatic Black Bears Influencing Female Mate Choice Preferences</b>

Chemosensory cues inform mammalian mate selection by signaling genetic quality, social status, and reproductive fitness. Gut microbiota impact host odor through the metabolic regulation of volatile organic compounds (VOCs), but effects on chemosensory-driven mate choice remain underexplored. Here we supplemented the diets of two groups of captive male Asiatic black bears (U. thibetanus) with either antibiotic (Anti-treated, n = 10) or dietary supplement yeast tablets (DS-treated, n = 10). Urine and fecal samples were collected concurrently before and after the treatment. Urinary VOCs were analyzed using GC-MS, while gut microbiome community structure was assessed via 16S rRNA sequencing. In male DS-treated bears, the relative abundance of Sarcina and Vagococcus bacteria declined significantly (P &lt; 0.05) compared to pre-treatment, although with no bacterial α-diversity change. Urinary VOCs exhibited distinct clustering between pre- and post-DS treatment groups (R²Y = 0.95, Q²= 0.422). Two-choice behavioral tests showed that estrous females sniffed urine samples from DS-treated males significantly more often (P = 0.018) and longer (P = 0.034) than pre-treatment samples, although no response was observed to pre- and post-treatment male urine in the Anti-treated group. The relative abundances of four urinary VOCs were significantly and positively correlated with females’ sniffing frequency and duration for both male groups (both P &lt; 0.05). Co-occurrence network analysis demonstrated robust covariation between gut microbiota and urinary VOCs (both P &lt; 0.05). Our results evidence the role of the gut microbiome in mediating mate choice via urinary odor cues, with implications for ex-situ captive breeding programs.