Effects of repeated ovarian hyperstimulation on gut microbiota composition in a mouse model.

OBJECTIVE: This study investigated whether repeated cycles of controlled ovarian hyperstimulation (COH) influ-ence the gut microbiota composition in female mice. METHODS: Eight-week-old female Swiss mice (n=20; 10 per group) underwent ten weekly COH treatments with ex-ogenous gonadotropins or received saline injections as con-trols. One week after the final treatment, fecal samples were collected for gut microbiota profiling using 16S rRNA gene sequencing (V4 region amplification; primers 515F/806R) on the Illumina MiSeq platform. Sequencing data were pro-cessed in QIIME2 with DADA2 for denoising and Green-genes-based taxonomic assignment. Alpha-diversity (Shan-non, Simpson, observed OTUs, Faith's PD) and beta-diversity (Bray-Curtis, Jaccard, unweighted/weighted UniFrac) metrics were calculated. Differential taxonomic abundance and bio-marker analyses were performed using Random Forest and LEfSe. Statistical significance was set at p<0.05. RESULTS: No significant differences were observed between COH and control groups for alpha-diversity in-dices (all p>0.05) or beta-diversity distances (PERMANO-VA p>0.05). Relative abundances across major taxonomic levels were similar, although the Firmicutes:Bacteroidetes ratio was lower in COH-treated mice. Biomarker analyses identified Bacteroidales, Erysipelotrichaceae, and Parabac-teroides as enriched in the COH group, while Rikenellaceae and Odoribacter were more abundant in controls. CONCLUSION: Repeated COH did not significantly alter overall gut microbial diversity in mice under the conditions tested. However, specific taxonomic shifts and biomark-er signatures were detected, indicating associations-not causality-between COH exposure and changes in microbi-ota composition. Further studies are needed to determine the functional and physiological implications of these as-sociations.