P-705 Non-Absorbable Antibiotic Exposure Impairs Ovarian Function and Fertility in Female Mice

Abstract Study question Does the gut microbiome homeostasis directly affect ovarian function and fertility in female mice? Summary answer By using non-absorbable antibiotic (nABX), we found absence of gut microbiome disrupts sex hormones and estrous cycles, triggers ovary dysfunction and fertility in female mice. What is known already The gut microbiome plays a crucial role in regulating host metabolism, endocrine function, and reproductive health. Antibiotic treatments have been shown to induce dysbiosis and metabolic imbalance. Our recent meta-analysis of over 1.2 million participants demonstrated that preconception antibiotic use is associated with increased risks of infertility. However, the mechanisms underlying the gut–ovary crosstalk remain poorly understood. Whether gut microbiome homeostasis directly governs ovarian function is still under-investigated. Study design, size, duration C57BL6/J female mice at 8 to 10 weeks old were assigned to either an nABX group (n = 8) treated with non-absorbable antibiotic cocktails (neomycin, 2.5 mg/mL; bacitracin, 2.5 mg/mL; and pimaricin, 1.25 μg/mL; adding in drinking water) or a control group receiving pure water (n = 10) for three weeks. We repeated two rounds of experiments to determine different outcomes. Participants/materials, setting, methods The estrous cycle was assessed through vaginal cytology. Both control and antibiotic-treated female mice were mated with proven fertile males to evaluate pregnancy rates. Fecal samples were collected for 16S rRNA gene sequencing to analyze gut microbiota composition. Following anesthesia, blood samples were obtained to determine circulating sex hormone levels. Ovaries were harvested for morphological assessment and bulk mRNA sequencing to investigate transcriptional changes associated with antibiotic-induced reproductive dysfunction. Main results and the role of chance Estrous cycle monitoring showed pronounced cycle irregularities in nABX group, as evidenced by prolonged diestrus. Moreover, serum level of Follicle-stimulating hormone (FSH) levels increased by 2-fold compared to controls (p < 0.01), indicating potential hypothalamic-pituitary-ovarian axis dysregulation. Importantly, nABX-treated mice were almost infertile and exhibited about 20% decrease in ovarian weight compared with controls (p < 0.01). Hematoxylin and eosin (H&E) staining of ovarian sections revealed a marked increase in the number of atretic follicles, a reduction in ovarian reserve, and fewer corpora lutea, suggesting impaired folliculogenesis and ovulatory dysfunction. mRNA sequencing analysis demonstrated that nABX treatment significantly altered in pathways related to sex hormone endocrinology, including aldosterone synthesis and secretion, thyroid hormone synthesis, and parathyroid hormone synthesis, secretion, and action (p < 0.05). These findings suggest that gut microbiota perturbation via antibiotic treatment significantly disrupted ovarian function and endocrine regulation in female mice. Limitations, reasons for caution This study is a preliminary investigation in animal models. Fecal microbiota transplantation from untreated mice to the nABX group is necessary to establish causality between gut microbiome disruption and ovarian dysfunction. Further explorations are required to identify the key effector and elucidate the specific microbial pathways and molecular mechanisms involved. Wider implications of the findings These findings suggest that gut microbiome perturbations can profoundly impact ovarian function and fertility in female mice. Of note, this study highlights a potential risk of antibiotic treatment on female fertility in clinical settings, underscoring the need for cautious antibiotic use and microbiome-targeted interventions for reproductive health. Trial registration number No