Gut Microbiome Variation Across Menstrual Phases: Distinct Roles of Diet and Physiological Hormonal Fluctuations in Healthy Women

Abstract Sex differences in the gut microbiome are well established, yet the contribution of reproductive hormone variations within women remains insufficiently characterized. Here, we investigated how hormonal fluctuations during a natural menstrual cycle shape gut microbiome in healthy women and whether these effects are distinct from dietary influences. Leveraging longitudinal shot-gun gut microbiome data from 160 Italian participants in the Women4Health cohort, combined with detailed dietary information, we examined relationships with five reproductive hormones hormones: 17β-estradiol, progesterone, follicle-stimulating hormone (FSH), luteinizing hormone (LH) and prolactin. Microbial alpha diversity was significantly lower at the follicular phase compared to ovulatory phase (p = 0.03), while beta diversity significantly increased from follicular to the late luteal phase (p = 8.7x10 − 6 ). Twenty-eight species and twenty-two microbial pathways were significantly associated with reproductive hormones, with the majority of associations observed for gonadotropins (FSH and LH) rather than gonadal steroids (17β-estradiol and progesterone). While we confirm strong relationships between diet and microbiome, mediation analysis showed no evidence for a mediating effect of hormones on those. Together, these findings reveal distinct hormonal and dietary influences on gut microbiome across the menstrual cycle in healthy women.