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Steroid hormones are one of the presumed modulators of Lactobacillus abundance in the vaginal epithelium. We set out to characterize the vaginal microbiome (VMB) and also provide an in-depth understanding of the relative contribution of estradiol (E2) and progesterone (P1) in shaping the vaginal microbiome of Nigerian women (n = 38) who experienced both uncomplicated term delivery and preterm delivery using samples longitudinally collected during pregnancy (17–21, 27–31, 36–41 weeks gestation) and 6 weeks postpartum. Vaginal swabs and blood samples were aseptically collected. Vaginal swabs were used for microbiome assessment using 16S ribosomal RNA (rRNA) gene sequencing. Blood samples were used for hormonal measurement using a competitive-based enzyme-linked immunosorbent assay (ELISA). Across several maternal covariates, maternal age, pregnancy status and delivery mode were not significantly associated with the vaginal microbiota whereas maternal E2 level (pE2 = 0.006, Omnibus), and P1 level (pP1 = 0.001, Omnibus) were significantly associated with the vaginal microbiome. E2 and P1 concentrations increased throughout pregnancy commensurately with increasing proportions of L. crispatus (pE2 = 0.036, pP1 = 0.034, Linear Mixed Model). An increasing trend of α-diversity was also observed as pregnancy progressed (pobserved ASV = 0.006, LMM). A compositional microbiome shift from Lactobacillus profile to non-Lactobacillus profile was observed in most postnatal women (pCST IV < 0.001, LMM). Analysis of our data shows a species-specific link between pregnancy steroid hormone concentration and L. crispatus abundance. [ABSTRACT FROM AUTHOR]