Multi-site characterization of reproductive tract and gut microbiota in unexplained recurrent pregnancy loss: a case control study

Abstract This cross-sectional case-control study aimed to characterize microbial communities across multiple anatomical sites in women with unexplained recurrent pregnancy loss compared with healthy controls. Forty women aged 18–40 years, including 20 with unexplained recurrent pregnancy loss and 20 healthy controls, were enrolled at Samson Assuta University Hospital in Ashdod, Israel. Swab samples from the cervix, posterior fornix, lower vagina, and feces were analyzed using 16S ribosomal ribonucleic acid amplicon sequencing. Microbial community composition, diversity, differences in taxon abundance, and concordance between anatomical sites were assessed. Microbial communities were dominated by Lactobacillus across all reproductive-tract sites in both groups. Faith’s phylogenetic diversity was significantly higher in women with recurrent pregnancy loss in the cervix (P=0.0139) and posterior fornix (P=0.0492), whereas Shannon diversity did not differ between groups. Beta-diversity analysis revealed significant compositional separation at the posterior fornix (R2=0.104, P=0.029). The Lactobacillus to Bifidobacterium log-ratio was significantly lower in women with recurrent pregnancy loss in the posterior fornix (P=0.037), with similar trends at other reproductive-tract sites. No individual taxa reached significance after multiple-testing correction. Microbial communities showed strong concordance among reproductive-tract sites and modestly greater gut–genital concordance in women with recurrent pregnancy loss. These findings suggest that unexplained recurrent pregnancy loss is associated with subtle, site-specific microbial differences, with the posterior fornix showing the clearest differences between groups.