Pregnancy-associated vaginal microbiome profiling detects Brucella reactivation in vaccinated pregnant ewes

The vaginal microbiome influences reproductive health and infection susceptibility in livestock, yet its dynamics during gestation, particularly regarding intracellular pathogens like Brucella , remain underexplored. Live attenuated vaccine candidates 16MΔ wzm and Rev1Δ wzm exhibit differing safety profiles in pregnant ewes. Here we longitudinally profiled vaginal microbiomes (VM) of pregnant ewes vaccinated with attenuated B. melitensis strains 16MΔwzm or Rev1Δwzm, using 16S rRNA (V3–V4) amplicon sequencing. Brucella was undetectable during pregnancy but reactivated at parturition in ewes vaccinated with 16MΔwzm, coinciding with significant vaginal microbiome disruption. In contrast, Rev1Δwzm-vaccinated ewes maintained stable VM similar to unvaccinated controls, with no Brucella detected. These findings provide a proof of concept that VM profiling accurately detects Brucella in shedder sheep, supporting the reactivation of 16M∆wzm infection at peripartum and the superior safety of Rev1Δwzm in pregnant ewes, as previously reported. Although studies under field conditions with larger cohorts are necessary, this non-invasive approach offers potential for monitoring Brucella infection dynamics in natural hosts, aiding brucellosis control within a sustainable One Health framework.