BACKGROUND

Despite advancements in artificial insemination, sheep fertility rates remain suboptimal. Recent studies in other species highlight the critical role of reproductive microbiota in influencing fertility outcomes. This research explores the relationship between ovine vaginal microbiota, associated functional pathways, and fertility using advanced nanopore long-reading metagenomic sequencing on 297 ewes from three Spanish breeds across four herds. The study aimed to describe a core vaginal microbiota, analyse the complex interactions with herd, breed, age, and parity factors, and identify taxa and genes associated with reproductive success by artificial insemination.

RESULTS

The study identified Staphylococcus, Escherichia, and Histophilus as the most abundant genera. Microbial communities varied considerably between breeds and herds, with high predictive accuracy (> 90%) in classification models. Differential abundance analysis revealed that the genera Histophilus, Fusobacterium, Bacteroides, Campylobacter, Streptobacillus, Gemella, Peptoniphilus, Helococcus, Treponema, Tissierella, and Phocaeicola were more abundant in non-pregnant ewes. Some of these taxa were also associated with four COG entries and one KEGG orthologue significantly linked to non-pregnancy, primarily involving carbohydrate metabolism, defence mechanisms, and structural resilience. Age and parity were also associated with microbiota composition, particularly in ewes older than five years or with more than three parturitions, suggesting that cumulative physiological changes may contribute to microbial shifts over time.

CONCLUSIONS

The ewe's vaginal microbiome appears to be mainly influenced by both herd and breed, though distinguishing genetic from environmental factors is challenging within our study design. While the overall microbiota showed a subtle effect on pregnancy, certain genera had a significant negative impact, likely due to pathogenic or inflammatory properties that disrupt reproductive health. The metagenomic approach used here enabled not only comprehensive taxonomic classification but also detailed functional analysis, providing deeper insights into the microbiome's role in reproductive outcomes. Video Abstract.