Limosilactobacillus reuteri enhances production performance and ovarian function via modulation of the gut-ovary axis in laying hens

BACKGROUND: Declines in nutrient absorption and reproductive function following peak laying significantly compromise egg production and quality in hens. Bacterial and enzymatic co-fermented feed is a promising intervention to improve gut health and production metrics. However, its mechanism of action remains unclear. RESULTS: Bacterial and enzymatic co-fermented feed supplementation in laying hens significantly enhanced egg production, egg weight, and feed intake while reducing the feed conversion ratio. Egg quality improved markedly, denoted by an increase in albumen height, the Haugh unit, and eggshell strength and thickness. The apparent digestibility of calcium in laying hens increased. Although the tibial growth indices remained unchanged, bone microarchitecture improved, evidenced by increased trabecular mineralisation and a reduction in the number of osteoclasts. Intestinal integrity was strengthened based on an elevated villus/crypt ratio, upregulation of barrier genes (ZO-1, Claudin-1, Occludin), increased IL-10 expression, and decreased IL-6 and TNF-α expression. Bacterial and enzymatic co-fermented feed reshaped the caecal microbiota, enriching Limosilactobacillus. This genus was also enriched in the oviduct and ovary. SourceTracker analysis revealed a caecum-oviduct-ovary microbial transmission route. In the ovary, bacterial and enzymatic co-fermented feed enhanced the expression of FSHR, AMH, and antioxidant genes (Nrf2, SOD1, CAT, GPX3, and HO-1); upregulated Bcl-2; and downregulated Caspase-3 and Caspase-8, supporting improved follicular health and antioxidant defences. CONCLUSIONS: This study is the first to clarify the regulatory mechanism of bacterial and enzymatic co-fermented feed on laying performance of laying hens from a microbial perspective. The co-fermented feed improved the laying performance, egg quality and tibial microarchitecture of laying hens by optimizing the microbial composition of the cecum, oviduct and ovary, thus delaying ovarian aging and enhancing ovarian function. Furthermore, we found the ovarian microbial composition of laying hens was affected by the oviduct microbiota, which was in turn influenced by the intestinal and cloacal microbiota, providing experimental evidence for the gut-oviduct-ovary axis in laying hens and a new perspective for exploring the correlation between feed nutrition and reproductive performance of laying hens. Video abstract.