Alleviation of ovarian ferroptosis and dysfunction induced by gut microbiota dysbiosis in laying hens treated with Lactobacillus paracasei R8

Declining egg production in hens is often accompanied by ovarian dysfunction. Emerging evidence suggests a link between intestinal abnormalities and reproductive endocrine disorders, yet whether gut dysbiosis contributes to this process via ferroptosis and mitochondria-associated pathways remains unclear. Here, we show that low-producing hens exhibit intestinal villus atrophy and downregulation of tight junction proteins in both ileum and ovary, accompanied by elevated inflammatory cytokines, impaired antioxidant defense, and aberrant expression of ferroptosis-related genes, indicating a coordinated disruption of intestinal–ovarian homeostasis in inflammation and iron metabolism. 16S rRNA sequencing revealed a markedly increased abundance of Lactobacillus species in high-producing hens, correlating positively with sex hormone levels and laying performance, whereas these bacteria were depleted in low-producing hens. Dietary supplementation with Lactobacillus paracasei R8 ( L. paracasei R8) improved laying performance, alleviated intestinal and ovarian injury, and attenuated ferroptosis-associated alterations. In LPS-induced intestinal epithelial and ovarian cell models, L. paracasei R8 improved barrier function, increased mitochondrial membrane potential, reduced lipid peroxidation and intracellular Fe2⁺ accumulation, exerting effects comparable to the ferroptosis inhibitor Fer-1. Co-treatment with the mitochondrial inhibitor CsA partially abrogated these effects, implicating mitochondrial regulation in the mechanism of action. Taken together, these findings identify an intestinal–ovarian injury pattern in low-producing hens associated with gut microbiota imbalance and iron-dependent cell death, and suggest that L. paracasei R8 may serve as a potential probiotic intervention to improve laying performance by alleviating gut–ovary-related pathological alterations through modulation of mitochondrial function and ferroptosis-related pathways.