The Gut-Ovarian Axis Regulates Clutch Persistence in Aging Laying Hens via Lipid Mediator Networks

Clutch persistence, the ability to sustain consecutive egg-laying cycles, is a critical determinant of profitability in the poultry industry, especially for aging laying hens (≥65 weeks). However, its underlying regulatory mechanisms remain poorly understood, largely due to limitations of the traditional “ovary-centric” research paradigm, which ignores the long-range regulatory effects of intestinal microbiota on ovarian metabolism. To address this knowledge gap, the present study aimed to dissect the comprehensive regulatory network governing clutch persistence using integrated multi-omics analyses. A total of 20 sixty-five-week-old Rhode Island Red (RIR) laying hens with cumulative egg production exceeding 300 eggs but distinct clutch persistence was stratified into high-persistence group (DLCD, ≥25 clutches, n=10) and low-persistence group (DLCS, ≤15 clutches, n=10). Multi-omics profiling, including ovarian transcriptomics, proteomics, and metabolomics, serum metabolomics and cecal microbiota 16S rRNA sequencing was performed. Data integration and association mining were conducted via Spearman correlation analysis with stringent thresholds (r > 0.6, P < 0.01). Integrated analyses revealed a “gut-ovary axis” regulatory model mediated by a lipid mediator network, operating through a three-tiered mechanism: 1) Gut Initiation: The DLCD group exhibited enriched cecal γ-Proteobacteria promoted biosynthesis of lipid precursors. 2) Serum Transport: Key serum lipid mediators, most notably LysoPC (22:6) (VIP = 4.5) and cholesterol ester CE (20:4)—served as critical carriers transducing gut-derived signals to the ovary. 3) Ovarian Execution: These lipid signals activated a core ovarian metabolic pathway centered on the PLA2G6-ALOX15B-AGPAT3 axis, which coordinated follicular development and ovulation by supplying steroid hormone synthesis substrates, exerting anti-inflammatory effects, and stabilizing membrane structures. Collectively, this study demonstrates that gut microbiota modulates clutch persistence in aging laying hens via lipid mediators, orchestrating a systemic “gut-serum-ovary” regulatory cascade. These findings provide a novel theoretical framework for extending the economic egg-laying cycle through targeted manipulation of intestinal microbiota or serum lipid metabolism.