Integrative multi-omics reveals comprehensive gut-liver-adipose metabolic changes in peak laying hens with high or low egg production.

Eggs serve as an indispensable global nutritional resource, sustaining the economic foundation of the commercial poultry industry. To meet this continuous demand, egg formation involves an exceptionally energy-intensive biological process requiring continuous yolk precursor synthesis, which imposes a massive metabolic burden on laying hens. However, the comprehensive metabolic differences across the gut-liver-adipose axis between peak laying hens with high or low egg production remain incompletely characterized. A total of 180 healthy Hy-Line Brown laying hens (45-week-old) were continuously fed and monitored for production performance over a 6-week period. Following the exclusion of individuals with extremely low egg production (≤10 eggs during the 6-week monitoring period; n = 5) and candidate hens that repeatedly produced unqualified eggs over multiple weeks (n = 20), the remaining hens were ranked by 6-week average laying rate and allocated into FH (high-production hens at 50 weeks of age, n = 15) and FL (low-production hens at 50 weeks of age, n = 14) groups. Initial body weight did not differ significantly between groups (FH: 1936 ± 29.0 g; FL: 1894 ± 48.2 g; P-value = 0.446). We integrated transcriptomic, untargeted metabolomic, targeted bile acid metabolomics, and microbiome (16S rRNA and metagenomic) profiles to characterize comprehensive metabolic changes across the gut-liver-adipose axis associated with divergent egg-production phenotypes. The results showed that: (1) FH hens exhibited higher serum APOB and lower conjugated bile acids (TCDCA, TCA, and THDCA), with hepatic upregulation of FASN, PPARA, CPT1A, and VTG1 along with downregulation of CYP7A1, CYP7B1, CYP8B1, and CYP27A1; (2) intersecting module hub genes (MHGs) with differentially expressed genes (DEGs) identified 354 upregulated and 299 downregulated core genes, with EEF2 identified as the primary hepatic downregulated hub gene; (3) in abdominal fat, GSEA revealed significant enrichment in fatty acid transport (NES = 1.54), long-chain fatty acid metabolic process (NES = 1.39), and steroid hormone biosynthesis (NES = 1.85), accompanied by significant downregulation of ANGPTL4 and upregulation of HSD3B1, VTG1, VTG2, and VTG3; (4) ileal mucosal transcriptomics identified 619 DEGs (502 upregulated), with GSEA highlighting enrichment in cell junction organization (NES = 1.52) and tube morphogenesis (NES = 1.39), which were further categorized into functional modules including enteric synaptic signaling, epithelial adhesion, mucosal vascularization, and tissue renewal; (5) the functional profile of the ileal microbiota in FH hens showed enrichment of functions related to complex carbohydrate degradation and carbohydrate-binding modules, with keystone taxa including Blautia and Bifidobacterium associated with production and lipid markers. Collectively, these findings suggest that high egg production during the peak laying period is associated with coordinated metabolic differences across the liver, abdominal fat tissue, and intestine. The observed profiles included reduced hepatic translation-related and primary bile acid synthesis-related signatures, adipose endocrine-related changes and microbial functional potential related to carbohydrate utilization and antioxidant-related functional potential. These findings provide candidate multi-omics features for precision nutritional strategies and genetic improvement in commercial poultry.