Comparative analysis of four compound stomachs development, fermentation dynamics, and microbiome structure in Hu sheep and its crossbred combinations

Objective:This study was conducted to investigate the differences in fermentation parameters, epithelial development, and microbial composition across the four stomach compartments between Hu sheep and its crossbreds, aiming to provide key references for targeted modulation of gastrointestinal tract function and improvement of ruminant feed conversion efficiency via crossbreeding.Methods: Forty-eight approximately 3-month-old male lambs, assigned to three genetic groups (HH:♂Hu×♀Hu, n=16; DH:♂Poll Dorset×♀Hu, n=16; SH:♂Southdown×♀Hu, n=16), were raised under uniform nutritional and management conditions for 95 d.Feed intake was determined daily and body weight were measured every 20 d.At the end of the trial, six sheep per group close to the average weight were slaughtered.Samples of digesta and tissue from the four stomachs were collected.Fermentation parameters, epithelial development, and microbial community structure were analyzed.Results: Compared with the HH group (acetate: 25.8, 21.8, and 6.36 mmol/kg; propionate: 6.8, 5.60, and 3.65 mmol/kg; butyrate: 3.66, 3.19, and 1.38 mmol/kg), the DH group exhibited significantly higher acetate concentrations (45.3, 33.7, and 12.9 mmol/kg), propionate (14.3, 11.1, and 5.94 mmol/kg), and butyrate (12.9, 6.69, and 4.52 mmol/kg) in the rumen, reticulum, and omasum, respectively, indicating enhanced fermentation capacity, a significantly thicker rumen mucosal epithelium suggesting a greater capacity for VFA absorption.Taxonomic analysis showed that Firmicutes and Bacteroidota were the predominant phyla in four stomachs.Differential analysis identified that the DH group had significantly enriched taxa associated with fiber degradation (e.g., Fibrobacterota, Spirochaetota) in the forestomachs.Conversely, the rumen of HH group was enriched with Anaerolineae, associated with complex plant fiber degradation.PICRUSt2 functional prediction indicated that differentially microbes in the DH group were primarily linked to the biosynthesis of lysine and arginine, whereas those in the HH group were enriched in degradation pathways such as amino acid degradation.Conclusion:The DH group markedly elevated VFA production by modulating the forestomach microbial structure.Combined with adaptive improvements in rumen epithelial, this effect potentially enhanced energy utilization and growth performance.