Ensiled diet improved the growth performance of Tibetan sheep by regulating the rumen microbial community and rumen epithelial morphology

Abstract The aim of this study was to investigate the effects of ensiled agricultural byproducts from Qinghai-Tibet plateau on growth performance, rumen microbiota, ruminal epithelium morphology and nutrient transport-related gene expression in Tibetan sheep. Fourteen male Tibetan sheep were randomly assigned to one of two diets: an untreated diet (without silage inoculum, CON, n=7) or an ensiled diet (with silage inoculum, ESD, n=7). The total experimental period lasted for 84 days, including early 14 days as adaption period and remaining 70 days for data collection. The ESD increased average daily gain (ADG, P = 0.046), dry matter intake (P < 0.001), ammonia nitrogen (NH3-N, P = 0.045), microbial crude protein (MCP, P = 0.034) and total volatile fatty acids (VFA) concentration (P < 0.001), and decreased ruminal pH value (P = 0.014). The proportion of propionate (P = 0.046) and the copy numbers of bacteria (P = 0.01) and protozoa (P = 0.002) were higher, while the proportion of acetate (P = 0.028) was lower in the sheep fed ESD compared to CON. Pyrosequencing of the 16S ribosomal RNA gene revealed that ESD increased the relative abundance of Firmicutes, Ruminococcus, Lachnospiraceae_AC2044_group, Lachnospiraceae_XPB1014_group, Christensenellaceae_R-7_group in the rumen (P < 0.05), while decreased the relative abundance of Bacteroidota, Prevotellaceae_UCG-003, and Veillonellaceae_UCG-001 (P < 0.05). Analyses with PICRUSt2 and STAMP indicated that the propionate metabolism pathway was enriched in the sheep fed ESD (P = 0.026). The ESD increased the rumen papillae height (P = 0.012), density (P = 0.036), and surface area (P = 0.001), and improved the thickness of the total epithelia (P = 0.018), stratum corneum (P = 0.040), stratum germinativum (P = 0.042), and stratum spinosum and basale (P = 0.004). The relative mRNA expression of Cyclin-dependent Kinase 2 (CDK2), CyclinA2, CyclinD2, zonula occludens (ZO-1), Occludin, monocarboxylate transporter isoform 1 (MCT1), monocarboxylate transporter isoform 4 (MCT4), sodium/potassium pump (Na+/K+-ATPase), and sodium/hydrogen antiporter 3 (NHE3) were higher in the rumen epithelial of sheep fed ESD than CON (P < 0.05). Conversely, the relative mRNA expressions of Caspase 3 and B-cell lymphoma-2 (Bcl-2) were lower in the sheep fed ESD than CON (P < 0.05). In conclusion, compared with untreated diet, feeding an ensiled diet altered the rumen microbial community, enhanced nutrient transport through rumen epithelium, and improved the growth performance of Tibetan sheep.