Dietary supplementation of concentrate improves lactation performance and immune function in grazing lactating yaks through regulating rumen microbes and metabolites

Grazing yaks often face protein deficiency due to the low nutritional quality of natural pasture, which limits milk production during lactation. This study aimed to investigate the effects of varying protein levels in concentrate supplementation on lactational performance, immune function, and rumen microbial and metabolites in grazing lactating yaks. Thirty-six lactating yaks were randomly assigned to four treatments: a natural grazing control group (CON) and three concentrate-supplemented groups receiving low- (CP15, 15.09% CP), medium- (CP17, 17.00% CP), or high-protein concentrate (CP19, 18.98% CP), with 1.5 kg/d of concentrate provided per yak, the experiment lasted 70 days, including a 10-day adaptation period and a 60-day experimental period. Concentrate supplementation significantly increased ADG and milk yield, and milk yield showed a linear increase with higher dietary protein levels (P < 0.001). Milk composition, including fat, protein, and lactose also improved across supplementation groups, with the greatest enhancement observed at the CP17 group (P < 0.05). Immune function, indicated by platelet count, exhibited both linear and quadratic responses to protein levels as reflected by platelet count, also showed significant improvements with increased protein levels, with both linear and quadratic effects (P = 0.014; P = 0.015).The microbial structure of rumen was affected by concentrate supplementation (P = 0.036), resulting in enrichment of genera Succinivibrionaceae_UCG-002, Fibrobacter, Ruminobacter, and Succinimonas in the CP17 cows (P < 0.05), whereas the abundance of Saccharofermentans decreased (P < 0.05). Metabolomics analysis showed that concentrate supplementation resulted in a significant modification in the ruminal metabolites profile. Compared to CON lactating yaks, ruminal calcium propionate, 2-nitrofuran, curvulalide, and 2,5-dihydroxybenzoic acid were greater in CP17 lactating yaks, whereas the ruminal 4-Pyridoxic acid, L-Carnitine, bitocholic acid, and taurodehydrocholic acid were less in lactating yaks. Predicted pathway analysis suggested that vitamin B6 metabolism was significantly enriched. Overall, these results indicated that moderate protein concentrate supplementation (CP17) could improve lactation performance by modulating rumen microbiome and metabolism in lactating cows.