Differential modulation of rumen function and bacterial communities in lambs by silage type and nitrogen source

The present study evaluated the effects of sweet sorghum silage as an alternative forage and slow-release urea as a non-protein nitrogen source on ruminal function and bacterial communities in lambs, as determined by enzymes activity, fermentation profiles, and microbial communities analysis of liquid and solid fractions of the collected digesta.

Forty-eight healthy male Hu lambs (24.11 ± 2.77 kg) were assigned to 4 groups (n = 12) based on a 2 × 2 factorial design: the type of dietary silage (corn silage, CS vs. sweet sorghum silage, SS) and nitrogen source (soybean meal, SM vs. slow-release urea, SRU). The treatments were identified as: CS-SM, CS-SRU, SS-SM, and SS-SRU. At the end of the trial, six lambs from each treatment group (n = 6) were selected by stratified random sampling for analysis of rumen fermentation, enzymes activity, and bacterial communities.

Compared to CS, SS significantly increased ruminal fibrolytic enzyme activities (carboxymethyl-cellulase, cellobiase, and xylanase) (P < 0.05). This enzymatic shift was associated with altered fermentation profiles, characterized by reduced total VFAs concentrations and an elevated acetate-to-propionate ratio (P < 0.05). In contrast, partial substitution of soybean meal (SM) with SRU increased fibrolytic activities (P < 0.05) and showed a tendency to elevate total VFAs (0.05 < P < 0.10), though it left microbial diversity unaffected (P > 0.05). SS significantly increased the Chao1 index in both solid and liquid fractions (P < 0.05), while the Shannon and Simpson indexes were significantly increased only in the solid fraction (P < 0.05). At the taxonomic level, 16 S rRNA sequencing revealed distinct patterns of modulation by the two dietary factors. The CS diet promoted Prevotella and Succiniclasticum but suppressed the Rikenellaceae RC9 gut group and Fibrobacter relative to SS. Interestingly, SRU further increased the relative abundance of Succiniclasticum compared to SM. Functional prediction suggested that this structural divergence corresponded to distinct metabolic potentials. Replacing CS with SS was predicted to increase the relative abundance of functional genes associated with catabolism and core metabolic processes (e.g., carbon metabolism and microbial metabolism in diverse environments) in both liquid and solid fractions (P < 0.05). Conversely, the CS diet was predicted to be enriched for anabolic functions, particularly within the solid fraction, where gene families related to amino acid biosynthesis, secondary metabolite production, and ABC transporters were significantly enriched (P < 0.05).

Ruminal responses to nitrogen appear to be substrate-dependent. While SRU positively influenced specific fermentation parameters, the structural remodeling of the microbial community was predominantly driven by substituting CS with SS. These results suggest distinct roles for dietary components: the forage base plays a dominant role in shaping microbial structure, whereas the nitrogen source modulates metabolic output.