Altering Dietary Soluble Protein Levels With Decreasing Crude Protein May Be a Potential Strategy to Improve Nitrogen Efficiency in Hu Sheep Based on Rumen Microbiome and Metabolomics

Abstract

Ruminants account for a relatively large share of global nitrogen (N) emissions. It has been reported that nutrition control and precise feeding can improve the N efficiency of ruminants. The objective of the study was to determine the effects of soluble protein (SP) levels in low-protein diets on growth performance, nutrient digestibility, rumen microbiota, and metabolites, as well as their associations of N metabolism in fattening Hu sheep. Approximately 6-month-old, 32 healthy fattening male Hu sheep with similar genetic merit and an initial body weight of 40.37 ± 1.18 kg were selected, and divided into four groups (n = 8) using the following completely randomized design: the control diet (CON) with a 16.7% crude protein (CP) content was prepared to meet the nutritional requirements of fattening sheep [body weight (BW): 40 kg, average daily gain (ADG): 200–250 g/d] according to the NRC recommendations; other three include low protein diets (LPA, LPB, and LPC) of CP decreased by ~10%, with SP proportion (%CP) of 21.2, 25.9, and 29.4 respectively. The feeding trial lasted for 5 weeks including the first week of adaptation. The results showed no difference in the growth performance (P > 0.05); DM and CP digestibility were higher in LPB and LPC, with maximum organic matter digestibility in LPB (P < 0.05). Low-protein diets decreased serum urea-N whereas urinary urea-N was lower in LPB and LPC (P < 0.05), while N retention and the biological value of N were higher in LPB and LPC (P < 0.05). Ruminal NH3-N concentration in LPA and LPB was low than CON (P < 0.05), while total volatile fatty acid (TVFA), acetate, propionate, and butanoate were all lowest in LPA (P < 0.05). In the rumen microbiome, LPB increased the community richness in Prevotellaceae and Prevotella_1 (P < 0.05); Metabolomics analysis revealed low-protein diets downregulated the amino acid metabolism pathways, while the biosynthesis of unsaturated fatty acids along with vitamin B6 metabolism were upregulated with increased SP. These findings could help us understand the role of different SP levels in the regulation of rumen microbial metabolism and N efficiency. Overall, low-protein diets (CP decreased by ~10%) can reduce serum urea-N and ruminal NH3-N without affecting the growth performance of fattening Hu sheep. Additionally higher N efficiency was obtained with an SP proportion of ~25–30%.

Regulation of Dietary Protein Solubility Improves Ruminal Nitrogen Metabolism In Vitro: Role of Bacteria–Protozoa Interactions

“…Regulation of dietary protein fractions is one of the important nutritional intervention strategies to improve nutritional metabolism in ruminants. Our previous in vivo experiments have shown that SP levels (% of CP) of ∼25–30% have higher N efficiency with dietary CP content decreased by∼10% in Hu sheep [ 10 ]. In this study, as time progressed, the pH value of each treatment gradually decreased and fluctuated from 6.85 to 5.90, which was mainly due to the decomposition and utilization of substrate nutrients by rumen microorganisms and the accumulation of organic acids [ 30 ].…”

“…All feeds were sampled by quartering and drying at 65 °C to a constant weight, and then they were ground to pass through a screen (1 mm) by a Retsch ZM 100 Wiley mill (Retsch GmbH, Haan, Germany) and stored. The SP content of six feeds was determined by following buffer-soluble N protocols in the standardization of procedures for N fractionation [ 18 ], which was described concretely in our previous study [ 10 ], and specific protein fractions (CP and SP values) are reported in Table S1 .…”

“…In total, 5 mL of each culture medium was collected, and the pH value was measured immediately with a pH meter (pHS-3C, Shanghai, China). Ammonia N (NH 3 -N) concentration was measured according to Broderick and Kang (phenol-hypochlorite system) [ 21 ], microbial protein (MCP) content was determined by the purine prediction method [ 22 ]; gas chromatography (GC-14B, Kyoto, Japan) was used for determination of volatile fatty acids (VFAs), with crotonic acid as the internal standard [ 23 ], and specific procedure settings referred to our previous publication [ 10 ]. The substrate residues at each time point were collected to detect dry matter digestibility (DMD) and N digestibility (ND).…”

“…Feed nutrient composition is considered an important reference strategy to improve N retention and reduce N emission in ruminant production, e.g., using low-protein diets [ 8 ] or adjusting the rumen degradable to non-degradable protein ratio with a non-soluble to soluble carbohydrates ratio in diets [ 9 ]. Our previous study on Hu sheep indicated that when reducing crude protein (CP) content as well as adjusting soluble protein (SP) to 25–30%, N efficiency was increased [ 10 ]. Stefanski et al, isolated different protein fractions from rapeseed meal and demonstrated a different efficiency of ruminal N metabolism of these fractions in vivo [ 11 ].…”

“…However, diet-microbiome interactions in the context of SP are rarely reported. Our previous research indicated that altering dietary SP levels to ~25–30% with decreasing CP increased the relative abundance of Prevotellaceae and Prevotella_1 in the rumen [ 10 ]. Ciliate protozoa accounting for an estimated 25% to 50% of microbial biomass in the rumen are also involved in the breakdown of nitrogenous substances, especially insoluble proteins [ 13 ].…”