Background: Intramuscular fat (IMF) is a key determinant of meat sensory quality, but the effects of rumen-protected betaine (RPB) on IMF deposition in Tibetan sheep remain unclear. Methods: Sixty male, 3-month-old Tibetan lambs (Initial weight 17.72 ± 1.36 kg) were randomly assigned to a control (basal diet, n = 30) or RPB supplementation (basal diet supplemented with 0.08% RPB, n = 30) for 90 d. Results: Transcriptomic and lipidomic analyses revealed that RPB modulated glycerophospholipid metabolism-related genes (DGKI and AGPAT1) and altered lipid profiles, including PC (16:0/20:5) and PC (16:0/16:0). RPB increased T-AOC and SOD activities and decreased MDA content (p < 0.05). Intramuscular adipocyte area and diameter increased, while density decreased (p < 0.05). LC-MS/MS analysis showed that unsaturated fatty acids C14:1, C17:1, and C22:4 were significantly increased (p < 0.05). Muscle fiber density in the longissimus dorsi increased, whereas shear force, cooking loss, hardness, and chewiness were reduced (p < 0.05). MYH1, MYH2, and MYH7 expression were upregulated by 2.05-, 1.89-, and 2.73-fold, respectively. Conclusions: In summary, dietary RPB supplementation was associated with increased IMF deposition, which may contribute to improved meat quality, potentially through modulation of glycerophospholipid metabolism.

Castration is a vital technical practice in Tibetan sheep farming on the Qinghai-Tibet Plateau. It reduces fighting among rams, making it easier to manage the herd while they are grazing. However, the impact of castration on the quality of Tibetan sheep meat and muscle metabolites is unclear at present. On this basis, this study examined differences in meat quality and metabolites between castrated (BTC) and non-castrated (BTN) Black Tibetan sheep using targeted and non-targeted metabolomics. Compared with the BTN group, the meat from the BTC group showed reduced drip loss and increased cooking loss, as well as higher levels of elasticity, chewiness and cohesiveness (P < 0.05). The levels of 11 amino acids, total amino acids, total umami amino acids, 23 fatty acids, SFA, PUFA and MUFA were significantly higher in the BTC group than in the BTN group (P < 0.05). A total of 34 differentially abundant metabolites were identified between the two groups, with uric acid, vitamin C, malate, glutamine and oleic acid being all upregulated in BTC. Purine metabolism, metabolic pathways, bile secretion, the phosphotransferase system, glyoxylate and dicarboxylate metabolism, pyrimidine metabolism, and ABC transporters were the key metabolic pathways associated with meat quality in the castrated group. In conclusion, there are certain differences in meat quality and muscle metabolites between BTC and BTN. Castration can affect meat quality characteristics to a certain extent.

IntroductionPreliminary research has identified that potassium and magnesium may influence the quality of Tibetan sheep meat, although the underlying mechanisms remain unclear.MethodsThis study investigated the effects of dietary supplementation with magnesium potassium sulfate (PMS) on carcass quality, meat quality, nutritional composition, and flavor metabolites of Tibetan sheep, and explored potential mechanisms using a metabolomics approach.ResultsDietary inclusion of 0.3% PMS significantly enhanced carcass quality, as shown by increases in abdominal wall thickness and backfat thickness by 13.13% and 7.41%, respectively, compared with the control group (p < 0.05). Supplementation with 0.25% PMS significantly elevated pH at 24 h, a, and intramuscular fat content, while reducing pH at 45 min and b (p < 0.05). The 0.25% PMS group also exhibited significantly higher concentrations of linoleic acid, linolenic acid, EPA, and DHA than the control group (p < 0.05). Volatile compound analysis showed that PMS was associated with the production of fruity and nutty compounds (e.g., methyl 3-methylpentanoate and methyl heptanoate) and with the suppression of malodorous substances such as 1-hydrazinopropan-2-ol. Non-volatile metabolomics revealed that PMS supplementation was associated with enhanced fatty acid biosynthesis, which correlated with the adipocyte cytokine signaling pathway.DiscussionPMS supplementation was further associated with modulation of the sphingolipid signaling pathway, which correlated with synergistic regulation of meat quality and contributed to an improved flavor profile of Tibetan sheep meat.

This study evaluated whether dietary guanidinoacetic acid (GAA) was associated with meat quality, jejunal microbiota and muscle metabolomic profiles in Tibetan sheep. A total of 120 healthy, 2-month-old, weaned male Tibetan sheep (17.33 ± 0.21 kg) were allocated to four groups: CON, basal diet (control group); LG, basal diet supplemented with 0.08% GAA; MG, basal diet supplemented with 0.10% GAA; HG, basal diet supplemented with 0.12% GAA. The feeding trial lasted 90 days after a 10-day adaptation. Compared with CON, 0.12% GAA was associated with higher relative abundances of selected jejunal taxa (Eubacterium_nodatum_group, Family_XIII_AD3011_group and Bacillus) and increased acetate and propionate concentrations. Untargeted metabolomics identified significantly altered muscle metabolites associated with glycerophospholipid metabolism, including PC 37:5, PC 38:4 and LPA 14:0. The HG group also showed higher glutathione peroxidase activity and total antioxidant capacity, a higher a* value and lower shear force. Correlation analyses indicated associations among jejunal microbiota, muscle metabolites and meat-quality traits. These findings provide further evidence supporting the potential use of dietary GAA as a feed additive for improving selected meat-quality traits in Tibetan sheep.

Optimizing the dietary nitrogen-to-sulfur (N:S) ratio is crucial for enhancing ruminal microbial fermentation and nutrient digestibility. This study investigated the effects of different dietary N:S ratios on ruminal antioxidant indices, digestive enzyme activities, immune parameters, volatile fatty acid (VFA) profiles, microbial communities, and metabolomic profiles in plateau-type Tibetan sheep. Ninety 2-month-old male Tibetan sheep (initial body weight, 15.55 ± 0.20 kg) were randomly assigned to three dietary treatments formulated with N:S ratios of 9.5:1 (HP-H), 8.5:1 (HP-M), and 7.5:1 (HP-L). Sheep fed the HP-H diet exhibited improved ruminal antioxidant indices compared with the other treatments, with CAT and T-AOC activities being higher than those of both the HP-M and HP-L groups, while SOD and GSH-Px activities were significantly higher only compared with those in the HP-L (p < 0.05). Furthermore, ruminal cellulase activity, as well as propionate and butyrate levels, were significantly higher in the HP-H group (p < 0.05). Microbiome analysis revealed that the HP-H diet enriched the relative abundances of Candidatus Saccharimonas, the Rikenellaceae RC9 gut group, and uncultured rumen bacteria. Correspondingly, untargeted metabolomics identified higher signal intensities of 3-hydroxyphenylacetic acid, nervonic acid, arachidonic acid (peroxide-free), and calciferol in the ruminal fluid of the HP-H group relative to the other treatments. In conclusion, when the nitrogen-to-sulfur ratio was 9.5:1, ruminal antioxidant enzyme activities were increased, and the concentrations of propionic acid and butyric acid were increased. The differential microorganisms and metabolites were mainly associated with lipid metabolism, as well as cofactor and vitamin metabolism.

Tibetan sheep is a primitive Chinese breed valued for its excellent edible quality and high nutritional value. Despite this, limited research has explore dlimited research has explored the relationship between oat silage supplementation level and its impact on the rumen microbiota, muscle metabolism, and subsequent meat quality. This experiment therefore investigated how varying levels of dietary oat silage affect the Tibetan sheep’s rumen: specifically, its morphological development, enzyme activity, fermentation parameters, microbial flora, and muscle metabolites.The study also examined the relationship between these factors and the meat’s edible quality and conventional nutritional traits. The results showed that compared to lower oat silage ratios a hay/oat silage ratio of 5:5 (OS3) significantly enhanced meat tenderness, color, and protein content. A hay/oat silage ratio of 2.5:7.5 (OS4) significantly increased rumen papillae length and butyrate concentration, thereby improving rumen antioxidant capacity and promoting overall rumen growth and development. Further analyses of rumen microbial communities, metabolomics, and correlation demonstrated that the level of oat silage primarily altered muscle glycolysis process, which subsequently affected conventional nutritional components, color, and tenderness. Additionally, the OS3 group increased the abundance of FD2005, UCG-002, Eubacterium_ruminantium_group, and Succinivibrionaceae_UCG-001. Overall, a hay-to-oat silage ratio of 5:5 was optimal, improving both the rumen microbiota composition and muscle metabolic pathways in Tibetan sheep, which consequently enhanced both rumen metabolism and the meat’s edible and conventional nutritional quality.

Tibetan sheep grazing on the Qinghai-Tibet Plateau require dietary protein supplementation; however, they face economic constraints due to the high cost of feed transportation in this region. Given that the leucine metabolite β-hydroxy-β-methyl butyrate (HMB) enhances both protein synthesis and intestinal nutrient absorption, this study employed metagenomics and untargeted metabolomics to systematically evaluate HMB's effects on antioxidant capacity, immune response, microbiota, metabolites, and the health of the small intestine in Tibetan sheep. A total of 120 healthy weaned 60-day-old male Tibetan lambs were assigned to diets containing 0 mg/kg (control group, CON), 430 mg/kg (low HMB, L-HMB), 715 mg/kg (medium HMB, M-HMB), or 1,000 mg/kg (high HMB, H-HMB) for 90 d. At the end of the experiment, 6 lambs from each group were slaughtered for intestinal tissue and content sampling. The M-HMB treatment significantly increased average daily gain of the lambs without affecting feed intake, thereby improving feed utilization efficiency. M-HMB promoted the development of small intestinal morphological and elevated villus height, while also enhancing the activities of digestive enzyme and disaccharidase activities. Furthermore, M-HMB enhanced the antioxidant capacity, immune response, and barrier function of the small intestine. Metagenomic analysis revealed that M-HMB supplementation improved the composition of the small intestinal microbiota in Tibetan sheep, specifically increasing the relative abundance of Ruminococcus bacterium P7 and R. bromii, and enhanced microbial carbohydrate degradation capacity. Metabolomic analysis demonstrated that M-HMB supplementation significantly altered the small intestinal metabolite profile, enhancing carbohydrate metabolic pathways and increased the production of short-chain fatty acids (SCFAs). M-HMB upregulated PLCβ1 and ERK1/2 protein expression levels in small intestinal tissue and elevated the proportion of Ki67-positive cells at the basal crypt region of small intestinal crypts, suggesting enhanced proliferative activity of intestinal epithelial cells. In summary, dietary supplementation with M-HMB (715 mg/kg) promoted small intestinal growth and development, enhanced digestive and absorptive functions, optimized the microbial composition, improved carbohydrate degradation, and increased the production of SCFAs, ultimately improving the growth performance of Tibetan sheep lambs.

IntroductionThis study seeks to examine the impact of supplementing different concentrations of valine (specifically, 0.1% and 0.15% treatment groups) on the rumen microbial community, muscle metabolites, and meat quality in Tibetan sheep.MethodsBoth targeted and untargeted metabolomics approaches were employed to conduct a thorough analysis of meat quality and muscle metabolites. Simultaneously, 16S rDNA sequencing was utilized to explore the composition of the rumen microbial community in Tibetan sheep and its potential regulatory mechanisms on meat quality.ResultsThe findings reveal that the 0.15% valine treatment group exhibited superior edible quality and flavor characteristics, along with an increased protein content in the meat. In comparison to the control group (0% valine supplementation), the 0.1% and 0.15% treatment groups demonstrated an increase in meat yield by 2.24% and 3.61%, respectively, and a reduction in shear force by 18.33% and 26.93%. Correlation analysis between non-targeted metabolomics and meat quality indicated that valine supplementation may influence amino acid metabolism in muscle tissue, thereby affecting meat pH and tenderness. Furthermore, the supplementation of 0.15% valine led to an increased abundance of Succiniclasticum, Christensenellaceae R-7 group, and NK4A214_group in the rumen of Tibetan sheep, while concurrently reducing the relative abundance of certain uncultured bacterial taxa (e.g., g__uncultured).DiscussionOverall, in comparison to the 0% (K group) and 0.10% (V1 group) treatments, the 0.15% (V2 group) treatment resulted in enhanced meat quality and metabolite content. Additionally, valine supplementation exerted a significant impact on the composition of rumen microbial communities in Tibetan sheep, thereby may influence muscle tenderness and protein deposition.

Research has demonstrated that resveratrol (RES) and β-hydroxy-β-methylbutyric acid (HMB) promote gastrointestinal health by altering the intestinal microbiome. The present study aimed to evaluate the impacts of RES and HMB on the bacterial community composition and metabolomic profiles of the jejunum in Tibetan sheep. A total of 120 male Tibetan lambs, with an average initial body weight of 16.87 ± 0.31 kg, were randomly assigned to four experimental groups according to a 2×2 factorial design. The experimental diets contained 11.19% protein diet, or 12.69% protein diet supplemented with RES/HMB (H-RES-HMB and L-RES-HMB, respectively), and 11.19% protein diet, or 12.69% protein diet without supplemented with RES/HMB (LCP and HCP, respectively). The findings indicated a significant increase in jejunal digestive enzyme activities (α-amylase and chymotrypsin), antioxidant capacity (catalase, glutathione peroxidase, and superoxide dismutase), and immune responses (including immunoglobulins A, G, and M) in the H-RES-HMB group, in contrast to a notable decrease in markers of oxidative stress and pro-inflammatory cytokines (interleukin 1β, interleukin 6, and tumor necrosis factor alpha) (P < 0.05). Morphological assessments revealed a significant reduction in crypt depth within the H-RES-HMB group (P < 0.05). Among the short-chain fatty acids, butyric acid concentration was significantly elevated in the H-RES-HMB group (P < 0.05). Additionally, mRNA expression levels of barrier-associated genes, including OCLN, Muc-2, and ZO-1 in the jejunum, were significantly enhanced in the H-RES-HMB group. Microbial community analysis demonstrated that the inclusion of RES and HMB in the 14% protein diet resulted in an increase in the relative abundance of Firmicutes, Bacillus, and Methanobrevibacter, while simultaneously reducing the abundance of Proteobacteria, Escherichia, and Shigella. Notably, the increased abundance of these beneficial bacteria was positively correlated with butyric acid concentrations. Furthermore, metabolomic analysis indicated higher levels of L-Arginine and His-Lys in the H-RES-HMB group, which also showed a positive correlation with butyric acid concentration, whereas isovaleric acid, D-xylose, and diacetyl concentrations were lower in this group. In conclusion, the findings suggest that a 14% protein diet is more effective in enhancing jejunal morphology and barrier function. Supplementation with RES and HMB in this diet resulted in elevated concentrations of volatile fatty acids, particularly butyric acid, by modulating microbial community composition (notably Firmicutes, Bacillus, and Methanobrevibacter) and metabolite profiles (including Gamma-aminobutyric acid, Succinate, L-Arginine, and His-Lys). A synergistic effect was found between the crude protein level (14%) and dietary RES/HMB on villus phenotype, digestive enzyme capacity, and antioxidant activity.IMPORTANCETo thrive in harsh, high-altitude environments, Tibetan sheep require efficient nutrient absorption. Our study shows that a diet with optimal protein (12.69%) supplemented with natural additives—resveratrol and β-hydroxy-β-methylbutyric acid—significantly enhances gut health. This dietary regimen strengthened the intestinal barrier, suppressed harmful inflammation, and boosted local immunity. Furthermore, it enriched the gut microbiome with beneficial butyric acid-producing bacteria, a key nutrient for intestinal cells. This research establishes a practical nutritional strategy to improve intestinal function and overall resilience in Tibetan sheep, supporting sustainable livestock production in challenging plateau regions.

IntroductionThe subcutaneous fat exerts energy storage and protective effects. However, excessive fat deposition results in waste of biological resources and compromises meat quality. According to the biological function, we hypothesized that GAA's could reduce subcutaneous fat deposition in Tibetan sheep.MethodsA total of 120 healthy weaned male lambs (2 months old) with similar body weight (17.33 ± 0.21 kg) were randomly divided into four groups, each receiving a basal diet or diet containing 0.08%, 0.10%, or 0.12% GAA. The experimental period lasted 90 days.ResultsOur results showed that 0.10% GAA significantly reduced subcutaneous fat thickness (P < 0.05). Antioxidant capacity was improved, as evidenced by a reduced MDA (P, P-linear, P-quadratic < 0.05), while increased SOD and T-AOC (P, P-linear, P-quadratic < 0.05). The contents of C18:3(n-3) (P, P-quadratic < 0.05) and C18:2(n-6) (P, P-linear < 0.05) was significantly increased when supplementing 0.10% GAA, while those of C16:0 (P < 0.05) and C22:2(n-6) (P, P-linear < 0.05) was significantly decreased. Pearson correlation analysis revealed that the differential genes including ANGPT2, PLA2G4A, PIK3CB, IDNK, MAPKAPK5 and the differential metabolites including DG (16:0/20:4), TG (16:0/16:0/18:1), DG (16:1/18:1), DG (16:0/18:2), Cer (d18:1/18:1) were closely associated with the antioxidant capacity and fat deposition.ConclusionIn summary, 0.10% GAA reduced subcutaneous fat thickness, improved the fatty acid profile, and maintained redox homeostasis in Tibetan sheep.

To optimize the enzymatic hydrolysis process of Tibetan sheep meat and to characterize the flavor profile of the resulting hydrolysates,using Tibetan sheep meat as the raw material,single-factor experiments combined with response surface methodology were employed to refine the hydrolysis conditions.Flavor compounds and taste characteristics of the hydrolysates were comprehensively analyzed.The optimal enzymatic hydrolysis conditions were determined to be:A solid-to-liquid ratio of 1:2,an enzyme ratio of 3:1,a hydrolysis time of 3.5 hours,an enzyme concentration of 0.9%,pH8.2,and a temperature of 56 °C.Under these conditions,the degree of hydrolysis,peptide yield,and sensory evaluation score reached 28.08%,81.14%,and 18.00,respectively.Gas chromatography-ion mobility spectrometry(GC-IMS)identified 58 volatile compounds,with multivariate statistical analysis highlighting hexanal monomer,hexanal dimer,1-octen-3-ol monomer,n-pentanal monomer,and n-octanal dimer as key contributors to the flavor profile.Umami and sweet amino acids accounted for 34.99%of total taste-active amino acids in the hydrolysate.Levels of umami nucleotides(5'-UMP,5'-GMP,and 5'-IMP)were significantly higher than in the control group(P<0.05),resulting in a markedly increased equivalent umami concentration(EUC),indicating enhanced umami taste.In contrast,the control group exhibited a significantly higher fatty acid content(63.74%)than the enzymatic hydrolysate group(P<0.05).Overall,the enzymatic hydrolysate showed a prominent umami and sweet flavor profile,complex aroma composition,and superior sensory characteristics,indicating its potential application in the development of meat-based seasonings.

As a source of non-protein nitrogen, ammonium chloride (NH4Cl) is widely utilized in ruminant diets to reduce feed costs. However, the impact of its supplementation level on the flavor of sheep meat remains unclear, despite the known influence of fat on meat flavor. This study aimed to investigate the effects of dietary NH4Cl supplementation levels on the lipidome and flavor compounds of subcutaneous adipose tissue in Tibetan sheep, providing a scientific basis for dietary optimization in Tibetan sheep farming. Eighty 2-month-old early-weaned Tibetan lambs were selected and randomly allocated into four groups, fed diets supplemented with 0% (N0 group), 1.49% (N1 group), 2.24% (N2 group), and 3.01% (N3 group) NH4Cl for an experimental period of 105 days. The study conducted histomorphological observations, lipidomics analysis, and determination of flavor compounds. The results showed that NH4Cl supplementation significantly reduced (p < 0.05) the contents of various unsaturated fatty acids and n-3 polyunsaturated fatty acids (n-3 PUFA) in the subcutaneous adipose tissue of Tibetan sheep. Specifically, the total saturated fatty acid (total SFA) content in the N3 group was significantly higher than that in the other groups, while the total monounsaturated fatty acid (total MUFA) content was significantly lower than that in the N1 and N2 groups (p < 0.05). The absolute contents of phosphatidylcholine (PC), phosphatidylethanolamine (PE), and the sum of triglycerides (TGs) and diglycerides (DGs) in the N3 group were significantly higher (p < 0.05) than those in the other groups. Regarding flavor compounds, the contents of ketone aroma compounds, such as 2-propanone and 2-butanone monomer, were significantly higher (p < 0.05) in the N0 group than in the other groups. The ammonia content in the N1 and N3 groups was significantly higher (p < 0.05) than that in the N0 and N2 groups, while the allyl sulfide content in the N2 group was significantly higher (p < 0.05) than that in the other groups. Correlation analysis revealed that the majority of TG and DG differential lipids were significantly positively correlated with allyl sulfide, and most differential lipids belonging to the PC, PE, and hexosylceramide (Hex1Cer) classes were significantly positively correlated with ammonia (|r| ≥ 0.80, p < 0.01). Conversely, PC (16:0_18:3) exhibited significant negative correlations with multiple beneficial aroma compounds (|r| ≥ 0.80, p < 0.01). The study indicates that dietary NH4Cl supplementation levels exceeding 2.24% are associated with alterations in lipid metabolism and reduced synthesis of unsaturated fatty acids and beneficial flavor compounds, such as 2-propanone and 2-butanone, in subcutaneous adipose tissue. This is also associated with the abnormal accumulation of phospholipids and ceramides, which correlate strongly with elevated ammonia concentrations in adipose tissue and the generation of oxidation products such as propanal, potentially compromising meat flavor quality.

IntroductionUrea shows substantial potential as a cost-effective alternative to soybean meal in ruminant production. Given the critical role of subcutaneous adipose tissue (SAT) in meat quality, this study evaluated the effects of different dietary urea levels on the histomorphology, fatty acid composition, lipidomic profiles, and volatile flavour compounds of SAT in Tibetan sheep.MethodsEighty female lambs (16.2 ± 1.01 kg, 2 months old) were randomly assigned to four dietary treatments supplemented with 0% (U0), 0.80% (U1), 1.19% (U2), and 1.61% (U3) urea. The 105-day trial comprised a 15-day adaptation period followed by a 90-day experimental period.ResultsThe results indicated that: high urea levels (U2 and U3) disrupted adipocyte integrity, resulting in disordered arrangement, cellular rupture, and a significant reduction in adipocyte diameter (p < 0.05). Increasing dietary urea levels significantly decreased the concentrations of trans monounsaturated fatty acids (e.g., C16:1 T) and n-3 polyunsaturated fatty acids (p < 0.05), while increasing the level of cis monounsaturated fatty acid C18:1n9c (p < 0.05). Lipidomic analysis identified 48 differential lipid molecules (|log2Fold change| ≥ 2, adjusted p < 0.05), primar ily comprising glycerolipids (Triglycerides, Triglycerides) and glycerophospho lipids (phosphatidylcholine, phosphatidylethanolamine, Phosphatidylglycerol). Urea supplementation significantly influenced the volatile compounds profile (p < 0.05); excessive urea (U3) led to ammonia accumulation and pungent off odors, whereas moderate supplementation imparted desirable milky and vanilla aromas.DiscussionIn conclusion, dietary urea levels affected lipid metabolism and reduced trans-fatty acid content; excessive dietary urea disrupted cell structure, increased ammonia accumulation and reduced aroma-contributing substances in the SAT of Tibetan sheep.

The research explored the effect of dietary rumen-protected betaine (RPB) supplementation on development and function of ileum in Tibetan sheep. Sixty male lambs (17.72 ± 0.19 kg, aged 2 months) were randomly divided into control group (Ctrl) and RPB group (0.08

This study aimed to investigate the effects of adding different levels of valine (0.1% and 0.15% treatment groups) on the edible quality, nutritional quality, amino acid profile, fatty acid composition, and flavor of Tibetan sheep foreleg muscle. Targeted metabolomics was employed to conduct a comprehensive analysis of meat quality and muscle metabolites. Additionally, GC-IMS was used to examine the effects of valine on the flavor of Tibetan sheep foreleg meat. The results indicated that the addition of valine significantly improved the quality of Tibetan sheep foreleg muscle (P<0.05). Compared with the control group (0% valine supplementation), the post-slaughter pH of foreleg muscle in the treatment groups (from 45 minutes to 24 hours) declined more slowly (with the slowest decline observed in the 0.15% treatment group), and the 0.15% treatment group exhibited the best color (L*43.06,a*33.57,b*16.66); the shear force of the foreleg meat from Tibetan sheep in the 0.1% and 0.15% treatment groups was reduced by 26.57% and 35.91% respectively, with tenderness significantly improved (P<0.05). In terms of nutritional quality, the 0.15% treatment group had the highest protein content (21.5 g/100 g) in the foreleg meat, whilst the 0.1% treatment group had the highest fat content (3.15 g/100 g), which was more than double that of the control group; The levels of essential amino acids such as isoleucine and threonine in the treatment groups were significantly higher than those in the control group (P<0.05). The 0.1% treatment group had the highest content of 5Z,8Z,11Z,14Z,17Z-Eicosapentaenoic acid, whilst the 0.15% treatment group had the highest content of 11Z,14Z-Eicosadienoic acid. The 0.1% treatment group had the highest levels of 1-octen-3-ol-M, 1-pentanol-D, 1-hexanol-D, 1-hexanol-M and 4-hydroxy-4-methyl-2-pentanone were highest in the 0.1% treatment group. ROAV analysis identified five key flavor compounds in the 0.1% treatment group (with 1-octene-3-ol monomer detected in greater quantities). Correlation analysis indicated that polyunsaturated fatty acids in Tibetan sheep forequarter meat may be more likely to promote the formation of aldehyde and ketone flavor compounds. In summary, the addition of valine to the diet can effectively improve the meat quality and flavor of Tibetan sheep foreleg muscle.

The experiment aimed to investigate the effects of dietary supplementation with different levels of potassium magnesium sulfate (PMS) on the fat content, fatty acid composition, and volatile flavor compounds in intramuscular fat of Tibetan sheep. A total of 120 healthy two-month-old weaned female Tibetan lambs were randomly divided into four groups, with three replicates per group and 10 lambs per replicate. The control group (group K) was fed a basal diet, while the experimental groups were fed the basal diet supplemented with 0.20% (group S1), 0.25% (group S2), and 0.30% (group S3) PMS in the concentrate supplement, respectively. The pre-test period was seven days, and the formal test period was 120 days. At the end of the experiment, intramuscular fat samples were collected. Fat content was determined by Soxhlet extraction, and fatty acid composition and volatile flavor compounds were analyzed by gas chromatography-mass spectrometry (GC-MS), followed by correlation analysis. The results showed that the crude fat content of intramuscular adipose tissue in group S2 was significantly higher than that in the other groups (P<0.05). Fatty acid analysis revealed that, compared with group K, the polyunsaturated fatty acid (PUFA) content in group S2 was significantly increased (P<0.05), while the content of arachidonic acid was significantly decreased (P<0.05). The n-6 PUFA/n-3 PUFA ratio showed no significant difference (P>0.05). A total of 39 volatile flavor compounds were detected, and the concentrations of the off-flavor substances 1, 9-tetradecadiene and 3-methyldodecanoic acid in group S2 were significantly reduced (P<0.05), while the concentrations of beneficial flavor compounds such as 2-butylacrolein, 2-nonanol and 2-undecanol were increased. Correlation analysis indicated that there were significant or extremely significant correlations between fatty acid composition and the concentrations of certain volatile flavor compounds (P<0.05 or P<0.01). The study shows that dietary supplementation with an appropriate level of PMS can improve the flavor quality of Tibetan sheep meat, with 0.25% being the optimal supplementation level.

The parturition season of grazing Tibetan ewes spans from October to March, a period that exacerbates the adverse impacts of nutrient-deficient herbage on milk yield, body condition, and postpartum recovery. To alleviate the weight loss of ewes during the cold seasons, we provided concentrate supplements at four levels (dry matter (DM) basis), 260 g (C1), 440 g (C2), 520 g (C3), and 610 g (C4), alongside a basal diet of grazed pasture. A total of 96 multiparous Tibetan ewes (third parity, body weight: 45.17 ± 3.69 kg (body weight (BW) were enrolled within 12–18 h postpartum and randomly allocated to four dietary groups (n = 24 ewes per group). We measured growth performance, ruminal histomorphology, fermentation parameters, and digestive enzymes. A multi-omics technique (16S rRNA gene sequencing and RNA-seq) was employed to investigate the mechanisms underlying alterations in ruminal function. The results showed that increasing the concentrate level decreased body weight loss and increased average dry matter intake (p < 0.05). Rumen morphology was significantly altered: papilla width and muscle layer thickness were greatest in the C4 group, whereas submucosal thickness was highest in the C1 group (p < 0.05). Cellulase activity was lowest in the C1 group (p < 0.05). Papilla width of lactating Tibetan ewes in the C4 group was higher (p < 0.05) than that in the C1 and C3 groups. Concentrate supplementation altered ruminal microbiota composition and diversity. Each group exhibited a distinct microbial signature: the C1 group was characterized by Lachnospiraceae_XPB1014_group, Candidatus_Omnitrophus, Paenibacillus, and unclassified_Oligoflexaceae; the C2 group was enriched in Papillibacter, Anaerovibrio, V9D2013_group, and unclassified_Peptococcaceae; the C3 group was characterized by unclassified_Bacteroidales_RF16_group; and the C4 group was characterized by Ruminococcus, Pseudobutyrivibrio, and Mitsuokella (p < 0.05). Transcriptomic analysis identified differentially expressed genes (TRPA1, EPHB1, GATA3, C4, ABCG2, THBS4, and TNFRSF11B) that are predominantly involved in immune regulation, signal transduction, and nutrient digestion. The results of Spearman correlation analysis showed that Anaerovibrio was negatively correlated with propionate (r = −0.565, p < 0.05). However, it was positively correlated with the ratio of acetate and propionate (r = 0.579, p < 0.05). Moreover, Lachnospiraceae_XPB1014_group was negatively correlated with cellulase (r = −0.699, p < 0.05) and α-amylase (r = −0.514, p < 0.05). These findings suggest that the increasing concentrate supplementation alleviates body weight loss in lactating Tibetan sheep by orchestrating improvements in rumen histomorphology, digestive function, altering bacteria composition, and ruminal immune and modulating host epithelial gene expression.

Subcutaneous fat deposition significantly influences animal growth, carcass quality, and meat characteristics. This study investigates the effects of varying dietary protein levels on backfat thickness, antioxidant capacity, fatty acid composition, differentially expressed genes (DEGs), and lipid molecules in Tibetan sheep. Sixty lambs were randomly assigned to two groups: a high-protein group (13.03% protein) and a low-protein group (11.58% protein), with each group containing 30 lambs (3 replicates per group, 10 lambs per replicate). Results showed that the low-protein group had significantly smaller fat cell diameters than the high-protein group ( P < 0.05, as determined by H&E staining). Additionally, the low-protein group exhibited significantly higher activities of GSH-Px and SOD, and lower MDA content compared to the high-protein group. Gas chromatography identified 33 fatty acids in the fat samples, with oleic, stearic, and palmitic acids being most abundant. The LP group had significantly lower C22:0 and higher C20:2, C20:3n6, C20:4n6, and C20:3n3 levels than the HP group ( P < 0.05). Transcriptomic analysis revealed 70 DEGs, of which 33 were upregulated and 37 were downregulated. KEGG analysis showed DEGs were enriched in 5 lipid metabolism pathways, including osteoclast differentiation, IL-17 signaling, and fluid shear stress/atherosclerosis. PPI analysis identified key lipid metabolism genes (FOS, FOSB, JUN, NR4A1, JUNB, PPARG). qRT-PCR validated RNA-Seq data accuracy. Lipid analysis detected 39 lipid classes and 2,605 lipid species, such as 856 TGs, 335 DGs, 279 Cer, 226 PCs, and 205 PEs. The LP group had higher DG and TG proportions, with significant increases in DG (40:4e), DG (32:1e), DG (34:4e), DG (20:5_18:2), and TG (16:18:1_18:3) levels. Correlation analysis showed that NR4A1 , FOS , JUN , and JUNB positively correlated with catalase (CAT) activity, while FOS , JUN , and JUNB were linked to fatty acid metabolism and adipocyte development. PPARG positively correlated with PUFAs (C20:2, C20:3n6, C20:4n6, C20:3n3, and C20:5n3). Lipid differential molecules (DG (40:4e) and DG (20:5_18:2)) positively correlated with CAT activity, and DG (32:1e) positively correlated with C22:0. Lipid differential molecules including DG (40:4e), DG (32:1e), DG (34:4e), DG (20:5_18:2), and TG (16:18_18:3) negatively correlated with adipocyte diameter. In conclusion, a diet with 11.58% protein regulates lipid-related gene expression, enhances antioxidant capacity in subcutaneous fat, and increases unsaturated fatty acid content.

The rumen is a balanced ecosystem that harbors a variety of microorganisms and plays a crucial role in the digestion and absorption of nutrients in ruminants. This study aimed to evaluate the effects of dietary crude protein on antioxidant activity, immunocompetence, and structural properties in the rumen of Tibetan sheep. 60 2-month-old male lambs with an average weight of 15.40 +/- 0.81 Kg were randomly assigned to either a low-protein diet (10.20% of dry matter, LP group) or a high-protein diet (11.58% of dry matter, HP group). The experiment was conducted over 97 days, including a 7-day adaptation period to the diets. The results indicated that the high-protein diet significantly increased rumen weight, rumen volume, papilla length, papilla width, and muscle layer (p < 0.05). Compared to the 10.20% protein group, supplementation with 11.58% crude protein significantly increased the activities of total antioxidant capacity (T-AOC) and superoxide dismutase (SOD) (p < 0.05). A total of 446 significant differentially expressed genes (DEGs) were identified in response to the high-protein diet (426 upregulated and 20 downregulated). Based on Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, six signaling pathways related to nutrient metabolism, antioxidant activity, and immunity were identified. Protein-protein interaction (PPI) analysis identified genes involved in fatty acid biosynthesis (ACACB and ACSF3), nutrient metabolism (ATP1B1, SLC5A1, IDH2, ATP1A2, NT5E, and NNT), and muscle development (MYH11, KCNMA1, and MYL9). In conclusion, an 11.58% crude protein diet enhanced papillary development and antioxidant activity in Tibetan sheep, likely by modulating the expression of functional genes.

Resveratrol (RES) and β-Hydroxy-β-methyl butyric acid (HMB) have been shown to exert antioxidant, anti-inflammatory effect as well as influence intestinal microbiota composition in monogastric animal. However, the mechanism by which RES and HMB regulate duodenal function in ruminants remains unclear. In this study, the effects of RES and HMB on the development and health of the duodenum in Tibetan sheep were investigated. A total of 120 early weaned Tibetan rams with similar initial body weight (15.5 ± 0.14 kg) were selected and randomly allocated into 4 groups. Each group received a basal diet supplemented with 1.5 g RES/d (RES group), 1.25 g HMB/d (HMB group), 1.5 g RES/d plus 1.25 g HMB/d (RES_HMB group), and without any additives (CON group). The results showed that RES and HMB supplementation significantly increased (p < 0.05) the villus height and the ratio of villus height to crypt depth in the duodenum. Meanwhile, the concentrations of cellulase and trypsin were increased (p < 0.05) when RES and HMB were supplemented in the basal diet. The RES group exhibited higher (p < 0.05) levels of anti-inflammatory markers (IgG, IL-6, and IL-1β), while combined supplementation of RES and HMB promoted (p < 0.05) the duodenal antioxidant capacity, as indicated by increased levels of GSH-Px and T-AOC. Furthermore, the RES group was characterized by a higher relative abundance of Butyrivibrio, while the HMB group was characterized by a higher relative abundance of Aeromonas, Rummeliibacillus, UCG-002, Ralstonia and Stenotrophomonas (p < 0.05 and LDA > 3.5). Metabolomics analysis showed that differential metabolites were significantly enriched (p < 0.05) in the cyanoamino acid metabolism, biosynthesis of secondary metabolites, protein digestion and absorption and ABC transporters. These results indicated that RES and HMB can be used as feed additives to maintain intestinal health by enhancing duodenal development, promoting digestive enzyme secretion, and improving antioxidant and anti-inflammatory capacity in Tibetan sheep.