Microbiome-metabolomics analysis of the effects of decreasing dietary crude protein content on goat rumen mictobiota and metabolites.

OBJECTIVE: The objective of this study was to investigate the effects of decreasing dietary crude protein content on rumen fermentation, mictobiota, and metabolites in goats. METHODS: In an 84-day feeding trial, a total of twelve male Anhui white goat kids with initial body weight 15.9±1.13 kg were selected and randomly classified into two groups, feeding a normal crude protein diet (14.8% CP, NCP) or a low crude protein diet (12.0% CP, LCP). At the end of the experimental trial (on day 84), six animals were randomly selected from each group and were slaughtered to collect rumen fluid samples for the analysis of rumen fermentation parameters, microbiome, and metabolome. RESULTS: The concentrations of ammonia-nitrogen, total volatile fatty acid, acetate, and propionate were decreased (p<0.05) in the LCP group in comparison with those in the NCP group. The abundances of genera Prevotella, Campylobacter, Synergistetes, and TG5, which were associated with nitrogen metabolism, were lower (p<0.05) in the LCP group compared with those in the NCP group. The levels of 78 metabolites (74 decreased, 4 increased) in the rumen fluid were altered (p<0.05) by the treatment. Most of the ruminal metabolites that showed decreased levels in the LCP group were substrates for microbial protein synthesis. Metabolic pathway analysis showed that vitamin B6 metabolism was significantly different (p<0.05) in rumen fluid between the two treatments. CONCLUSION: Decreased dietary protein level inhibited rumen fermentation through microbiome and metabolome shifts in goat kids. These results enhance our understanding of ruminal bacteria and metabolites of goat fed a low protein diet.

Prognostic potential of pre-partum blood biochemical and immune variables for postpartum mastitis risk in dairy cows.

BACKGROUND: Mastitis is the most frequent diseases for transition cows. Identification of potential biomarkers for diagnosis of mastitis is important for its prevention. Thus, this study was conducted to investigate blood variables related to lipid metabolism, oxidative stress and inflammation, and serum variables that are related to health in postpartum cows. RESULTS: Seventy-six healthy Holstein dairy cows at week 4 before calving were selected to collect blood samples from weeks - 4 to 4 weekly relative to calving, respectively. Milk yield and composition were recorded weekly. According to the cut-off of somatic cell counts (SCC) for diagnosis of mastitis, 33 cows with SCC ≥ 500,000 cells ml- 1, 20 cows with 200,000 cells ≤ SCC < 500,000 cells ml- 1, and 23 cows with SCC < 200,000 cells ml- 1 were defined as high, middle, and low SCC, respectively. Serum concentrations of ß-hydroxybutyrate were higher (P < 0.01) during all weeks, and non-esterified fatty acids were higher in high SCC than in low SCC cows from weeks - 3 to 2 relative to calving. Higher serum concentrations of superoxide dismutase (P < 0.01) and lower malondialdehyde levels (P < 0.01) in low SCC than in high SCC cows indicate that the latter suffered from oxidative stress. The difference analysis of the three groups suggested that none of the above-mentioned variables can be used as potential prognostic candidates. On the other hand, high SCC cows exhibited higher blood neutrophil to lymphocyte ratio (NLR, P < 0.01) and platelet to lymphocyte ratio (PLR, P < 0.01) than low SCC cows, with a higher NLR (P < 0.01) in middle SCC than in low SCC cows. The high SCC cows had lower levels of anti-inflammatory factors including IL-10 (P = 0.05), but higher levels of proinflammatory factors such as IL-6 (P < 0.01), TNF-α (P < 0.05), and PSGL-1 (P < 0.01) than low SCC cows. CONCLUSIONS: The significantly different NLR and PLR pre-partum between the middle and low SCC cows suggest their prognostic potential for postpartum mastitis risk.

Effects of stocking density on oxidative stress status and mammary gland permeability in early lactating dairy cows.

The current study was conducted to investigate the effect of stocking density (SD) on oxidative stress status and mammary gland permeability in early lactating dairy cows. Thirty-two dairy cows were allocated into 16 blocks, basing on parity, previous milk yield, and body weight, and were then randomly assigned into one of the two treatments as follows: 75% (75SD) and 100% (100SD) SD. The cows were fed with same diet throughout the 8-week experimental period. The milk yield and milk sample were collected on two consecutive days during the 8-week experimental period weekly. Plasma samples were collected on fourth and eighth experimental weeks. Raw, energy-corrected, and 4% fat-corrected milk yield were significantly higher in 75SD-cows than that of 100SD-animals, respectively. The milk somatic cell count was lower in 75SD-cows than that of 100SD-animals. The levels of Na+ , Na+ /K+ , bovine serum albumin and plasmin were lower in 75SD-cows than those of 100SD-cattle, respectively. The 75SD-cows had reduced insulin and insulin/glucagon levels but higher prolactin and growth hormone concentrations, compared with those of 100SD-animals, respectively. In conclusion, compared with low SD-animals, early lactating cows with higher SD had higher oxidative stress status, which further led to a greater mammary gland permeability.

Lactation performance and rumen fermentation in dairy cows fed a diet with alfalfa hay replaced by corn stover and supplemented with molasses.

OBJECTIVE: The objective of current study was to investigate the lactation performance and rumen fermentation characteristics of dairy cows fed a diet with alfalfa hay replaced by corn stover but supplemented with molasses. METHODS: Sixteen Holstein cows in mid-lactation were randomly assigned to 1 of 2 dietary treatments: i) alfalfa based diet (AH), and ii) corn stover based diet supplemented with molasses (CSM). The experiment was conducted according to a 2×2 crossover design with 22-d each period, consisting of 17 d for adaptation and 5 d for data and samples collection. RESULTS: Dry matter intake and milk yield were higher for cows fed AH than CSM (p<0.01). Milk protein content and nitrogen conversion were higher (p<0.05), while milk urea nitrogen was lower (p<0.01) for cows fed AH than CSM-fed cows. Contents of milk total solids, fat and lactose were not different between two groups (p>0.10). Total rumen volatile fatty acid concentration tended to be higher (p = 0.06) for cows fed AH than CSM-fed cows. Molar proportion of acetate was lower (p = 0.04), but valerate was higher (p = 0.02) in cows fed AH than CSM-fed cows. Rumen concentration of propionate, and isobutyrate, and ratio of acetate to propionate tended to be different (p<0.10) between two groups. The feed cost per kilogram of milk was lower in CSM than AH (p<0.01). No differences were found in feed efficiency and most plasma parameters tested (p>0.10). CONCLUSION: In comparison with AH diet, CSM diet could be fed to dairy cows without negative effect on feed efficiency, ruminal fermentation, but economically beneficial, indicating that CSM could be an alternative choice for dairy farms instead of AH to feed mid-lactation dairy cows.

Local Mammary Glucose Supply Regulates Availability and Intracellular Metabolic Pathways of Glucose in the Mammary Gland of Lactating Dairy Goats Under Malnutrition of Energy.

As glucose is the regulator of both the milk yield and mammary oxidative status, glucose supply is considered to play important nutritional and physiological role on mammary gland (MG) metabolism. However, inconsistent results were observed from different infusion methods to evaluate the effect of glucose on MG glucose metabolism. Thus, precise method should be developed to learn how availability and intracellular metabolic pathways of glucose in the MG are altered by the direct mammary glucose supply. In addition, limited information is available on the role of mammary glucose supply in milk synthesis in lactating ruminants under an energy-deficient diet. Direct glucose supply to the MG was implemented in the current study through the external pudendal artery infusion under an energy-deficient diet. Six doses of glucose (0, 20, 40, 60, 80, and 100 g/d) were infused through the external pudendal arteries, which is the main artery to the MG, to six lactating goats fed with basal diet meeting 81% energy requirement in a 6 × 6 Latin square design. Milk and lactose yields were both quadratically increased with increased glucose infusion, whereas the milk yield changed inconsistently with the increased energy balance (EB), indicating local glucose supply, rather than EB, improved milk production. Glucose fluxes in the MG were significantly increased and correlated with mammary plasma flow. However, the ratio of lactose yield to glucose absorbed by the MG was significantly decreased. The increased glucose fluxes in the MG and changed glucose-related metabolites in milk indicated that the glucose availability and intracellular metabolic pathways was regulated by local mammary glucose. Acute glycolysis consumed the superfluous glucose and induced accumulation of oxygen radicals in the MG during over-supplied glucose conditions. The present study provided insight to optimal glucose supply to the MG during the lactation.

Metabolomics Integrated with Transcriptomics Reveals a Subtle Liver Metabolic Risk in Dairy Cows Fed Different Crop By-products.

Ruminants make large contributions to sustainable agriculture by converting crop by-products into agricultural food. Multi-omics integrative analysis helps to uncover the underlying molecular mechanisms. The liver metabolome-transcriptome interface (LMTI) in dairy cows, including 3938 significant correlations (p < 0.01 and |ρ| > 0.6) among 772 genes, 306 metabolites, and 305 microRNAs, is first demonstrated. How different crop by-products, corn stover (CS) and rice straw (RS), affect the liver metabolic functions based on the LMTI is further analyzed. Compared to the CS-fed cows, 13 out of 24 metabolites have lower relative concentrations (variable importance projection > 1.0 and p < 0.05), and 51 out of 68 genes are downregulated in the RS group (p < 0.01 and fold change < -2). Integrated analysis of metabolomics and transcriptomics reveal that lipid metabolism is most enriched including 14 subpathways. The altered metabolites and genes revealed the enriched ketogenesis induced by the linoleic acid pathways (p = 0.017, topology value = 1), which is supported by blood and histomorphometric phenotypes. The above results indicate the foreseeable liver metabolic disorders when RS is fed to cows. These findings provide new insights into the liver metabolic mechanism and into crop by-products utilization using integrative omics technologies.

Metabolomics of four biofluids from dairy cows: potential biomarkers for milk production and quality.

The fundamental understanding of the mechanisms regulating milk protein synthesis is limited. This study aimed to elucidate the metabolic mechanisms of milk production affected by forage quality through studying metabolites from four biofluids (rumen fluid, milk, serum, and urine) collected from 16 lactating cows fed alfalfa hay (AH, high-quality, n = 8) and corn stover (CS, low-quality, n = 8) using gas chromatography-time-of-flight/mass spectrometry. The cows fed AH exhibited higher milk yield (P < 0.01), milk protein yield (P = 0.04), and milk efficiency (P < 0.01) than those fed CS. A total of 165, 195, 218, and 156 metabolites were identified in the rumen fluid, milk, serum, and urine, respectively, while 29 metabolites were found in all four biofluids. In addition 55, 8, 28, and 31 metabolites in each biofluid were significantly different (VIP > 1 and P < 0.05) between the AH- and CS-fed animals. These metabolites were involved in glycine, serine, and threonine metabolism; tyrosine metabolism; and phenylalanine metabolism. Further integrated key metabolic pathway analysis showed that the AH-fed cows may have more comprehensive amino acid metabolisms, suggesting that these metabolite-associated pathways may serve as biomarkers for higher milk yield and better milk protein quality.