Huizeng Sun

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.

Effects of Replacement of Soybean Meal by Fermented Cottonseed Meal on Growth Performance, Serum Biochemical Parameters and Immune Function of Yellow-feathered Broilers

The study was conducted to examine the effects of partially replacing soybean meal (SBM) by solid-state fermented cottonseed meal (FCSM) on growth performance, serum biochemical parameters and immune function of broilers. After inoculated with Bacillus subtilis BJ-1 for 48 h, the content of free gossypol in cottonseed meal was decreased from 0.82 to 0.21 g/kg. A total of 600, day-old male yellow-feathered broilers were randomly divided into four groups with three replicates of 50 chicks each. A corn-SBM based control diet was formulated and the experimental diets included 4, 8 or 12% FCSM, replacing SBM. Throughout the experiment, broilers fed 8% FCSM had higher (p<0.05) body weight gain than those fed 0, 4 and 12% FCSM. The feed intake in 8% FCSM group was superior (p<0.05) to other treatments from d 21 to 42. On d 21, the concentration of serum immunoglobin M in the 4% and 8% FCSM groups, as well as the content of complements (C3, C4) in 8% FCSM group were greater (p<0.05) than those in the SBM group. Besides, birds fed 8% FCSM had increased (p<0.05) serum immunoglobin M, immunoglobulin G and complement C4 levels on d 42 compared with bird fed control diet. No differences (p>0.05) were found between treatments regarding the serum biochemical parameters and the relative weights of immune organs. In conclusion, FCSM can be used in broiler diets at up to 12% of the total diet and an appropriate replacement of SBM with FCSM may improve growth performance and immunity in broilers.

Molecular analysis of intestinal bacterial microbiota of broiler chickens fed diets containing fermented cottonseed meal

The study was conducted to investigate the effects of dietary inclusion of fermented cottonseed meal (FCM) on the ileal and cecal bacterial microbiota of broiler chickens. A total of 300 newborn yellow-feathered broiler chickens were randomly divided into 2 treatments with 3 replicates each (50 birds per replicate): control and 80 g/kg of FCM group. The feeding trial lasted for 42 d. Ileal and cecal digesta samples were collected from 8 chicks per replicate at 21 and 42 d of age to determine the composition of bacterial microbiota using denaturing gradient gel electrophoresis, cloning, sequencing, and real-time quantitative PCR analysis. The results demonstrated that the microbial composition in the ileum and cecum were considerably affected by the diet. The similarity dendrogram of banding profiles showed a more rapid stabilization of intestinal bacterial microbiota in broilers fed diets supplemented with FCM, compared with that of the birds fed the control diet. No significant difference was observed in total number of bands and Shannon-Weaver index, indicating that FCM had no effects on bacterial diversity. However, enumeration of bacteria in the ileal and cecal contents by quantitative PCR showed an increased (P < 0.05) population of lactobacilli, as well as a decreased (P < 0.05) Escherichia coli number by the dietary inclusion of FCM. In summary, dietary inclusion of FCM did not affect the intestinal microbial diversity but shifted intestinal microbiota, with a more homogenous population and an increased colonization of lactobacilli. The results also support the concept that dietary FCM inclusion could promote the beneficial bacteria in the intestinal tract.

Systematic microRNAome profiling reveals the roles of microRNAs in milk protein metabolism and quality: insights on low-quality forage utilization

In this study, we investigated the molecular regulatory mechanisms of milk protein production in dairy cows by studying the miRNAomes of five key metabolic tissues involved in protein synthesis and metabolism from dairy cows fed high- and low-quality diets. In total, 340, 338, 337, 330, and 328 miRNAs were expressed in the rumen, duodenum, jejunum, liver, and mammary gland tissues, respectively. Some miRNAs were highly correlated with feed and nitrogen efficiency, with target genes involved in transportation and phosphorylation of amino acid (AA). Additionally, low-quality forage diets (corn stover and rice straw) influenced the expression of feed and nitrogen efficiency-associated miRNAs such as miR-99b in rumen, miR-2336 in duodenum, miR-652 in jejunum, miR-1 in liver, and miR-181a in mammary gland. Ruminal miR-21-3p and liver miR-2285f were predicted to regulate AA transportation by targeting ATP1A2 and SLC7A8, respectively. Furthermore, bovine-specific miRNAs regulated the proliferation and morphology of rumen epithelium, as well as the metabolism of liver lipids and branched-chain AAs, revealing bovine-specific mechanisms. Our results suggest that miRNAs expressed in these five tissues play roles in regulating transportation of AA for downstream milk production, which is an important mechanism that may be associated with low milk protein under low-quality forage feed.

The Effects of Dietary Phosphorus on the Growth Performance and Phosphorus Excretion of Dairy Heifers

The objective of this study was to investigate the effects of reducing dietary phosphorus (P) on the frame size, udder traits, blood parameters and nutrient digestibility coefficient in 8- to 10-month-old Holstein heifers. Forty-five heifers were divided into 15 blocks according to the mo of age and were randomly assigned one of three dietary treatments: 0.26% (low P [LP]), 0.36% (medium P [MP]), or 0.42% (high P [HP]) (dry matter basis). Samples were collected at the wk 1, 4, 8. The results show that low dietary P had no effect on body measurement. The blood P concentration decreased with decreasing dietary P (p<0.05), while the blood calcium content of LP was higher than that of the MP and HP groups (p<0.05), though still in the normal range. The serum contents of alkalinephosphatase, potassium, and magnesium were similar among the treatments. No differences were found in all nutrients’ apparent digestibility coefficients with varied dietary P. However, with P diet decreased from HP to LP, the total fecal P and urine P concentration declined significantly, as did fecal water soluble P (p<0.05). In conclusion, reducing the dietary P from 0.42% to 0.26% did not negatively affect the heifers’ growth performance but did significantly lessen manure P excretion into the environment.

Lactation-related metabolic mechanism investigated based on mammary gland metabolomics and 4 biofluids’ metabolomics relationships in dairy cows

BackgroundLactation is extremely important for dairy cows; however, the understanding of the underlying metabolic mechanisms is very limited. This study was conducted to investigate the inherent metabolic patterns during lactation using the overall biofluid metabolomics and the metabolic differences from non-lactation periods, as determined using partial tissue-metabolomics. We analyzed the metabolomic profiles of four biofluids (rumen fluid, serum, milk and urine) and their relationships in six mid-lactation Holstein cows and compared their mammary gland (MG) metabolomic profiles with those of six non-lactating cows by using gas chromatography-time of flight/mass spectrometry.ResultsIn total, 33 metabolites were shared among the four biofluids, and 274 metabolites were identified in the MG tissues. The sub-clusters of the hierarchical clustering analysis revealed that the rumen fluid and serum metabolomics profiles were grouped together and highly correlated but were separate from those for milk. Urine had the most different profile compared to the other three biofluids. Creatine was identified as the most different metabolite among the four biofluids (VIP = 1.537). Five metabolic pathways, including gluconeogenesis, pyruvate metabolism, the tricarboxylic acid cycle (TCA cycle), glycerolipid metabolism, and aspartate metabolism, showed the most functional enrichment among the four biofluids (false discovery rate < 0.05, fold enrichment >2). Clear discriminations were observed in the MG metabolomics profiles between the lactating and non-lactating cows, with 54 metabolites having a significantly higher abundance (P < 0.05, VIP > 1) in the lactation group. Lactobionic acid, citric acid, orotic acid and oxamide were extracted by the S-plot as potential biomarkers of the metabolic difference between lactation and non-lactation. The TCA cycle, glyoxylate and dicarboxylate metabolism, glutamate metabolism and glycine metabolism were determined to be pathways that were significantly impacted (P < 0.01, impact value >0.1) in the lactation group. Among them, the TCA cycle was the most up-regulated pathway (P < 0.0001), with 7 of the 10 related metabolites increased in the MG tissues of the lactating cows.ConclusionsThe overall biofluid and MG tissue metabolic mechanisms in the lactating cows were interpreted in this study. Our findings are the first to provide an integrated insight and a better understanding of the metabolic mechanism of lactation, which is beneficial for developing regulated strategies to improve the metabolic status of lactating dairy cows.

Effects of Corn and Soybean Meal Types on Rumen Fermentation, Nitrogen Metabolism and Productivity in Dairy Cows

Twelve multiparous Holstein dairy cows in mid-lactation were selected for a replicated 4×4 Latin square design with a 2 ×2 factorial arrangement to investigate the effects of corn and soybean meal (SBM) types on rumen fermentation, N metabolism and lactation performance in dairy cows. Two types of corn (dry ground [DGC] and steam-flaked corn [SFC]) and two types of SBM (solvent-extracted and heat-treated SBM) with different ruminal degradation rates and extents were used to formulate four diets with the same basal ingredients. Each period lasted for 21 days, including 14 d for adaptation and 7 d for sample collection. Cows receiving SFC had a lower dry matter (DM) and total N intake than those fed DGC. However, the milk yield and milk protein yield were not influenced by the corn type, resulting in higher feed and N utilization efficiency in SFC-fed cows than those receiving DGC. Ruminal acetate concentrations was greater and total volatile fatty acids concentrations tended to be greater for cows receiving DGC relative to cows fed SFC, but milk fat content was not influenced by corn type. The SFC-fed cows had lower ruminal ammonia-N, less urea N in their blood and milk, and lower fecal N excretion than those on DGC. Compared with solvent-extracted SBM-fed cows, cows receiving heat-treated SBM had lower microbial protein yield in the rumen, but similar total tract apparent nutrient digestibility, N metabolism measurements, and productivity. Excessive supply of metabolizable protein in all diets may have caused the lack of difference in lactation performance between SBM types. Results of the present study indicated that increasing the energy degradability in the rumen could improve feed efficiency, and reduce environmental pollution.

Arteriovenous blood metabolomics: An efficient method to determine the key metabolic pathway for milk synthesis in the intra-mammary gland

The present study aimed to identify metabolic signature changes of the arteriovenous metabolome and the new metabolites that involved in mammary biological process during milk synthesis. GC/MS-based metabolomics profiling of arteriovenous plasma from 30 lactating dairy cows fed three diets identified a total of 144 metabolites. Phenylalanine and tyrosine, involved in aminoacyl-tRNA biosynthesis and phenylalanine metabolism, were shown higher expression in the artery than in the vein based on both GC/MS and targeted analysis for cows fed both alfalfa hay diet and rice straw diet. Mammary uptake or clearance of citric acid, stearic acid, oleic acid, fructose, β-mannosylglycerate, 4-hydroxybutyrate, and D-talose were significantly correlated with milk performance or feed intake, indicating that these metabolites might be newly identified precursors or indicators of milk synthesis. This comprehensive assessment of metabolic changes in the arteriovenous metabolome will provide a fundamental understanding of the key metabolites involved in milk synthesis and shows implications of how metabolites from arteriovenous plasma across MG are involved in biological processes or physiological functions for milk synthesis. The newly identified metabolites from the present study provide potential new targeted insights into the study of physiological process for milk synthesis in the MG.

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.

Effects of Aspergillus Oryzae Culture and 2-Hydroxy-4-(Methylthio)-Butanoic Acid on In vitro Rumen Fermentation and Microbial Populations between Different Roughage Sources

An in vitro experiment was conducted to evaluate the effects of Aspergillus oryzae culture (AOC) and 2-hydroxy-4-(methylthio)-butanoic acid (HMB) on rumen fermentation and microbial populations between different roughage sources. Two roughage sources (Chinese wild rye [CWR] vs corn silage [CS]) were assigned in a 2×3 factorial arrangement with HMB (0 or 15 mg) and AOC (0, 3, or 6 mg). Gas production (GP), microbial protein (MCP) and total volatile fatty acid (VFA) were increased in response to addition of HMB and AOC (p<0.01) for the two roughages. The HMB and AOC showed inconsistent effects on ammonia-N with different substrates. For CWR, neither HMB nor AOC had significant effect on molar proportion of individual VFA. For CS, acetate was increased (p = 0.02) and butyrate was decreased (p<0.01) by adding HMB and AOC. Increase of propionate was only occurred with AOC (p<0.01). Populations of protozoa (p≤0.03) and fungi (p≤0.02) of CWR were differently influenced by HMB and AOC. Percentages of F. succinogenes, R. albus, and R. flavefaciens (p<0.01) increased when AOC was added to CWR. For CS, HMB decreased the protozoa population (p = 0.01) and increased the populations of F. succinogenes and R. albus (p≤0.03). Populations of fungi, F. succinogenes (p = 0.02) and R. flavefacien (p = 0.03) were increased by adding AOC. The HMB×AOC interactions were noted in MCP, fungi and R. flavefacien for CWR and GP, ammonia-N, MCP, total VFA, propionate, acetate/propionate (A/P) and R. albus for CS. It is inferred that addition of HMB and AOC could influence rumen fermentation of forages by increasing the number of rumen microbes.