We applied chemical group-based metabolomics to identify blood metabolic signatures associated with residual feed intake in beef cattle. A group of 56 crossbred growing beef steers (average BW = 261.3 ± 18.5 kg) were adapted to a high-forage total mixed ration in a confinement dry lot equipped with GrowSafe intake nodes for period of 49 d to determine their residual feed intake classification (RFI). After RFI determination, weekly blood samples were collected three times from beef steers with the lowest RFI [most efficient (HFE); n = 8] and highest RFI and least-efficient [least efficient (LFE); n = 8]. Plasma was prepared by centrifugation and composited for each steer. Metabolome analysis was conducted using a chemical isotope labeling (CIL)/liquid chromatography–mass spectrometry, which permitted the analysis of metabolites containing amine/phenol-, carboxylic acid-, and carbonyl-chemical groups, which are metabolites associated with metabolisms of amino acids, fatty acids, and carbohydrates, respectively. A total number of 495 amine/phenol-containing metabolites were detected and identified; pathway analysis of all these metabolites showed that arginine biosynthesis and histidine metabolism were enriched (P < 0.10) in HFE, relative to LFE steers. Biomarker analyses of the amine/phenol-metabolites identified methionine, 5-aminopentanoic acid, 2-aminohexanedioic acid, and 4-chlorolysine as candidate biomarkers of RFI [false discovery rate ≤ 0.05; Area Under the Curve (AUC) > 0.90]. A total of 118 and 330 metabolites containing carbonyl- and carboxylic acid-chemical groups, respectively were detected and identified; no metabolic pathways associated with these metabolites were altered and only one candidate biomarker (methionine sulfoxide) was identified. These results identified five candidate metabolite biomarkers of RFI in beef cattle which are mostly associated with amino acid metabolism. Further validation using a larger cohort of beef cattle of different genetic pedigree is required to confirm these findings.

We applied whole blood transcriptome analysis and gene set enrichment analysis to identify pathways associated with divergent selection for low or high RFI in beef cattle. A group of 56 crossbred beef steers (average BW = 261 ± 18.5 kg) were adapted to a high-forage total mixed ration in a confinement dry lot equipped with GrowSafe intake nodes for period of 49 d to determine their residual feed intake (RFI). After RFI determination, whole blood samples were collected from beef steers with the lowest RFI (most efficient; low-RFI; n = 8) and highest RFI (least efficient; high-RFI; n = 8). Prior to RNA extraction, whole blood samples collected were composited for each steer. Sequencing was performed on an Illumina NextSeq2000 equipped with a P3 flow. Gene set enrichment analysis (GSEA) was used to analyze differentially expressed gene sets and pathways between the two groups of steers. Results of GSEA revealed pathways associated with metabolism of proteins, cellular responses to external stimuli, stress, and heat stress were differentially inhibited (false discovery rate (FDR) < 0.05) in high-RFI compared to low-RFI beef cattle, while pathways associated with binding and uptake of ligands by scavenger receptors, scavenging of heme from plasma, and erythrocytes release/take up oxygen were differentially enriched (FDR < 0.05) in high-RFI, relative to low-RFI beef cattle. Taken together, our results revealed that beef steers divergently selected for low or high RFI revealed differential expressions of genes related to protein metabolism and stress responsiveness.

We examined the effects of dietary supplementation of a multi-component blend of prebiotics and probiotics on health, immune status, metabolism, and performance of newly weaned beef steers during a 35-d receiving period. Eighty newly weaned crossbred steers (12-hour post-weaning; 206 ± 12 kg of BW) from a single source were stratified by BW into 4 pens (20 steers per pen) such that each pen had similar BW at the beginning of the experiment. The pens were randomly assigned to receive a corn silage-based diet with no additive (CON; 2 pens; n = 40 steers) or a basal diet supplemented with SYNB feed additive at an average of 28 g/steer/d (SYNB; 2 pens; n = 40 steers). The SYNB additive is a blend of live S. cerevisiae and the fermentation products of S. cerevisiae, Enterococcus lactis, Bacillus licheniformis, and B. subtilis and was supplemented for the first 21 d only. Percentage of steers treated for bovine respiratory disease (BRD) was calculated for each dietary treatment. Daily DMI and meal events (meal frequency and duration) were measured. Weekly body weights were measured to calculate average daily gain (ADG). Blood samples collected on days 0, 14, 21, 28, and 35 were used for ex vivo TNF-α release assay following LPS stimulation, plasma metabolome analysis, and mRNA expression analysis of 84 innate and adaptive immune-related genes. Compared with CON, supplemental SYNB increased (P ≤ 0.05) ADG, DMI, and meal events during the first 7 d. At d 21, there was no treatment effect (P > 0.05) on final BW, DMI, ADG, and meal events; however, beef steers fed supplemental SYNB had greater (P = 0.02) meal duration. Over the entire 35-d receiving period, beef steers fed supplemental SYNB had greater (P = 0.01) ADG and feed efficiency, tended to have greater (P = 0.08) meal duration, and had lower percentage (35 vs. 50%) of animals treated for BRD and lower percentage of sick animals treated for BRD more than once (7.15 vs. 45%). Whole blood expression of pro-inflammatory genes was downregulated while that of anti-inflammatory genes was upregulated in beef steers fed supplemental SYNB. Beef steers fed supplemental SYNB had lower (P = 0.03) plasma concentration of TNF-α after LPS stimulation. Six nutrient metabolic pathways associated with health benefits were enriched (FDR ≤ 0.05) in beef steers fed supplemental SYNB. This study demonstrated that dietary supplementation of SYNB during the first 21 d of arrival reduced BRD morbidity, improved the performance, immune, and metabolic status of beef steers over a 35-d receiving period thereby extending the SYNB effect by a further 14 days post supplementation.

We evaluated the mRNA expression of genes involved in hepatic fatty acid, amino acid, and mitochondrial energy metabolism in crossbred beef steers with divergent low and high residual feed intake (RFI). Low-RFI beef steers (n = 8; RFI = - 1.93 kg/d) and high-RFI beef steers (n = 8; RFI = + 2.01kg/d) were selected from a group of 56 growing crossbred beef steers (average BW = 261 ± 18.5 kg) fed a high-forage total mixed ration after a 49-d performance testing period. At the end of the 49-d performance testing period, liver biopsies were collected from the low-RFI and high-RFI beef steers for RNA extraction and cDNA synthesis. The mRNA expression of 84 genes each related to fatty acid metabolism, amino acid metabolism, and mitochondrial energy metabolism were analyzed using pathway-focused PCR-based arrays. The mRNA expression of 8 genes (CRAT, SLC27A5, SLC27A2, ACSBG2, ACADL, ACADSB, ACAA1, and ACAA2) involved fatty acid transport and β-oxidation were upregulated (FC ≥ 2.0, FDR ≤ 0.05) in low-RFI, compared to high-RFI steers. Among those involved in amino acid metabolism, hepatic mRNA expression of a gene encoding for aminoadipate aminotransferase, an enzyme related to lysine degradation, was downregulated (FC = -5.45, FDR = 0.01) in low-RFI steers, whereas those of methionine adenosyltransferase I and aspartate aminotransferase 2, which both link amino acid and lipid metabolism, were upregulated (FC ≥ 2.0, FDR ≤ 0.05). Two mitochondrial energy metabolism genes (UQCRC1 and ATP5G1) involved in ATP synthesis via oxidative phosphorylation were upregulated (FC ≥ 2.0, FDR ≤ 0.05) in low-RFI beef steers, compared to high-RFI beef steers. The results of this study demonstrated that low-RFI beef steers exhibit upregulation of molecular mechanisms related to fatty acid transport, fatty acid β-oxidation, and mitochondrial ATP synthesis, which suggest that low-RFI beef steers have enhanced metabolic capacity to maximize capture of energy and nutrients from feeds consumed.

We investigated whole blood and hepatic mRNA expressions of immune genes and rumen microbiome of crossbred beef steers with divergent residual feed intake phenotype to identify relevant biological processes underpinning feed efficiency in beef cattle. Low-RFI beef steers (n = 20; RFI = − 1.83 kg/d) and high-RFI beef steers (n = 20; RFI = + 2.12kg/d) were selected from a group of 108 growing crossbred beef steers (average BW = 282 ± 30.4 kg) fed a high-forage total mixed ration after a 70-d performance testing period. At the end of the 70-d testing period, liver biopsies and blood samples were collected for total RNA extraction and cDNA synthesis. Rumen fluid samples were also collected for analysis of the rumen microbial community. The mRNA expression of 84 genes related to innate and adaptive immunity was analyzed using pathway-focused PCR-based arrays. Differentially expressed genes were determined using P-value ≤ 0.05 and fold change (FC) ≥ 1.5 (in whole blood) or ≥ 2.0 (in liver). Gene ontology analysis of the differentially expressed genes revealed that pathways related to pattern recognition receptor activity, positive regulation of phagocytosis, positive regulation of vitamin metabolic process, vascular endothelial growth factor production, positive regulation of epithelial tube formation and T-helper cell differentiation were significantly enriched (FDR < 0.05) in low-RFI steers. In the rumen, the relative abundance of PeH15, Arthrobacter, Moryella, Weissella and Muribaculaceae was enriched in low-RFI steers, while Methanobrevibacter, Bacteroidales_BS11_gut_group, Bacteroides and Clostridium_sensu_stricto_1 were reduced. In conclusion, our study found that low-RFI beef steers exhibit increased mRNA expression of genes related to immune cell functions in whole blood and liver tissues, specifically those involved in pathogen recognition and phagocytosis regulation. Additionally, these low-RFI steers showed differences in relative abundance of some microbial taxa which may partially account for their improved feed efficiency compared to high-RFI steers.

Graphical Abstract Summary: We determined the effect of a direct-fed microbial supplement containing a mixture of Lactobacillus animalis, Propionibacterium freudenreichii, Bacillus subtilis , and Bacillus licheniformis on the milk lipidome of dairy cows in early lactation. Our results showed that the supplemental additive altered the milk lipidome toward increased relative concentrations of polyunsaturated fatty acids (PUFA), which would increase PUFA consumption in place of saturated fatty acid, resulting in potential health benefits such as reduced risk of coronary heart disease in consumers of milk.

The amine/phenol-metabolome of rumen fluid was analyzed to identify amino acid metabolism-related biomarkers associated with phenotypic selection for low or high residual feed intake (RFI) in beef cattle. Fourteen beef steers (most feed-efficient (HFE; RFI = −1.89 kg/d, n = 7) and least feed-efficient (LFE; RFI = +2.05 kg/d, n = 7)) were selected from a total of 56 crossbred growing beef steers (average BW = 261 ± 18.5 kg) after a 49-d feeding period in a dry lot equipped with two GrowSafe intake nodes. Rumen fluid samples were collected 4 h after feeding on d 56, 63, and 70 from the HFE and LFE beef steers. Metabolome analysis of the rumen fluid was performed using chemical isotope labeling/liquid chromatography-mass spectrometry to identify all metabolites containing amine/phenol chemical groups, which are mostly amino acid metabolites. A total of 493 metabolites were detected and identified in the rumen fluid. The partial least squares discriminant scores plot showed a slight separation between the two groups of steers, and a total of eight metabolites were found to be differentially abundant (FDR ≤ 0.05). Out of the eight differentially abundant metabolites, four metabolites (isomer 1 of cadaverine, baeocystin, 6-methyladenine, and N(6)-methyllysine) qualified as candidate biomarkers of divergent RFI phenotype based on area under the curve ≥ 0.70. The results of this study revealed that divergent RFI phenotype is associated with alteration in rumen amine/phenol-metabolome of beef steers.

We applied ruminal and plasma metabolomics and ruminal 16S rRNA gene sequencing to determine the metabolic pathways and ruminal bacterial taxa associated with divergent residual body weight gain phenotype in crossbred beef steers. A group of 108 crossbred growing beef steers (average BW = 282.87 ± 30kg) were fed a forage-based diet for a period of 56 d in a confinement dry lot equipped with GrowSafe intake nodes to determine their residual body weight gain (RADG) phenotype. After RADG identification, blood and rumen fluid samples were collected from beef steers with the highest RADG (most efficient; n = 16; 0.76 kg/d) and lowest RADG (least efficient; n = 16; -0.65 kg/d). Quantitative untargeted metabolome analysis of the plasma and rumen fluid samples was conducted using chemical isotope labelling/liquid chromatography–mass spectrometry. Differentially abundant metabolites in each of the plasma and rumen fluid samples between the two groups of beef steers were determined using a false discovery rate (FDR)-adjusted P-values ≤ 0.05 and area under the curve (AUC) &gt; 0.80. Rumen and plasma metabolic pathways that were differentially enriched or depleted (P ≤ 0.05) in beef steers with positive RADG compared to those with negative RADG were determined by the quantitative pathway enrichment analysis. A total of 1629 metabolites were detected and identified in the plasma of the beef steers; eight metabolites including alanyl-phenylalanine, 8-hydroxyguanosine, and slaframine were differentially abundant (FDR ≤ 0.05; AUC &gt; 0.80) in beef steers with divergent RADG; five metabolic pathways including steroid hormone biosynthesis, thiamine metabolism, propanoate metabolism, pentose phosphate pathway, and butanoate metabolism were enriched (P ≤ 0.05) in beef steers with positive RADG, relative to negative RADG steers. A total of 1908 metabolites were detected and identified in the rumen of the beef steers; results of the pathway enrichment analysis of all the metabolites revealed no metabolic pathways in the rumen were altered (P &gt; 0.05). The rumen fluid samples were also analyzed using 16S rRNA gene sequencing to assess the bacterial community composition. We compared the rumen bacterial community composition at the genus level using a linear discriminant analysis effect size (LEfSe) to identify the differentially abundant taxa between the two groups of beef steers. The LEfSe results showed greater relative abundance of Bacteroidetes_vadinHA17 and Anaerovibrio in steers with positive RADG compared to the negative RADG group, while steers in the negative RADG group had greater relative abundance of Candidatus_Amoebophilus, Clostridium_sensu_stricto_1, Pseudomonas, Empedobacter, Enterobacter, and Klebsiella compared to the positive RADG group. Our results demonstrate that beef steers with positive or negative RADG exhibit differences in plasma metabolic profiles and some ruminal bacterial taxa which probably explain their divergent feed efficiency phenotypes.