Effects of rumen-bypass protein supplement on growth performance, hepatic mitochondrial protein complexes, and hepatic immune gene expression of beef steers with divergent residual feed intake.

We investigated the impact of a rumen-bypass protein (RBP) supplement on growth performance, plasma and urinary N (UN) concentration, hepatic mitochondrial protein complexes, and hepatic mRNA expression of immune genes of beef steers with negative or positive residual feed intake (RFI) phenotype. Forty crossbred beef steers with an average body weight (BW) of 492 ± 36 kg were subjected to a generalized randomized block design over a 42-day experimental period. This study followed a 2 × 2 factorial arrangement of treatments. The factors evaluated were: 1) RFI classification (low-RFI (-2.12 kg/d) vs. high-RFI (2.02 kg/d), and 2) rumen-bypass protein supplement: RBP supplement (RBP; 227 g/steer/d) vs. control diet (CON; 0 g/d), resulting in four distinct treatments: LRFI-CON (n = 10), LRFI-RBP (n = 10), HRFI-CON (n = 10), and HRFI-RBP (n = 10). The RBP supplement (84% crude protein) is a mixture of hydrolyzed feather meal, porcine blood meal, and DL-methionine hydroxy analogue. The beef steers were stratified by BW, randomly assigned to treatments, and housed in four pens (1 treatment/pen) equipped with two GrowSafe feed bunks each to measure individual dry mater intake (DMI). Body weight was measured every 7 d. Liver tissue samples were collected on d 42 from all the beef steers. These samples were used for mRNA expression analysis of 16 immune-related genes and for evaluating the mitochondrial protein complexes I - V. No significant effects due to RBP supplementation or RFI × RBP interactions (P > 0.05) were observed for average daily gain (ADG) and DMI. However, compared to high-RFI steers, low-RFI steers showed a trend towards reduced DMI (12.9 vs. 13.6 kg/d; P = 0.07) but ADG was similar for the two RFI groups. Regardless of RFI status, supplemental RBP increased blood urea nitrogen (BUN) (P = 0.01), with a lower BUN concentration in low-RFI steers compared to high-RFI ones. A tendency for interaction (P = 0.07) between RFI and RBP was detected for the UN concentrations; feeding the dietary RBP increased the UN concentration in high-RFI beef steers (209 vs. 124 mM), whereas the concentration was lower than that of the CON group for low-RFI beef steers (86 vs. 131 mM). Interactions of RBP and RFI were observed (P ≤ 0.05) for mitochondrial activities of complexes IV, V, and mRNA expressions of some immune genes such as TLR2, TLR3, and IL23A. In conclusion, while RBP supplementation did not alter growth performance, its observed effects on hepatic immune gene expression, mitochondrial protein complexes, BUN, and UN depended on the beef steers' RFI phenotype. Therefore, the RFI status of beef steers should be considered in future studies evaluating the effects of dietary protein supplements.

Effects of dietary supplementation of peanut skins (Arachis hypogaea) on performance, digestibility, and rumen fermentation of cattle: a meta-analysis.

This study aimed to estimate the magnitude of the effects of dietary inclusion of peanut skins (PS) byproduct (Arachis hypogea L.) on intake, total-tract digestibility, and rumen fermentation of cattle via meta-analysis. Data were collected following the PRISMA methodology. Nine manuscripts and a graduate thesis met the inclusion criteria from 1983 to 2010. The effect size was estimated by calculating the weighted raw mean differences (RMD) between PS vs. control diets. The RMD was compared with a robust variance estimation method followed by a meta-regression and a dose-response analysis fitting the diet characteristics like crude protein content (CP), NDF content, ether extract content (EE), tannin content, and PS level in diet (0 to 40%) as covariates. Dietary PS decreased (P < 0.01) total-tract CP digestibility (52.0 vs. 64.3%), final body weight (371.5 vs. 397.9 kg), and average daily gain (1.14 vs. 1.44 kg/day) among treatment comparisons. Likewise, PS decreased total VFA (92.6 vs. 107.6 mmol/L) and NH3-N (8.22 vs. 12.1 mg/dL), but no effects were observed on rumen pH (6.47 vs. 6.14) and VFA molar proportions. Despite the between-cluster variance, dietary PS increased the ether extract digestibility (77.5 vs. 70.2%) among treatment comparisons. The subset and dose-response analysis revealed that PS should not exceed 8% (DM basis) in the diet to prevent negative effects on CP digestibility and animal performance. In conclusion, the results of this meta-analysis do not support the dietary inclusion of PS in cattle diets beyond 8%.

The differential plasma and ruminal metabolic pathways and ruminal bacterial taxa associated with divergent residual body weight gain phenotype in crossbred 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 ± 30 kg) 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 were 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) > 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 1,629 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 > 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 1,908 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 > 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.

Effects of dietary inclusion of dry distillers grains with solubles on performance, carcass characteristics, and nitrogen metabolism in meat sheep: a meta-analysis.

We conducted a meta-analysis in this scientific study to determine the effects of feeding meat sheep dry distillers grains with solubles (DDGS). Thirty-three peer-reviewed articles that met our inclusion requirements and were published between 1997 and 2021 were examined. To calculate the variation in performance, fermentation, carcass features, and nitrogen efficiency between the DDGS and control (no DDGS) treatments, we used 940 sheep weighing an average of 29.1 ± 1.5 kg. We used a hierarchical mixed model to conduct a meta-regression, subset, and dose-response analysis, while taking into consideration categorical variables like breed (pure or cross-breed), and continuous factors, like CP, NDF, and DDGS inclusion rate. Our findings indicate that sheep fed DDGS had higher (p < 0.05) final body weight (51.4 vs. 50.4 kg), neutral detergent fiber digestibility (55.9 vs. 53.8%), and total-tract ether extract digestibility (81.7 vs. 78.7%) than sheep on a control diet. No effects were observed on DMI, CP, and rumen fermentation, but dietary DDGS tended to increase (p = 0.07) HC weight (25.53 vs. 24.6 kg) and meat (redness) color (16.6 vs. 16.3) among treatment comparisons. Dietary DDGS was associated with higher N intake (29.9 vs. 26.8 g/d), fecal N (8.2 vs. 7.8 g/d), and digestibility (71.9 vs. 68.5%). Urinary nitrogen was significantly (p < 0.05) affected linearly by increasing the intake of DDGS in the diet. Based on the dose-response analysis, dietary DDGS inclusion should not exceed 20% to avoid negative effects on performance, nitrogen metabolism, and meat color. Dietary protein from DDGS should not exceed 17% to prevent reduced TVFA concentrations. Breed strongly influenced (p < 0.05) RMD in performance, and inconsistent responses were observed between crossbreed and purebred sheep comparisons. Despite these inconsistencies, no publication bias was observed, but a high variance (Ω2) among comparisons-between-studies was detected. This meta-analysis showed evidence in support of the hypothesis that feeding meat sheep DDGS at a rate of 20% can improve their performance, digestibility, carcass weight, and meat color.

The Effects of Incremental Doses of Aflatoxin B1 on In Vitro Ruminal Nutrient Digestibility and Fermentation Profile of a Lactating Dairy Cow Diet in a Dual-Flow Continuous Culture System.

Aflatoxin B1 (AFB1) is a mycotoxin known to impair human and animal health. It is also believed to have a deleterious effect on ruminal nutrient digestibility under in vitro batch culture systems. The objective of this study was to evaluate the effects of increasing the dose of AFB1 on ruminal dry matter and nutrient digestibility, fermentation profile, and N flows using a dual-flow continuous culture system fed a diet formulated for lactating dairy cows. Eight fermenter vessels were used in a replicated 4 × 4 Latin square design with 10 d periods (7 d adaptation and 3 d sample collection). Treatments were randomly applied to fermenters on diet DM basis: (1) 0 µg of AFB1/kg of DM (Control); (2) 50 µg of AFB1/kg of DM (AF50); (3) 100 µg of AFB1/kg of DM (AF100); and (4) 150 µg of AFB1/kg of DM (AF150). Treatments did not affect nutrient digestibility, fermentation, and N flows. Aflatoxin B1 concentration in ruminal fluid increased with dose but decreased to undetectable levels after 4 h post-dosing. In conclusion, adding incremental doses of AFB1 did not affect ruminal fermentation, digestibility of nutrients, and N flows in a dual-flow continuous culture system fed diets formulated for lactating dairy cows.

Copy number variant-based genome wide association study reveals immune-related genes associated with parasite resistance in a heritage sheep breed from the United States.

Florida Native is a heritage sheep breed in the United States and expresses superior ability to regulate gastrointestinal nematodes. The objective of the present study was to investigate the importance of copy number variants (CNVs) on resistance to natural Haemonchus contortus infections. A total of 300 Florida Native sheep were evaluated. Phenotypic records included fecal egg count (FEC, eggs/gram), FAMACHA© score, percentage cell volume (PCV, %), body condition score (BCS) and average daily gain (ADG, kg). Sheep were genotyped using the GGP Ovine 50K single nucleotide polymorphism (SNP) chip. Log ratios from 45.2 k SNP markers spanning the entire genome were utilized for CNV detection. After quality control, 261 animals with CNVs and phenotypic records were used for the association testing. Association tests were carried out using correlation-trend test and principal component analysis correction to identify CNVs associated with FEC, FAMACHA©, PCV, BCS and ADG. Significant CNVs were detected when their adjusted p-value was <.05 after FDR correction. A total of 8124 CNVs were identified, which gave 246 non-overlapping CNVs. Fourteen CNVs were significantly associated with FEC and PCV. CNVs associated with FEC overlapped 14 Quantitative Trait Locus previously associated with H. contortus resistance. Our study demonstrated for the first time that CNVs could be potentially involved with parasite resistance in Florida Native sheep. Immune-related genes such as CCL1, CCL2, CCL8, CCL11, NOS2, TNF, CSF3 and STAT3 genes could play an important role for controlling H. contortus resistance. These genes could be potentially utilized as candidate markers for selection of parasite resistance in this breed.

Effects of a multicomponent microbial feed additive containing prebiotics and probiotics on health, immune status, metabolism, and performance of newly weaned beef steers during a 35-d receiving period.

We examined the effects of dietary supplementation of a multicomponent 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 postweaning; 206 ± 12 kg of body weight [BW]) from a single source were stratified by BW into four 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; two pens; n = 40 steers) or a basal diet supplemented with SYNB feed additive at an average of 28 g/steer/d (SYNB; two pens; n = 40 steers). The SYNB additive is a blend of live Saccharomyces cerevisiae and the fermentation products of S. cerevisiae, Enterococcus lactis, Bacillus licheniformis, and Bacillus 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 dry matter intake (DMI) and meal events (meal frequency and duration) were measured. Weekly BWs were measured to calculate average daily gain (ADG). Blood samples collected on days 0, 14, 21, 28, and 35 were used for ex-vivo tumor necrosis factor alpha (TNF-α) release assay following lipopolysaccharides (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 (false discovery rate ≤ 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.

Meta-analysis of the effects of the dietary application of exogenous alpha-amylase preparations on performance, nutrient digestibility, and rumen fermentation of lactating dairy cows.

Several studies have evaluated the effects of the dietary application of exogenous alpha-amylase preparations (AMA) as a strategy to increase total tract starch digestibility (TTSD) and milk yield (MY) in dairy cows, but the results have been inconsistent. Thus, the objective of this study was to evaluate the effects of the dietary application of AMA on the performance, digestibility, and rumen fermentation of lactating dairy cows using a meta-analytic method. A total of 18 peer-reviewed manuscripts (N = 32 treatment comparisons) from 2003 to 2019 were systematically identified following the PRISMA method. The weighted raw mean differences between dietary AMA and control treatments were compared with a robust variance estimation. Likewise, diet characteristics like crude protein (CP) content, NDF content, starch content, days in milk (DIM), experimental design (Latin square and continuous), and AMA dose (0 to 732 Kilo Novo units [KNU]/kg TMR) were used as covariates in a meta-regression, subgrouping, and dose-response analysis. Compared to the control, dietary AMA increased (P < 0.05) DM digestibility (69.32% vs. 68.30%), TTSD (94.62% vs. 94.10%), milk protein concentration and yield (3.11% vs. 3.08%; 1.14 vs. 1.10 kg/d) and tended to increase (P = 0.09) fat-corrected milk (35.96 vs. 35.10 kg/d), but no effects were observed on DM intake (22.99 vs. 22.90 kg/d) and feed efficiency (1.50 vs. 1.48). Dietary AMA tended (P = 0.10) to reduce rumen pH (6.27 vs. 6.30). Both the enzyme dose and DIM strongly influenced (P < 0.05) the effects of AMA on digestibility and performance. The dose-response analysis revealed that feeding 600 KNU/kg to high-producing early lactation (< 70 DIM) dairy cows increased FCM and milk protein. Accounting for the type of experimental design was associated with a lower between-studies-variance among comparisons. Overall, this meta-analysis supports the hypothesis that dietary AMA supplementation is associated with a better lactational performance in dairy cows. However, these effects are only suitable for high-producing early lactation dairy cows.

For more than a decade, starch-degrading enzymes (amylolytic enzymes) have been used as a strategy to increase total-tract starch degradation to increase milk yield of dairy cows. Therefore, we conducted a meta-analysis to evaluate the effectiveness of starch-degrading enzymes on starch digestion and milk yield in dairy cows. Collectively, results across the literature suggest that feeding starch-degrading enzymes increased the degradation of starch in the rumen of dairy cows and tended to increase milk yield. Our results suggest that starch-degrading enzymes could increase milk yield in high-producing early lactation dairy cows.

Effect of Lactobacillus hilgardii, Lactobacillus buchneri, or their combination on the fermentation and nutritive value of sorghum silage and corn silage.

The objective of this study was to determine the effect of inoculation with Lactobacillus hilgardii with or without Lactobacillus buchneri on the fermentation, chemical composition, and aerobic stability of sorghum and corn silage after 2 ensiling durations. Sorghum forage was harvested at 27% dry matter (DM; experiment 1), and different corn hybrids were harvested at late (43.8% DM; experiment 2) or normal maturity (34% DM; experiment 3). All harvested forages were chopped and ensiled in quadruplicate in vacuum-sealed nylon-polyethylene bags (40 × 61 cm) for 30 and 90 d after treatment with (1) deionized water (uninoculated) or (2) L. buchneri (1.5 × 105 cfu/g of fresh weight; LB); (3) L. hilgardii (1.5 × 105 cfu/g of fresh weight; LH); or (4) L. buchneri and L. hilgardii (1.5 × 105 cfu/g of fresh weight of each inoculant). Data for each experiment were analyzed separately accounting for the 2 × 2 × 2 factorial treatment arrangement. Inoculating sorghum forage with LB or LH separately increased acetate and 1,2 propanediol concentration, tended to increase DM loss, reduced lactate concentration and the lactate-to-acetate ratio, and increased aerobic stability after 90 but not after 30 d of ensiling. Inoculating late-harvested corn silage with LB or LH separately increased and decreased DM loss, respectively, increased 1,2 propanediol concentration, reduced lactate-to-acetate ratio and yeast counts but did not affect aerobic stability. Inoculating normal-harvested corn silage with LH reduced DM loss and increased 1,2 propanediol concentration and yeast counts; LB reduced lactate concentration, lactate-to-acetate ratio, and total acids. Either inoculant alone increased aerobic stability after 30 or 90 d. The main benefit of combining LB with LH was prevention of increases in DM losses by LH or LB separately. No improvement in aerobic stability resulted from applying LH instead of LB separately or from combining them. Application of LB or LH separately improved aerobic stability of sorghum silage after 90 d and normal-harvested corn silage after 30 or 90 d but did not affect that of late-harvested corn silage.

Meta-analysis of effects of inoculation with Lactobacillus buchneri, with or without other bacteria, on silage fermentation, aerobic stability, and performance of dairy cows.

A meta-analysis of 158 peer-reviewed articles was conducted to examine effects of inoculation with Lactobacillus buchneri (LB)-based inoculants (LBB) that did or did not include homolactic or obligate heterolactic bacteria on silage fermentation and aerobic stability. A complementary meta-analysis of 12 articles examined LBB inoculation effects on dairy cow performance. Raw mean differences between inoculant and control treatment means weighted by inverse variance were compared with a hierarchical effects model that included robust variance estimation. Meta-regression and subgrouping analysis were used to identify effects of covariates including forage type, application rate (≤104, 105, 106, or ≥ 107 cfu/g as fed), bacteria type (LB vs. LB plus other bacteria), enzyme inclusion, ensiling duration, and silo type (laboratory or farm scale). Inoculation with LBB increased acetate (62%), 1, 2 propanediol (364%) and propionate (30%) concentration and aerobic stability (73.8%) and reduced lactate concentration (7.2%), yeast counts (7-fold) and mold counts (3-fold). Feeding inoculated silage did not affect milk yield, dry matter intake, and feed efficiency in lactating dairy cows. However, forage type, inoculant composition, and dose effects on silage quality measures were evident. Inoculation with LBB increased aerobic stability of all silages except tropical grasses. Adding obligate homolactic or facultative heterolactic bacteria to LB prevented the small increase in DM losses caused by LB alone. The 105 and 106 cfu/g rates were most effective at minimizing DM losses while aerobic stability was only increased with 105, 106, and ≥ 107 cfu/g rates. Inoculation with LBB increased acetate concentration, reduced yeast counts and improved aerobic stability but did not improve dairy cow performance.

Effects of a blend of mannan and glucan on growth performance, apparent nutrient digestibility, energy status, and whole-blood immune gene expression of beef steers during a 42-d receiving period.

We examined the effects of dietary supplementation of a blend of mannan and glucan on the growth performance, energy status, and whole-blood immune gene expression of newly weaned beef steers during a 42-d receiving period. Forty-eight newly weaned Angus crossbred steers (2-d post-weaning; 199 ± 13 kg of initial body weight [BW]) from a single source were stratified by BW and randomly assigned to one of the two treatments: basal diet with no additive (CON; n = 24) or a basal diet top-dressed with 5 g of a blend of mannan and glucan (MANGLU; n = 24). Average daily gain (ADG) and feed efficiency (FE) from days 1 to 14, 15 to 42, and 1 to 42 were calculated from daily dry matter intake (DMI) and weekly BW. Blood samples were collected on days 0, 14, and 42 for measurement of plasma glucose and nonesterified fatty acids (NEFA). Blood samples collected on days 14 and 42 were composited for each steer for untargeted carbonyl-metabolome analysis (measurement of carbonyl-containing metabolites). Expression of 84 immune-related genes was analyzed on blood samples collected on day 42. Beginning on days 37 to 42, total mixed ration, refusals, and fecal samples were collected once daily to determine apparent total tract digestibility of DM, CP, NDF, and ADF using indigestible NDF as an internal marker. Over the 42-d feeding trial, supplemental MANGLU tended to increase final BW (P = 0.07) and ADG (P = 0.06). Compared to CON, beef steers fed supplemental MANGLU had greater (P = 0.01) DMI during the first 14 d, greater DM digestibility (P = 0.03), and tended to have greater NDF digestibility (P = 0.09). No treatment effects (P > 0.10) on plasma glucose and NEFA on days 14 and 42 were detected; however, carbonyl-metabolome analysis revealed increased (FDR ≤ 0.05) plasma concentrations of galactose and glyceraldehydes, and altered (FDR ≤ 0.05) concentrations of some microbiome-derived metabolites in beef steers fed MANGLU. Compared with CON, MANGLU increased (P ≤ 0.05) the expression of five immune-related genes involved in recognition of and mounting immune defense against microbial pathogens. In conclusion, the results of this study demonstrated that supplemental MANGLU enhances beef cattle immunocompetence and productivity during feedlot receiving period.

Effects of Multi-Species Direct-Fed Microbial Products on Ruminal Metatranscriptome and Carboxyl-Metabolome of Beef Steers.

We examined the effects of two direct-fed microbial (DFM) products containing multiple microbial species and their fermentation products on ruminal metatranscriptome and carboxyl-metabolome of beef steers. Nine ruminally-cannulated Holstein steers were assigned to 3 treatments arranged in a 3 × 3 Latin square design with three 21-d periods. Dietary treatments were (1) Control (CON; basal diet without additive), (2) Commence (PROB; basal diet plus 19 g/d of Commence), and (3) RX3 (SYNB; basal diet plus 28 g/d of RX3). Commence and RX3 are both S. cerevisiae-based DFM products containing several microbial species and their fermentation products. Mixed ruminal contents collected multiple times after feeding on day 21 were used for metatranscriptome and carboxyl-metabolome analysis. Partial least squares discriminant analysis revealed a distinct transcriptionally active taxonomy profiles between CON and each of the PROB and SYNB samples. Compared to CON, the steers fed supplemental PROB had 3 differential (LDA ≥ 2.0; p ≤ 0.05) transcriptionally active taxa, none of which were at the species level, and those fed SYNB had eight differential (LDA > 2.0, p ≤ 0.05) transcriptionally active taxa, but there was no difference (p > 0.05) between PROB and SYNB. No functional microbial genes were differentially expressed among the treatments. Compared with CON, 3 metabolites (hydroxylpropionic acid and 2 isomers of propionic acid) were increased (FC ≥ 1.2, FDR ≤ 0.05), whereas 15 metabolites, including succinic acid and fatty acid peroxidation and amino acid degradation products were reduced (FC ≤ 0.83, FDR ≤ 0.05) by supplemental PROB. Compared with CON, 2 metabolites (2 isomers of propionic acid) were increased (FC ≥ 1.2, FDR ≤ 0.05), whereas 2 metabolites (succinic acid and pimelate) were reduced (FC ≤ 0.83, FDR ≤ 0.05) by supplemental SYNB. Compared to SYNB, supplemental PROB reduced (FC ≤ 0.83, FDR ≤ 0.05) the relative abundance of four fatty acid peroxidation products in the rumen. This study demonstrated that dietary supplementation with either PROB or SYNB altered the ruminal fermentation pattern. In addition, supplemental PROB reduced concentrations of metabolic products of fatty acid peroxidation and amino acid degradation. Future studies are needed to evaluate the significance of these alterations to ruminal fatty acid and amino acid metabolisms, and their influence on beef cattle performance.

An expansin-like protein expands forage cell walls and synergistically increases hydrolysis, digestibility and fermentation of livestock feeds by fibrolytic enzymes.

Bacterial expansin-like proteins have synergistically increased cellulose hydrolysis by cellulolytic enzymes during the initial stages of biofuel production, but they have not been tested on livestock feeds. The objectives of this study were to: isolate and express an expansin-like protein (BsEXLX1), to verify its disruptive activity (expansion) on cotton fibers by immunodetection (Experiment 1), and to determine the effect of dose, pH and temperature for BsEXLX1 and cellulase to synergistically hydrolyze filter paper (FP) and carboxymethyl cellulose (CMC) under laboratory (Experiment 2) and simulated ruminal (Experiment 3) conditions. In addition, we determined the ability of BsEXLX1 to synergistically increase hydrolysis of corn and bermudagrass silages by an exogenous fibrolytic enzyme (EFE) (Experiment 4) and how different doses of BsEXLX1 and EFE affect the gas production (GP), in vitro digestibility and fermentation of a diet for dairy cows (Experiment 5). In Experiment 1, immunofluorescence-based examination of cotton microfiber treated without or with recombinant expansin-like protein expressed from Bacillus subtilis (BsEXLX1) increased the surface area by > 100% compared to the untreated control. In Experiment 2, adding BsEXLX1 (100 µg/g FP) to cellulase (0.0148 FPU) increased release of reducing sugars compared to cellulase alone by more than 40% (P < 0.01) at optimal pH (4.0) and temperature (50°C) after 24 h. In Experiment 3 and 4, adding BsEXLX1 to cellulase or EFE, synergistically increased release of reducing sugars from FP, corn and bermudagrass silages under simulated ruminal conditions (pH 6.0, 39°C). In Experiment 5, increasing the concentration of BsEXLX1 linearly increased (P < 0.01) GP from fermentation of a diet for dairy cows by up to 17.8%. Synergistic effects between BsEXLX1 and EFE increased in vitro NDF digestibility of the diet by 23.3% compared to the control. In vitro digestibility of hemicellulose and butyrate concentration were linearly increased by BsEXLX1 compared to the control. This study demonstrated that BsEXLX1 can improve the efficacy of cellulase and EFE at hydrolyzing pure substrates and dairy cow feeds, respectively.

DI/LC-MS/MS-Based Metabolome Analysis of Plasma Reveals the Effects of Sequestering Agents on the Metabolic Status of Dairy Cows Challenged with Aflatoxin B1.

The study applied a targeted metabolomics approach that uses a direct injection and tandem mass spectrometry (DI-MS/MS) coupled with a liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based metabolomics of plasma to evaluate the effects of supplementing clay with or without Saccharomyces cerevisiae fermentation product (SCFP) on the metabolic status of dairy cows challenged with aflatoxin B1. Eight healthy, lactating, multiparous Holstein cows in early lactation (64 ± 11 DIM) were randomly assigned to one of four treatments in a balanced 4 × 4 duplicated Latin square design with four 33 d periods. Treatments were control, toxin (T; 1725 µg aflatoxin B1 (AFB1)/head/day), T with clay (CL; 200 g/head/day), and CL with SCFP (YEA; 35 g of SCFP/head/day). Cows in T, CL, and YEA were dosed with aflatoxin B1 (AFB1) from days 26 to 30. The sequestering agents were top-dressed from day 1 to 33. On day 30 of each period, 15 mL of blood was taken from the coccygeal vessels and plasma samples were obtained from blood by centrifugation and analyzed for metabolites using a kit that combines DI-MS/MS with LC-MS/MS-based metabolomics. The data were analyzed using the GLIMMIX procedure of SAS. The model included the effects of treatment, period, and random effects of cow and square. Significance was declared at p ≤ 0.05. Biomarker profiles for aflatoxin ingestion in dairy cows fed no sequestering agents were determined using receiver-operator characteristic (ROC) curves, as calculated by the ROCCET web server. A total of 127 metabolites such as amino acids, biogenic amines, acylcarnitines, glycerophospholipids, and organic acids were quantified. Compared with the control, T decreased (p < 0.05) plasma concentrations of alanine, leucine, and arginine and tended to decrease that of citrulline. Treatment with CL had no effects on any of the metabolites relative to the control but increased (p ≤ 0.05) concentrations of alanine, leucine, arginine, and that of citrulline (p = 0.07) relative to T. Treatment with YEA resulted in greater (p ≤ 0.05) concentrations of aspartic acid and lysine relative to the control and the highest (p ≤ 0.05) plasma concentrations of alanine, valine, proline, threonine, leucine, isoleucine, glutamic acid, phenylalanine, and arginine compared with other treatments. The results of ROC analysis between C and T groups revealed that the combination of arginine, alanine, methylhistidine, and citrulline had sufficient specificity and sensitivity (area under the curve = 0.986) to be excellent potential biomarkers of aflatoxin ingestion in dairy cows fed no sequestering agents. This study confirmed the protective effects of sequestering agents in dairy cows challenged with aflatoxin B1.

Metatranscriptomic Analysis of Sub-Acute Ruminal Acidosis in Beef Cattle.

Subacute ruminal acidosis (SARA) is a metabolic disease of ruminants characterized by low pH, with significant impacts on rumen microbial activity, and animal productivity and health. Microbial changes during subacute ruminal acidosis have previously been analyzed using quantitative PCR and 16S rRNA sequencing, which do not reveal the actual activity of the rumen microbial population. Here, we report the functional activity of the rumen microbiota during subacute ruminal acidosis. Eight rumen-cannulated Holstein steers were assigned randomly to acidosis-inducing or control diet. Rumen fluid samples were taken at 0, 3, 6, and 9 h relative to feeding from both treatments on the challenge day. A metatranscriptome library was prepared from RNA extracted from the samples and the sequencing of the metatranscriptome library was performed on Illumina HiSeq4000 following a 2 × 150 bp index run. Cellulolytic ruminal bacteria including Fibrobacter succinogenes, Ruminococcus albus, and R. bicirculans were reduced by an induced acidotic challenge. Up to 68 functional genes were differentially expressed between the two treatments. Genes mapped to carbohydrate, amino acid, energy, vitamin and co-factor metabolism pathways, and bacterial biofilm formation pathways were enriched in beef cattle challenged with sub-acute acidosis. This study reveals transcriptionally active taxa and metabolic pathways of rumen microbiota during induced acidotic challenge.