Bioactive metabolites of Asparagopsis stabilized in canola oil completely suppress methane emissions in beef cattle fed a feedlot diet.

Asparagopsis taxiformis (Asparagopsis) has been shown to be highly efficacious at inhibiting the production of methane (CH4) in ruminants. To date, Asparagopsis has been primarily produced as a dietary supplement by freeze-drying to retain the volatile bioactive compound bromoform (CHBr3) in the product. Steeping of Asparagopsis bioactive compounds into a vegetable oil carrier (Asp-Oil) is an alternative method of stabilizing Asparagopsis as a ruminant feed additive. A dose-response experimental design used 3 Asp-Oil-canola oil blends, low, medium, and high Asp-Oil which provided 17, 34, and 51 mg Asparagopsis derived CHBr3/kg dry matter intake (DMI), respectively (in addition to a zero CHBr3 canola oil control), in a tempered-barley based feedlot finisher diet, fed for 59 d to 20 Angus heifers (five replicates per treatment). On four occasions, live weight was measured and CH4 emissions were quantified in respiration chambers, and blood, rumen fluid, and fecal samples were collected. At the end of the experiment, all animals were slaughtered, with carcasses graded, and samples of meat and edible offal collected for testing of consumer sensory qualities and residues of CHBr3, bromide, and iodide. All Asp-Oil treatments reduced CH4 yield (g CH4/kg DMI, P = 0.008) from control levels, with the low, medium, and high Asp-Oil achieving 64%, 98%, and 99% reduction, respectively. Dissolved hydrogen increased linearly with increasing Asp-Oil inclusion, by more than 17-fold in the high Asp-Oil group (P = 0.017). There was no effect of Asp-Oil treatment on rumen temperature, pH, reduction potential, volatile fatty acid and ammonia production, rumen pathology, and histopathology (P > 0.10). There were no differences in animal production and carcass parameters (P > 0.10). There was no detectable CHBr3 in feces or any carcass samples (P > 0.10), and iodide and bromide residues in kidneys were at levels unlikely to lead to consumers exceeding recommended maximum intakes. Overall, Asp-Oil was found to be safe for animals and consumers of meat, and effective at reducing CH4 emissions and yield by up to 99% within the range of inclusion levels tested.

Red seaweed, Asparagopsis taxiformis (Asparagopsis), has been shown to be highly effective at inhibiting the production of methane (CH4) in ruminants. An alternative to feeding whole, freeze-dried Asparagopsis is steeping the biomass in vegetable oil to stabilize the bioactive compounds (Asp-Oil) and feeding Asp-Oil to ruminants as a component of their dietary intake. This experiment measured the CH4 reduction potential and safety of Asp-Oil in a trial with 20 Angus heifers, fed iso-fat feedlot diets containing one of the three levels of Asp-Oil, or a control oil. Compared to the control, bromoform inclusion levels of 17, 34, and 51 mg/kg of dry matter (DM; low, medium, high) reduced CH4 yield (g CH4/kg DM intake) by 64%, 98%, and 99%, respectively. There were no effects on animal production or carcass characteristics. There were no impacts on animal health, welfare, or rumen function. Carcasses were safe for human consumption, and there was no bromoform detected in any carcass samples. Overall, Asp-Oil was found to effectively reduce CH4 emissions and is safe for animals and consumers of meat and edible offal.

Metabolomics and proteomics insights into subacute ruminal acidosis etiology and inhibition of proliferation of yak rumen epithelial cells in vitro.

BACKGROUND: Untargeted metabolomics and proteomics were employed to investigate the intracellular response of yak rumen epithelial cells (YRECs) to conditions mimicking subacute rumen acidosis (SARA) etiology, including exposure to short-chain fatty acids (SCFA), low pH5.5 (Acid), and lipopolysaccharide (LPS) exposure for 24 h. RESULTS: These treatments significantly altered the cellular morphology of YRECs. Metabolomic analysis identified significant perturbations with SCFA, Acid and LPS treatment affecting 259, 245 and 196 metabolites (VIP > 1, P < 0.05, and fold change (FC) ≥ 1.5 or FC ≤ 0.667). Proteomic analysis revealed that treatment with SCFA, Acid, and LPS resulted in differential expression of 1251, 1396, and 242 proteins, respectively (FC ≥ 1.2 or ≤ 0.83, P < 0.05, FDR < 1%). Treatment with SCFA induced elevated levels of metabolites involved in purine metabolism, glutathione metabolism, and arginine biosynthesis, and dysregulated proteins associated with actin cytoskeleton organization and ribosome pathways. Furthermore, SCFA reduced the number, morphology, and functionality of mitochondria, leading to oxidative damage and inhibition of cell survival. Gene expression analysis revealed a decrease the genes expression of the cytoskeleton and cell cycle, while the genes expression associated with inflammation and autophagy increased (P < 0.05). Acid exposure altered metabolites related to purine metabolism, and affected proteins associated with complement and coagulation cascades and RNA degradation. Acid also leads to mitochondrial dysfunction, alterations in mitochondrial integrity, and reduced ATP generation. It also causes actin filaments to change from filamentous to punctate, affecting cellular cytoskeletal function, and increases inflammation-related molecules, indicating the promotion of inflammatory responses and cellular damage (P < 0.05). LPS treatment induced differential expression of proteins involved in the TNF signaling pathway and cytokine-cytokine receptor interaction, accompanied by alterations in metabolites associated with arachidonic acid metabolism and MAPK signaling (P < 0.05). The inflammatory response and activation of signaling pathways induced by LPS treatment were also confirmed through protein interaction network analysis. The integrated analysis reveals co-enrichment of proteins and metabolites in cellular signaling and metabolic pathways. CONCLUSIONS: In summary, this study contributes to a comprehensive understanding of the detrimental effects of SARA-associated factors on YRECs, elucidating their molecular mechanisms and providing potential therapeutic targets for mitigating SARA.

The addition of curcumin to the diet of post-weaning dairy calves: effects on ruminal fermentation, immunological, and oxidative responses.

Incorporating Curcumin into animal diets holds significant promise for enhancing both animal health and productivity, with demonstrated positive impacts on antioxidant activity, anti-microbial responses. Therefore, this study aimed to determine whether adding Curcumin to the diet of dairy calves would influence ruminal fermentation, hematologic, immunological, oxidative, and metabolism variables. Fourteen Jersey calves were divided into a control group (GCON) and a treatment group (GTRA). The animals in the GTRA received a diet containing 65.1 mg/kg of dry matter (DM) Curcumin (74% purity) for an experimental period of 90 days. Blood samples were collected on days 0, 15, 45, and 90. Serum levels of total protein and globulins were higher in the GTRA group (P < 0.05) than the GCON group. In the GTRA group, there was a reduction in pro-inflammatory cytokines (IL-1ß and IL-6) (P < 0.05) and an increase in IL-10 (which acts on anti-inflammatory responses) (P < 0.05) when compared to the GCON. There was a significantly higher (P < 0.05) concentration of immunoglobulin A (IgA) in the serum of the GTRA than the GCON. A Treatment × Day interaction was observed for haptoglobin levels, which were higher on day 90 in animals that consumed Curcumin than the GCON (P < 0.05). The catalase and superoxide dismutase activities were significantly higher (P < 0.05) in GTRA, reducing lipid peroxidation when compared to the GCONT. Hematologic variables did not differ significantly between groups. Among the metabolic variables, only urea was higher in the GTRA group when compared to the GCON. Body weight and feed efficiency did not differ between groups (meaning the percentage of apparent digestibility of dry matter, crude protein, and acid detergent fiber (ADF) and neutral detergent fiber (NDF). There was a tendency (P = 0.09) for treatment effect and a treatment x day interaction (P = 0.05) for levels of short-chain fatty acids in rumen fluid, being lower in animals that consumed curcumin. There was a treatment vs. day interaction (P < 0.05) for the concentration of acetate in the rumen fluid (i.e., on day 45, had a reduction in acetate; on day 90, values were higher in the GTRA group when compared to the GCON). We conclude that there was no evidence in the results from this preliminary trial that Curcumin in the diet of dairy calves interfered with feed digestibility. Curcumin may have potential antioxidant, anti-inflammatory, and immune effects that may be desirable for the production system of dairy calves.

Trace mineral source and chromium propionate supplementation affect performance and carcass characteristics in feedlot steers.

Angus-crossbred steers (n = 400; 369.7 ±â€…7.6 kg) were used to determine the influence of trace mineral (TM) source and chromium propionate (Cr Prop) supplementation on performance, carcass characteristics, and ruminal and plasma variables in finishing steers. Steers were blocked by body weight (BW) and randomly assigned within block to treatments in a 2 × 2 factorial arrangement, with factors being: 1) TM source (STM or HTM) and 2) Cr supplementation (0 or 0.25 mg Cr/kg DM, -Cr or + Cr, respectively). Treatments consisted of the addition of: 1) sulfate TM (STM; 90, 40, and 18 mg/kg DM of Zn, Mn, and Cu, respectively), 2) STM and 0.25 mg Cr/kg DM from Cr Prop, 3) hydroxychloride TM (HTM; 90, 40, and 18 mg/kg DM of Zn, Mn, and Cu, respectively), and 4) HTM and 0.25 mg Cr/kg DM from Cr Prop. Each treatment consisted of 10 replicate pens with 10 steers per pen. Body weights were obtained on consecutive days at the initiation and termination of the 154-d study. Steers were fed a steam-flaked corn-based finishing diet. Ractopamine hydrochloride was fed for the last 31 d of the study. Ruminal fluid and blood samples were obtained from one steer per pen on days 28 and 84 for ruminal volatile fatty acids (VFA) and plasma TM and glucose analysis. Steers were slaughtered at the end of the study and individual carcass data were collected. No Cr × TM source interactions (P = 0.48) were detected. Steers supplemented with HTM had greater (P = 0.04) hot carcass weight (HCW), dressing percentage (DP), longissimus muscle (LM) area, and USDA yield grade (YG), and tended (P = 0.12) to have greater average daily gain (ADG) than those receiving STM. Average daily gain, gain:feed, dressing percentage, and longissimus muscle area were greater (P = 0.04) for + Cr steers compared to-Cr steers. Hot carcass weight tended (P = 0.06) to be greater for + Cr steers. Ruminal acetate concentrations at 28 d were lesser (P = 0.01) for HTM vs. STM steers, and greater (P = 0.04) for + Cr steers compared to-Cr steers. Plasma concentrations of Zn, Cu, and Mn were not affected by TM source or Cr supplementation. Steers supplemented with Cr had greater (P = 0.05) plasma glucose concentrations than-Cr steers at 28 but not at 84 d. Results of this study indicate replacing STM with HTM improved carcass characteristics in finishing steers, and Cr Prop supplementation improved steer performance and carcass characteristics.

Trace minerals (TM) are supplemented to finishing cattle diets to prevent TM deficiencies. Sources of TM differ in their bioavailability and effect on rumen fermentation. Chromium is a TM required in low concentrations to enhance insulin activity. We tested the effect of TM source (hydroxychloride; HTM vs. sulfate; STM) and supplemental Cr propionate (Cr Prop) on performance and carcass characteristics of finishing steers. Providing 0.25 mg of supplemental Cr/kg DM, from Cr Prop, improved gain, feed efficiency, and carcass characteristics in steers. Steers supplemented with HTM tended to gain faster and had improved carcass characteristics of economic importance compared to those supplemented with STM.

Effects of biotin and coated cobalamin on lactation performance, nutrient digestion and rumen fermentation in Holstein dairy cows.

Biotin (BI) and cobalamin (CA) are essential for rumen propionate production and hepatic gluconeogenesis. The study evaluated the influence of BI or/and coated CA (CCA) on milk performance and nutrient digestion in cows. Sixty Holstein dairy cows were assigned in a 2 × 2 factorial arrangement and randomised block design to four groups. The factors were BI at 0 or 20 mg/day and CCA at 0 or 9 mg CA/day. Dry matter intake increased with BI addition but was unchanged with CCA supply. Addition of BI or CCA increased fat-corrected milk, milk fat and milk protein yields and feed efficiency. Moreover, lactose yield was increased by CCA addition. Dry matter, organic matter, crude protein and acid detergent fibre total-tract digestibility increased for BI or CCA supply. When CCA was supplemented, positive response of neutral detergent fibre digestibility to BI addition was enhanced. Supplementing BI did not affect pH, propionate content and acetate to propionate ratio, but increased total volatile fatty acids (VFA) and acetate contents. Supplementing CCA decreased pH and acetate to propionate ratio, but increased total VFA, acetate and propionate contents. Rumen protease and carboxymethyl-cellulase activities and fungi, bacteria and Butyrivibrio fibrisolvens numbers increased for BI or CCA supply. In addition, protozoa increased for BI addition, and protease activity and Prevotella ruminicola increased for CCA supply. When CCA was supplemented, positive responses of R. albus and Ruminobacter amylophilus numbers to BI addition were enhanced. Blood glucose concentration was unchanged with BI supply, but increased for CCA supply. Blood nonesterified fatty acids and ß-hydroxybutyrate contents reduced with BI or CCA supply. Supplementation with BI or CCA increased blood BI or CA content. The results showed that supplementing BI or/and CCA improved lactation performance and nutrient digestion, and CCA supply did not enhance the lactation performance response to BI supply.

Niacin mitigates rumen epithelial damage in vivo by inhibiting rumen epithelial cell apoptosis on a high concentrate diet.

To investigate the effects of niacin on rumen fermentation, rumen epithelial antioxidant activity, and rumen epithelial cell apoptosis on high concentrate (HC) diets, nine male Hu sheep were randomly fed one of three diets: low concentrate diet (LC; concentrate: forage (C:F) = 20:80, high concentrate diet (HC; C:F = 80:20), and HCN diet (HC diet + niacin at 800 mg/kg diet air-dry matter). Compared with the LC group, the HC group had a lower rumen pH, higher volatile fatty acids and lactic acid in the rumen, reduced activity of antioxidant enzymes and total antioxidant capacity, and increased malondialdehyde content in the rumen epithelium (P < 0.05). Rumen epithelial papilla morphology was decreased, and apoptosis-related indicators and serum inflammatory cytokines were increased in the HC group over the LC group (P < 0.05). Compared with the HC diet, the HCN diet increased rumen pH, rumen epithelium antioxidant capacity, and rumen epithelial papilla morphology, decreased rumen lactate content, serum inflammatory cytokines, and apoptosis-related indicators (P < 0.05). Therefore, adding 800 mg/kg niacin helped protect against rumen epithelial damage by avoiding drastic changes in the rumen environment and improved rumen epithelial antioxidant capacity to inhibit rumen epithelial cell apoptosis in sheep on a HC diet.

Sulfur, fresh cassava root and urea independently enhanced gas production, ruminal characteristics and in vitro degradability.

BACKGROUND: Total fresh cassava root (FCR) production was 275 million tonnes in 2018 which equals 61.1 % of the total production, and Thailand produced 10.7 % FCR of the total production. FCR is one of the main energy source for ruminant. The limitation of FCR utilization is due to the presence of hydrogen cyanide (HCN). The study aimed to evaluate the effect of sulfur, urea and FCR at various levels on in vitro gas production, ruminal fermentation and in vitro degradability. The study hypothesized that: (1) sulfur, urea and FCR have no interaction effect and (2) effect of FCR and urea is related to sulfur addition. RESULTS: The study aimed to elucidate the optimum level of elemental sulfur, fresh cassava root (FCR) and urea and their effect on in vitro gas production, ruminal fermentation, thiocyanate concentration, and in vitro degradability. A 3 × 2 × 4 in a completely randomized design were conducted. Factor A was level of sulfur at 0 %, 1 and 2 % of concentrate dry matter (DM), factor B was level of urea at 2 and 4 % of concentrate DM, and factor C was level of the FCR at 0, 200, 300 and 400 mg DM of the total substrate. The study found that elemental sulfur, urea and FCR had no interaction effect on the kinetics of in vitro gas, ruminal fermentation, HCN and in vitro degradability. Elemental sulfur supplementation (P < 0.05) significantly increased the in vitro gas produced from an insoluble fraction (b), in vitro DM degradability and either neutral detergent fiber (NDF) or acid detergent fiber (ADF) degradability and propionate (C3) concentration while decreased the ruminal HCN concentration. Urea levels showed a (P < 0.05) significant increase of the potential extent of in vitro gas production, ruminal ammonia nitrogen (NH3-N) and total volatile fatty acid (TVFA). Fresh cassava root supplementation (P < 0.05) significantly increased the in vitro gas produced from an immediate soluble fraction (a), in vitro gas produced from insoluble fraction, in vitro gas production rate constant, total VFA, C3 concentration and HCN while decreased ruminal pH, acetate and butyrate concentration. It could be concluded that 2 % elemental sulfur, 4 % urea and 300 mg FCR showed a greater effect on in vitro gas production, ruminal fermentation and HCN reduction. CONCLUSIONS: The study found that elemental sulfur, urea, and FCR had no interaction effect on the kinetics of in vitro gas, total in vitro gas, ruminal fermentation, and HCN concentration. It could be concluded that 2 % elemental sulfur, 4 % urea, and 300 mg FCR showed a greater effect on in vitro gas production, ruminal fermentation, and HCN reduction.

Anti-lipopolysaccharide antibody mitigates ruminal lipopolysaccharide release without acute-phase inflammation or liver transcriptomic responses in Holstein bulls.

Anti-lipopolysaccharide (LPS) antibody administration has the potential benefits of neutralizing and consequently controlling rumen-derived LPS during subacute ruminal acidosis. Four Holstein bulls were used in this crossover study with a 2-week wash-out period. Anti-LPS antibody (0 or 4 g) was administered once daily for 14 days. Significantly lower ruminal LPS and higher 1-h mean ruminal pH were identified in the 4 g group. However, blood metabolites, acute-phase proteins, cytokines, and hepatic transcriptomes were not different between the two groups. Therefore, anti-LPS antibody administration mitigated ruminal LPS release and pH depression without accompanying responses in acute-phase inflammation or hepatic transcriptomic expression.

Crude saponin extract from Sesbania grandiflora (L.) Pers pod meal could modulate ruminal fermentation, and protein utilization, as well as mitigate methane production.

The aim of the study was to conduct a basic evaluation of the in vitro effect of crude protein (CP) levels in concentrate and a saponin extract from Sesbania graniflora pods meal (SES) on the kinetics of gas, nutrient digestibility, ruminal fermentation, protein efficiency uses, and methane (CH4) mitigation. Eight treatments were formed according to a 2 × 4 factorial design in a completely randomized design (CRD). The first factor referred to the levels of CP at 14 and 16% on dry matter (DM) basis in the concentrate diet, and the second factor referred to the levels of SES supplementation at 0, 0.2, 0.4, and 0.6% of the total substrate on a DM basis. The results showed that S. graniflora pod meal contained 21.73% CP, 10.87% condensed tannins, and 16.20% crude saponins, respectively. Most kinetics of gas as well as cumulative gas were not influenced by the CP levels or SES addition (P > 0.05) except gas production from immediately soluble fraction (a) was significantly different by CP levels. Ammonia-nitrogen concentration of incubation at 4 h was significantly difference based on the CP levels and SES supplementation (P < 0.05). Increasing SES levels significantly (P < 0.05) decreased protozoal population. In vitro digestibility of DM and organic matter was not changed by CP levels or SES addition. Butyrate and acetate to propionate ration were decreased, and propionate was increased when increasing SES dose (P < 0.05), while CP levels did not change total volatile fatty acids and molar portions. The ruminal CH4 concentration was reduced by 44.12% when 0.6% SES was added after 8 h of incubation. Therefore, SES supplementation could enhance protein utilization and improve rumen fermentation particularly lowering CH4 production.

Estimation of ruminal outflow in buffaloes fed diets with different energy and protein sources by use of reticular and omasal sampling.

This study aimed to evaluate the effects of different traditional or alternative energy and protein sources, associated or not, on feeding behavior, ruminal kinetics, and post-ruminal flow of nutrients. Besides, it was assessed diets' effects on different sites (reticulum and omasum) of buffaloes. Four ruminally cannulated male Murrah buffaloes (average initial weight of 637 ± 66.37 kg) were randomly distributed in a 4 × 4 Latin square design. Treatments were arranged as 2 × 2 factorial arrangement. The first factor evaluated was the inclusion of energy sources (ground corn and crude glycerin), and the second factor was the inclusion of protein sources (soybean meal and cottonseed cake). Buffaloes fed cottonseed cake had a higher content of neutral detergent fiber (NDF) and potentially digestible detergent fiber (pdNDF) in the rumen environment than buffaloes fed soybean meal. There was a sampling site effect on rumen digestion rates of pdNDF, passage rates of indigestible neutral detergent fiber (iNDF), and pdNDF, and flow of iNDF. In this study, omasal collections were more representative. Total replacement of ground corn by crude glycerin promoted less NDF ruminal digestibility, and care should be taken to include this energy source. The cottonseed cake does not cause a difference in rumen dynamics and can totally replace soybean meal in feedlot buffaloes' diet.

Effects of pomegranate peel extract on ruminal and post-ruminal in vitro degradation of rumen inoculum of the dairy cow.

This experiment was carried out to study the effect of water extracted pomegranate peel extract (PE) on ruminal protein degradation and post-ruminal digestion in the dairy cow. PE was added at six levels of total phenolics (g/kg of the basal diet); 3.75 (PE1); 4.4 (PE2); 5.05 (PE3); 5.70 (PE4); and 6.35 (PE5). Rumen degradable crude protein (rdCP) decreased with PE addition (L < 0.0001), but total CP degradability (tdCP) was not affected. Compared to PE0, PE2, and PE3 diets showed higher (L = 0.054, Q = 0.029) digestibility of bypass CP (dBCP). Increasing levels of PE resulted in a decrease in proteolytic bacteria numbers (p < 0.0001). At PE4 and PE5 levels, total VFA and acetate concentrations linearly decreased compared to PE0. PE inclusion lowered the acetate:propionate ratio (L = 0.0001) and Ammonia-N production after 24 h (L = 0.0008) of incubation. The total number of protozoa, genera Dasytricha and Isotricha, and subfamilies Entodiniinae, Diplodiniinae, and Ophrioscolecinae decreased with increasing dietary PE concentration (p < 0.0001). The results suggest that all levels of PE addition reduce the protozoal population and Ammonia-N concentration. All PE levels slowed down protein degradation in the rumen but PE2 and PE3 showed the greatest effect.

Ruminal Degradation of Rumen-Protected Glucose Influences the Ruminal Microbiota and Metabolites in Early-Lactation Dairy Cows.

Rumen-protected glucose (RPG) plays an important role in alleviating the negative energy balance of dairy cows. This study used a combination of rumen microbes 16S and metabolomics to elucidate the changes of rumen microbial composition and rumen metabolites of different doses of RPG's rumen degradation part in early-lactation dairy cows. Twenty-four multiparous Holstein cows in early lactation were randomly allocated to control (CON), low-RPG (LRPG), medium-RPG (MRPG), or high-RPG (HRPG) groups in a randomized block design. The cows were fed a basal total mixed ration diet with 0, 200, 350, and 500 g of RPG per cow per day, respectively. Rumen fluid samples were analyzed using Illumina MiSeq sequencing and ultrahigh-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry. MRPG supplementation increased bacterial richness and diversity, including increasing the relative abundance of cellulolytic bacteria, such as Ruminococcus, Lachnospiraceae_NK3A20_group, Ruminiclostridium, and Lachnospiraceae_UCG-008 MRPG significantly increased the concentrations of acetate, propionate, butyrate, and total volatile fatty acid in the rumen. Ruminal fluid metabolomics analysis showed that RPG supplementation could significantly regulate the synthesis of amino acids digested by protozoa in the rumen. Correlation analysis of the ruminal microbiome and metabolome revealed some potential relationships between major bacterial abundance and metabolite concentrations. Our analysis found that RPG supplementation of different doses can change the diversity of microorganisms in the rumen and affect the rumen fermentation pattern and microbial metabolism and that a daily supplement of 350 g of RPG might be the ideal dose.IMPORTANCE Dairy cows in early lactation are prone to a negative energy balance because their dry matter intake cannot meet the energy requirements of lactation. Rumen-protected glucose is used as an effective feed additive to alleviate the negative energy balance of dairy cows in early lactation. However, one thing that is overlooked is that people often think that rumen-protected glucose is not degraded in the rumen, thus ignoring its impact on the microorganisms in the rumen environment. Our investigation and previous experiments have found that rumen-protected glucose is partially degraded in the rumen. However, there are few reports on this subject. Therefore, we conducted research on this problem and found that rumen-protected glucose supplementation at 350 g/day can promote the development and metabolism of rumen flora. This provides a theoretical basis for the extensive application of rumen bypass glucose at a later stage.

Effects of a proinflammatory response on metabolic function of cultured, primary ruminal epithelial cells.

Inflammation of ruminal epithelium may occur during ruminal acidosis as a result of translocation and interaction of ruminal epithelial cells (REC) with molecules such as lipopolysaccharide (LPS). Such inflammation has been reported to alter cellular processes such as nutrient absorption, metabolic regulation, and energy substrate utilization in other cell types but has not been investigated for REC. The objectives of this study were to investigate the effects of LPS on metabolism of short-chain fatty acids by primary REC, as well as investigating the effects of media containing short-chain fatty acids on the proinflammatory response. Ruminal papillae from 9 yearling Speckle Park beef heifers were used to isolate and culture primary REC. Cells were grown in minimum essential medium (MEM) for 12 d before use and then reseeded in 24-well culture plates. The study was conducted as a 2 × 2 factorial, where cells were grown in unaltered MEM (REG) or medium containing 2 mM butyrate and 5 mM propionate (SCFA) with (50,000 EU/mL; +LPS) or without LPS (-LPS) for 24 h. Supernatant samples were collected for analysis of glucose and SCFA consumption. Cells were collected to determine the expression of mRNA for genes associated with inflammation (TNF, IL1B, CXCL2, CXCL8, PTGS2, and TLR4), purinergic signaling (P2RX7, ADORAB2, and CD73), nutrient transport [SLC16A1 (MCT1), SLC16A3 (MCT4), SLC5A8, and MCU], and cell metabolism [ACAT1, SLC2A1 (GLUT1), IGFBP3, and IGFBP5]. Protein expression of TLR4 and ketogenic enzymes (BDH1 and HMGCS1) were also analyzed using flow cytometry. Statistical analysis was conducted with the MIXED model of SAS version 9.4 (SAS Institute Inc., Cary, NC) with medium, LPS exposure, and medium × LPS interaction as fixed effects and animal within plate as a random effect. Cells tended to consume more glucose when exposed to LPS as opposed to no LPS exposure (31.8 vs. 28.7 ± 2.7), but consumption of propionate and butyrate was not influenced by LPS. Expression of TNF and IL1B was upregulated when exposed to LPS, and expression of CXCL2 and CXCL8 increased following LPS exposure with SCFA (medium × LPS). For cells exposed to LPS, we found a downregulation of ACAT1 and IGFBP5 and an upregulation of SLC2A1, SLC16A3, MCU, and IGFBP3. Medium with SCFA led to greater expression of MCU. SLC16A1 was upregulated in cells incubated with SCFA and without LPS compared with the other groups. Protein expression of ketogenic enzymes was not affected; however, BDH1 mean fluorescence intensity (MFI) expression tended to be less in cells exposed to LPS. These data are interpreted to indicate that when REC are exposed to LPS, they may increase glucose metabolism. Moreover, transport of solutes was affected by SCFA in the medium and by exposure to LPS. Overall, the results suggest that metabolic function of REC in vitro is altered by a proinflammatory response, which may lead to a greater glucose requirement.

Soybean oil supplementation and starter protein content: Effects on growth performance, digestibility, ruminal fermentation, and urinary purine derivatives of Holstein dairy calves.

This study investigated the effects of feeding dairy calves starter diets containing 19% or 22% crude protein (CP) content on a dry matter basis and either supplemented or not with soybean oil (SBO, 0 vs. 3%, dry matter basis) on growth performance, digestibility, urinary nitrogen, and purine derivatives (PD) excretion. A total of 48 female Holstein dairy calves (mean 39.8 kg of body weight) were randomly distributed to experimental diets in a 2 × 2 factorial arrangement of treatments. The 4 dietary treatments were (1) starter diet without SBO supplement and 19% CP (NSBO-19CP), (2) starter diet without SBO supplement and 22% CP (NSBO-22CP), (3) starter diet with 3% SBO and 19% CP (SBO-19CP), and (4) starter diet with 3% SBO and 22% CP (SBO-22CP). Milk feeding value was similarly based on a constant protocol across experimental treatments and calves had ad libitum access to water and starter diets throughout the study. All calves were weaned on d 63 of age and remained in the study until d 83 of age. Calves supplemented with SBO had lower starter feed intake and average daily gain (ADG) and lower feed efficiency (FE) but had a higher fecal score indicating a higher likelihood of diarrhea occurrence compared with unsupplemented calves. Wither heights, digestibilities of organic matter, CP, and neutral detergent fiber were decreased, and ruminal volatile fatty acids tended to be reduced, and the molar proportion of ruminal butyrate (preweaning) and acetate (postweaning) reduced by supplemental SBO. The urinary allantoin and total PD excretion were reduced; however, urinary nitrogen excretion was increased when calves were supplemented with SBO. The CP amount did not affect starter feed intake, FE, or diarrhea occurrence rate, whereas the 22CP diets increased neutral detergent fiber digestibility, improved ADG (tendency), and increased allantoin and urinary PD excretion compared with the 19CP diets. The starter feed intake, ADG, FE, diarrhea occurrence rate, nutrient digestibility, and ruminal fermentation were not affected by the interaction between starter SBO and CP level; however, hip height and total PD in calves that received the SBO-22CP diets were higher than those fed the SBO-19CP diets. In conclusion, based on our experimental conditions, supplemental SBO could not be recommended for dairy calves. Furthermore, our findings indicate that SBO has negative effects on performance more attributed to reducing starter intake, digestibility, and ruminal volatile fatty acid concentration rather than because of a limitation of starter metabolizable protein supply and intestinal amino acid availability. Therefore, our results indicate that feeding the higher starter CP content is not a viable strategy to compensate for the negative effects of SBO supplementation on the growth performance of dairy calves.

Effects of diet fermentability and supplementation of 2-hydroxy-4-(methylthio)-butanoic acid and isoacids on milk fat depression: 2. Ruminal fermentation, fatty acid, and bacterial community structure.

The experiment was conducted to understand ruminal effects of diet modification during moderate milk fat depression (MFD) and ruminal effects of 2-hydroxy-4-(methylthio)-butanoic acid (HMTBa) and isoacids on alleviating MFD. Five ruminally cannulated cows were used in a 5 × 5 Latin square design with the following 5 dietary treatments (dry matter basis): a high-forage and low-starch control diet with 1.5% safflower oil (HF-C); a low-forage and high-starch control diet with 1.5% safflower oil (LF-C); the LF-C diet supplemented with HMTBa (0.11%; 28 g/d; LF-HMTBa); the LF-C diet supplemented with isoacids [(IA) 0.24%; 60 g/d; LF-IA]; and the LF-C diet supplemented with HMTBa and IA (LF-COMB). The experiment consisted of 5 periods with 21 d per period (14-d diet adaptation and 7-d sampling). Ruminal samples were collected to determine fermentation characteristics (0, 1, 3, and 6 h after feeding), long-chain fatty acid (FA) profile (6 h after feeding), and bacterial community structure by analyzing 16S gene amplicon sequences (3 h after feeding). Data were analyzed using the MIXED procedure of SAS (SAS Institute Inc., Cary, NC) in a Latin square design. Preplanned comparisons between HF-C and LF-C were conducted, and the main effects of HMTBa and IA and their interaction within the LF diets were examined. The LF-C diet decreased ruminal pH and the ratio of acetate to propionate, with no major changes detected in ruminal FA profile compared with HF-C. The α-diversity for LF-C was lower compared with HF-C, and ß-diversity also differed between LF-C and HF-C. The relative abundance of bacterial phyla and genera associated indirectly with fiber degradation was influenced by LF-C versus HF-C. As the main effect of HMTBa within the LF diets, HMTBa increased the ratio of acetate to propionate and butyrate molar proportion. Ruminal saturated FA were increased and unsaturated FA concentration were decreased by HMTBa, with minimal changes detected in ruminal bacterial diversity and community. As the main effect of IA, IA supplementation increased ruminal concentration of all branched-chain volatile FA and valerate and increased the percentage of trans-10 C18 isomers in total FA. In addition, α-diversity and the number of functional features were increased for IA. Changes in the abundances of bacterial phyla and genera were minimal for IA. Interactions between HMTBa and IA were observed for ruminal variables and some bacterial taxa abundances. In conclusion, increasing diet fermentability (LF-C vs. HF-C) influenced rumen fermentation and bacterial community structure without major changes in FA profile. Supplementation of HMTBa increased biohydrogenation capacity, and supplemental IA increased bacterial diversity, possibly alleviating MFD. The combination of HMTBa and IA had no associative effects in the rumen and need further studies to understand the interactive mechanism.

Rumen and Serum Metabolomes in Response to Endophyte-Infected Tall Fescue Seed and Isoflavone Supplementation in Beef Steers.

Fescue toxicosis impacts beef cattle production via reductions in weight gain and muscle development. Isoflavone supplementation has displayed potential for mitigating these effects. The objective of the current study was to evaluate isoflavone supplementation with fescue seed consumption on rumen and serum metabolomes. Angus steers (n = 36) were allocated randomly in a 2 × 2 factorial arrangement of treatments including endophyte-infected (E+) or endophyte-free (E-) tall fescue seed, with (P+) or without (P-) isoflavones. Steers were provided a basal diet with fescue seed for 21 days, while isoflavones were orally administered daily. Following the trial, blood and rumen fluid were collected for metabolite analysis. Metabolites were extracted and then analyzed by UPLC-MS. The MAVEN program was implemented to identify metabolites for MetaboAnalyst 4.0 and SAS 9.4 statistical analysis. Seven differentially abundant metabolites were identified in serum by isoflavone treatment, and eleven metabolites in the rumen due to seed type (p < 0.05). Pathways affected by treatments were related to amino acid and nucleic acid metabolism in both rumen fluid and serum (p < 0.05). Therefore, metabolism was altered by fescue seed in the rumen; however, isoflavones altered metabolism systemically to potentially mitigate detrimental effects of seed and improve animal performance.

ß-glucan from Saccharomyces cerevisiae is involved in immunostimulation of ovine ruminal explants.

In this study, we investigated whether ß-glucan from Saccharomyces cerevisiae exerts beneficial effects on mucosal immunity in an ovine ruminal explant (ORE) model. Once the ORE model was established, viability was assessed through histological change, E-cadherin expression, CK-18 and Ki-67 distribution. Then, the OREs were co-cultured with ß-glucan, following which, gene and protein expression levels of sheep ß-defensin-1 (SBD-1), pro-inflammatory interleukin (IL)-6, and anti-inflammatory IL-10 were detected using quantitative real-time polymerase chain reaction (qPCR) and enzyme-linked immunosorbent assay (ELISA). Hematoxylin & eosin staining, qPCR, and immunohistochemistry showed that the overall ORE structure was intact after 96 hours in culture, but explants cultured for more than 24 hours showed epithelial degradation. Therefore, we performed the follow-up test within 24 hours. qPCR and ELISA revealed that the gene and protein expression levels of SBD-1, IL-6, and IL-10 in the OREs significantly increased (P < 0.05) after treatment with ß-glucan compared with controls. This study identified the feasibility and optimal conditions of ORE culture and demonstrated that ß-glucan activates SBD-1, IL-6, and IL-10 secretion in OREs to promote mucosal immunity.

Dans la présente étude nous avons examiné si le ß-glucane de Saccharomyces cerevisiae amène des effets bénéfiques sur l'immunité mucosale dans un modèle d'explant ruminal ovin (ORE). Une fois que le modèle ORE fut établi, la viabilité fut évaluée via les changements histologiques, l'expression d'E-cadhérine et la distribution de CK-18 et Ki-67. Puis, les OREs furent co-cultivés avec du ß-glucane, après quoi, les degrés d'expression des gènes et des protéines ß-défensine-1 ovine (SBD-1), interleukine (IL)-6 pro-inflammatoire et IL-10 anti-inflammatoire furent détectés en utilisant une réaction d'amplification en chaîne par la polymérase quantitative en temps réel (qPCR) et une épreuve immuno-enzymatique (ELISA). Une coloration à l'hématoxyline et éosine, le qPCR et l'immunohistochimie ont montré que la structure globale d'ORE était intacte après 96 heures en culture, mais des explants cultivés pour plus de 24 heures présentaient une dégradation épithéliale. Par conséquent, nous avons effectué les tests de suivi en dedans de 24 heures. Les analyses par qPCR et ELISA ont révélé que les degrés d'expression des gènes et des protéines SBD-1, IL-6 et IL-10 dans les OREs augmentèrent de manière significative (P < 0,05) après un traitement avec du ß-glucane comparativement aux témoins. Cette étude a identifié la faisabilité et les conditions optimales pour la culture d'ORE et a démontré que le ß-glucane active la sécrétion de SBD-1, IL-6 et IL-10 dans les OREs afin de promouvoir l'immunité mucosale.(Traduit par Docteur Serge Messier).

Antimethanogenic effects of nitrate supplementation in cattle: A meta-analysis.

Supplementing a diet with nitrate is regarded as an effective and promising methane (CH4) mitigation strategy by competing with methanogens for available hydrogen through its reduction of ammonia in the rumen. Studies have shown major reductions in CH4 emissions with nitrate supplementation, but with large variation in response. The objective of this study was to quantitatively investigate the effect of dietary nitrate on enteric CH4 production and yield and evaluate the variables with high potential to explain the heterogeneity of between-study variability using meta-analytical models. A data set containing 56 treatments from 24 studies was developed to conduct a meta-analysis. Dry matter (DM) intake, nitrate dose (g/kg of DM), animal body weight, roughage proportion of diet, dietary crude protein and neutral detergent fiber content, CH4 measurement technique, and type of cattle (beef or dairy) were considered as explanatory variables. Average DM intake and CH4 production for dairy cows (16.2 ± 2.93 kg/d; 311 ± 58.8 g/d) were much higher than for beef cattle (8.1 ± 1.57 kg/d; 146 ± 50.9 g/d). Therefore, a relative mean difference was calculated and used to conduct random-effect and mixed-effect model analysis to eliminate the large variations between types of animal due to intake. The final mixed-effect model for CH4 production (g of CH4/d) had 3 explanatory variables and included nitrate dose, type of cattle, and DM intake. The final mixed-effect model for CH4 yield (g of CH4/kg of DM intake) had 2 explanatory variables and included nitrate dose and type of cattle. Nitrate effect sizes on CH4 production (dairy: -20.4 ± 1.89%; beef: -10.1 ± 1.52%) and yield (dairy: -15.5 ± 1.15%; beef: -8.95 ± 1.764%) were significantly different between the 2 types of cattle. When data from slow-release nitrate sources were removed from the analysis, there was no significant difference in type of cattle anymore for CH4 production and yield. Nitrate dose enhanced the mitigating effect of nitrate on CH4 production and yield by 0.911 ± 0.1407% and 0.728 ± 0.2034%, respectively, for every 1 g/kg of DM increase from its mean dietary inclusion (16.7 g/kg of DM). An increase of 1 kg of DM/d in DM intake from its mean dietary intake (11.1 kg of DM/d) decreased the effect of nitrate on CH4 production by 0.691 ± 0.2944%. Overall, this meta-analysis demonstrated that nitrate supplementation reduces CH4 production and yield in a dose-dependent manner, and that elevated DM intake decreases the effect of nitrate supplementation on CH4 production. Furthermore, the stronger antimethanogenic effect on CH4 production and yield in dairy cows than in beef steers could be related to use of slow-release nitrate in beef cattle.

Dietary Supplementation with Sodium Sulfate Improves Rumen Fermentation, Fiber Digestibility, and the Plasma Metabolome through Modulation of Rumen Bacterial Communities in Steers.

Six steers were used to study the effects of dietary supplementation with sodium sulfate (Na2SO4) on rumen fermentation, nutrient digestion, rumen microbiota, and plasma metabolites. The animals were fed a basal ration with Na2SO4 added at 0 g/day (sulfur [S] content of 0.115% dry matter [DM]), 20 g/day (S at 0.185% DM), or 40 g/day (S at 0.255% DM) in a replicate 3-by-3 Latin square design. The results indicated that supplementing with Na2SO4 increased the ruminal concentration of total volatile fatty acids, the molar proportions of acetate and butyrate, the ruminal concentrations of microbial protein, SO42--S, and S2--S, and the digestibility of fiber, while it decreased the molar proportion of propionate and the ruminal concentration of ammonia nitrogen. Supplementing with Na2SO4 increased the diversity and the richness of rumen microbiota and the relative abundances of the phylum Firmicutes and genera Ruminococcus 2, Rikenellaceae RC9 gut group, and Desulfovibrio, whereas it decreased the relative abundances of the phylum Bacteroidetes and genera Prevotella 1, Prevotellaceae UCG-001, and Treponema 2 Supplementing with Na2SO4 also increased the plasma concentrations of amino acids (l-arginine, l-methionine, l-cysteine, and l-lysine), purine derivatives (xanthine and hypoxanthine), vitamins (thiamine and biotin), and lipids (acetylcarnitine and l-carnitine). It was concluded that supplementing the steer ration with Na2SO4 was beneficial for improving the rumen fermentation, fiber digestibility, and nutrient metabolism through modulating the rumen microbial community.IMPORTANCE Essential elements like nitrogen and sulfur greatly affect rumen fermentation and metabolism in ruminants. However, little knowledge is available on the effects of sulfur on the rumen microbiota and plasma metabolome. The results of the present trial demonstrated that supplementing the steer ration with sodium sulfate markedly improved rumen fermentation, fiber digestibility, and metabolism of amino acids, purine derivatives, and vitamins through effects on the ruminal microbiome. The facts obtained from the present trial clarified the possible mechanisms of the positive effects of sulfur on rumen fermentation and nutrient utilization.

Dietary starch concentration alters reticular pH, hepatic copper concentration, and performance in lactating Holstein-Friesian dairy cows receiving added dietary sulfur and molybdenum.

To test the hypothesis that Cu metabolism in dairy cows is affected by dietary starch concentration and additional sulfur S and Mo, 60 Holstein-Friesian dairy cows that were [mean ± standard error (SE)] 33 ± 2.5 days postcalving and yielding 41 ± 0.9 kg of milk/d were fed 1 of 4 diets in a 2 × 2 factorial design experiment over a 14-wk period. The 4 diets had a Cu concentration of approximately 15 mg/kg of dry matter (DM), a grass silage-to-corn silage ratio of 1:1, a dietary starch concentration of either 150 g/kg of DM (low starch, LS) or 220 g/kg of DM (high starch, HS), and were either unsupplemented (-) or supplemented (+) with an additional 0.8 g of S/kg of DM and 4.4 mg of Mo/kg of DM. We found an effect of dietary starch concentration on mean reticular pH, which was 0.15 pH units lower in cows fed the high starch diets. The addition of S and Mo decreased intake by 1.8 kg of DM/d, an effect that was evident beginning in wk 1 of the study. Mean milk and fat yields were 37.0 and 1.51 kg/d, respectively, and were not affected by dietary treatment. We found an effect of dietary starch concentration on milk protein concentration, protein yield, and urea nitrogen, which were increased by 2.8 g/kg, 0.09 kg/d, and 2.1 mg/dL, respectively, in cows fed the high starch diets. We found no effect of dietary treatment on either cow live weight or body condition. Mean plasma Cu, Fe, and Zn concentrations were 15.3, 42.1, and 14.4 µmol/L, respectively, and were not affected by dietary treatment. In contrast, we found an interaction between dietary starch concentration and Cu antagonists on plasma Mo, where feeding additional S and Mo increased plasma Mo to a greater extent when cows were offered the high versus low starch diet. We also found that increasing dietary starch concentration increased serum ceruloplasmin activity, but serum haptoglobin concentration was not affected by dietary treatment. The addition of S and Mo decreased hepatic Cu concentration, whereas in cows fed the higher dietary starch concentration, hepatic Cu concentration was increased over the period of our study. We concluded that increasing dietary starch concentration decreases rumen pH and increases milk protein yield and hepatic Cu concentration, whereas feeding additional S and Mo decreases intake and hepatic Cu concentration.