Branched-chain Volatile Fatty Acids Research Articles

Supplementing branched-chain volatile fatty acids in dual-flow cultures varying in dietary forage and corn oil concentrations. I: Digestibility, microbial protein, and prokaryotic community structure

Branched-chain amino acids are deaminated by amylolytic bacteria to branched-chain volatile fatty acids (BCVFA), which are growth factors for cellulolytic bacteria. Our objective was to determine the dietary conditions that would increase the uptake of BCVFA by rumen bacteria. We hypothesized that increased forage would increase cellulolytic bacterial abundance and incorporation of BCVFA into their structure. Supplemental polyunsaturated fatty acids, supplied via corn oil (CO), should inhibit cellulolytic bacteria growth, but we hypothesized that additional BCVFA would alleviate that inhibition. Further, supplemental BCVFA should increase neutral detergent fiber degradation and efficiency of bacterial protein synthesis more with the high forage and low polyunsaturated fatty acid dietary combination. The study was an incomplete block design with 8 dual-flow continuous cultures used in 4 periods with 8 treatments (n = 4 per treatment) arranged as a 2 × 2 × 2 factorial. The factors were: high forage (HF) or low forage (LF; 67 or 33%), without or with supplemental CO (3% dry matter), and without or with 2.15 mmol/d (which included 5 mg/d of 13C each of BCVFA isovalerate, isobutyrate, and 2-methylbutyrate). The isonitrogenous diets consisted of 33:67 alfalfa:orchardgrass pellet, and was replaced with a concentrate pellet that mainly consisted of ground corn, soybean meal, and soybean hulls for the LF diet. The main effect of supplementing BCVFA increased neutral detergent fiber (NDF) degradability by 7.6%, and CO increased NDF degradability only in LF diets. Supplemental BCVFA increased bacterial N by 1.5 g/kg organic matter truly degraded (6.6%) and 0.05 g/g truly degraded N (6.5%). The relative sequence abundance decreased with LF for Fibrobacter succinogenes, Ruminococcus flavefaciens, and genus Butyrivibrio compared with HF. Recovery of the total 13C dose in bacterial pellets decreased from 144 µg/ mg with HF to 98.9 µg/ mg with LF. Although isotope recovery in bacteria was greater with HF, BCVFA supplementation increased NDF degradability and efficiency of microbial protein synthesis under all dietary conditions. Therefore, supplemental BCVFA has potential to improve feed efficiency in dairy cows even with dietary conditions that might otherwise inhibit cellulolytic bacteria.

Supplementing branched-chain volatile fatty acids in dual-flow cultures varying in dietary forage and corn oil concentrations. II: Biohydrogenation and incorporation into bacterial lipids

To maintain membrane homeostasis, ruminal bacteria synthesize branched-chain fatty acids (BCFA) or their derivatives (vinyl ethers) that are recovered during methylation procedures as branched-chain aldehydes (BCALD). Many strains of cellulolytic bacteria require 1 or more branched-chain volatile fatty acid (BCVFA). Therefore, the objective of this study was to investigate BCVFA incorporation into bacterial lipids under different dietary conditions. The study was an incomplete block design with 8 continuous culture fermenters used in 4 periods with treatments (n = 4) arranged as a 2 × 2 × 2 factorial. The factors were high (HF) or low forage (LF, 67 or 33% forage, 33:67 alfalfa:orchardgrass), without or with supplemental corn oil (CO; 3% dry matter, 1.5% linoleic fatty acid), and without or with 2.15 mmol/d (5 mg/d 13C each of isovalerate, isobutyrate, and 2-methylbutyrate). After methylation of bacterial pellets collected from each fermenter's effluent, fatty acids and fatty aldehydes were separated before analysis by gas chromatography and isotope ratio mass spectrometry. Supplementation of BCVFA did not influence biohydrogenation extent. Label was only recovered in branched-chain lipids. Lower forage inclusion decreased BCFA in bacterial fatty acid profile from 9.45% with HF to 7.06% with LF and decreased BCALD in bacterial aldehyde profile from 55.4% with HF to 51.4% with LF. Supplemental CO tended to decrease iso even-chain BCFA and decreased iso even-chain BCALD in their bacterial lipid profiles. The main 18:1 isomer was cis-9 18:1, which increased (P < 0.01) by 25% from CO (data not shown). Dose recovery in bacterial lipids was 43.3% lower with LF than HF. Supplemental CO decreased recovery in the HF diet but increased recovery with LF (diet × CO interaction). Recovery from anteiso odd-chain BCFA and BCALD was the greatest; therefore, 2-methylbutyrate was the BCVFA primer most used for branched-chain lipid synthesis. Recovery in iso odd-chain fatty acids (isovalerate as primer) was greater than label recovery in iso even-chain fatty acids (isobutyrate as primer). Fatty aldehydes were less than 6% of total bacterial lipids, but 26.0% of 13C recovered in lipids were recovered in BCALD because greater than 50% of aldehydes were branched-chain. Because BCFA and BCALD are important in the function and growth of bacteria, especially cellulolytics, BCVFA supplementation can support the rumen microbial consortium, increasing fiber degradation and efficiency of microbial protein synthesis.

The effect of sainfoin (Onobrychis viciifolia Scop.) drying temperature and pelleting on in vitro fermentation characteristics using equine faecal inocula

This study was conducted to determine the effects of drying temperature and pelleting of the tanniniferous forage legume sainfoin (Onobrychis viciifolia Scop.) on the in vitro fermentation characteristics by equine faecal inocula. The sainfoin used for the experiment was air dried on the land and oven dried at 30, 70 and 130 °C for 2.5 h. Part of the dried sainfoin samples was left as dried hay, another part was pelleted, resulting in 6 different samples. The hay and pellets were tested for particle size distribution and pellets were tested for their pellet quality in terms of hardness, durability and bulk density. The gas production technique was used to simulate fermentation and faecal samples of eight randomly selected horses were used to create four independent inocula mixtures that were examined in two subsequent gas production runs. Per inocula mixture, the hay and pellet samples were incubated in triplicate as is, or in combination with polyethylene glycol to inactivate the tannins. Cumulative gas production, a proxy for the organic matter (OM) fermentation, was measured for 72 h, whereafter fermentation fluids were sampled and analysed for volatile fatty acids (VFA) and pH. The results of the pellet quality measurements showed that the particle sizes of the sainfoin particles in the pellets were decreased by a higher drying temperature or by the pelleting process, indicating a loss of fibre physical effectiveness, a factor that may be indicative for chewing activity and hence saliva production. The gas production results showed a significant (P < 0.05) effect of drying temperature and the addition of polyethylene glycol on the cumulative gas production, the VFA production and the percentage of branched chain VFA produced. Hay dried at 130 °C had a lower (P < 0.05) gas production compared to hay dried at 30 and 70 °C, which indicates a lower OM fermentability. The addition of polyethylene glycol increased the gas and total VFA production, which indicates that tannins in sainfoin decrease the production of gas and VFA in hay pellets compared to the hay product. The pelleting process had no effect on the fermentation characteristics, but pelleting conditions strongly decreased the level of the amino acid lysine and its reactivity. The results indicate the increase in drying temperature to cause a decrease in the OM hindgut fermentation and the total and reactive lysine contents of sainfoin. The positive effects of the tannins (a higher proportion of propionic acid relative to acetic acid and a lower protein fermentation) remain, despite of technological treatments.

Assessing milk response to different combinations of branched-chain volatile fatty acids and valerate in Jersey cows

Some cellulolytic bacteria require 1 or more branched-chain volatile fatty acids (BCVFA) for the synthesis of branched-chain AA and branched-chain long-chain fatty acids because they are not able to uptake branched-chain AA or lack 1 or more enzymes to synthesize branched-chain AA de novo. Supplemental BCVFA and valerate were included previously as a feed additive that was later removed from the market; these older studies and more current studies have noted improvements in neutral detergent fiber digestibility and milk efficiency. However, most studies provided a single BCVFA or else isobutyrate (IB), 2-methylbutyrate (MB), isovalerate, and valerate altogether without exploring optimal combinations. Our objective was to determine a combination of isoacids that is optimal for milk production. Sixty (28 primiparous and 32 multiparous) lactating Jersey cows (106 ± 54 days in milk) were blocked and assigned randomly to either a control (CON) treatment without any isoacids, MB [12.3 mmol/kg dry matter (DM)], MB + IB (7.7 and 12.6 mmol/kg DM of MB and IB, respectively), or all 4 isoacids (6.2, 7.3, 4.2, and 5.1 mmol/kg DM of MB, IB, isovalerate, and valerate, respectively). Cattle were fed the CON treatment for a 2-wk period, then were assigned randomly within a block to treatments for 8 wk (n = 15). There was a trend for an interaction of supplement and parity for milk components. There were no differences in components for primiparous cows, whereas MB + IB tended to increase protein concentration by 0.04 and 0.08 percentage units in multiparous cows compared with the CON and MB treatments, respectively. Feeding MB + IB increased fat concentration by 0.23 to 0.31 percentage units compared with all other treatments in multiparous cows. Milk yield and dry matter intake (DMI) did not change with treatment. Treatment interacted with week for milk net energy for lactation/DMI; MB + IB tended to increase milk net energy of lactation/DMI by 0.10 Mcal/kg compared with MB and approached a trend for CON, mainly during the early weeks of the treatment period, whereas differences decreased during the last 2 wk of the treatment period. Cows fed MB had the highest 15:0 anteiso fatty acids in the total milk fatty acid profile, which was greater than that for CON or MB + IB cows, but not cows supplemented with isoacids. Cows fed MB alone had the numerically lowest milk net energy for lactation/DMI. The combination of MB + IB appeared optimal for increasing feed efficiency in our study and was not at the expense of average daily gain. Further research is needed for evaluating how potential changes in supplemental isoacid dosage should vary under differing dietary conditions.

Evaluation of a Natural Phytogenic Formulation as an Alternative to Pharmaceutical Zinc Oxide in the Diet of Weaned Piglets.

A natural phytogenic formulation (NPF) was tested as an alternative to pharmaceutical zinc oxide (ZnO) in weaned piglets with respect to growth performance, apparent total tract digestibility and faecal microbiota composition and metabolic activity. Two dietary NPF levels (NPF: 1000 and 2000 mg/kg diet) were compared to a positive control (ZnO: 3000 mg ZnO/kg diet) and a negative control (CON: no added ZnO or NPF) using 84 weaned piglets from 29 d to 78 d (days of age). Feed conversion ratio was improved (p < 0.05) in ZnO and NPF piglets were compared to CON at 50 d. Dry matter, organic matter and crude protein (p < 0.05) digestibility was improved in NPF piglets compared to CON at 57 d. Compared to CON, NPF inclusion reduced E. coli (p < 0.05) and increased C. leptum subgroup (p < 0.01) at 57 d and 78 d, and reduced C. perfringens subgroup (p < 0.05; at 78 d). The ZnO reduced (p < 0.001) E. coli and C. perfringens subgroup (p < 0.01) compared to CON at 78 d. Moreover, ZnO and NPF reduced molar ratios of branched chain volatile fatty acids (p < 0.05) compared to CON, while NPF also increased butyric acid (p < 0.05) at 78 d. In conclusion, the NPF appeared to be a promising alternative to pharmaceutical doses of ZnO.

Biotin and Leucine Alone or in Combination Promoted the Synthesis of Odd- and Branched-Chain Fatty Acids in the Rumen In Vitro

The odd- and branched-chain fatty acids (OBCFA) accumulated in ruminant products are a class of beneficial fatty acids for human health. Since biotin and leucine are involved in OBCFA synthesis, this study aimed to evaluate their effect on OBCFA synthesis in vitro. There were four treatments: the control group that only provided the basal diet, or the basal diet supplemented with biotin (4 mg/kg dry matter, DM), leucine (4 g/kg DM), or a combination of biotin (4 mg/kg DM) and leucine (4 g/kg DM). The results showed that biotin promoted the degradation of DM (p &lt; 0.10), while leucine significantly increased the concentration of branched-chain volatile fatty acids and valerate (p &lt; 0.05). The concentrations of total odd-chain fatty acids, total iso, total anteiso, total branched-chain fatty acids, total OBCFA, and total fatty acids were significantly increased by the supplementation of biotin or leucine (p &lt; 0.05). Biotin and leucine significantly stimulated the activities of acetyl-CoA carboxylase, fatty acid synthase, and malonyl-CoA, with a significant interaction effect (p &lt; 0.05). In conclusion, the results of this study suggested that biotin and leucine can be used as effective nutrition strategies to promote OBCFA synthesis.

Effects of exogenous amylolytic or fibrolytic enzymes inclusion on in vitro fermentation of lactating dairy cow diets in a dual-flow continuous-culture system

The objective of this study was to determine the effects of including exogenous amylolytic or fibrolytic enzymes in a diet for high-producing dairy cows on in vitro ruminal fermentation. Eight dual-flow continuous-culture fermentors were used in a replicated 4 × 4 Latin square. The treatments were control (CON), a xylanase and glucanase mixture (T1), an α-amylase mixture (T2), or a xylanase, glucanase, and α-amylase mixture (T3). Treatments were included at a rate of 0.008% of diet dry matter (DM) for T1 and T2 and at 0.02% for T3. All treatments replaced the equivalent amount of soybean meal in the diet compared with CON. All diets were balanced to have the same nutrient composition [30.2% neutral detergent fiber (NDF), 16.1% crude protein (CP), and 30% starch; DM basis], and fermentors were fed 106 g/d divided into 2 feedings. At each feeding, T2 was pipetted into the respective fermentor and an equivalent amount of deionized water was added to each fermentor to eliminate potential variation. Experimental periods were 10 d (7 d for adaptation and 3 d for sample collection). Composite samples of daily effluent were collected and analyzed for volatile fatty acids (VFA), NH3-N, and lactate concentrations, degradability of DM, organic matter, NDF, CP, and starch, and flow and metabolism of N. Samples of fermentor contents were collected from each fermentor at 0, 1, 2, 4, 6, and 8 h after feeding to determine kinetics of pH, NH3-N, lactate, and VFA concentrations over time. All data were analyzed using PROC GLIMMIX of SAS (SAS Institute Inc.), and the repeated variable of time was included for kinetics measurements. Treatment did not affect mean pH, degradability, N flow and metabolism, or the concentrations of VFA, NH3-N, or lactate in the effluent samples. Treatment did not affect pH, acetate:propionate ratio, or the concentrations of lactate, NH3-N, total VFA, acetate, propionate, butyrate, isobutyrate, valerate, or caproate. However, the concentration of total VFA tended to change at each time point depending upon the treatment, and T2 tended to have a greater proportion of 2-methylbutyrate and isovalerate than CON, T1, or T3. As 2-methylbutyrate and isovalerate are branched-chain VFA that are synthesized from branched-chain amino acids, T2 may have an increased fermentation of branched-chain amino acids or decreased uptake by fibrolytic microorganisms. Although we did not observe changes in N metabolism due to the enzymes, there could be changes in microbial populations that utilize branched-chain VFA. Overall, the tested enzymes did not improve in vitro ruminal fermentation in the diet of high-producing dairy cows.

Dynamic Variations in Rumen Fermentation Characteristics and Bacterial Community Composition during In Vitro Fermentation

This study aimed to explore the dynamic variations of rumen fermentation characteristics and bacterial community composition during a 24 h in vitro fermentation. A total of twenty-three samples were collected from original rumen fluid (ORF, n = 3), fermentation at 12 h (R12, n = 10), and fermentation at 24 h (R24, n = 10). Results showed that gas production, concentrations of microbial crude protein, ammonia nitrogen, and individual volatile fatty acids (VFA), as well as total VFA and branched-chain VFA concentrations, were higher in R24 when compared with R12 (p &lt; 0.05). However, no significant differences were observed in acetate to propionate ratio and fermentation efficiency between R12 and R24 (p &gt; 0.05). Bacterial diversity analysis found that Shannon index and Simpson index were higher in R24 (p &lt; 0.05), and obvious clusters were observed in rumen bacterial community between R12 and R24. Taxonomic analysis at the phylum level showed that the abundances of Proteobacteria and Fibrobacteres were higher in R12 than that in R24, and inverse results were observed in Bacteroidetes, Firmicutes, Cyanobacteria, Verrucomicrobia, Lentisphaerae, and Synergistetes abundances. Taxonomic analysis at the genus level revealed that the abundances of Rikenellaceae RC9 gut group, Succiniclasticum, Prevotellaceae UCG-003, Christensenellaceae R-7 group, Ruminococcaceae UCG-002, Veillonellaceae UCG-001, and Ruminococcaceae NK4A214 group were higher in R24, whereas higher abundances of Succinivibrionaceae UCG-002, Ruminobacter, and Fibrobacter, were found in R12. Correlation analysis revealed the negative associations between gas production and abundances of Proteobacteria, Succinivibrionaceae UCG-002, and Ruminobacter. Moreover, the abundances of Firmicutes, Rikenellaceae RC9 gut group, Christensenellaceae R-7 group, and Ruminococcaceae UCG-002 positively correlated with VFA production. These results indicate that both rumen fermentation characteristics and bacterial community composition were dynamic during in vitro fermentation, whereas the fermentation pattern, efficiency, and bacterial richness remained similar. This study provide insight into the dynamics of rumen fermentation characteristics and bacterial composition during in vitro fermentation. This study may also provide a reference for decision-making for the sampling time point when conducting an in vitro fermentation for bacterial community investigation.

Effect of commercial slow-release urea product on in vitro rumen fermentation and ruminal microbial community using RUSITEC technique

BackgroundThe objectives of this study were to determine the effect of commercial slow-release urea (SRU) on in vitro fermentation characteristics, nutrient digestibility, gas production, microbial protein synthesis and bacterial community using a rumen simulation technique (RUSITEC). The experiment was a completely randomized design with four treatments and four replications of each treatment. Treatments were: control diet (no SRU addition), control diet plus 0.28% SRU (U28), or plus 0.56% SRU (U56), and control diet that was modified substituting a part of soybean meal equivalent to 0.35% SRU (MU35; dry matter [DM] basis). The experiment consisted of 8 d of adaptation and 7 d of data and sample collection. Rumen inoculum was obtained from three ruminally fistulated Angus cows fed the same diet to the substrate incubated.ResultsDigestibility of DM, organic matter (OM), crude protein (CP), fibre and starch was not affected, but daily production of gas (P < 0.07) and methane (P < 0.05) was quadratically increased with increasing SRU supplementation. The increase of SRU addition did not affect fermentation pH and total volatile fatty acid (VFA) production, whereas linearly (P < 0.01) decreased proportion of propionate, and linearly (P < 0.01) increased acetate to propionate ratio and ammonia nitrogen (N) concentration. The microbial N efficiency was also linearly (P < 0.03) improved with increasing supplementation of SRU. In comparison with control diet, the dietary substitution of SRU for part of soybean meal increased (P < 0.05) the digestibility of DM, OM and CP and decreased (P < 0.02) the total gas production. The total VFA production and acetate to propionate ratio did not differ between control and MU35, whereas the proportion of butyrate was lower (P < 0.05) and that of branched-chain VFA was greater (P < 0.05) with MU35 than control diet. Total and liquid-associated microbial N production as well as ammonia N concentration were greater (P < 0.03) with MU35 than control diet. Observed operational taxonomic units (OTUs), Shannon diversity index, and beta diversity of the microbial community did not differ among treatments. Taxonomic analysis revealed no effect of adding SRU on the relative abundance of bacteria at the phylum level, while at the genus level, the beneficial impact of SRU on relative abundance of Rikenellaceae and Prevotellaceae in feed particle-associated bacteria, and the abundance of Roseburia in liquid associate bacteria was greater (P < 0.05) with MU35.ConclusionsSupplementation of a dairy cow diet with SRU showed potential of increase in ammonia N concentration and microbial protein production, and change fermentation pattern to more acetate production. Adding SRU in dairy cow diet also showed beneficial effect on improving digestibility of OM and fibre. The results suggest that SRU can partially substitute soybean meal in dairy cow diet to increase microbial protein production without impairing rumen fermentation.

Use of Camelina sativa and By-Products in Diets for Dairy Cows: A Review.

Simple SummaryCamelina sativa, also known as false flax, is an oilseed adaptable to wide agronomic conditions and is an environment-friendly crop utilized by human beings for decades. Camelina seed and its derivatives have a nutritional value comparable to other products fed to dairy cows. However, similar to other oil crops, it has anti-nutritional factors which have brought some regulations from authorities on the inclusion levels of camelina seed and oil-extracted products. Many studies have been conducted, and effects on feed intake, digestion, and metabolism are controversial at higher inclusion levels, Yet, there has still been no effect noticed on the metabolic hormones of dairy cows when included at an appropriate level (2 kg/animal/day seeds or 10% inclusion on a dry matter basis of oil extracted products). Although inclusion of the seed and expeller on milk fat suppression is prominent, milk fat produced by a diet including camelina seed and its by-products is rich in health-beneficial fatty acids. In addition, with processing, its anti-nutritional factors can be reduced, and solvent-extracted meals can be used in higher inclusions than current recommended levels, without affecting the digestion and metabolism of the animal.Camelina sativa, belonging to the Brassicaceae family, has been grown since 4000 B.C. as an oilseed crop that is more drought- and cold-resistant. Increased demand for its oil, meal, and other derivatives has increased researchers’ interest in this crop. Its anti-nutritional factors can be reduced by solvent, enzyme and heat treatments, and genetic engineering. Inclusion of camelina by-products increases branched-chain volatile fatty acids, decreases neutral detergent fiber digestibility, has no effect on acid detergent fiber digestibility, and lowers acetate levels in dairy cows. Feeding camelina meal reduces ruminal methane, an environmental benefit of using camelina by-products in ruminant diets. The addition of camelina to dairy cow diets decreases ruminal cellulolytic bacteria and bio-hydrogenation. This reduced bio-hydrogenation results in an increase in desirable fatty acids and a decrease in saturated fatty acids in milk obtained from cows fed diets with camelina seeds or its by-products. Studies suggest that by-products of C. sativa can be used safely in dairy cows at appropriate inclusion levels. However, suppression in fat milk percentage and an increase in trans fatty acid isomers should be considered when increasing the inclusion rate of camelina by-products, due to health concerns.

Use of Fecal Indices as a Non-Invasive Tool for Ruminal Activity Evaluation in Extensive Grazing Sheep.

For a simple, non-invasive evaluation of nutritional status of sheep kept under extensive grazing conditions on annual rangeland, fecal indices (2,6 diaminopimelic acid, nitrogen, and phosphorus) obtained during the vegetative, reproductive, and dry grassland phenological stages, were correlated with ruminal physiological biomarkers (volatile fatty acids and ruminal ammonia). Through correlation analysis and linear regressions, the degree of association between the variables studied was established. The fecal indices that presented the highest degree of association with ruminal variables were FN and FP, being highly correlated with the production of branched-chain volatile fatty acids (isobutyrate and isovalerate) and with ruminal ammonia (r ≥ 0.65), establishing simple linear regression equations of high significance (p ≤ 0.05). Therefore, fecal indices, especially fecal concentrations of N and P, could reflect the metabolism at the ruminal level and with it the availability of compounds for microbial growth, which would help to establish the nutritional status of sheep herds under extensive grazing conditions.

Substitution of molasses for corn grain at two levels of degradable protein. II. Effects on ruminal fermentation, digestion, and nitrogen metabolism

Our objective was to evaluate cow N metabolism and ruminal measures with diets containing 3 different levels of molasses or finely ground dry corn grain with 2 levels of ruminally degradable protein (RDP). Twelve lactating, ruminally cannulated Holstein cows (parity 2.25 ± 0.62; 185 ± 56 DIM; 41.3 ± 6.3 kg of milk initially) were individually fed in an experiment designed as a split-plot, replicated 3 × 3 Latin square, where each period lasted 28 d. Six diets were formulated according to a 2 × 3 factorial arrangement of treatments, where 2 levels of RDP (+RDP and -RDP) were fed throughout the experiment as the whole plot, and 3 levels of molasses (0, 5.25, or 10.50% of dry matter replacing finely ground dry corn grain) were fed in sequences of the Latin squares. Dry matter intake did not differ by diet, although ash intake increased linearly with increasing molasses. Ruminal pH, organic acid concentration, and ammonia concentration were not affected by diet. Molar percentages of ruminal acetate decreased and butyrate increased linearly with increasing levels of molasses. Ruminal free amino acid concentration was greater for +RDP, whereas branched-chain volatile fatty acids declined linearly with increasing molasses. Rumen content mass, ruminal liquid, and ruminal acetate pool size was greater for -RDP, although ruminal lactate pool size tended to be greater with +RDP. Increased ruminal lactate when increasing molasses with high RDP should be explored further, to optimize microbial efficiency and rumen health. Total-tract apparent dry matter digestibility based on spot sampling was not affected by diet; however, ash digestibility increased linearly with increasing levels of molasses. Calculated urine output was greater for cows fed diets with increasing levels of molasses and for cows fed +RDP. Grams of N distributed to excretion pools were not different across diets, although, as molasses increased, a lower proportion of N intake was excreted in urine. Overall, the results from this experiment showed that dairy cows used dietary carbohydrates differently during ruminal fermentation, with increasing molasses resulting in increased butyrate molar proportions at the expense of acetate. Additionally, RDP tended to modify the effects of carbohydrate fermentation, resulting in a tendency for increasing lactate molar pool size only in diets with greater RDP, although this did not ultimately affect ruminal pH.

Intake, digestibility rumen fermentation and nitrogen balance in lambs offered alfalfa and tall fescue-mixtures harvested and ensiled after a frost

The objective of this study was to determine the effect of harvesting alfalfa (Medicago sativa L.) and tall fescue [Schedonorus arundinaceus (Schreb.) Dumort] after frost and ensiling mixtures of these forages on silage fermentation characteristics, intake, digestibility and ruminal fermentation by ewe lambs. Forages were harvested in October of 2017, wilted, and blended as either alfalfa alone, 67% alfalfa + 33% novel endophyte-infected tall fescue, 33% alfalfa + 67% fescue, or 100% fescue. Twenty bins were lined with 2 plastic bags, packed (n = 5/trt) with the respective forages, stored for 3 months, and then assigned randomly to Dorper ewe lambs (n = 20; mean BW = 34.7 ± 6.65 kg) and offered for ad libitum consumption. Lambs were allowed 14 d of adaptation followed by 5 d of total feces and urine collection. Data were analyzed using PROC-MIXED of SAS and orthogonal polynomial contrasts were used to identify trends associated with different proportions of fescue to alfalfa. Silage total acids, lactate, and acetate concentrations increased linearly (P < 0.01), while silage ammonia decreased linearly and quadratically (P < 0.01) with increasing proportions of fescue in the silage. The proportions of lactate to total acids (g/kg total acids) increased linearly (P < 0.01) and quadratically (P = 0.02) with increasing fescue-to-alfalfa ratio. Dry matter (DM) and organic matter (OM) intake (g/kg BW), digestibility, and intake of digestible DM and OM (g/kg BW) decreased linearly (P < 0.01) as fescue proportion increased in the silages. Total ruminal volatile fatty acids (VFA), as well as concentrations of isovalerate and total branched-chain VFA decreased (P < 0.05) linearly and quadratically, butyrate and valerate decreased linearly (P < 0.01), and ruminal acetate concentrations increased (P < 0.01) linearly with increasing fescue concentration in the silage. All N utilization measurements (intake, absorption and retention) decreased with increasing fescue concentration in the silage (P < 0.01). Ensiling alfalfa with fescue that was harvested after a frost may improve forage fermentation characteristics, but may reduce intake and digestibility by lambs compared to feeding undiluted alfalfa silage.

The In Vitro Digestion and Fermentation Characteristics of Feedstuffs Inoculated With Cecal or Colic Fluid of Dezhou Donkey

The present study was conducted to evaluate the in vitro dry matter disappearance (IVDMD) and fermentation characteristics of 6 fibrous feedstuffs incubated with donkey cecal or colic microorganisms. The fibrous feeds were corn straw, wheat straw, peanut vine, peanut shell, wheat shell and wheat bran (WB), which are commonly applied in large-scale donkey farms in China. After 48 hours fermentation, the highest IVDMD occurred in WB, and the lowest occurred in peanut shell (P < .05) regardless what inoculum applied. The IVDMD was positively correlated with OM (R = 0.42, P < .01), CP (R = 0.76, P < .01) and EE (R = 0.56, P < .01), while it was negatively correlated with NDF (R = 0.75, P < .01) and ADF (R = 0.79, P < .01). In terms of volatile fatty acids (VFAs), the greatest net VFA production also occurred in WB (P < .05). The acetate and branched-chain volatile fatty acid proportions were greater in the colon culture fluids than in caecum (P < .05), but the propionate proportions were lower in colon than in caecum (P < .05). This resulted in an increase in the ratio of acetate to propionate in colon culture fluids. In summary, based on the highest IVDMD, net VFA production and CP content occurring with the lowest contents of both NDF and ADF, WB had the highest nutritive value compared to other fibrous substrates. The extent of feed degradation and fermentation depended mainly on the nature of the incubated feedstuffs. Nevertheless, further study is required to investigate these fibrous feeds on hindgut fermentation and in vivo nutrient digestibility.

Porcine in vitro fermentation characteristics of canola co-products in neutral and acidic fermentation medium pH

An in vitro study was conducted to determine effects of reducing pH of porcine in vitro fermentation medium on compositions of glucosinolate degradation products and porcine in vitro fermentation characteristics for solvent-extracted canola meal (SECM) and cold-pressed canola cake (CPCC). Two canola co-products were subjected to porcine in vitro fermentation for 72 h. Accumulated gas production during microbial fermentation was recorded and modeled to estimate kinetics of gas production. Glucosinolate degradation products after microbial fermentation were determined and fermentation medium pH after incubation was recorded. Total and individual volatile fatty acids (VFA) concentrations per unit weight of dry matter (DM) of feedstuffs were determined. On DM basis, SECM and CPCC contained 6.15 and 11.1 µmol/g of glucosinolates, respectively. Goitrin concentration for CPCC was lower (P < 0.05) than that for SECM. Isothiocyanate and indole-3-acetonitrile concentrations for CPCC were lower (P < 0.05) than those for SECM, whereas thiocyanate concentration for CPCC was greater (P < 0.05) than that for SECM. A reduction in fermentation medium pH from 6.2 to 5.2 increased (P < 0.05) goitrin and indole-3-acetonitrile concentrations and decreased (P < 0.05) isothiocyanates concentration for SECM, but did not affect concentration of these 2 glucosinolate degradation products for CPCC. Fermentation medium pH after in vitro fermentation for SECM tended to be greater (P = 0.081) than that for CPCC. Canola co-product type and fermentation medium pH did not interact on gas production parameters. However, a reduction in fermentation medium pH from 6.2 to 5.2 resulted in increased (P < 0.05) lag time and reduced (P < 0.05) fractional rate of degradation and total gas production. Canola co-product type and fermentation medium pH did not interact on total or individual VFA production per gram of DM of feedstuff fermented. However, reducing fermentation medium pH from 6.2 to 5.2 increased (P < 0.05) total VFA and acetate productions, and tended to reduce (P = 0.083) branched-chain VFA production SECM and CPCC. In conclusion, a reduction in fermentation medium pH from 6.2 to 5.2 resulted in increased production of goitrin and indole-3-acetonitriles from SECM-derived glucosinolates, but did not affect production of thiocyanate from SECM-derived glucosinolates. Glucosinolates in CPCC were less affected by the fermentation medium pH used in the current study. It appears that there are other factors other than pH that affect the degradation of canola-derived glucosinolates by microorganisms from hindgut of pigs.

Intake, ruminal fermentation parameters, and apparent total-tract digestibility by beef steers consuming Pensacola bahiagrass hay treated with calcium oxide.

To determine the effect of CaO-treated Pensacola bahiagrass (Paspalum notatum) hay on intake, ruminal fermentation parameters, and apparent total-tract digestibility of nutrients, nine ruminally cannulated Angus-crossbred steers were used in a triplicated 3×3 Latin square design. Steers had ad libitum access to either 1) untreated dry hay (DH; n = 8); 2) hay at 50% DM treated with 8.9% CaCO3 (dry matter [DM] basis; CC; n = 9); or 3) hay at 50% DM treated with 5% CaO (DM basis; CO; n = 8). Water was added to reach 50% DM in the CC and CO diets. Ruminal fluid and blood samples were collected every 3h for 24h. Ruminal fluid was analyzed for pH, volatile fatty acids (VFA), and ammonia-nitrogen (NH3-N). Blood was analyzed for plasma urea nitrogen (PUN). Hay and fecal samples were collected for 4 d, four times daily for hay and twice daily for feces, to determine apparent total-tract digestibility of nutrients. The hay provided to steers during the digestibility period was analyzed for in vitro organic matter digestibility (IVOMD) for 48h. Data were analyzed as repeated measures for blood and ruminal fermentation parameters. Total DM intake was not affected (P ≥ 0.674) by treatment. A treatment effect (P < 0.001) was observed for average ruminal pH, where steers consuming CO had the greatest pH (P < 0.001). Ruminal concentration of NH3-N tended (P = 0.059) to be reduced in steers consuming CO. There was a treatment × time interaction (P = 0.023) on concentrations of PUN, where at 3h DH and CO were lesser than CC (P ≤ 0.050) and at 21h DH was lesser than CC (P = 0.020). Total VFA, acetate, propionate, butyrate, branched-chain VFA, and valerate concentrations were affected by treatment (P ≤ 0.035), where a reduction (P ≤ 0.034) occurred in steers consuming CO. No treatment differences were observed for total-tract digestibility of DM (P = 0.186), organic matter (P = 0.169), or crude protein (P = 0.152); however, steers consuming DH had greater neutral detergent fiber (P = 0.038) than CC and tended to be greater than CO (P = 0.082). The CO hay had greater (P = 0.005) IVOMD compared with DH and tended (P = 0.100) to be greater than CC. Bahiagrass hay treated with CaO may reduce ruminal fermentation, as indicated by decreased total VFA concentration without altering DM intake. The addition of CaO did not improve the digestibility of bahiagrass hay in vivo; however, in vitro results are contradictory and warrant further elucidation.

Effect of dietary inclusion of tea residue and tea leaves on ruminal fermentation characteristics and methane production

The aim of this study was to compare the differences of dietary tea leaves (TL) and tea residue (TR) inclusion on rumen fermentation characteristics and to explore whether TR could be an alternative feedstuff of ruminants. For these purposes, seven treatments consisted of two inclusion types (TL vs. TR) and three inclusion levels (g/g of dry matter basis) of 10% (TL10/TR10), 20% (TL20/TR20), and 30% (TL30/TR30) in each inclusion type, plus control group with inclusion of 0% (CON) were designed, with four replicates in each group, to conduct an in vitro ruminal fermentation test. Results showed that the contents of crude protein, neutral detergent fiber, and acid detergent fiber were higher in TR than TL, while TL contained more ether extract and crude ash than TR. Interaction effects between inclusion type and inclusion level were observed in concentrations of isobutyrate and microbial crude protein (MCP), as well as in gas production and digestibility of organic matter. Fermentation characteristics were significantly influenced by TL and TR depending on the inclusion level, except for the concentration of total branched-chain volatile fatty acid. These significant differences of fermentation characteristics due to inclusion level mainly focused on CON and tea inclusion, with higher values in CON than TR or TL groups. The total gas production during the 48-h incubation showed no differences among CON, TL10, and TR10. The inclusion of TR and TL decreased the production of methane. The concentration of MCP in CON, TR10 and TR30 was lower than TR20 and all TL groups. In conclusion, dietary inclusion of TR and TL possessed equivalent effect on rumen fermentation characteristics and methane production, substituting diet with TR or TL for over 10% would inhibit rumen fermentation despite positive effects in TR20 and all TL groups regarding more MCP and less methane production. This study indicates that special attention should be paid to the inclusion level of TR and TL when considering them as alternative feedstuffs of ruminants. Further in vivo study is needed to evaluate the applicability of tea residue as a feedstuff for production of ruminants.

Effects of L-glutamine supplementation on degradation rate and rumen fermentation characteristics in vitro.

ObjectiveTwo follow-up studies (exp. 1 and 2) were conducted to determine the effects of L-glutamine (L-Gln) supplementation on degradation and rumen fermentation characteristics in vitro.MethodsFirst, rumen liquor from three cannulated cows was used to test L-Gln (50 mM) degradation rate and ammonia-N production at 6, 12, 24, 36, and 48 h after incubation (exp. 1). Second, rumen liquor from two cannulated steers was used to assess the effects of five levels of L-Gln including 0% (control), 0.5%, 1%, 2%, and 3% at 0, 3, 6, 12, 24, 36, and 48 h after incubation on fermentation characteristics, gas production, and degradability of nutrients (exp. 2).ResultsIn exp. 1, L-Gln degradation rate and ammonia-N concentrations increased over time (p<0.001). In exp. 2, pH was reduced significantly as incubation time elapsed (p<0.001). Total gas production tended to increase in all groups as incubation time increased. Acetate and propionate tended to increase by increasing glutamine (Gln) levels, whereas levels of total volatile fatty acids (VFAs) were the highest in 0.5% and 3% Gln groups (p<0.001). The branched-chain VFA showed both linear and quadratic effects showing the lowest values in the 1% Gln group particularly after 6 h incubation (p<0.001). L-Gln increased crude protein degradability (p<0.001), showing the highest degradability in the 0.5% Gln group regardless of incubation time (p<0.05). Degradability of acid detergent fiber and neutral detergent fiber showed a similar pattern showing the highest values in 0.5% Gln group (p<0.10).ConclusionAlthough L-Gln showed no toxicity when it was supplemented at high dosages (2% to 3% of DM), 0.5% L-Gln demonstrated the positive effects on main factors including VFAs production in-vitro. The results of this study need to be verified in further in-vivo study.

Method of diet delivery to dairy cows: Effects on nutrient digestion, rumen fermentation, methane emissions from enteric fermentation and stored manure, nitrogen excretion, and milk production

This study was conducted to examine the effect of method of diet delivery to dairy cows on enteric CH4 emission, milk production, rumen fermentation, nutrient digestion, N excretion, and manure CH4 production potential. Sixteen lactating cows were used in a crossover design (35-d period) and fed ad libitum twice daily a diet [52:48, forage:concentrate ratio; dry matter (DM) basis] provided as forages and concentrates separately (CF) or as a total mixed ration (TMR). For the CF treatment, concentrates were offered first followed by mixed forages 45 min afterward. Method of diet delivery had no effect on DM intake, but neutral detergent fiber (NDF) intake was greater when the diet was delivered as TMR as compared with CF. Apparent total-tract digestibility of DM, crude protein, and gross energy was slightly (1 percentage unit) lower when the diet was offered as TMR than when offered as CF. In contrast, NDF digestibility was greater when the cows were fed TMR versus CF. Although average daily ruminal pH was not affected by method of diet delivery, daily duration of ruminal pH <5.6 was less when the diet was delivered as TMR as compared with CF (0.9 h/d versus 3.7 h/d). Delivering the diet as TMR increased ruminal total volatile fatty acid and NH3 concentrations, but had no effect on acetate, propionate, or branched-chain volatile fatty acid molar proportions. Yields of milk, milk fat, or milk protein, and milk production efficiency (kg of milk/kg of DM intake or g of N milk/g of N intake) were not affected by the method of diet delivery. Daily production (g/d), yield (% gross energy intake), and emission intensity (g/kg of energy-corrected milk) of enteric CH4 averaged 420 g/d, 4.9%, and 9.6 g/kg and were not affected by diet delivery method. Fecal N output was greater when the diet was delivered as TMR versus CF, whereas urinary N excretion (g/d, % N intake) was not affected. Manure volatile solids excretion and maximal CH4 production potential were not affected by method of diet delivery. Under the conditions of this study, delivering the diet as concentrates and forages separately versus a total mixed ration had no effect on milk production, enteric CH4 energy losses, urinary N, or maximal manure CH4 emission potential. However, feeding the diet as total mixed ration compared with feeding concentrates and forages separately attenuated the extent of postprandial decrease in ruminal pH, which has contributed to improving NDF digestibility.

Conditions stimulating neutral detergent fiber degradation by dosing branched-chain volatile fatty acids. III: Relation with solid passage rate and pH on prokaryotic fatty acid profile and community in continuous culture

Our objectives were to evaluate potential interactions in culture conditions that influence how exogenously dosed branched-chain VFA (BCVFA) would be recovered as elongated fatty acids (FA) or would affect bacterial populations. A 2 × 2 × 2 factorial arrangement of treatments evaluated 3 factors: (1) without versus with BCVFA (0 vs. 2 mmol/d each of isobutyrate, isovalerate, and 2-methylbutyrate; each dose was partially substituted with 13C-enriched tracers before and during the collection period); (2) high versus low pH (ranging diurnally from 6.3 to 6.8 vs. 5.7 to 6.2); and (3) low versus high particulate-phase passage rate (kp; 2.5 vs. 5.0%/h) in continuous cultures administered a 50:50 forage:concentrate diet twice daily. Samples of effluent were collected and composited before harvesting bacteria from which FA and DNA were extracted. Profiles and enrichments of FA in bacteria were evaluated by gas chromatography and isotope-ratio mass spectrometry. The 13C enrichment in bacterial FA was calculated as percentage recovery of dosed 13C-labeled BCVFA. Dosing BCVFA increased the even-chain iso-FA, preventing the reduced concentration at higher kp and potentially as a physiological response to decreased pH. However, decreasing pH decreased recovery of 13C in these even-chain FA, suggesting greater reliance on isobutyrate produced from degradation of dietary valine. The iso-FA were decreased, whereas anteiso-FA and 16:0 increased with decreasing pH. Thus, 2-methylbutyrate still appeared to be important as a precursor for anteiso-FA to counter the increased rigidity of bacterial membranes that had more saturated straight-chain FA when pH decreased. Provision of BCVFA stimulated the relative sequence abundance of Fibrobacter and Treponema, both of which require isobutyrate and 2-methylbutyrate. Numerous bacterial community members were shifted by low pH, including increased Prevotella and genera within the phylum Proteobacteria, at the expense of members within phylum Firmicutes. Because of relatively few interactions with pH and kp, supplementation of BCVFA can stimulate neutral detergent fiber degradability via key fibrolytic bacteria across a range of conditions. Decreasing pH shifted bacterial populations and their FA composition, suggesting that further research is needed to distinguish pH from dietary changes.