Effect of fish oil on ruminal biohydrogenation of C18 unsaturated fatty acids in steers fed grass or red clover silages

Red clover and fish oil (FO) are known to alter ruminal lipid biohydrogenation leading to an increase in the polyunsaturated fatty acid (PUFA) and conjugated linoleic acid (CLA) content of ruminant-derived foods, respectively. The potential to exploit these beneficial effects were examined using eight Hereford × Friesian steers fitted with rumen and duodenal cannulae. Treatments consisted of grass silage or red clover silage fed at 90% of ad libitum intake and FO supplementation at 0, 10, 20 or 30 g/kg diet dry matter (DM). The experiment was conducted with two animals per FO level and treatments formed extra-period Latin squares. Flows of fatty acids at the duodenum were assessed using ytterbium acetate and chromium ethylene diamine tetra-acetic acid as indigestible markers. Intakes of DM were higher (P < 0.001) for red clover silage than grass silage (5.98 v. 5.09 kg/day). There was a linear interaction effect (P = 0.004) to FO with a reduction in DM intake in steers fed red clover silage supplemented with 30 g FO/kg diet DM. Apparent ruminal biohydrogenation of C18:2n-6 and C18:3n-3 were lower (P < 0.001) for red clover silage than grass silage (0.83 and 0.79 v. 0.87 and 0.87, respectively), whilst FO increased the extent of biohydrogenation on both diets. Ruminal biohydrogenation of C20:5n-3 and C22:6n-3 was extensive on both silage diets, averaging 0.94 and 0.97, respectively. Inclusion of FO in the diet enhanced the flow of total CLA leaving the rumen with an average across silages of 0.22, 0.31, 0.41 and 0.44 g/day for 0, 10, 20 or 30 g FO/kg, respectively, with a linear interaction effect between the two silages (P = 0.03). FO also showed a dose-dependent increase in the flow of trans-C18:1 intermediates at the duodenum from 4.6 to 15.0 g/day on grass silage and from 9.4 to 22.5 g/day for red clover silage. Concentrations of trans-C18:1 with double bonds from Δ4-16 in duodenal digesta were all elevated in response to FO in both diets, with trans-11 being the predominant isomer. FO inhibited the complete biohydrogenation of dietary PUFA on both diets, whilst red clover increased the flow of C18:2n-6 and C18:3n-3 compared with grass silage. In conclusion, supplementing red clover silage-based diets with FO represents a novel nutritional strategy for enhancing the concentrations of beneficial fatty acids in ruminant milk and meat.     

Red clover polyphenol oxidase and lipid metabolism

Increasing the polyunsaturated fatty acid (PUFA) composition of milk is acknowledged to be of benefit to consumer health. Despite the high PUFA content of forages, milk fat contains only about 3% of PUFA and only about 0.5% of n-3 fatty acids. This is mainly due to intensive lipid metabolism in the rumen (lipolysis and biohydrogenation) and during conservation (lipolysis and oxidation) such as drying (hay) and ensiling (silage). In red clover, polyphenol oxidase (PPO) has been suggested to protect lipids against degradation, both in the silage as well as in the rumen, leading to a higher output of PUFA in ruminant products (meat and milk). PPO mediates the oxidation of phenols and diphenols to quinones, which will readily react with nucleophilic binding sites. Such binding sites can be found on proteins, resulting in the formation of protein-bound phenols. This review summarizes the different methods that have been used to assess PPO activity in red clover, and an overview on the current understanding of PPO activity and activation in red clover. Knowledge on these aspects is of major importance to fully harness PPO's lipid-protecting role. Furthermore, we review the studies that evidence PPO-mediated lipid protection and discuss its possible importance in lab-scale silages and further in an in vitro rumen system. It is demonstrated that high (induction of) PPO activity can lead to lower lipolysis in the silage and lower biohydrogenation in the rumen. There are three hypotheses on its working mechanism: (i) protein-bound phenols could directly bind to enzymes (e.g. lipases) as such inhibiting them; (ii) binding of quinones in and between proteins embedded in a lipid membrane (e.g. in the chloroplast) could lead to encapsulation of the lipids; (iii) direct binding of quinones to nucleophilic sites in polar lipids also could lead to protection. There is no exclusive evidence on which mechanism is most important, although there are strong indications that only lipid encapsulation in protein-phenol complexes would lead to an effective protection of lipids against ruminal biohydrogenation. From several studies it has also become apparent that the degree of PPO activation could influence the mode and degree of protection. In conclusion, this review demonstrates that protein-bound phenols and encapsulation in protein-phenol complexes, induced by PPO-mediated diphenol oxidation, could be of interest when aiming to protect lipids against pre-ruminal and ruminal degradation.     

Influence of damaging and wilting red clover on lipid metabolism during ensiling and in vitro rumen incubation

This paper describes the relationship between protein-bound phenols in red clover, induced by different degrees of damaging before wilting and varying wilting duration, and in silo lipid metabolism. The ultimate effect of these changes on rumen biohydrogenation is the second focus of this paper. For this experiment, red clover, damaged to different degrees (not damaged (ND), crushing or frozen/thawing (FT)) before wilting (4 or 24 h) was ensiled. Different degrees of damaging and wilting duration lead to differences in polyphenol oxidase (PPO) activity, measured as increase in protein-bound phenols. Treatment effects on fatty acid (FA) content and composition, lipid fractions (free FAs, membrane lipids (ML) and neutral fraction) and lipolysis were further studied in the silage. In FT, red clover lipolysis was markedly lower in the first days after ensiling, but this largely disappeared after 60 days of ensiling, regardless of wilting duration. This suggests an inhibition of plant lipases in FT silages. After 60 days of ensiling no differences in lipid fractions could be found between any of the treatments and differences in lipolysis were caused by reduced FA proportions in ML of wilted FT red clover. Fresh, wilted (24 h) after damaging (ND or FT) and ensiled (4 or 60 days; wilted 24 h; ND or FT) red clover were also incubated in rumen fluid to study the biohydrogenation of C18:3n-3 and C18:2n-6 in vitro. Silages (both 60 days and to a lower degree 4 days) showed a lower biohydrogenation compared with fresh and wilted forages, regardless of damaging. This suggests that lipids in ensiled red clover were more protected, but this protection was not enhanced by a higher amount of protein-bound phenols in wilted FT compared with ND red clover. The reduction of rumen microbial biohydrogenation with duration of red clover ensiling seems in contrast to what is expected, namely a higher biohydrogenation when a higher amount of FFA is present. This merits further investigation in relation to strategies to activate PPO toward the embedding of lipids in phenol-protein complexes.     

Solubility and flow of calcium,magnesium and phosphorus in the digestive tract of sheep given maize or alfalfa silages

Four sheep, each fitted with a rumen cannula and a re-entrant cannula in the proximal duodenum and terminal ileum, were used for collection of rumen, duodenal and ileal digesta. Four normal sheep were used for collection of faeces. They were given maize silage, maize silage ensiled with urea, wilted alfalfa silage or formic acid-treated alfalfa silage. Although the solubility of the calcium (Ca) in the duodenal digesta was higher with sheep on maize silages than with those on alfalfa silages, sheep fed on maize silage showed negative apparent absorption (?32.7 and ?16.5%) owing to the low concentration of Ca in whole-plant maize. The apparent absorption of magnesium (Mg) was 40.6–42.3% for maize silages and 24.2–26.8% for alfalfa silages. The differences between silages with respect to apparent absorption of phosphorus (P) were not significant and ranged between 1.5 and 10.9%. There were no appreciable differences in soluble proportions, flow and apparent absorption of Ca, Mg and P between the two maize silages or the two alfalfa silages. There was a net absorption of Ca and Mg and a net secretion of P in the stomach of animals on all silages.     

Effects of mixtures of red clover and maize silages on the partitioning of dietary nitrogen between milk and urine by dairy cows

Eight multiparous lactating Holstein-Friesian cows were used to evaluate the partitioning of dietary nitrogen (N) from diets based on mixtures of red clover and maize silages in comparison with diets based on ryegrass silage. All cows received 4 kg/day of a standard dairy concentrate with one of four forage treatments in an incomplete changeover design with three 4-week periods. Three treatments were based on mixtures of red clover and maize silage. N intake was altered both by varying the ratio of these silages (40/60 and 25/75 on a dry matter (DM) basis) and by an additional treatment for which the DM intake of the 40/60 mixture was restricted to the level achieved with grass silage. Rumen passage rates were estimated from faecal excretion curves following a pulse oral dose of Dysprosium-labeled silage and urinary excretion of purine derivatives (PD) was used as an index of rumen microbial protein synthesis. Red clover silage mixtures led to significantly increased feed intake (21.5, 20.7 and 15.2 kg DM/day for 40/60 and 25/75 red clover/maize silage mixtures and grass silage, respectively), milk production (25.8, 27.8 and 20.0 kg/day for the same treatments, respectively) and milk component yields, but were without effect on milk fat and protein concentrations. The large increase in the yield of milk (24.5 kg/day) and milk components for the restricted red clover/maize silage treatment, in comparison with the grass silage treatment, was proportionately greater than the increase in DM intake (16.6 kg DM/day). There were no significant treatment effects on diet digestibility, while the higher intakes of red clover silage mixtures were associated with higher rumen passage rates (5.82%, 6.24% and 4.55%/h, respectively). There were significant effects of both N intake and forage source on the partitioning of dietary N between milk and urine. When dietary protein was diluted by the inclusion of maize silage, red clover silage led to increased milk N and reduced urinary N in comparison with grass silage. Improvements in N utilisation may be related to increased dietary starch and/or rumen passage rates leading to increased microbial protein synthesis for these treatments. Urinary excretion of PD was significantly higher for all diets based on mixtures of red clover and maize silages, in comparison with grass silage. Urinary N output was close to literature predictions based on N intake for the diet based on ryegrass silage, but 40 to 80 g/day (25% to 30%) less than predicted for the diets based on the mixtures of red clover and maize silages.     

The effect of N-fertilisation rate or inclusion of red clover to timothy leys on fatty acid composition in milk of dairy cows fed a commercial silage : concentrate ratio

The aim of this experiment was to, under typical Swedish production conditions, evaluate the effects of grass silages subjected to different N-fertilisation regimes fed to dairy cows on the fatty acid (FA) composition of their milk, and to compare the grass silages in this respect to red clover-dominated silage. Grass silages made from first year Phleum pratense L. leys subjected to three N-fertilisation regimes (30, 90 and 120 kg N/ha, designated G-30, G-90 and G-120, respectively) and a mixed red clover–grass silage (Trifolium pratense L. and P. pratense L.; 60/40 on dry matter (DM) basis, designated RC–G) were produced. The experiment was conducted as a change-over design, including 24 primiparous and multiparous dairy cows of the Swedish Red breed, each of which was allocated to three of the four diets. The cows were offered 11 kg DM of silage and 7 kg concentrates. The silages had similar DM and energy concentrations. The CP concentration increased with increase in N-fertilisation level. There was a linear increase in DM intake of the different silages with increased N fertilisation. There were also differences in concentrations of both individual and total FAs amongst silages. The daily milk production (kg/day) did not significantly differ between treatments, but G-30 silage resulted in higher concentrations of 18:2n-6 in the milk compared with the other two grass silages. The highest concentrations of 18:3n-3 and cis-9, trans-11 18:2 were found in milk from cows offered the RC–G silage. The G-30 diet resulted in higher concentration of 18:2n-6 and the same concentration of 18:3n-3 in the milk as the other grass silages, despite lower intake levels of these FAs. The apparent recoveries of 18:3n-3 from feed to milk were 5.74%, 4.27%, 4.10% and 5.31% for G-30, G-90, G-120 and RC–G, respectively. A higher recovery when red clover is included in the diet confirms previous reports. The higher apparent recovery of 18:3n-3 on the G-30 treatment may be related to the lower silage DM intake, which led to a higher relative proportion of ingested FAs originating from concentrates compared with the G-90 and G-120 diets. With the rates and types of concentrates used in this study, the achieved differences in FA composition among the silages were not enough to influence the concentrations of unsaturated FAs in milk.     

Temperate forages ensiled with molasses or fresh cheese whey: Effects on conservation quality, effluent losses and ruminal degradation

Effects of ensiling and adding molasses or increasing levels of fresh cheese whey on the conservation and rumen degradability of temperate pasture silages were evaluated. Forage from three paddocks of mixed grass and legume pastures was used to make 45 silages (15 silages per paddock) with 5 treatments, including silage without additives (control), silage with 15 g/kg dehydrated molasses and silage with 20, 50 and 100 g/kg fresh cheese whey. The chemical composition and fermentation quality (i.e., pH, ammonia N, loss of dry matter (DM) and neutral detergent fibre (NDF), effluent production) of the silages were determined. Fresh and ensiled materials were evaluated for in situ rumen degradability. Ensiling reduced DM and NDF rumen degradability (P<0.01). When additives were employed, the reduction of DM degradability of the silages decreased (P≤0.03). Addition of molasses led to the lowest pH (P<0.01) and DM losses (P<0.01), and highest DM degradability (P<0.01). The conservation and DM degradation results of dried molasses silage was superior to those of fresh whey silages. In general, an increase in the level of whey increased DM degradability (P≤0.03), but linearly increased effluent production (P<0.01) and losses (P<0.01).     

Rumen fermentation,nutrient flow to the duodenum and digestibility in bulls fed calcium soaps of rapeseed fatty acids and soya bean meal coated with calcium soaps

Three Polish Friesian bulls fitted with rumen and duodenal canulas were used in a 3 × 3 Latin square experiment to study the effect of Ca soaps of rapeseed fatty acids (CSRFA) on rumen fermentation, nutrient and fatty acid (FA) flow to the duodenum, and to investigate in vivo the possibility of using CSRFA as a means of protecting soya bean meal protein against degradation in the rumen. Treatments were (1) control, no fat (2) CSRFA, at 2% of dietary DM and (3) SBM protein coated with CSRFA (CSRFA:SBM, 1:1, wt/wt), at 4% of dietary DM. The animals were fed isonitrogenous diets, containing 45% meadow hay, 10% fodder beet and 45% concentrate mixture on a DM basis. Intake was limited to 80 g DM d⁻¹ kg^−0.75. There were no treatment effects on rumen fermentation, rumen liquid turnover rate or volume, suggesting that CSRFA were inert in the rumen. A slight decrease in ammonia concentration in the rumen fluid from the CSRFA:SBM diet and the degradability data do not confirm the protection of SBM protein by CSRFA. Treatments did not affect apparent OM digestibility in the rumen, total N and NAN (non-ammonia nitrogen) flow into the duodenum or microbial protein synthesis. However, rumen degradability of protein was increased by feeding CSRFA. Feeding CSRFA significantly increased the duodenal flows of C18:0, C18:1 and total FA, but when expressed as a percentage of intake, there were no significant treatment differences. The average duodenal flow of total FA was 81.7% of intake. Extents of biohydrogenation of unsaturated FA were decreased in diets containing CSRFA. Estimated net biohydrogenation of C18:2, C18:3 and total unsaturated C18 FA of CSRFA was 43.3, 60.4 and 59.4%, respectively. Postrumen and total tract digestibility of total and individual FA were not different between treatments     

Effect of botanical composition of silages on rumen fatty acid metabolism and fatty acid composition in longissimus muscle and subcutaneous fat of lambs

To study the effect of feeding silages with different botanical composition, on rumen and lamb fat, 30 male lambs were assigned to five different silage groups for 11 weeks: botanically diverse silage (BDS); white clover silage (WCS); red clover silage (RCS), intensive English ryegrass silage (IRS) and crushed linseed and maize silage (MSL). Besides the silages, animals received organic wheat and barley and the MSL group additionally received bicarbonate (15 g/day). Silages were sampled when the bales were opened and analysed for fatty acid (FA) content and chemical composition. At slaughter, ruminal contents were sampled and 24 h after slaughter, longissimus muscle and subcutaneous (SC) fat were sampled. All samples were analysed for FA composition. The MSL group ingested the highest amount of FA (35.8 g/day v. 13.5, 19.4, 17.2 and 30.4 g/day for MSL v. BDS, WCS, RCS and IRS, respectively) and the sum of the major polyunsaturated FA, C18:2 n-6 and C18:3 n-3, was similar for groups BDS, WCS, RCS and MSL (61.3 g/100 g, 62.3 g/100 g, 62.3 g/100 g, 63.7 g/100 g of FA methylesters (FAME), respectively), while group IRS ingested higher proportions of these FA (74.5 g/100 g of FAME). Rumen data showed that animals fed BDS presented higher proportions of biohydrogenation intermediates, particularly C18:1 t11 and CLA c9t11, suggesting partial inhibition of rumen biohydrogenation. In the MSL group, the content of C18:3 n-3 in the rumen was highest, most probably due to reduced lipolysis and hence biohydrogenation through the combined effect of esterified C18:3 n-3 and seed protection. Additionally, C18:3 n-3 proportions were higher in rumen contents of RCS animals compared with WCS animals, which could be due to the activity of the polyphenol oxidase enzyme in the RC silages. Proportions of C18:3 n-3 were similar between treatments both for SC and intramuscular (IM) fat, whereas CLA c9t11 content was higher in the SC fat of BDS animals and lower in the IM fat of IRS animals compared with the other forage groups. No differences were found for C20:4 n-6, C20:5 n-3, C22:5 n-3 and C22:6 n-3 in the IM fat of the animals. Nevertheless, indices for desaturation and elongation activity in muscle of BDS animals suggest some stimulation of the first three steps of desaturation and elongation (Δ6-desaturase, elongase and Δ5-desaturase) of long-chain FA.     

Subcutaneous Adipose Fatty Acid Profiles and Related Rumen Bacterial Populations of Steers Fed Red Clover or Grass Hay Diets Containing Flax or Sunflower-Seed

Steers were fed 70∶30 forage∶concentrate diets for 205 days, with either grass hay (GH) or red clover silage (RC), and either sunflower-seed (SS) or flaxseed (FS), providing 5.4% oil in the diets. Compared to diets containing SS, FS diets had elevated (P<0.05) subcutaneous trans (t)-18:1 isomers, conjugated linoleic acids and n-6 polyunsaturated fatty acid (PUFA). Forage and oilseed type influenced total n-3 PUFA, especially α-linolenic acid (ALA) and total non-conjugated diene biohydrogenation (BH) in subcutaneous fat with proportions being greater (P<0.05) for FS or GH as compared to SS or RC. Of the 25 bacterial genera impacted by diet, 19 correlated with fatty acids (FA) profile. Clostridium were most abundant when levels of conjugated linolenic acids, and n-3 PUFA''s were found to be the lowest in subcutaneous fat, suggestive of their role in BH. Anerophaga, Fibrobacter, Guggenheimella, Paludibacter and Pseudozobellia were more abundant in the rumen when the levels of VA in subcutaneous fat were low. This study clearly shows the impact of oilseeds and forage source on the deposition of subcutaneous FA in beef cattle. Significant correlations between rumen bacterial genera and the levels of specific FA in subcutaneous fat maybe indicative of their role in determining the FA profile of adipose tissue. However, despite numerous correlations, the dynamics of rumen bacteria in the BH of unsaturated fatty acid and synthesis of PUFA and FA tissue profiles require further experimentation to determine if these correlations are consistent over a range of diets of differing composition. Present results demonstrate that in order to achieve targeted FA profiles in beef, a multifactorial approach will be required that takes into consideration not only the PUFA profile of the diet, but also the non-oil fraction of the diet, type and level of feed processing, and the role of rumen microbes in the BH of unsaturated fatty acid.     

The effects of barley, unmolassed sugar-beet pulp and molasses supplements on organic matter, nitrogen and fibre digestion in the rumen of cattle given a silage diet

In a 4 × 4 Latin-square experiment with a 2 × 2 factorial arrangement of treatments, 4 cattle fitted with a rumen and duodenal cannula were given four grass-containing diets [480 g kg⁻¹ of the total dry matter (DM) intake] and barley (BU), barley + molasses (2:1) (BM), sugar-beet pulp (SU) or sugar-beet pulp + molasses (SM). Duodenal flow was estimated using Cr-mordanted straw and CoEDTA as markers, and microbial nitrogen entering the small intestine using purine bases of nucleic acids.

Molasses-containing diets had a higher (P < 0.01) organic matter (OM) digestibility. The proportion of digestible OM apparently disappearing in the rumen averaged 0.72 and was not significantly affected by the diet. When cattle received molasses, the quantity of microbial N entering the small intestine was higher (P < 0.05) and there was a trend towards a higher efficiency of microbial N synthesis (28.8 vs. 25.6 g N kg⁻¹ OM apparently digested in the rumen). When S diets were consumed, total non-ammonia N flow at the duodenum exceeded N intake by 7.0 g day⁻¹ and when B diets were consumed, it was 0.7 g day⁻¹ less than N intake. Feed N degradability in the rumen and apparent N digestibility of S diets were lower (P < 0.05; P < 0.001) than those of B diets.

Rumen (P < 0.05) and total (P < 0.001) digestibility of neutral detergent fibre (NDF) and acid detergent fibre (ADF) was higher when S diets were given. The proportion of digestible fibre disappearing in the rumen was not affected by the diet. The rate and extent of silage and hay DM degradation were not significantly affected by the diet. However, dietary inclusion of molasses decreased (P < 0.05) the lag time of both hay and silage DM degradation.

The rumen dilution rate of liquid averaged 0.097 and that of particles, 0.049; neither was significantly different for either B and S diets or U and M diets. Duodenal liquid flow was higher (P < 0.05) for M diets.

Average rumen pH was not affected by the diet, but the molasses diets increased (P < 0.05) the range in rumen pH. The BM diet was associated with higher (P < 0.01) rumen ammonia concentration than the other diets. Low rumen ammonia concentrations (< 2 mM) were observed for long periods between feeds. The molar proportion of butyrate was higher on B diets and there was a trend towards a higher proportion of acetate and propionate on S diets. Molasses tended to increase the molar proportion of propionate and butyrate.     

The effects of concentrate energy source on feed intake and rumen fermentation parameters of dairy cows offered a range of grass silages

The effects of concentrate energy source on feed intake and rumen fermentation parameters of lactating dairy cattle, offered one of three grass silages differing in fermentation and intake characteristics, were evaluated in a partially balanced changeover design experiment involving four rumen fistulated dairy cows. Three silages were harvested using different management practices prior to and at ensiling. Silages A and C and silage B were harvested from primary or secondary regrowths either untreated or treated with a bacterial inoculant. For silages A, B and C, dry matter (DM) concentrations were 334, 197 and 183 g/kg (S.E. 3.1), pH values 4.00, 4.79 and 4.80 (S.E. 0.042) and ammonia nitrogen (N) concentrations were 123, 319 and 295 g/kg total N (S.E. 20.0), respectively. Two concentrates were formulated to contain similar crude protein, effective rumen degradable protein, digestible undegradable protein and metabolisable energy concentrations but using different carbohydrate sources to achieve a wide range of starch concentrations. For the low and high starch concentrates starch concentrations were 17 and 304 g/kg DM and acid detergent fibre concentrations were 170 and 80 g/kg DM, respectively. The silages were offered ad libitum, supplemented with 10 kg fresh concentrate daily. For silages A, B and C, DM intakes were 10.9, 7.2 and 8.6 kg/day and concentrate energy sources did not alter (P>0.05) intake. Increasing the level of starch in the concentrate decreased the molar concentration of acetate (P<0.05) and tended to increase the molar concentration of propionate (P<0.1). Silage type altered the molar concentration of acetate (P<0.01) and the acetate:propionate ratio (P<0.05). There were no silagetype×concentrate interactions (P>0.05) on silage intake or rumen fermentation parameters. It is concluded that when concentrate and silage form equal proportions of the diet, the composition of the silage has an over-riding influence on rumen fermentation parameters. Furthermore, the changes in milk fat concentration, observed in a concurrent production study, due to changes in silage and concentrate types can be accounted for by changes in the ratio of lipogenic to glucogenic precursors in the rumen fluid.     

The response of sheep to big bale grass silage ensiled with,or supplemented separately with,molassed sugar beet feed

Second cut big bale grass silage was prepared with grass ensiled alone or with added pelleted, molassed sugar beet feed (MSBF) at 32 or 64 g MSBF kg⁻¹ fresh weight of grass. Addition of MSBF at ensilage increased silage dry matter (DM), water soluble carbohydrate and lactic acid concentrations and decreased ammonia concentration. When fed to wether lambs, MSBF addition, whether at ensilage or given separately, variably improved DM intake, live-weight gain and feed conversion efficiency. Ruminal concentrations of ammonia were reduced and acetate molar proportion increased in diets supplemented with MSBF separately compared with the silage only control diet. Animal performance was generally improved by MSBF supplementation with responses to silages made with added MSBF comparable to those from silage supplemented with MSBF at feeding. There was little difference in animal performance between silages made with 32 or 64 g kg⁻¹ MSBF added at ensiling.     

Grass maturity effects on cattle fed silage-based diets. 1. Organic matter digestion,rumen fermentation and nitrogen utilization

Four silages were harvested at approximately one-week intervals from the same timothymeadow fescue sward. Advanced maturity of the herbage was evidenced by increased neutral detergent fibre [409, 497, 579 and 623 g in 1 kg dry matter (DM)] and decreased nitrogen (N; 29.9, using four ruminally and duodenally cannulated young cattle in a 4 × 4 Latin square experiment. On DM basis (g kg⁻¹), the diet comprised grass silage (700), rolled barley (240) and rapeseed meal (60) and it was given at a rate of 70 g DM (kg live weight)^−0.75 per day.Organic matter digestibility decreased in a curvilinear manner (P_{LINEAR (L)} < 0.001, P_{CUBIC (C)} < 0.01) the values being 0.821, 0.816, 0.758 and 0.747 for the diets based on the four silages in the order of harvest date. Rumen pH increased linearly (P_L < 0.05) and ammonia N concentration decreased curvilinearly (P_L < 0.01, P_C < 0.05) as the grass matured. The molar proportion of acetate in the rumen VFA increased (P_L < 0.001) and the proportion of butyrate decreased (P_L < 0.001) with increased grass maturity. The silage harvest date did not affect the proportion of propionate. The changes in rumen fermentation pattern were associated with a decrease (P_L < 0.05) in rumen protozoal number with increasing maturity of grass.N intake decreased significantly (P_L < 0.001, P_C < 0.01) with the maturity of grass from 167.5 to 118.0 g per day, but duodenal non-ammonia N decreased only from 111.3 to 97.3 g per day indicating greater N losses from the rumen with early-cut silages. The efficiency of microbial protein synthesis in the rumen was not affected by the maturity of grass ensiled. Apparent digestibility of N decreased (P_L < 0.001, P_C < 0.01) and the degradability of N in the rumen decreased (P_L < 0.05) as the grass matured.     

Microbial inoculant effects on silage and in vitro ruminal fermentation, and microbial biomass estimation for alfalfa, bmr corn, and corn silages

Whole crop third cut alfalfa, brown mid-rib (bmr) corn, and corn were chopped and inoculated with one of four microbial inoculants used. Uninoculated silage was the control treatment. Each crop was ensiled in four mini-silos (1 L glass jars) per treatment. All silos were fermented for 60 days at room temperature (22 °C), and then they were opened and analyzed for fermentation products, fiber constituents and N fractions. A fraction of wet silage was ground with a blender for 30 s. In vitro gas production was measured in 160 ml sealed serum vials at 3, 6, 9, 24, and 48 h using the wet ground silage. At 9 and 48 h, rumen fluid was analyzed for volatile fatty acids (VFA) and microbial biomass yield (MBY). In all the three crops, the four inoculants produced only minor changes in pH and fermentation products during ensiling. Of the variables measured, soluble nonprotein N fractions were the characteristics most often affected by some inoculants. At 9 h incubation, in vitro gas production and VFA did not differ between control and inoculated silages, but MBY did. Among crops, alfalfa and corn silages had higher MBY than did bmr corn silage. Among inoculants, three of the four inoculated silages produced more MBY than did control. At 48 h, alfalfa silage produced higher MBY than did corn or bmr silage, and two of the inoculated silages had more MBY than did the control. There was no inoculant by crop interaction. Results suggest that some silage inoculants are capable of altering rumen fermentation, even in cases where effects on silage fermentation are small, and that this effect may be linked to better preservation of crop protein during ensiling.     

Lipid metabolism in mixtures of red clover (Trifolium repens) and perennial ryegrass (Lolium perenne) in lab scale silages and in vitro rumen incubations

Most often, farmers consider red clover an unattractive forage because of its low ensilability. Nevertheless, several in vivo and in vitro experiments also showed advantages of red clover silages such as decreased rumen biohydrogenation of polyunsaturated fatty acids. This has been attributed to a possible protective role of protein-bound phenols, with polyphenol oxidase playing a key role in their formation. This enzyme is active in red clover, but not in other green forages, such as, for example, perennial ryegrass. Therefore, the aim was to study the lipid metabolism within red clover/ryegrass mixtures in lab scale silages and during in vitro rumen batch incubations. Ensilability of red clover increased with higher proportions of ryegrass in the silage mixture. However, the lipid-protecting mechanism of red clover does not seem to occur in the co-ensiled ryegrass as lipolysis of polar lipids linearly increased with increasing proportions of ryegrass (86.0%, 91.6%, 89.9%, 93.1% and 95.6% in 60-day-old silages with 100/0, 75/25, 50/50, 25/75 and 0/100 red clover/ryegrass, respectively). Rumen lipolysis and biohydrogenation of C18:3n-3 and C18:2n-6 were negatively related to red clover proportions in the silage mixtures. The lipid-protective mechanism in red clover silages is confirmed, but it seems not to be transferred to lipids in co-ensiled forages.     

Ensiled sugar beets as dietary component and their effect on preference and dry matter intake by goats

This study examines the influence of ensiled sugar beets (Beta vulgaris ssp. vulgaris) on preference and short-time dry matter (DM) intake of goats. Whole sugar beets were ensiled either without any silage additive (sugar beet silage (SBS)) or with a chemical additive (6 l/t) containing 85% formic and propionic acids (treated sugar beet silage (TBS)) and were chopped after ensiling. The influence of different proportions of SBS (0%, 6.25%, 12.5%, 18.75% and 25% of dietary DM) and TBS (0%, 5%, 10%, 15% and 20% of DM) in partial mixed rations on short-time DM intake by goats (adult Saanen type wethers; n = 10) was studied in two separate, independent trials. Preference behaviour was investigated by offering the goats two different experimental diets per day and offer each possible combination of them (n = 10) once throughout the trial. On this basis, the mean DM intake (within 30 min and 3 h) of each experimental diet was determined. Both sugar beet silages showed good fermentation quality with low pH. By using the chemical silage additive, the production of ethanol was reduced and considerable amounts of sugar were preserved. In the short-term preference experiment, increasing concentrations of sugar beet silage in the ration increased DM intake (p < 0.05) in a linear way with a strong preference for those diets containing high amounts of sugar beet silage. Initial DM intake after 30 min was 4–5 fold increased for diets containing the highest beet concentration. It is assumed that the preference was positively influenced by sugar and sweet taste or the associated higher concentration of metabolisable energy.     

Silages containing buckwheat and chicory: quality, digestibility and nitrogen utilisation by lactating cows

The suitability of silages containing buckwheat (Fagopyrum esculentum) and chicory (Cichorium intybus) for the nutrition of dairy cows was determined. Buckwheat and chicory were sown in mixture with ryegrass (Lolium multilorum), and a pure ryegrass culture served as a control forage. Swards were harvested 55 d after sowing and were ensiled after wilting, without additives in small round bales. Finally, buckwheat and chicory made up the dietary dry matter (DM) proportions of 0.46 and 0.34, respectively. Concentrates were restricted to 2 kg/d. Diets were fed to 3 x 6 late-lactating cows for 15 d at ad libitum access. During the collection period (days 10-15) amounts of feed intake and faeces, urine and milk were recorded and samples were taken. Ensilability was good for buckwheat and ryegrass swards, but was so less for the chicory sward, which was rich in total ash. The buckwheat silage was rich in acid detergent fibre (445 g/kg DM) and lignin (75.7 g/kg DM) and contained less crude protein (135 g/kg DM) and ether extract (15.8 g/kg DM) than the other silages. Consistent with that, the apparent digestibility of the organic matter and fibre were lowest when feeding this silage. The potassium concentrations in the chicory and ryegrass silages were high (61 g/ kg) and lower in buckwheat (47 g/kg). No significant treatment effects on intake, body weight, milk yield or milk composition as well as plasma beta-hydroxybutyrate and non-esterified fatty acids occurred. Being lowest in nitrogen (N) content, the buckwheat silage resulted in the lowest urine N losses and the most efficient N utilisation for milk protein synthesis, but this at cost of body N retention. The results show that silages containing buckwheat and chicory may be used as components of the forage part of dairy cows' diets even though they were found to have a lower feeding value than ryegrass silage.     

Effect of organic grass-clover silage on fiber digestion in dairy cows

There are differences in grass-clover proportions and chemical composition between herbage from primary growth (PG) and regrowth (RG) in grass-clover leys. Mixing silages made from PG and RG may provide a more optimal diet to dairy cows than when fed separately. We tested the hypotheses that increasing dietary proportions of grass-clover silage made from RG compared with PG would increase digestion rate of potentially degradable NDF (pdNDF), and increase ruminal accumulation of indigestible NDF (iNDF). Eight rumen cannulated Norwegian Red cows were used in two replicated 4×4 Latin squares with 21-day periods. Silages were prepared from PG and RG of an organically cultivated ley, where PG and RG silages were fed ad libitum in treatments with RG replacing PG in ratios of 0, 0.33, 0.67 and 1 on dry matter basis in addition to 8 kg concentrate. We evaluated the effect of the four diets with emphasis on rumen- and total tract fiber digestibility. Increasing RG proportions decreased silage intake by 7%. Omasal flow of pdNDF decreased, whereas iNDF flow increased with increasing RG proportions. Increasing RG proportions decreased rumen pool sizes of NDF and pdNDF, whereas pool sizes of iNDF and CP increased. Increasing RG proportions increased digestion rate of NDF, which resulted in greater total tract digestion of NDF. Pure PG diet had the highest calculated energy intake, but the improved rumen digestion of NDF by cows offered 0.33 and 0.67 of RG leveled out milk fat and protein yields among the three PG containing diets.     

Characterization of the rumen lipidome and microbiome of steers fed a diet supplemented with flax and echium oil

Developing novel strategies for improving the fatty acid composition of ruminant products relies upon increasing our understanding of rumen bacterial lipid metabolism. This study investigated whether flax or echium oil supplementation of steer diets could alter the rumen fatty acids and change the microbiome. Six Hereford × Friesian steers were offered grass silage/sugar beet pulp only (GS), or GS supplemented either with flax oil (GSF) or echium oil (GSE) at 3% kg⁻¹ silage dry matter in a 3 × 3 replicated Latin square design with 21-day periods with rumen samples taken on day 21 for the analyses of the fatty acids and microbiome. Flax oil supplementation of steer diets increased the intake of polyunsaturated fatty acids, but a substantial degree of rumen biohydrogenation was seen. Likewise, echium oil supplementation of steer diets resulted in increased intake of 18:4n-3, but this was substantially biohydrogenated within the rumen. Microbiome pyrosequences showed that 50% of the bacterial genera were core to all diets (found at least once under each dietary intervention), with 19.10%, 5.460% and 12.02% being unique to the rumen microbiota of steers fed GS, GSF and GSE respectively. Higher 16S rDNA sequence abundance of the genera Butyrivibrio, Howardella, Oribacterium, Pseudobutyrivibrio and Roseburia was seen post flax feeding. Higher 16S rDNA abundance of the genus Succinovibrio and Roseburia was seen post echium feeding. The role of these bacteria in biohydrogenation now requires further study.