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.     

Digestion of fatty acids in ruminants: a meta-analysis of flows and variation factors. 1. Total fatty acids

A database built from 95 experiments with 303 treatments was used to quantify the ruminal biohydrogenation (BH) of fatty acids (FA), efficiency of microbial protein synthesis (EMPS), duodenal flow and intestinal absorption of total FA and of FA with 12 to 18 C units, in response to variations in dietary FA content, source or technological treatment of fat supplement. Flows of FA were expressed relative to dry matter intake (DMI) to compile data from bovine and ovine species. BH tended to increase curvilinearly with FA intake, whereas dietary FA did not affect EMPS. A linear relationship between FA intake and duodenal flow of total FA was obtained, with a coefficient of 0.75 ± 0.06 g duodenal FA/kg DMI for each g FA intake/kg DMI. Between experiments, positive balances of total FA (intake - duodenum) were related to low EMPS. Relationships between duodenal flows of FA with 12 to 18 C units and their respective intakes were linear, with a coefficient that increased with the number of C units. Duodenal flow of bacterial FA was linearly related to FA intake (coefficient 0.33 ± 0.13), whereas contribution of bacterial lipid to duodenal flow decreased as FA intake increased. For each FA with 12 to 16 C units, prediction of FA absorption from its respective duodenal flow was linear. For total FA and FA with 18 C units, apparent absorption levelled off at high duodenal flows. All these relationships were discussed according to current knowledge on microbial metabolism in the rumen and on the intestinal digestibility of FA in the intestine.     

Effect of soybean oil availabilities on rumen biohydrogenation and duodenal flow of fatty acids in beef cattle fed a diet with crude glycerine

Soybean oil with different ruminal availability (whole soybeans (WS), soybean oil (SO) and calcium salts (CS)) was used to evaluate the fatty acid (FA) intake, rumen biohydrogenation (BH) and duodenal flow of FA in Nellore steers fed diets with crude glycerine (CG). Eight castrated Nellore steers were fitted with a ruminal and duodenal silicone cannula, and distributed in a double, simultaneous, Latin square 4 × 4 design with four diets and four experimental periods. Concentrates contained ground maize, urea, mineral salts, CG (100 g/kg DM) and soybean products with different availability of soybean oil: (1) no additional fat (CO), (2) WS, (3) SO or (4) CS. Fat supplementation was fixed to obtain 50 g ether extract/kg DM. Experimental treatments had no effect on DM intake, DM duodenal flow or ruminal turnover rate of C:16 FA. However, fat addition increased C:18 and turnover rates of total FA rumen (p < 0.05). CS resulted in lower C:18 turnover rates and lower ruminal BH of monounsaturated and unsaturated FA (UFA) than WS (p < 0.05). SO resulted in a greater duodenal flow of C18:0 (stearic acid), C18:1t-11 (vaccenic acid) and saturated FA than the WS and CS diets (p < 0.05). CS resulted in a higher duodenal flow of C18:3n-3 (linolenic acid) than WS (p < 0.05). The association of CG and calcium salts in Nellore steers was the best nutritional strategy to increase duodenal flow of healthier UFA, which may increase the deposition of these FA in meat. However, SO associated with CG association increased the duodenal flow of vaccenic acid, which is main precursor of endogenous synthesis of conjugated linoleic acids in tissues.     

In situ evaluation of the ruminal and intestinal degradability of extruded whole horse beans

Four rumen and proximal duodenum fistulated non-lactating Holstein cows were used to determine the effect of extrusion at 120 degrees C of whole horse beans (Vicia faba cv Talo) on in vitro nitrogen (N) solubility and in situ degradation of dry matter (DM) and crude protein (CP) in the rumen and intestine. Cows were fed a ration of 30% whole horse beans (WHB) and 70% Italian rye-grass hay. The degradation of DM and CP was estimated using nylon bags suspended in the rumen for 2, 4, 7, 16, 24 and 48 h; the effective ruminal degradability of DM and CP was evaluated assuming a ruminal outflow rate of 0.06/h. Bags incubated in the rumen for 16 h were introduced into the small intestine through the duodenal cannula and subsequently recovered in the feces. Extrusion of WHB reduced N-solubility in buffer solution (21.1 vs 74.9%). Processing diminished the effective rumen degradability of DM (74.6 vs 80.4%) and CP (70.2 vs 89.2%). Meanwhile, the amounts of DM and CP digested in the intestine increased: 9.6 vs 1.4% and 25.2 vs 3.0% respectively. Therefore, feeds containing extruded WHB increase the availability of dietary proteins in the intestine compared with diets containing raw WHB.     

Effect of chestnut and quebracho tannins on fatty acid profile in rumen liquid- and solid-associated bacteria: an in vitro study

Tannins are phenolic compounds that interfere with biohydrogenation (BH) of polyunsaturated fatty acids (FAs). The aim of the present in vitro study was to investigate the effects of two different sources of tannins on FA profiles of rumen bacteria, with particular reference to rumenic and vaccenic acid. A control diet (C; composed of 300 g/kg of wheat straw, 132 g/kg of soyabean meal, 96 g/kg of barley meal, 152 g/kg of maize meal, 300 g/kg of maize gluten and 20 g/kg of mineral vitamin premix, all expressed on dry matter (DM)) and four diets, obtained by adding to C two different types of tannins from chestnut (TC) and from quebracho (TQ) at two concentration levels (49 and 82 g/kg DM), were compared. The content of the main unsaturated FAs (C18:1 cis9, C18:1 trans11, C18:2 cis9, cis12 and C18:3 cis9, cis12, cis15) from solid-associated bacteria (SAB) and liquid-associated bacteria (LAB) was affected by the presence of tannins in the diets. In particular, C18:1 trans11 content was significantly increased, especially with TC1, whereas the decreasing of C18:1 cis9 was unaffected, regardless of the presence or the kind of tannins added to feeds. SAB contained higher amounts of intermediates of polyunsaturated FA BH (as C18:1 trans11 and C18:2 cis9, trans11) than LAB that were characterized by a higher amount of C18:0. In the concentration range adopted in this study, the effect of TC and TQ on changes of bacterial FA profile was comparable. Tannins seem to be a good means to modulate the FA profile of rumen bacteria, favouring the accumulation of C18:1 trans11 during in vitro rumen fermentation.     

Ruminal fatty acid outflow in dry cows fed different sources of linoleic acid: reticulum and omasum as alternative sampling sites to abomasum

This study was designed to determine the rumen outflow of fatty acids (FA) and biohydrogenation (BH) extent using alternative sampling sites (reticulum and omasum) to abomasum in dry cows fed different sources of FA. Four Holstein non-pregnant dry cows (≥3 parturitions, and 712 ± 125 kg BW), cannulated in the rumen and abomasum, were randomly assigned to a 4 × 4 Latin square design experiment, containing the following treatments: 1) control (CON); 2) soya bean oil (SO), dietary inclusion at 30 g/kg; 3) whole raw soya beans (WS), dietary inclusion at 160 g/kg; and 4) calcium salts of FA (CSFA), dietary inclusion at 32 g/kg. Rumen outflow of nutrients was estimated using the three markers reconstitution system (cobalt-EDTA, ytterbium chloride, and indigestible neutral detergent fibre [NDF]). Diets with FA sources decreased feed intake and increased FA intake. No differences in nutrient intake and digestibility were detected among cows fed diets supplemented with different FA sources. Diets with FA sources reduced the rumen outflow of DM and NDF, hence decreasing their passage rates. In addition, SO diet reduced the ruminal outflow of DM and NDF in comparison with WS and CSFA. Omasal sampling yielded the highest values of rumen outflow of NDF and potentially degradable NDF (pdNDF), whereas the reticular and abomasal samplings yielded intermediate and least values, respectively. The interaction effect between diet and sampling site was observed for rumen outflow of majority FA (except for C16:0, C18:0, and C18:2 trans-10, cis-12) and BH extension of C18:1 cis, C18:2, and C18:3. Calculations derived from abomasal sampling revealed that WS and CSFA diets had lower BH extent of C18:1 cis and C18:2 in comparison with SO, whereas cows fed CSFA had greater BH extent of C18:3 and lower BH extent of C18:1 cis compared to those fed WS. However, the latter results were not similar when calculations were performed based on the reticular and omasal samplings. Thus, there is evidence that neither reticular nor omasal samplings are suitable for estimating rumen outflow of FA in dry cows. In addition, WS and CSFA diets can increase the abomasal flow of polyunsaturated FA in dry cows.     

Evaluation of different markers for determination of the microbial nitrogen flow into the duodenum of dairy cows

2.6‐Diaminopimelic acid (DAPA), ribonucleic acid (RNA), ¹⁵N, D‐alanine (D‐ALA) and the amino acid profiles (AAP) were compared as microbial markers for determination of the microbial protein synthesis in the rumen. Three dairy cows (Schwarzbuntes Milchrind, LW 602 kg), each fitted with a rumen cannula and a re‐entrant cannula in the proximal duodenum, were offered four isoenergetic and isonitrogenous diets (mean daily intake 15.0 ± 0.45 kg DM; forage: concentrate = 50:50) in a periodic experiment. The diets contained soyabean extracted meal, meat and bone meal, pea meal and dried clover as major sources of protein. On the 4th day after administration of 9 g ¹⁵N‐labelled urea (95 atom‐% ¹⁵N‐excess) per day, samples of rumen fluid and duodenal digesta were obtained 3 h after feeding. The bacteria were isolated by differential centrifugation. Bacteria harvested from the rumen had significantly higher ¹⁵N enrichment and D‐ALA: N ratio than ‘duodenal’ bacteria. However, DAPA: N ratio was higher in ‘duodenal’ bacteria compared to rumen bacteria. There were no differences in RNA: N ratio between rumen and ‘duodenal’ bacteria. The source of the bacteria in the digestive tract has an influence on the ratio of microbial N: total N, especially when ¹⁵N, AAP, DAPA and D‐ALA but not RNA were used as markers. The most reproducible method was D‐ALA (C.V. 4.7 for rumen and 6.8 for ‘duodenal’ bacteria) followed by ¹⁵N (10.8 resp. 4.8) and RNA (9.7 resp. 8.2). The results obtained with ¹⁵N and D‐ALA agreed closely at the same source of bacteria. The RNA method reached the level of these markers (¹⁵N, D‐ALA) when the bacteria were isolated from the duodenum. It is concluded that D‐ALA (bacteria isolated from rumen and duodenum) and also ¹⁵N (bacteria isolated from duodenum) were the best markers for estimation of the microbial protein synthesis.     

Predicting duodenal flows and absorption of fatty acids from dietary characteristics in ovine and bovine species: a meta-analysis approach

Dietary and ruminal factors modify the ruminal biohydrogenation (RBH) of polyunsaturated fatty acids (FA), with duodenal FA flows being quantitatively and qualitatively different from FA intake. Using a meta-analysis approach from a database on duodenal flows of FA in ruminants, this study aimed to determine predictive equations for duodenal and absorbed flows of saturated FA, C18:1, C18:2 and C18:3 isomers, odd- and branched-chain FA (OBCFA), C20:5n-3, C22:5n-3 and C22:6n-3 and to quantify the effects of dietary and digestive factors on those equations. The database was divided into four subsets: forage, seed, vegetable oils or animal fats (oil/fat), and fish products (fish) subsets. Models of duodenal and absorbed FA flows were obtained through variance–covariance analysis. Effects of potential interfering factors (conservation mode and botanical families of forages, lipid source, technological processing of lipid supplements, diet composition and animal characteristics) were analysed. We obtained 83 models for duodenal FA flows as a function of FA intake for saturated FA (C14:0, C16:0 and C18:0), C18:1, C18:2 and C18:3 isomers and seven other models for OBCFA. For the seed/oil/fat subset, intakes of total C18:3, C18:2 and starch content increased the duodenal t11-C18:1 flow with 0.08, 0.16 and 0.005 g/kg of dry matter intake (DMI), respectively, whereas intake level [(DMI×100)/BW] decreased it. The c9c12c15-C18:3 RBH was higher for oil/fat than seed (96.7% v. 94.8%) and a protective effect of Leguminosae v. Gramineae against RBH for that FA appeared in the forage subset. The duodenal C17:0 flow increased with starch content and decreased with ruminal pH, respectively, whereas duodenal iso-C16:0 flow decreased with dietary NDF content for the seed/oil/fat subset. The duodenal C20:5n-3, C22:5n-3 and C22:6n-3 flows depended on their respective intake and the inhibitory effect of C22:6n-3 on duodenal C18:0 flow was quantified. Thirteen models of absorbed FA flows were performed depending on their respective duodenal flows. This study determined the effects of different qualitative and quantitative dietary and digestive factors, allowing for improved predictions of duodenal and absorbed FA flows.     

A simplified management of the in situ evaluation of feedstuffs in ruminants: Application to the study of the digestive availability of protein and amino acids corrected for the ruminal microbial contamination

The ruminal effective degradability (RED) and intestinal effective digestibility (IED) for dry matter, crude protein (CP) and amino acids (AA) were estimated by a simplified in situ method using pooled samples from rumen-incubated residues, which represented the ruminal outflow of undegraded feed. The effect of microbial contamination in the rumen was corrected using ¹⁵N infusion techniques. Studies were carried out for soybean meal (SBM), barley grain (BG) and lucerne hay (LH) in three wethers cannulated in the rumen and the duodenum. Uncorrected values of RED for CP obtained either by mathematical integration or our simplified method were similar in all feeds. Microbial N in the pooled samples of SBM, BG and LH were 2%, 11% and 24% of total N, respectively. However, intestinal incubation eliminated this microbial charge by 100%, 99% and 88%, respectively. With microbial corrections, RED showed an increase, and IED showed a decrease, except for SBM. With this correction, intestinal digested CP was reduced by 2% in SBM, 13% in BG and 34% in LH. Corrected IED of AA was relatively similar in SBM (97–99%). However, large variations were observed in BG (74–93%) and in LH (10–88%). Digestion in the rumen and intestine changed the essential AA pattern. Overall, our results support that AA digestion is affected by the characteristics of their radicals and their contents in plant cell wall proteins. The accurate estimation of feed metabolisable AA or protein requires effective measures that are corrected by ruminal microbial contamination. The proposed in situ method largely simplifies these tasks and allows a more complete and less expensive feed evaluation.     

Effect of silage type and concentrate level on conjugated linoleic acids, trans-C18:1 isomers and fat content in milk from dairy cows

The objective of the study was to examine how the fatty acid composition of milk especially concentrations of conjugated linoleic acids (CLA) and trans-C18:1 isomers and milk fat percentage were affected by silage type and concentrate level. Forty dairy cows were blocked and randomly assigned to one of four diets in a 2 x 2 factorial arrangement of treatments and a six week experimental period. Treatments were total mixed rations with maize (M) or grass (G) silage differing in polyunsaturated fatty acid (PUFA) profile and starch content, combined with a high (H) or a low (L) level of concentrate (with or without grain). Treatments had no significant effect on milk, protein and lactose yield, but energy corrected milk yield, milk fat percentage and fat yield was lower and protein percentage higher for maize compared with grass silage diets. Overall, maize silage diets resulted in higher concentrations of CLA isomers compared with grass silage diets, but there was a significant interaction between silage type and concentrate level for concentrations of cis9,trans11-CLA; trans10,cis12-CLA; trans11-C18:1 and trans10-C18:1. A high level of concentrate increased trans10,cis12-CLA and trans10-C18:1 and reduced cis9,trans11-CLA and trans11-C18:1 when maize but not grass silage was provided. The results suggest that high levels of concentrate (grain) do not significantly alter the pattern of PUFA biohydrogenation in the rumen, the concentration of CLA and trans-C18:1 isomers in milk or cause milk fat depression unless combined with forage naturally high in starch and C18:2n-6 such as maize silage.     

Rumen degradability and intestinal digestibility of brewers' grains as affected by origin and heat treatment and of barley rootlets

The rumen degradability and intestinal digestibility of dry matter (DM) and nitrogen (N) of three samples of brewers' grains (BG) and three of barley rootlets (BR) were determined. Rumen degradability was determined by the nylon bag technique in three rumen fistulated wethers. Intestinal digestibility was determined by the mobile nylon bag technique in two duodenal fistulated wethers. N content ranged from 41.2 to 46.4 g/kg DM for BG, and from 42.8 to 53.7 g/kg DM for BR. N effective degradability (NED), calculated for rumen outflow rates determined in each sheep, ranged from 57.2% to 70.9% for BG and from 79.0% to 84.0% for BR. N intestinal digestibility (NID) determined on 8 h-rumen incubated residues, ranged from 84.9% to 89.8% for BG and from 67.3% to 81.3% for BR. Lower rumen degradability was partially compensated by higher intestinal digestibility, resulting in a smaller variation in the estimated amount of digestible bypass N, which ranged from 24.7% to 36.7% for BG and from 10.8% to 17.1% for BR. One BG sample was selected to study the effects of heat treatment (HT) on its chemical composition, rumen degradability and intestinal digestibility. The BG sample was either freeze-dried (UBG) or dried at 50°C (50BG), 100°C (100BG), 135°C (135BG) and 175°C (175BG). Total N content was not affected by HT, but the acid–detergent insoluble N (expressed as percentage of total N content) increased from 13.7% to 54.1%. HT reduced the NED (from 76.5% to 25.6%) and, as a consequence, the supply of undegraded N to the duodenum was increased by 1.2, 1.8, 2.4 and 3.2 times for 50BG, 100BG, 135BG and 175BG, respectively. Drying at 50°C and 100°C had no adverse effects on the NID determined on 8 h-rumen incubation residues (mean value of 84.3%), but drying at 135 and 175°C decreased it to values of 80.1 and 51.6%, respectively. As a consequence, the estimated amount of digestible bypass N was increased by 1.2, 1.8, 2.3 and 1.9 times when drying at 50°C, 100°C, 135°C and 175°C, respectively.     

Effects of capric acid on rumen methanogenesis and biohydrogenation of linoleic and α-linolenic acid

Capric acid (C10:0), a medium chain fatty acid, was evaluated for its anti-methanogenic activity and its potential to modify the rumen biohydrogenation of linoleic (C18:2n-6) and α-linolenic acids (C18:3n-3). A standard dairy concentrate (0.5 g), supplemented with sunflower oil (10 mg) and linseed oil (10 mg) and increasing doses of capric acid (0, 10, 20 and 30 mg), was incubated with mixed rumen contents and buffer (1 : 4 v/v) for 24 h. The methane inhibitory effect of capric acid was more pronounced at the highest (30 mg) dose compared to the medium (20 mg) (-85% v. -34%), whereas the lower dose (10 mg) did not reduce rumen methanogenesis. A 23% decrease in total short-chain fatty acid (SCFA) production was observed, accompanied by shifts towards increased butyrate at 20 mg and increased propionate at 30 mg of capric acid (P < 0.001). Capric acid linearly decreased the extent of biohydrogenation of C18:2n-6 and C18:3n-3, by up to 60% and 86%, respectively. This reduction was partially due to a lower extent of lipolysis when capric acid was supplemented. Capric acid at 20 and 30 mg completely inhibited the production of C18:0 (P < 0.001), resulting in an accumulation of biohydrogenation intermediates, mainly C18:1t10 + t11 and C18:2t11c15. In contrast to effects on rumen fermentation (methane production and proportions of SCFA), 30 mg of capric acid did not induce major changes in rumen biohydrogenation as compared to the medium (20 mg) dose. This study revealed the dual action of capric acid, being inhibitory to both methane production and biohydrogenation of C18:2n-6 and C18:3n-3.     

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.     

Role of the protozoan Isotricha prostoma, liquid-, and solid-associated bacteria in rumen biohydrogenation of linoleic acid

From the simultaneous accumulation of hydrogenation intermediates and the disappearance of Isotricha prostoma after algae supplementation, we suggested a role of this ciliate and/or its associated bacteria in rumen biohydrogenation of unsaturated fatty acids. The experiments described here evaluated the role of I. prostoma and/or its associated endogenous and exogenous bacteria in rumen biohydrogenation of C18:2n-6 and its main intermediates CLA c9t11 and C18:1t11. Fractions of I. prostoma and associated bacteria, obtained by sedimentation of rumen fluid sampled from a monofaunated sheep, were used untreated, treated with antibiotics or sonicated to discriminate between the activity of I. prostoma and its associated bacteria, the protozoan or the bacteria, respectively. Incubations were performed in triplicate during 6 h with unesterified C18:2n-6, CLA c9t11 or C18:1t11 (400 μg/ml) and 0.1 g glucose/cellobiose (1/1, w/w). I. prostoma did not hydrogenate C18:2n-6 or its intermediates whereas bacteria associated with I. prostoma converted a limited amount of C18:2n-6 and CLA c9t11 to trans monoenes. C18:1t11 was not hydrogenated by either I. prostoma or its associated bacteria but was isomerized to C18:1c9. A phylogenetic analysis of clones originating from Butyrivibrio-specific PCR product was performed. This indicated that 71% of the clones from the endogenous and exogenous community clustered in close relationship with Lachnospira pectinoschiza. Additionally, the biohydrogenation activity of solid-associated bacteria (SAB) and liquid-associated bacteria (LAB) was examined and compared with the activity of the non-fractioned I. prostoma monofaunated rumen fluid (LAB + SAB). Both SAB and LAB were involved in rumen biohydrogenation of C18:2n-6. SAB fractions performed the full hydrogenation reaction to C18:0 while C18:1 fatty acids, predominantly C18:1t10 and C18:1t11, accumulated in the LAB fractions. SAB and LAB sequence analyses were mainly related to the genera Butyrivibrio and Pseudobutyrivibrio with 12% of the SAB clones closely related to the C18:0 producing B. proteoclasticus branch. In conclusion, this work suggests that I. prostoma and its associated bacteria play no role in C18:2n-6 biohydrogenation, while LAB convert C18:2n-6 to a wide range of C18:1 fatty acids and SAB produce C18:0, the end product of rumen lipid metabolism.     

Evaluation of the effects of different diets on microbiome diversity and fatty acid composition of rumen liquor in dairy goat

Fat supplementation plays an important role in defining milk fatty acids (FA) composition of ruminant products. The use of sources rich in linoleic and α-linolenic acid favors the accumulation of conjugated linoleic acids isomers, increasing the healthy properties of milk. Ruminal microbiota plays a pivotal role in defining milk FA composition, and its profile is affected by diet composition. The aim of this study was to investigate the responses of rumen FA production and microbial structure to hemp or linseed supplementation in diets of dairy goats. Ruminal microbiota composition was determined by 16S amplicon sequencing, whereas FA composition was obtained by gas-chromatography technique. In all, 18 pluriparous Alpine goats fed the same pre-treatment diet for 40±7 days were, then, arranged to three dietary treatments consisting of control, linseed and hemp seeds supplemented diets. Independently from sampling time and diets, bacterial community of ruminal fluid was dominated by Bacteroidetes (about 61.2%) and Firmicutes (24.2%) with a high abundance of Prevotellaceae (41.0%) and Veillonellaceae (9.4%) and a low presence of Ruminococcaceae (5.0%) and Lachnospiraceae (4.3%). Linseed supplementation affected ruminal bacteria population, with a significant reduction of biodiversity; in particular, relative abundance of Prevotella was reduced (−12.0%), whereas that of Succinivibrio and Fibrobacter was increased (+50.0% and +75.0%, respectively). No statistically significant differences were found among the average relative abundance of archaeal genera between each dietary group. Moreover, the addition of linseed and hemp seed induced significant changes in FA concentration in the rumen, as a consequence of shift from C18 : 2n-6 to C18 : 3n-3 biohydrogenation pathway. Furthermore, dimethylacetal composition was affected by fat supplementation, as consequence of ruminal bacteria population modification. Finally, the association study between the rumen FA profile and the bacterial microbiome revealed that Fibrobacteriaceae is the bacterial family showing the highest and significant correlation with FA involved in the biohydrogenation pathway of C18 : 3n-3.     

Biohydrogenation of linoleic acid by rumen fungi compared with rumen bacteria

AIMS: To investigate biohydrogenation of linoleic acid by rumen fungi compared with rumen bacteria, and to identify the fungus with the fastest biohydrogenation rate. METHODS AND RESULTS: Biohydrogenation of linoleic acid by mixed rumen fungi and mixed rumen bacteria were compared in vitro. With mixed rumen bacteria, all biohydrogenation reactions were finished within 100 min of incubation and the end product of biohydrogenation was stearic acid. With mixed rumen fungi, biohydrogenation proceeded more slowly over a 24-h period. Conjugated linoleic acid (CLA; cis-9, trans-11 C18 : 2) was an intermediate product, and vaccenic acid (VA; trans-11 C18 : 1) was the end product of biohydrogenation. Fourteen pure fungal isolates were tested for biohydrogenation rate. DNA sequencing showed that the isolate with the fastest rate belonged to the Orpinomyces genus. CONCLUSIONS: It is concluded that rumen fungi have the ability to biohydrogenate linoleic acid, but biohydrogenation is slower in rumen fungi than in rumen bacteria. The end product of fungal biohydrogenation is VA, as for group A rumen bacteria. Orpinomyces is the most active biohydrogenating fungus. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study to demonstrate that rumen fungi can biohydrogenate fatty acids. Fungi could influence CLA content of ruminant products.     

Effects of linseed lipids fed as rolled seeds, extruded seeds or oil on organic matter and crude protein digestion in cows

Effects of fatty acids of linseed in different forms, on ruminal fermentation and digestibility were studied in dry cows fitted with ruminal and duodenal cannulas. Four diets based on maize silage, lucerne hay and concentrates (65/10/25 dry matter (DM)) were compared in a 4 × 4 Latin square design experiment where the diets were: control diet (C), diet RL supplied 75 g/kg DM rolled linseeds, diet EL supplied 75 g/kg DM extruded linseeds, and diet LO supplied 26 g/kg DM linseed oil and 49 g/kg DM linseed meal. The diets did not differ in total organic matter (OM) and fibre digestibility, in forestomach and intestinal OM digestibility, and in duodenal N flow. Microbial N duodenal flow tended to be lower for RL versus C diet (P<0.1). Extrusion did not reduce ruminal crude protein (CP) degradation in vivo and in situ. Volatile fatty acid concentration and pattern, and protozoa concentration in the rumen, did not vary among diets. Results confirm the absence of a negative effect of a moderate supply of linseed on rumen function, as well as no effect of extrusion on its ruminal CP degradability.     

Utilisation of fish oil in ruminants: I. Fish oil metabolism in sheep

Five sheep with rumen and abomasal cannulae were offered three diets sequentially in the order: control (C) pellets (lucerne hay-oat grain: 60/40, w/w), control plus unprotected tuna oil (UTO pellets), and control plus tuna oil protected (casein-formaldehyde matrix) against ruminal biohydrogenation (PTO pellets). In supplemented diets, tuna oil constituted 3% (w/w) of total dry matter (DM), and each supplement was fed for 12 days, with 9 days allowed between the two fish oil feeding periods to minimise carry-over effects. Daily DM intake was 785±38 g/head during the control period. It was significantly reduced by UTO feeding (6.2%, P<0.05) but not PTO feeding. The level of EPA in the abomasum during PTO feeding was double that measured during UTO feeding (1.30 versus 0.61% of FA, P<0.05). The level of DHA in the abomasum did not significantly differ between UTO and PTO feeding periods. Both tuna oil supplements significantly increased the levels of 18:1 trans and that of a fatty acid derivative identified as 10-hydroxystearic acid (10-HSA) in both the rumen and abomsum. Tuna oil supplementation also altered the fatty acid composition of plasma lipid fractions and 10-HSA was solely incorporated into plasma free fatty acids. This study indicates that substantial protection of tuna oil against ruminal hydrogenation inhibited reduced feed intake, but increased the proportion of 18:1 trans isomer and fatty acids derivatives (10-HSA), which indicate interference with metabolism in the rumen.     

Effect of heat or formaldehyde treatment on the rumen degradability and intestinal tract apparent digestibility of protein in soya-bean meal and in rapeseed meals of different glucosinolate content

Three Hereford × Friesian/Holstein heifers were each fitted with rumen and simple T-piece duodenal cannulae. They were used in an experiment of latin square design to study the rumen degradability and intestinal apparent digestibility (using the mobile dacron bag technique) of protein in soya-bean meal (SBM) and in low and high glucosinolate rapeseed meals (LgsRSM and HgsRSM, respectively) each in untreated (UT), heat treated (HT) and formaldehyde treated (FT) forms. For rumen incubation times of 24 h there were no significant protein source × processing treatment interactions for either degrability of protein in the rumen (dgN) or for the proportion of food protein undegradable in the rumen and digestible in the small intestine (DUDN). At this time dgN values for FT, HT and UT were 0.263, 0.374 and 0.418 (SEM 0.0078; P < 0.001) respectively and DUDN values for SBM, LgsRSM and HgsRSM were 0.320, 0.348 and 0.386 (SEM 0.0078; P < 0.001) respectively. Compared with UT, and to a lesser extent HT, FT at all times after 0 h incubation significantly decreased dgN values and increased DUDN values for all three protein sources. Compared with UT, HT significantly decreased dgN. Apparent digestibility of the protein of SBM in the intestines was greater than that of LgsRSM and HgsRSM, and overall the values of available undegraded protein for FTLgs and HgsRSM, but not of FTSBM, were similar to or greater than for the UT sources. FT tended to decrease the total tract apparent digestibility of the protein sources.     

Effects of Nitrogen fertilisation and regrowth period on fatty acid concentrations in perennial ryegrass (Lolium perenne L.)

Information on lipids in forages is scarce, and effects of nitrogen (N) application level and regrowth period on the fatty acid (FA) concentration and composition of perennial ryegrass (Lolium perenne L.) were studied. N was applied at 0, 45 and 100 kg ha^?1, and swards were cut after various regrowth periods, resulting in six treatments designed as randomised blocks with three replicates. The stages of development ranged from vegetative to elongating swards, with herbage yield levels from 1.9 to 4.2 t dry matter (DM) ha^?1. Concentrations of individual FA were determined by gas chromatography, and canopy characteristics and herbage quality were assessed. The treatments resulted in canopies with contrasting DM yields and protein concentrations. Five FAs, representing 98% of total FAs, were studied in detail. On an average, the concentration of these major FAs in fresh grass was 15.1 g kg^?1 DM, and 69% of the major FAs consisted of C18:3. Regrowth period affected (P < 0.05) the total FA concentration, and significantly (P < 0.01) lower concentrations of C18:3 and C16:1 were found after a longer period of regrowth. N application resulted in higher (P < 0.001) concentrations of all FAs. The FA composition was not affected by N application, but a longer regrowth period significantly (P < 0.001) decreased the proportion of C18:3 and increased those of C18:2 and C16:0. A strong, positive overall linear relation was found between the concentrations of total FAs and C18:3 with the crude protein concentration in the herbage. These studies demonstrate opportunities to affect the FA concentration and composition of FA in herbage through management strategies, which could affect milk FA composition.