Identification of lucerne stem cell wall traits related to in vitro neutral detergent fibre digestibility

Genetic improvement of forage digestibility, especially utilizing marker assisted selection and recombinant DNA techniques, requires identification of specific biochemical traits and associated genes that impact digestibility. We undertook a study to identify cell wall (CW) traits of lucerne (Medicago sativa L.) stems that were consistently and strongly correlated with in vitro neutral detergent fibre (NDF) digestibility, a measurement that has been shown to correlate with animal performance. Spring and summer harvested lucerne stem material, for 2 years, from 24 individual plants in each of two germplasm sources were analyzed for 16 and 96 h in vitro NDF digestibility, and cell wall concentration and composition (monosaccharide constituents of cellulose, hemicellulose, and pectin; and Klason lignin (KL)) by the Uppsala dietary fibre method using near-infrared reflectance spectroscopy (NIRS). Pearson correlation coefficients were calculated for the relationships among these cell wall traits and with in vitro NDF digestibility. Concentrations of the pectin monosaccharide components were all negatively correlated (r=−0.73 to −0.94) with total cell wall concentration. In contrast, the three most abundant cell wall components glucose (Glc), xylose (Xyl) and Klason lignin were not correlated, or only weakly positively correlated (r<0.35), with cell wall concentration. Cell wall concentration was consistently negatively correlated (r=−0.60 to −0.94) with both 16 and 96 h in vitro NDF digestibility. In contrast, Klason lignin concentration was only marginally correlated (r<0.30) with 16 h in vitro NDF digestibility, but strongly negatively correlated (r=−0.71 to −0.74) with 96 h in vitro NDF digestibility. This is consistent with previous reports which show that lignin affects potential extent of digestion, but not rate. Cell wall glucose and xylose concentrations were inconsistently correlated with fibre digestibility. The monosaccharide components of pectin were consistently positively correlated (r=0.54–0.90) with in vitro NDF digestibility, except for 96 h in vitro NDF digestibility of spring harvested stems. Growth environment (year) and germplasm source had only minor impacts on the preceding correlation patterns, whereas spring versus summer harvests accounted for the inconsistencies observed among correlations for cell wall traits. The results of this study indicate that genetic improvement of fibre digestibility of lucerne stems should target genes that reduce total cell wall concentration, perhaps by reducing the rate of xylem tissue deposition during maturation, and reduce Klason lignin and increase pectin concentrations in the cell wall to improve potential extent and rate of fibre digestibility, respectively.     

Ruminal degradability and in vitro intestinal digestibility of sunflower meal and in vitro digestibility of olive by-products supplemented with urea or sunflower meal: Comparison between goats and sheep

The in vitro digestibility of two-stage dried olive cake (TSDOC) and olive leaves (OL) unsupplemented or supplemented with increasing amounts of urea (U) or sunflower meal (SM) (0, 1.5, 2 and 2.5 g/100 g organic matter (OM) of the by-product) was determined. Chemical and amino acid composition, in vitro digestibility, in situ rumen degradability of crude protein and amino acids, and in situ–in vitro intestinal digestibility of SM CP and amino acids was determined. The in sacco rumen degradability and in vitro intestinal availability of CP and individual amino acids were also determined. Results obtained in Granadina goats and Segureña wethers were compared. SM provides arginine, glycine and aspartic and glutamic acids. The addition of increasing amounts of U or SM improved (P<0.001) the IVDMD and IVOMD of both TSDOC and OL. There was no effect (P>0.05) of the rumen inoculum origin on in vitro TSDOC digestibility. In contrast, values for OL were higher (P<0.001) for goats versus sheep. In sacco ruminal CP degradability of SM was relatively high, and similar in sheep and goats (ED=0.78 and 0.75 for sheep and goats). Individual amino acid ruminal degradability had different values, being lowest for methionine, leucine, proline, tyrosine and cysteine. Values obtained for individual amino acids differed from those of CP. Apparent intestinal digestibility of undegraded protein (AIDUP) of SM was high (0.86 and 0.98, respectively, for sheep and goats). The intestinally absorbable protein (IADP) was low (18.9 and 24.0 for sheep and goats, respectively). Results indicate that goats and sheep have the same capacity for TSDOC digestion, but goats showed a better capacity than sheep for OL utilisation. Although the amino acids supply to the intestine from SM is not important it could be a good supplement for low degradable protein feedstuffs such as TSDOC and OL.     

Validation of in sacco method: influence of sampling site,nylon bag or rumen contents,on fibrolytic activity of solid-associated microorganisms

Three ruminally cannulated cows, fed twice daily with a 70:30 forage:concentrate diet, were used to investigate the differences in fibrolytic activity of solid-associated microorganisms between nylon bags and rumen contents. Two different grass hays (regrowth and late harvested) were incubated in ruminal nylon bags. After 2 h or 23 h incubation time, pH was measured in bags and rumen contents, and enzymes of solid-associated microorganisms were extracted from bag residues and surrounding digesta by grinding, freezing, defrosting and sonication. Xylanase, avicelase, β-D-xylosidase and β-D-glucosidase activities were measured. Activities were lower in bag residues than in rumen digesta, and differences were greater after 2 h than after 23 h incubation time. Causes of these differences are discussed. For each incubation time and each enzyme, the differences in solid-associated microorganisms activities between rumen and bags contents were independent of the quality of hay in the bag. Thus the lower fibrolytic activity inside the bags may account for an underestimation of in vivo ruminal fiber degradation by the in sacco method, but this underestimation may be similar whatever the nature and content of forage cell walls.     

Effect of tea saponin on rumen fermentation in vitro

The present study was conducted to investigate the effect of tea saponins (TS) on ruminal fermentation in vitro using gas syringes as incubators. The TS were added at levels of 0, 2, 4, 6 and 8 mg against 200 mg mixture of corn meal and grass meal (1/1, w/w) in rumen fluid. In vitro gas production (GP) was recorded and methane concentration was determined at 3, 6, 9, 12 and 24 h incubation. After 24 h, the incubation was stopped and the inoculants were determined for pH, ammonia-N, volatile fatty acids (VFAs), protozoa counts and microbial protein yield. The GP was increased with the increasing level of TS except 8 mg at 24 h, which kept little change from that of the control. Methane concentration was decreased at all levels of TS at each incubation time. At 24 h incubation, inclusion with 2, 4, 6 and 8 mg of TS decreased methane concentration by 13, 22, 25 and 26%, respectively. The pH of ruminal fluid was slightly lower at 4 and 6 mg TS, but all values were in the normal range. Ammonia-N concentrations decreased significantly (P < 0.01) when the TS were included. Concentrations of individual and total VFAs were not significantly effected by TS addition. The TS significantly inhibited the protozoa growth in ruminal fluid (P < 0.01). At 24 h incubation, protozoa counts were reduced by 19, 25, 45 and 79%, respectively at levels of 2, 4, 6 and 8 mg of TS compared to that in control. The microbial protein was enhanced with the TS addition except 2 mg level, and reached 1.92, 2.36 and 2.61 mg/mL with addition of 4, 6 and 8 mg TS, compared to 1.50 mg/mL in control. It is suggested that TS could modify the rumen fermentation and inhibit the release of methane and ammonia, which may be beneficial for improving nutrient utilization and animal growth.     

Potential of carvacrol to modify in vitro rumen fermentation as compared with monensin

The aim of this study was to assess the effect of carvacrol supplement as a dietary additive to rumen fermentors, fed a barley seed:alfalfa hay (70:30) ration and to compare its effect with monensin supplementation. The material was incubated with goat ruminal fluid and four different treatments were included: no additive (C), 7.5 mg/l monensin (M), 250 mg/l carvacrol (C250) and 500 mg/l carvacrol (C500). The addition of carvacrol reduced in vitro dry matter (DM), crude protein (CP) and neutral-detergent fibre (NDF) digestion. The effects induced by C250 on DM digestion at 72 h of incubation were comparable with those of M, whereas a greater reduction was obtained when carvacrol was supplemented at 500 mg/l concentration (68.9, 68.5 and 53.0 v. 76.1% for M, C250 and C500 v. C, respectively). The reduced CP potential degradability by supplements (51.2, 53.9 and 51.5 v. 72.8% for M, C250 and C500 v. C, respectively) was mainly caused by a reduction of the slowly degradable fraction. Volatile fatty acid (VFA) profiles determined after 48 h of incubation showed C250 increased butyrate and decreased acetate proportions, whereas M mainly stimulated propionate proportions, suggesting that the mechanism of action of carvacrol and M differs. C500 significantly reduced total VFA production. Carvacrol could be of great interest for its usage as a potential modulator of ruminal fermentation. Future research, including in vivo studies, in order to understand the factors that contribute to its antimicrobial activity and the selection of the optimal dose is required.     

Plasmalogens of microbial communities associated with barley straw and clover in the rumen

Abstract: Microbial plasmalogen aldehydes (detected as dimethyl acetals, DMA) have been used to compare microbial populations associated with clover and barley straw incubated in nylon mesh bags in the rumen of a cow. The results suggest that the populations involved in the digestion of these substrates differ substantially and that population changes occur as digestion proceeds: these conclusions were supported by electron-microscopic observations. Analysis of DMA suggested that populations associated with the particles of straw and clover differed more markedly than the corresponding populations in the liquid phase. When straw was pre-incubated with the rumen cellulolytic bacterium Ruminococcus flavefaciens strain 17, the DMA characteristic of this bacterium were present at increased levels during subsequent incubation of the straw in the rumen, though the R. flavefaciens DMA tended to contribute a smaller proportion of the total DMA as the incubation time in the rumen was increased from 24 to 72h.     

Einfluß der Gerbsäure in Kombination mit Strohmehl auf die Verdaulichkeit der Rohnährstoffe und Aminosäuren bei Legehennen der Mastrichtung

Four fungi varieties (Wild variety, Chaetomium cellulolyticum, Trichoderma harzianum and Penicillium colony 10) were cultivated in a solid‐state fermentation on a wheat straw‐wheat bran‐sugar beet pulp mixture (20 : 40: 40%).

The fermented substrates contained xylanase (22.1 to 78.6 Units g^‐1 DM) and cellulase (0.47 to 14.0 Units g^‐1 DM). Fermented substrate was homogeneously mixed with ground wheat straw (1: 2; 1: 5 and 1 : 10), moistened to 30 or 50% DM, airtightly stored for 24 h at 50°C and deep frozen. All samples were given in nylon bags and incubated for 48 h in the rumen of 4 sheep.

In sacco DMD decreased with higher straw proportions in the mixture (1: 2; 1 : 5 and 1: 10: 52.0; 46.8 and 44.2% resp.), the DM‐content did not significantly influence the in sacco DMD. The Wild variety and Chaetomium cellulolyticum did not significantly influence the in sacco DMD, Trichoderma harzianum and Penicillium colony 10 decreased DMD in some cases.

In general added fungi did not improve the rumen degradability of fibre.     

Effect of the proportions of neutral detergent fibre and starch, and their degradation rates, on in vitro ruminal fermentation

Effects of proportions of neutral detergent fibre (aNDFom) and starch, as well as their degradation rates, on rumen fermentation were tested using an in vitro rumen simulation system (SIMCO). The in vitro system was designed to simulate selective particle retention and had an average fluid volume of 1150 ml with a liquid dilution rate of approximately 0.07 h⁻¹. Two types of hay (aNDFom sources) and two types of starch were each included at two different levels in the diet and were examined in an experiment following a 2×2×2 factorial arrangement of treatments (eight diet combinations). The hay was either late-cut timothy (Phleum pretense L.) or early cut meadow grass (Poa pratensis L.), with ruminal in situ aNDFom digestion rates of 0.03–0.04 and 0.07–0.08 h⁻¹, respectively. The two starch types were raw (R) and cooked (C) potato starch with previously determined in vitro ruminal digestion rates of 0.04 and 0.20 h⁻¹, respectively. The starch levels were 300 and 600 g/kg diet dry matter (DM) with the remaining being hay (282–682 g/kg DM) and peptone (14–111 g/kg DM). The aNDFom level varied among the diets with different starch levels and hay types. The peptone acted as a source of peptides and, together with ammonia salts from buffer, was used to balance the N contents of the diets. The feeding level for each of the eight vessels was 28 g DM/d. Two 10-day simulations were made with the system. The average pH was higher (P<0.05) for all treatments with raw potato starch (6.19) versus cooked starch (6.07). Protozoa scores, on a qualitative scale, declined faster at the higher starch level. The aNDFom digestibility was, as expected, higher (P<0.001) for meadow hay (0.57) than timothy (0.32), and was also higher (P<0.001) at the lower starch level (0.54) versus the higher (0.35). Microbial protein production efficiency (mg microbial N/g organic matter truly digested) was higher for the faster degrading aNDFom (P<0.01) and starch (P<0.05) sources, but was not affected by starch level. Cooked starch resulted in a lower acetate proportion (449 mmol/mol versus 591 mmol/mol VFA; P<0.001) but higher proportions of propionate (297 mmol/mol versus 236 mmol/mol VFA; P<0.001), and butyrate (169 mmol/mol versus 127 mmol/mol VFA; P<0.01). Butyrate increased with starch level (127 mmol/mol versus 169 mmol/mol VFA; P<0.01), and was also higher for meadow hay versus timothy (168 mmol/mol versus 128 mmol/mol VFA; P<0.01). Interactions between the treatments demonstrate that the response in VFA pattern to starch level is dependent on starch and aNDFom sources. Substrates such as starch and aNDFom are fermented differently depending on their rates of ruminal degradation.     

Effect of glyphosate contaminated feed on rumen fermentation parameters and in sacco degradation of grass hay and corn grain

Abstract Four rumen fistulated wethers were used to investigate the effect of glyphosate contaminated feed on rumen fermentation. The rations were based on corn silage, urea and a vitamin-mineral premix, either in the absence or presence of 0.77 g glyphosate per kg DM. Furthermore, rations were fed either with or without aromatic amino acid supplementation. During four periods of 28 days, sheep received each of the four dietary treatments according to a Latin square. After 14 days of adaptation rumen fermentation parameters (pH, ammonia, volatile fatty acids) were measured on day 15 over a five-hour period after the morning feeding. The remaining 13 days served for in sacco degradation studies with grass hay and corn grain. Ammonia (NH3) and pH of rumen fluid were within the normal range for all dietary treatments (NH3: 9.1-32.3 mmol x l(- l), pH: 6.2-6.7). Neither rumen fermentation parameters nor in sacco DM and NDF degradation of incubated feedstuffs were significantly affected by glyphosate, with or without aromatic amino acid supplementation. Kinetic profiles of the in sacco dry matter and NDF degradation of grass hay were almost identical for the dietary treatments.     

Chemical defense in birch: Inhibition of digestibility in ruminants by phenolic extracts

Summary The biological activity of phenolic extracts originating from winter twigs of birch (Betula pendula Roth.) was measured using the ruminant in vitro method and the nylon bag technique. Different extracts were prepared by extraction with organic solvents, removing phenols of corresponding solubility. The extract of birch twigs (diameter <1.5 mm) contained about 19% phenol equivalents, corresponding to 6% of twig dry matter (DM). Coarse birch twigs (diameter 1.5–5 mm) contained about 3% in the DM Phenolic extracts from the fine birch twigs were added to coarse birch twigs and common timothy (Phleum pratense L.) to mimic natural concentrations of fine birch twigs. Controls and the plant material with phenolic extract added were incubated for different times with rumen inocula taken from a sheep fed browse and a goat fed hay. Nylon bags containing phenolic treated hay were incubated in the rumen of the goat for 6 and 48 h. Phenolic extracts had a considerable negative effect on the organic matter (OM), protein and cell wall (neutral detergent fiber, NDF) digestibility in vitro. The nylon bag OM disappearance was also depressed by the extract. The effects were measurable after 6 h of digestion both in vitro and in sacco.The high inhibitory effect by the extracts on digestibility persisted even after removal of lipophilic fractions. This suggests that some or several water-soluble phenolic substances are responsible for the depression of digestibility. The depression of OM digestibility is linearly related to the concentration of phenols added. However, the inhibition of nylon bag digestibility plateaus at high phenol concentrations, suggesting that some fraction of the substances undergo complex formation with macromolecules of the plant.The results strongly indicate that water-soluble phenols of birch make up an important part of its chemical defense in winter by possessing antinutritional properties. Thus their potential importance in the nutrition of wild herbivores must not be ignored.     

Relationships between kernel vitreousness and dry matter degradability for diverse corn germplasm: II. Ruminal and post-ruminal degradabilities

Correlations between kernel vitreousness and ruminal in situ (RDMD) and total tract dry matter (TDMD; sum of ruminal in situ and post-ruminal in vitro measurements) degradabilities were determined for 33 diverse corn germplasm sources. These included a wide range of endosperm characteristics from opaque 2 (o2) types to densely packed flint types, and a number of intermediates. Harvests were done at two growth stages; 1/2 milk-line (ML) and black-layer (BL). Kernels from middle portion of ears were oven dried at 40 °C for 72 h and ground through a Wiley mill (6 mm screen) for measurement of in situ RDMD after 0 and 14 h of incubation using two steers (1.5 g/bag × 8 replicates per time point per steer in 5 cm × 5 cm bags of 50 μm pore size). Residue from the 14 h bags proceeded to an 8 h in vitro enzymatic post-ruminal digestion after which the residue was oven dried at 62 °C for 48 h and dry matter content determined. Inbred by harvest-stage interactions were observed for 0-h disappearance and TDMD. Vitreousness had strong negative correlations with degradability measurements, particularly for more mature (BL) samples (−0.728, −0.770 and −0.603) versus ML (−0.569, −0.541 and −0.338) for 0 h disappearance, RDMD and TDMD, respectively. Vitreousness was highly correlated with corn degradability, especially at a black-layer stage of harvest, in this diverse corn germplasm.     

Effect of monensin, fish oil or their combination on in vitro fermentation and conjugated linoleic acid (CLA) production by ruminal bacteria

An in vitro study was conducted to examine the effect of adding monensin, fish oil, or their combination on rumen fermentation and conjugated linoleic acid (CLA) production by mixed ruminal bacteria when incubated with safflower oil. Concentrate (1 g/100 ml) with safflower oil (0.2 g/100 ml) was added to a mixed solution (600 ml) of strained rumen fluid and buffer (control). Monensin (10 ppm), fish oil (0.02 g/100 ml), or monensin plus fish oil was also added into control mixture. All the culture solutions prepared were incubated anaerobically at 39 °C for 12 h. A higher pH and ammonia concentration were observed from the culture solution containing monensin at 12 h of incubation than those from the control or the culture containing fish oil. Monensin increased (P < 0.007) the C₃ content over all the collection times of culture solution while reducing the C₄ content at 6 h (P < 0.018) and 12 h (P < 0.001) of incubations. Supplementation of monensin, fish oil or their combination changed the content of C₁₈-fatty acids of ruminal culture. Monensin alone reduced (P < 0.021) the content of cis-9, trans-11 CLA compared to fish oil at all sampling times, but increased (P < 0.041) the trans-10, cis-12 CLA production compared to fish oil addition and the control which were similar at incubation for 12 h. The combination of monensin and fish oil increased the content of cis-9, trans-11 CLA (P < 0.023) and transvaccenic acid (TVA, P < 0.018) significantly compared to the control or monensin alone at incubation for 12 h.     

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.     

Grass maturity effects on cattle fed silage-based diets. 2. Cell wall digestibility,digestion and passage kinetics

Four silages were harvested at approximately 1-week intervals from the same timothy-meadow fescue sward and studied in a 4 × 4 Latin square experiment with four ruminally and duodenally cannulated young cattle. The diets comprised silage and concentrate (7:3 dry matter (DM) basis) and were fed at a rate of 70 g DM kg^−0.75 liveweight in two equal meals per day.Neutral detergent fibre (NDF) digestibility was 0.757, 0.765, 0.692 and 0.686 on diets based on the four silages in order of harvest date. Increasing maturity of grass ensiled showed linear (P_L < 0.001) and cubic (P_C < 0.01) trends. NDF was separated into digestible (DNDF) and indigestible (INDF) fractions, which differed clearly in their rate of passage from the rumen (on average 0.0141 vs. 0.0258 h⁻¹). The rate of digestion (k_d) of DNDF was on average 0.076 h⁻¹ when derived from the rumen evacuations but only 0.036 h⁻¹ when calculated from the disappearance from nylon bags incubated in the rumen. Both methods detected decreased k_d of NDF with increasing maturity of grass ensiled.Rate of passage from the rumen increased with increasing maturity of grass both when determined for NDF with rumen evacuation technique and from the faecal excretion of ytterbium calculated with a two-pool model. Mean retention time (MRT) in the non-escapable pool of particles increased (P_L < 0.01) with increasing grass maturity, the opposite being true for the escapable pool (P_L < 0.05), resulting in no change in the total ruminal MRT. Pool sizes of ruminal DM P_L < 0.01) and NDF (P_L < 0.001) increased with increasing maturity of grass. Ruminal NDF digestibility was calculated by different methods. When digestion kinetic parameters were derived from rumen evacuations and two-pool models used for passage kinetics, estimated digestibilities were very close to the observed ones.     

An alternative oven method combined with different detergent strengths in the analysis of neutral detergent fibre

Two methods for preparing neutral detergent fibre (NDF) – the standard method of Van Soest, P.J., Robertson, G.B., Lewis, B.A., `Symposium: Carbohydrate methodology, metabolism, and nutritional implications in dairy cattle.' J. Dairy Sci. 74 3583–3597 (1991), and an alternative 16 h oven method at 90°C. Neutral detergent solution (100%) was compared with solutions diluted to 50, 25 and 0% of normal strength. Sodium sulfite and a heat-stable amylase were used in all treatments. Forty-four samples consisting of 11 dried and ensiled forages, three straws, nine starchy concentrates, 10 high-protein concentrate feeds, three samples of tropical browse leaves and eight manures from both ruminants and non-ruminants were used. The 0% treatment was the only treatment giving significantly higher NDF values compared to the others (p<0.05). Sensitivity to detergent strength seemed to be sample-specific in the high protein group as meat meal and maize gluten had higher NDF values at reduced detergent strengths. We recommend using the oven method and a detergent strength of 25% of the current standard in combination with sodium sulfite for low protein feeds. For starchy feeds a heat-stable amylase should be used and high lipid samples should be acetone-extracted before isolating NDF. For high protein feed samples only the standard method can yet be recommended. The oven method is labour-saving and more convenient and arguments for reduced detergent strength suggest lower cost and reduced waste water pollution.     

The suitability of a faecal suspension of sheep as inocula for the estimation of utilizable crude protein of feeds by in vitro incubation

The objective of the experiments was to study the suitability of using a faecal suspension of sheep for the estimation of the utilizable crude protein (uCP) of feeds for sheep by an in vitro incubation. Twenty-four single feeds and eight feed mixtures were used as incubation substrates. In Experiment 1, the gas production after the in vitro incubation with rumen fluid or with a faecal suspension of a sheep were compared using the Hohenheim gas test. It was found that there were significant linear regression between the 24, 48 and 72 h gas production with rumen fluid and those with faecal suspensions of 35, 50, 100 and 150 g wet faeces of sheep (which were 18.6, 23.5, 52.0 and 70.5 g faeces DM, respectively) per litre McDougall's buffer (P < 0.0001). The highest regression coefficient (r2) was calculated between the gas production after inoculation with a suspension of 100 g wet faeces per litre McDougall's buffer (x, ml x 200 mg (-1) feed DM) for 48 h and the gas production after inoculation with rumen fluid (y, ml x 200 mg (-1) feed DM) for 24 h: y = 0.82 (+/- 0.07)x + 9.87 (+/-3.83), r2 = 0.82, n = 32, P < 0.0001. Based on these results, in Experiment 2 the estimation of utilizable crude protein (uCP) of feeds was compared by using the in vitro incubation technique of Zhao and Lebzien (2000), where feeds were inoculated either with rumen fluid or with a faecal suspension (100 g wet faeces of sheep, i.e. 52 g faeces DM per litre McDougall's buffer). The results indicated that there were no significant differences of the estimated uCP after inoculation with rumen fluid or the faecal suspension (P > 0.05). A significant regression was found between the uCP after incubation for 48 h with 100 g wet faeces (x, g x kg (-1) DM) and the uCP after incubation for 24 h with rumen fluid (y, g x kg(-1) DM): y = 0.95 (+/-0.10)x - 4.90 (+/-26.70), r2 = 0.75, n = 32, Although this regression was significant, the coefficient r2 was not high. Therefore, further research is needed before sheep faeces could replace rumen fluid as an inocula for the estimation of uCP by the in vitro incubation technique.     

In vitro estimations of the rate and extent of ruminal digestion of starch-rich feed fractions compared to in vivo data

An experiment was conducted to study the rumen digestion characteristics of whole feeds (WF) and the neutral detergent fibre (aNDF) and neutral detergent soluble (NDS) fractions of a range of starch-rich feeds using an automated in vitro gas production (GP) technique. In addition, the ruminal digestibility values predicted from the GP data were compared to previously acquired in vivo data. Nine feeds with starch concentrations ranging from 389 to 712 g/kg dry matter and with known in vivo digestibilities were subjected to neutral detergent extraction. The GP for each WF and the corresponding aNDF fractions were measured in duplicate in buffered rumen fluid during 72 h on two occasions. The fermentation residues were collected and analyzed for aNDF concentration to estimate their true organic matter (OM) and NDF digestibility. The GP from the NDS fraction was calculated by subtracting the GP from the aNDF fraction from the GP of the WF. A three-pool Gompertz model was fitted to the GP profiles (R² = 0.99) and a two compartment, mechanistic and dynamic rumen model was used to predict the digestibility of the potentially digestible feed fraction and the effective digestion rate (k_d). The true OM and NDF digestibility determined for the WF ranged from 0.804 to 1.011 and from 0.362 to 1.107, respectively. The NDF digestibility determined for the aNDF fraction ranged from 0.410 to 0.985. The effective k_d values estimated using GP data varied from 0.118 to 0.282/h for the WF and from 0.123 to 0.301/h for the NDS fraction, and were less (P<0.05) for maize compared to small grains (SG) but did not differ between barley and wheat (P>0.05). The effective k_d values for the aNDF fraction ranged from 0.039 to 0.082/h and did not differ (P>0.05) either between maize and SG or between barley and wheat. The predicted ruminal NDS digestibility determined using GP data closely matched the in vivo data describing starch digestion (R² = 0.81). The effective k_d values for the WF were strongly related (R² = 0.94) to those for the NDS fractions. The results indicate that when measured with the GP technique, the differences in the digestion characteristics of maize and small grains are less than those previously reported in studies using the in situ method. It is concluded that the predicted NDS digestibility determined using GP data corresponded well to the in vivo starch digestibility. Our results also suggest that the first order digestion rates of NDS (starch) in starch-rich feeds can be accurately determined by incubating WF samples in the GP system and using the GP kinetic data in a dynamic, mechanistic rumen model.     

Resistance of soil-bound prions to rumen digestion

Before prion uptake and infection can occur in the lower gastrointestinal system, ingested prions are subjected to anaerobic digestion in the rumen of cervids and bovids. The susceptibility of soil-bound prions to rumen digestion has not been evaluated previously. In this study, prions from infectious brain homogenates as well as prions bound to a range of soils and soil minerals were subjected to in vitro rumen digestion, and changes in PrP levels were measured via western blot. Binding to clay appeared to protect noninfectious hamster PrP(c) from complete digestion, while both unbound and soil-bound infectious PrP(Sc) proved highly resistant to rumen digestion. In addition, no change in intracerebral incubation period was observed following active rumen digestion of unbound hamster HY TME prions and HY TME prions bound to a silty clay loam soil. These results demonstrate that both unbound and soil-bound prions readily survive rumen digestion without a reduction in infectivity, further supporting the potential for soil-mediated transmission of chronic wasting disease (CWD) and scrapie in the environment.     

Rumen Fungi and Forage Fiber Degradation

The role of anaerobic rumen fungi in in vitro forage fiber degradation was determined in a two forage × two inoculum source × five treatment factorial design. Forages used as substrates for rumen microorganisms were Coastal bermuda grass and alfalfa; inoculum sources were rumen fluid samples from a steer fed Coastal bermuda grass hay or alfalfa hay; treatments were whole rumen fluid (WRF), WRF plus streptomycin (0.2 mg/ml of rumen fluid) and penicillin (1.25 mg/ml of fluid), WRF plus cycloheximide (0.5 mg/ml of fluid), WRF plus streptomycin, penicillin, and cycloheximide, and McDougall buffer. Populations of fungi as shown by sporangial development were greater on bermuda grass leaves than on alfalfa leaflets regardless of inoculum source. However, endogenous fungal populations were greater from the alfalfa hay inoculum. Cycloheximide inhibited the fungi, whereas streptomycin and penicillin, which inhibit bacterial populations, resulted in an increase in numbers of sporangia in the alfalfa inoculum, suggesting an interaction between bacteria and fungi. Bacteria (i.e., WRF plus cycloheximide) were equal to the total population in degrading dry matter, neutral-detergent fiber (NDF), acid-detergent fiber (ADF), and cellulose for both inocula and both forages. Degradation of dry matter, NDF, ADF, and cellulose by anaerobic fungi (i.e., WRF plus streptomycin and penicillin) was less than that due to the total population or bacteria alone. However, NDF, ADF, and cellulose digestion was 1.3, 2.4, and 7.9 percentage units higher, respectively, for bermuda grass substrate with the alfalfa versus bermuda grass inoculum, suggesting a slight benefit by rumen fungi. No substantial loss of lignin (72% H₂SO₄ method) occurred due to fungal degradation. The most active fiber-digesting population in the rumen was the bacteria, even when streptomycin and penicillin treatment resulted in an increase in rumen fungi over untreated WRF. The development of large numbers of sporangia on fiber may not indicate a substantial role as digesters of forage.     

Trace mineral source influences digestion,ruminal fermentation,and ruminal copper,zinc, and manganese distribution in steers fed a diet suitable for lactating dairy cows

High solubility of certain trace minerals (TM) in the rumen can alter nutrient digestibility and fermentation. The objectives of the present studies were to determine the effects of TM source on 1) nutrient digestibility and ruminal fermentation, 2) concentrations of soluble Cu, Zn, and Mn in the rumen following a pulse dose of TM, and 3) Cu, Zn, and Mn binding strength on ruminal digesta using dialysis against a chelating agent in steers fed a diet formulated to meet the requirements of a high producing dairy cow. Twelve Angus steers fitted with ruminal cannulae were adapted to a diet balanced with nutrient concentrations similar to a diet for a high producing lactating dairy cow for 21 d. Steers were then randomly assigned to dietary treatments consisting of 10 mg Cu, 40 mg Mn, and 60 mg Zn/kg DM from either sulfate (STM), hydroxychloride (HTM) or complexed trace minerals (CTM). The experimental design did not include a negative control (no supplemental Cu, Mn, or Zn) because the basal diet did not meet the National Research Council requirement for Cu and Zn. Copper, Mn, and Zn are also generally supplemented to lactating dairy cow diets at concentrations approximating those supplied in the present study. Following a 14-d adaptation period, total fecal output was collected for 5-d. Following the fecal collection period, rumen fluid was collected for Volatile fatty acid (VFA) parameters. On the following day, the same diet was provided for 14 d, without supplemental Cu, Zn, and Mn. This period served as a wash-out period. A pulse dose of 100, 400, and 600 mg of Cu, Zn, Mn, respectively, from either STM, HTM, or CTM, was administered via ruminal cannulae to the steers on day 15. Over a 24-h period ruminal samples were obtained every 2-h. Following centrifugation, the supernatant was analyzed for Cu, Mn, and Zn. Ruminal solid digesta samples from times 0, 12, and 24 h after bolus dosing were exposed to dialysis against Tris-EDTA. Digestibility of NDF and ADF were lesser in STM vs. HTM and vs. CTM supplemented steers. Steers receiving HTM and CTM had greater total VFA concentrations than STM, and molar proportions of individual VFA were not affected by treatment. Ruminal soluble Cu and Zn concentrations were greater post dosing in STM and CTM supplemented steers at 2, 4, and 6 h for Cu and 4, 6, 8, 10 and 12 h for Zn when compared to HTM supplemented steers. The release of Cu and Zn from ruminal solid digesta following dialysis against Tris-EDTA at 12 and 24 h postdosing was greater for steers receiving HTM compared to those receiving STM or CTM. Results indicate trace mineral source impacts: 1) how tightly bound Cu and Zn are to ruminal solid digesta; 2) fiber digestion; 3) and ruminal total VFA concentrations.