Evaluation of the effects of dietary particle fractions on fermentation profile and concentration of microbiota in the rumen of dairy cows fed grass silage-based diets

The study evaluated the effects of three different theoretical particle lengths (TPL) of grass silage on the distribution of particle fractions of the diet and the resulting effects on fermentation profile and concentrations of protozoa and mixed bacterial mass in the rumen of three lactating Holstein cows fed total mixed rations (45% grass silage, 5% grass hay and 50% concentrate) ad libitum. Decreasing TPL of grass silage (long, medium, short) reduced particles retained on the 19-mm sieve of the Penn State Particle Separator, while particle fractions from 8 mm to 19 mm and smaller than 8 mm were increased. Different TPL did not affect pH and the concentration of volatile fatty acids in the rumen. However, lowering the TPL from long to medium increased significantly the bicarbonate concentration, acetate proportion and protozoal number in the rumen, whereas the proportion of bacterial protein in ruminal digesta and its amino acid concentration were significantly increased by the short TPL. For the current feeding conditions, it can be concluded that increasing the fraction of particles between 8 and 19 mm and probably even the fraction below 8 mm by decreasing TPL of grass silage do not adversely affect rumen conditions and can be beneficial in terms of optimising concentration and activity of ruminal microbiota in high-yielding dairy cows.     

Ruminal large and small particle kinetics in dairy cows fed red clover and grass silages harvested at two stages of growth

Passage, comminution and digestion rates of large and small particles were estimated using a rumen evacuation technique and total faecal collection with five lactating dairy cows in a 5 × 5 Latin square experiment. Two grass and two red clover silages harvested at early and late primary growth stages and a 1:1 mixture of late harvest grass and early harvest red clover were the dietary treatments. Cows received 9.0 kg supplementary concentrate per day. Ruminal contents and faeces were divided into large (>1.25 mm) and small (1.25–0.038 mm) particles by wet sieving. Indigestible neutral detergent fibre (iNDF) was determined by 12 days ruminal in situ incubation followed by neutral detergent extraction. Plant species did not affect ruminal particle size distribution, whereas advancing forage maturity decreased the proportion of large particles for both grass and red clover silage diets. Ruminal pool size of iNDF was higher (P<0.001) with red clover compared to grass silage diets. Ruminal passage rates of iNDF and potentially digestible NDF (pdNDF) increased with decreasing particle size (P<0.01). Passage rate of iNDF for small particles was slower (P<0.01) when red clover compared to grass silage diets were fed. Particle comminution rate in the rumen was slower (P<0.001) with red clover compared to grass silage diets and it increased (P<0.01) with advancing forage maturity. The contribution of particle comminution to ruminal mean retention time of iNDF in the ruminal large particle pool was smaller (P<0.01) in red clover compared to grass silage diets and it increased (P<0.05) with the mixed silage compared to the separate silages. Passage rate of pdNDF for both large and small particles was not affected by dietary treatments. Digestion rate of pdNDF for large particles was faster (P<0.001) with red clover compared to grass silage diets. Differences in ruminal passage and digestion rates of the large and small particles, in addition to differences in the passage and digestion rates of red clover compared to grass silage diets, emphasize the need to consider particle size and forage type in metabolic models predicting feed intake and fibre digestibility in ruminants.     

Effect of zeolite A on rumen fermentation and phosphorus metabolism in dairy cows

The aim of the present study was to determine the effect of zeolite A on several physiological parameters and on mineral metabolism in the rumino-intestinal-tract of cows. Eight double fistulated (rumen and proximal duodenum) cows were fed maize silage, grass silage and concentrate. Zeolite A was added to the ration over a period of three weeks at 0, 10 and 20 g/kg dry matter (DM). The daily feed amounts were adjusted to the current performance and varied between 3.9 and 15.5 kg/d. Rumen fluid, duodenal chyme and faeces were sampled to characterise the nutrient digestibility. Blood samples were taken to analyse the concentration of inorganic phosphate. Zeolite A supplementation led to a significantly reduced ruminal DM digestibility and fermentation of organic matter. The molar proportion of acetate in the rumen increased, and propionate as well as valerate decreased significantly after zeolite A supplementation. The concentration of the total fatty acids and ruminal pH were not affected. No effect on faecal digestion of DM, organic matter nor on calcium and magnesium digestion was observed. Otherwise the phosphorus (P) concentration in rumen fluid correlated negatively with the mean zeolite A intake (r ² = 0.75; p = 0.0003). Further, the faecal excretion of P increased significantly for cows with the highest zeolite A dosage (36.9 g P/d) compared to the control group (29.9 g P/d). The lower digestibility of P resulted in a significantly decreased concentration of inorganic P in serum from a basal value of 2.05–1.16 mmol/l six days after starting zeolite A supplementation. The zeolite A treated cows showed a significantly higher Al concentration already in rumen fluid (14.31 and 13.84 mmol/l) compared to the control cows (6.33 mmol/l). The Al flow in the duodenum was also higher for zeolite A treated cows.     

Changes over time in the bacterial communities associated with fluid and food particles and the ruminal parameters in the bovine rumen before and after a dietary change

This work aimed to study the changes over time in the bacterial communities associated with the fluid and food particle fractions of the cow rumen following a change in diet. Four cannulated cows were fed a hay-based diet for 21?days and were then switched to a corn-silage-based diet for 33?days. The bacterial communities were regularly characterized by capillary electrophoresis?- single-strand conformation polymorphism (CE-SSCP) and qPCR, and the main ruminal parameters were determined. The dietary change led to slight reductions in the diversity index, bacterial concentration, pH, and NH(3)-N concentration, and to an increase in the redox potential and volatile fatty acid concentrations. CE-SSCP profiles were not significantly affected by the dietary change but did change over time, with frequent fluctuations in both fluid and food particle fractions before and after the dietary change. The food particle fraction had a higher diversity index of bacterial community (+1.2 points, P?< 0.001) and slightly more total bacteria than the fluid fraction of the rumen.     

Studies on the biotin flow at the duodenum of dairy cows fed diets with different concentrate levels and types of forages

Abstract

Biotin is involved in many vital metabolic pathways and must be provided for an efficient fermentation in the rumen, as well as for the intermediary metabolism of the host animal. Factors influencing ruminal biotin metabolism and output are widely unknown at present. Therefore, dairy cows fitted with permanent cannulas in the dorsal rumen and in the proximal duodenum were fed differently composed diets, and the biotin flow at the proximal duodenum was measured. The diets (on DM basis) consisted of 8.9 kg grass hay (Diet 1), 8.9 kg corn silage plus 2.0 kg concentrate (Diet 2), or 7.3 and 7.4 kg grass silage plus 10.0 kg concentrate (Diets 3 and 4). The concentrate in Diets 3 and 4 contained 87% wheat and corn grain, respectively. The cows were pre-fed the rations for 21 days. Thereafter duodenal digesta was sampled every two h for 5 days. Cr₂O₃ served as a flow marker and the microbial proportion of total nitrogen at the duodenum was estimated by near infrared spectroscopy (NIRS). The duodenal flow of biotin was not related to biotin intake, but to the amount of fermented organic matter (FOM) and the amount of microbial protein (Biotin [mg/d] = 0.518 · kg FOM?0.300; r = 0.85 and biotin [mg/d] = 0.012 · g microbial protein + 1.478; r = 0.84), irrespective of the composition of the diet fed. Mean daily biotin flow was 0.48 ± 0.11 mg/kg FOM without any systematic effect of diet composition. The ruminal biotin balance, calculated as the difference between biotin flow at the duodenum and biotin intake, was positive (1.4 – 2.0 mg/d) in cows fed the mixed roughage/concentrate diets and negative (?0.71 mg/d) when the pure hay diet was fed.     

Effect of maize silage to grass silage ratio and feed particle size on ruminal fermentation in vitro

The effect of the forage source on ruminal fermentation in vitro was investigated for fine (F) and coarse (C) milled diets, using a modified Hohenheim gas production test and a semi-continuous rumen simulation technique (Rusitec). It was hypothesised that the replacement of maize silage by grass silage might lead to associative effects and that interactions related to particle size variation could occur. Five diets with a maize silage to grass silage ratio of 100 : 0, 79 : 21, 52 : 48, 24 : 76 and 0 : 100 differed in their content of CP and carbohydrate fractions, as well as digestible crude nutrients, derived from a digestibility trial with wether sheep. For in vitro investigations, the diets were ground to pass a sieve of either 1 mm (F) or 4 mm (C) perforation. Cumulative gas production was recorded during 93 h of incubation and its capacity decreased with increasing proportion of grass silage in the diet. Across all diets, gas production was delayed in C treatments compared with F treatments. Degradation of crude nutrients and detergent fibre fractions was determined in a Rusitec system. Daily amounts of NH3-N and short-chain fatty acids (SCFA) were measured in the effluent. Degradation of organic matter (OM) and fibre fractions, as well as amounts of NH3-N, increased with stepwise replacement of maize silage by grass silage. Degradability of CP was unaffected by diet composition, as well as total SCFA production. In contrast to the results of the gas production test, degradation of OM and CP was higher in C than in F treatments, accompanied by higher amounts of NH3-N and SCFA. Interactions of silage ratio and particle size were rare. It was concluded that the stepwise replacement of maize silage by grass silage might lead to a linear response of most fermentation characteristics in vitro. This linear effect was also supported by total tract digestibility data. However, further investigations with silages of variable quality seem to be necessary.     

Effects of dietary forage particle size and concentrate level on fermentation profile, in vitro degradation characteristics and concentration of liquid- or solid-associated bacterial mass in the rumen of dairy cows

This study investigated effects of dietary forage particle size (PS) and concentrate level (CL) on fermentation profiles of particle-associated rumen liquid (PARL) and free rumen liquid (FRL), in vitro degradation characteristics and concentration of bacterial mass attached to the solid or fluid rumen digesta phase in dairy cows. The experiment was a 4 × 4 Latin square design with four late-lactation dairy cows in four 23 day periods. Cows were restrictively fed (17 kg dry matter (DM)/d) one of four diets varying in the theoretical PS (6 and 30 mm) of grass hay and in the levels (approximately 200 and 550 g/kg, DM basis) of a cereal-based concentrate. Proportion of large particles (>6 mm) and the content of structural fibre in the diet increased by reducing dietary CL and, particularly, by increasing hay PS. This effect was not reflected by changes in mean total volatile fatty acid concentration or pH in the rumen. However, cows fed high concentrate diets had pH of 5.28 and 5.37 in PARL at 3 h after the last meal, when fine or long chopped hay was offered. The low pH may indicate a depression of the capacity of PARL to degrade fibre in vitro. Gas production in vitro of concentrate increased with the high concentrate diet at 12 h, suggesting that amylolytic capacity was affected only in early phases of fermentation. In addition, elevating dietary CL appeared to shift ruminal fermentation outputs from propionate to butyrate and valerate. Inclusion of coarsely chopped hay to a high concentrate diet does not appear to bring advantages due to increased structure in restrictively fed dairy cows. In addition, results suggest that the response of pH in PARL is more sensitive to dietary changes (i.e., forage PS and CL) than the response in FRL, and so PARL might be better to evaluate the risk of ruminal disfunction in dairy cows.     

Influence of the maize silage to grass silage ratio and feed particle size of rations for ruminants on the community structure of ruminal Firmicutes in vitro

Aims: To investigate the effect of the forage source and feed particle size (FPS) in ruminant rations on the composition of the ruminal Firmicutes community in vitro. Methods and Results: Three diets, varying in maize silage to grass silage ratio and FPS, were incubated in a rumen simulation system. Microbial samples were taken from the liquid fermenter effluents. Microbial community analysis was performed by 16S rRNA‐based techniques. Clostridia‐specific single‐strand conformation polymorphism profiles revealed changes of the community structure in dependence on both factors tested. The coarse grass silage–containing diets seemed to enhance the occurrence of different Roseburia species. As detected by real‐time quantitative PCR, Ruminococcus albus showed a higher abundance with decreasing FPS. A slightly lower proportion of Bacilli was found with increasing grass silage to maize silage ratio by fluorescence in situ hybridization (FISH). In contrast, a slightly higher proportion of bacterial species belonging to the Clostridium‐clusters XIV a and b was detected by FISH with increasing grass silage contents in the diet. Conclusions: The ruminal Firmicutes community is affected by the choice of the forage source and FPS. Significance and Impact of the Study: This study supplies fundamental knowledge about the response of ruminal microbial communities to changing diets. Moreover, the data suggest a standardization of grinding of feeds for in vitro studies to facilitate the comparison of results of different laboratories.     

Effect of harvest time and physical form of alfalfa silage on chewing time and particle size distribution in boli,rumen content and faeces

The study examined the effects of physical form and harvest time of alfalfa silage on eating and ruminating activity and particle size distribution in feed boli, rumen content and faeces in dry cows. The alfalfa crop was harvested at two stages of growth (early: NDF 37%, late: NDF 44% in dry matter (DM)), and from each harvest, a chopped (theoretical cutting length: 19 mm) and an unchopped crop was ensiled in bales. The silages were fed restrictively to four rumen cannulated non-lactating Jersey cows (391 ± 26 kg) in a 4 × 4 Latin square design. The cows were fed restrictively 80% of their ad libitum intake twice daily. Chewing activity was recorded for 96 h continuously. Swallowed boli, rumen content, rumen fluid and faeces samples were collected, washed in nylon bags (0.01 mm pore size) and freeze-dried before dry sieving through 4.750, 2.360, 1.000, 0.500 and 0.212 mm pore sizes into six fractions. The length (PL) and width (PW) of particles within each fraction was measured by the use of image analysis. The eating activity (min/kg dry matter intake (P < 0.01) and min/kg NDF (P < 0.05)) was affected by harvest time. The mean ruminating time (min/kg DM) was affected by harvest time (P < 0.01), physical form (P < 0.05) and NDF intake per kg BW (P < 0.01). The proportion of washed particle DM of total DM in boli, rumen content, rumen fluid and faeces was affected by harvest time (P < 0.01) and highest by feeding late-harvested alfalfa silage. Two peaks on the probability density distribution function (PDF) of PW and PL values of boli, rumen content and faeces were identified. Chopping of the silage decreased the mean PL and PW, the most frequent PL (mode) and 95% percentile PL and PW values in boli. In the rumen content, chopping increased the mean PW (P < 0.05). The dimension sizes of faeces particles were not significantly affected by chopping. The mode PW value was lower in rumen content and faeces than in boli (P < 0.001), and the mode PL value was higher in boli and lower in faeces compared with rumen contents (P < 0.001). In conclusion, the mean total chewing activity per kg NDF decreased due to chopping and early harvest time. The mean PL and PW in boli decreased due to chopping and late harvest. The two peak values on the PDF (PL) and PDF (PW) of boli, rumen content and faeces particles are most likely related to the leaf and the stem residues.     

Degradability characteristics of dry matter and crude protein of forages in ruminants

Twelve corn silages, 22 grass silages and 14 grass hays, obtained from various farms located in the lower Fraser Valley region of British Columbia, and 16 alfalfa hays, grown primarily in the Columbia basin of central Washington State, were evaluated using both the rumen and the mobile nylon bag in situ techniques. Nylon bags containing each forage were incubated in duplicate for 0, 2, 4, 8, 12, 24, 48, 72, or 96 h in two of six non-lactating Holstein cows fitted with rumen and duodenal cannulae. All forage types were evaluated in terms of the following dry matter (DM) and crude protein (CP) digestion characteristics: soluble fraction A, degradable fraction B, degradation rate, lag phase, and effective degradability. The mobile nylon bag technique was used to determine intestinal disappearance of DM and CP from the forages following pre-incubation in the rumen for 12 h. Significant (P < 0.05) differences in degradation characteristics occurred within all forages with regard to the soluble and potentially degradable DM and CP fractions. Soluble CP content in the rumen varied from 44.08 to 75.37% and from 18.74 to 65.38% in the corn and grass silages, respectively, and from 48.27 to 75.43% and from 30.13 to 65.95% in the alfalfa and grass hays, respectively. Significant differences within each forage type were also observed for the degradable CP in fraction B: 10.89 to 45.28% for corn silage, 20.72 to 82.77% for grass silage, 16.67 to 44.88% for grass hay and 25.44 to 62.93% for alfalfa hays. Significant differences (P > 0.05) were observed in fractional rates of ruminal DM degradation of the grass hays and corn silages. Significant differences did exist in the fractional rates of ruminal CP degradation within all forage types with the exception of alfalfa hays. Effective degradabilities of DM and CP were also significantly different between samples of a particular forage type. The mobile nylon bag data indicated that approximately 20% of the original CP in the grass silage, grass hay and alfalfa hay samples disappeared in the intestine and that there was significant variation between individual samples. On average, in the corn silage samples more than 10% of the original nitrogenous material disappeared in the intestine. The results presented in this study clearly demonstrate that the use of tabulated values for describing individual batches of forages in terms of their degradability characteristics is inaccurate since they may not reflect the particular forage being used in the ration and thus may lead to errors in diet formulation.     

Effects of expander-treating a barley-based concentrate on ruminal fermentation, bacterial N synthesis, escape of dietary N, and performance of dairy cows

Three primiparous dairy cows in early lactation with cannulas in rumen, duodenum and ileum were used in a 3×3 Latin square design to study effects of expander treatment of a barley-based concentrate. The concentrate was either pelleted at 75–80°C or expander treated at 125–130°C prior to pelleting. The diets consisted of 6.7 kg DM of grass silage and 10 kg DM of (1) 100% pelleted, (2) 50% pelleted and 50% expanded or (3) 100% expanded concentrate. The diets were offered as a mixed ration in four equal meals daily. Ruminal fermentation, bacterial N synthesis, duodenal, ileal and faecal flow of nutrients, and animal performance were monitored. Expander treatment numerically increased ruminal digestion of starch, which explained the observed increase in ruminal VFA concentration and the lowered ruminal pH (P<0.05). The proportion of butyrate in rumen liquid increased, whereas the proportion of propionate decreased in the expanded compared to the pelleted treatment (P<0.05). Expander treatment tended to increase rumen volume and rumen NDF pool size. Ruminal digestion of NDF was numerically lower in the expanded than in the pelleted treatment. No differences in bacterial N synthesis or efficiency of synthesis were observed among treatments. Expander treatment numerically increased the duodenal flow of non-ammonia N (NAN) and amino acid N (AAN), and seemed to increase the flow of non-ammonia non-bacterial N (NANBN) to the duodenum to a similar extent as was indicated by nylon bag studies. Milk production and milk fat and protein content were increased by the expander treatment (P<0.05), indicating that expander treatment increased the supply of nutrients for milk production.     

Impact of subacute ruminal acidosis on the diversity of liquid and solid-associated bacteria in the rumen of goats

This study was aimed to investigate the impact of subacute ruminal acidosis (SARA) on the diversity of liquid (LAB) and solid-associated bacteria (SAB) following high-grain feeding. Six ruminally cannulated goats were divided into two groups: one group was fed a hay diet (COD), and the other group was fed a high grain diet (SAID). Rumen liquids and rumen solids were sampled after 2 weeks adaption. SARA was diagnosed with a pH below 5.8 for 8 h. SAID decreased ruminal pH (P < 0.001) and increased the acetate (P = 0.017), propionate (P = 0.001), butyrate (P < 0.001) and total volatile fatty acid (P < 0.001) concentration in rumen compared with the COD. Denaturing gradient gel electrophoresis fingerprints analysis revealed a clear separation between both the diet and the fraction of rumen digesta in bacterial communities. Pyrosequencing analysis showed that the proportion of phylum Bacteroidetes in the SAID-LAB and SAID-SAB communities was less than in the COD group, whereas the SAID group had a greater percentage of Firmicutes in both the LAB and SAB libraries. UniFrac analyses and a Venn diagram revealed a large difference between the two diets in the diversity of rumen bacterial communities. Overall, our findings revealed that SARA feeding did alter the community structure of rumen liquids and rumen solids. Thus, manipulation of dietary factors, such as ratio of forage to concentrate may have the potential to alter the microbial composition of rumen liquid and rumen solid.     

Urea and short-chain fatty acids metabolism in Holstein cows fed a low-nitrogen grass-based diet

Three ruminally cannulated and multicatheterised lactating dairy cows were used to investigate the effect of different supplement strategies to fresh clover grass on urea and short-chain fatty acid (SCFA) metabolism in a zero-grazing experiment with 24-h blood and ruminal samplings. Fresh clover grass was cut every morning and offered from 0800 to 1500 h. Maize silage was fed at 1530 h. The three treatments, arranged in a Latin square, differed by timing of feeding rolled barley and soya-bean hulls relative to fresh clover grass. All diets had the same overall composition. Treatments were soya-bean hulls fed at 0700 h and barley fed at 1530 h (SAM), barley fed at 0700 h and soya-bean hulls fed at 1530 h (BAM), and both soya-bean hulls and barley fed at 1530 h (SBPM). The grass had an unexpectedly low content of crude protein (12.7%) and the cows were severely undersupplied with rumen degradable protein. The treatment effects were numerically small; greater arterial ammonia concentration, net portal flux of ammonia and net hepatic flux of urea during part of the day were observed when no supplementary carbohydrate was fed before grass feeding. A marked diurnal variation in ruminal fermentation was observed and grass feeding increased ruminal concentrations of propionate and butyrate. The net portal fluxes of propionate, butyrate, isovalerate and valerate as well as the net hepatic uptake of propionate, butyrate, valerate and caproate increased after feeding at 0700 h. The hepatic extraction of butyrate showed a relatively large depression with grass feeding with nadir at 1200 to 1330 h. The increased net portal absorption and the decreased hepatic extraction resulted in an approximately six-fold increase in the arterial blood concentration of butyrate. The gut entry rate of urea accounted for 70 ± 10% of the net hepatic production of urea. Saliva contributed to 14% of the total amount of urea recycled to the gut. Urea recycling to the gut was equivalent to 58% of the dietary nitrogen intake. Despite the severe undersupply of rumen degradable protein, the portal-drained viscera did not extract more than 4.3% of the urea supplied with arterial blood. This value is in line with the literature values for cows fed diets only moderately deficient in rumen degradable protein and indicates that cows maximise urea transfer across gut epithelia even when the diet is moderately deficient in rumen degradable protein.     

Effect of harvest time of red and white clover silage on chewing activity and particle size distribution in boli,rumen content and faeces in cows

The study examined the effects of harvest time of red and white clover silage on eating and ruminating activity and particle size distribution in feed boli, rumen content and faeces in cows. The clover crops were harvested at two stages of growth and ensiled in bales. Red clover crops had 36% and 45% NDF in dry matter (DM) at early (ER) and late (LR) harvest, respectively, and the white clover crops had 19% and 29% NDF in DM at the early (EW) and late (LW) harvest, respectively. The silages were fed restrictively (80% of ad libitum intake) twice daily to four rumen cannulated non-lactating Jersey cows (588 ± 52 kg) in a 4 × 4 Latin square design. Jaw movements (JM) were recorded for 96 h continuously. Swallowed boli, rumen mat, rumen fluid and faeces samples were collected, washed in nylon bags (0.01 mm pore size) and freeze-dried before dry sieving through 4.750, 2.360, 1.000, 0.500, 0.212 and 0.106 mm into seven fractions. The length (PL) and width (PW) values of rumen and faeces particles within each fraction were measured by use of image analysis. The eating activity (min/kg DM intake; P < 0.05) was higher in LR compared with the other treatments. The eating activity (min/kg NDF intake; P < 0.05) was affected by clover type with highest values for white clover silage. The mean ruminating time (min/kg DM), daily ruminating cycles (P < 0.001) and JM during ruminating (P < 0.05) were affected by treatment with increasing values at later harvest time. The proportion of washed particle DM of total DM in boli (P < 0.001), rumen mat (P < 0.001), rumen fluid (P < 0.01) and faeces was (P < 0.001) highest by feeding LR. There were identified two peaks (modes 1 and 2) on the probability density distribution (PDF) of PW values of rumen mat and faeces, but only one peak (mode 1) for PL values. There was no difference in the mean and mode 1 PW and PL value in rumen mat between the four treatments. The mean PL, mode PL, mode 2 PW and mean PW in faeces were highest for LR (P < 0.05). The mean particle size in boli measured by sieving was higher at white clover compared with red clover treatments (P < 0.001) and the highest value in faeces was found in LR (P < 0.01). The two peaks on PDF for width values of rumen mat and faeces particles are most likely related to the leaves and the stems/petioles. In conclusion, the mean total chewing activity per kg DM was lowest for the white clover silage and increased for both silages due to later harvest time. The mean particle size in boli was smallest for LR, whereas the mean PL and PW in faeces were highest for the LR.     

Oral Samples as Non-Invasive Proxies for Assessing the Composition of the Rumen Microbial Community

Microbial community analysis was carried out on ruminal digesta obtained directly via rumen fistula and buccal fluid, regurgitated digesta (bolus) and faeces of dairy cattle to assess if non-invasive samples could be used as proxies for ruminal digesta. Samples were collected from five cows receiving grass silage based diets containing no additional lipid or four different lipid supplements in a 5 x 5 Latin square design. Extracted DNA was analysed by qPCR and by sequencing 16S and 18S rRNA genes or the fungal ITS1 amplicons. Faeces contained few protozoa, and bacterial, fungal and archaeal communities were substantially different to ruminal digesta. Buccal and bolus samples gave much more similar profiles to ruminal digesta, although fewer archaea were detected in buccal and bolus samples. Bolus samples overall were most similar to ruminal samples. The differences between both buccal and bolus samples and ruminal digesta were consistent across all treatments. It can be concluded that either proxy sample type could be used as a predictor of the rumen microbial community, thereby enabling more convenient large-scale animal sampling for phenotyping and possible use in future animal breeding programs aimed at selecting cattle with a lower environmental footprint.     

Effect of the corn silage to grass silage ratio and feed particle size of diets for ruminants on the ruminal Bacteroides-Prevotella community in vitro

This study examined whether different corn silage to grass silage ratios in ruminant rations and different grinding levels of the feed affect the composition of the ruminal Bacteroides-Prevotella community in vitro. Three diets, composed of 10% soybean meal as well as of different corn silage and grass silage proportions, were ground through 1 mm or 4 mm screened sieves and incubated in a semi-continuous rumen simulation system. On day 14 of the incubation microbes were harvested by centrifugation from the liquid effluent of fermenter vessels. Microbial DNA was extracted for single strand conformation polymorphism (SSCP) analysis of 16S rRNA genes followed by sequencing of single SSCP bands. Fluorescence in situ hybridization (FISH) and real-time quantitative (q) PCR were used to quantify differences in the relative abundance of Bacteroides-Prevotella and Prevotella bryantii. SSCP profiles revealed a significant influence of the forage source as well as of the feed particle size on the community structure of the Bacteroides-Prevotella group. Different, phylogenetically distinct, so far uncultured Prevotella species were detected by sequence analysis of several treatment-dependent occurring SSCP bands indicating different nutritional requirements of these organisms for growth. No quantitative differences in the occurrence of Bacteroides-Prevotella-related species were detected between diets by FISH with probe BAC303. However, real-time qPCR data revealed a higher abundance of P. bryantii with increasing grass silage to corn silage ratio, thus again indicating changes within the community composition of the Bacteroides-Prevotella group. As P. bryantii possesses high proteolytic activity its higher abundance may have been caused by the higher contents of crude protein in the grass silage containing diets. To conclude, results of this study show an influence of the forage source on the ruminal community of Bacteroides-Prevotella. Furthermore, they suggest an effect of the feed particle size on this bacterial group.     

Magnesium absorption as influenced by the rumen passage kinetics in lactating dairy cows fed modified levels of fibre and protein

The potassium sensitive magnesium absorption through the rumen wall may be influenced by additional dietary properties, such as diet type, forage type or forage to concentrate ratio. These properties are likely associated to rumen passage kinetics modified by dietary fibre content. The study aimed to assess the effects of rumen passage kinetics on apparent Mg absorption and retention in lactating dairy cows fed modified levels of fibre. Six lactating Red-Holstein and Holstein cows, including four fitted with ruminal cannulas were randomly assigned to a 3 × 3 cross-over design. The experimental diets consisted of early harvested low NDF (341 g NDF/kg DM) and late harvested high NDF (572 g NDF/kg DM) grass silage (80% DM) and of concentrates (20% of DM). As the low-fibre diet was excessive in protein, a third high-fibre diet was formulated to be balanced in digestible protein with the low-fibre diet to avoid any eventual confounding effects of NDF and protein excess. All diets were formulated to contain iso-Ca, -P, -Mg, -K and -Na. Passage kinetics of solid and liquid phase of rumen digesta were evaluated using ruminal marker disappearance profiles. Cows fed the low-fibre diet had compared to the other diets, an up to 40% lower solid and 26% lower liquid phase volume of rumen digesta and a 10% numerically higher fractional rumen liquid passage rate. Rumen pH lost 0.6 units and Mg concentration in the rumen liquid phase tripled when cows were fed the low-fibre diet. Faecal Mg excretion was up to 14% higher in cows fed the low-fibre diet and Mg absorbability was 12% compared to up to 19% in other diets. Urinary Mg excretion in cows fed the low-fibre diet was half of the ones in the other treatments, but Mg retention was not affected. Dietary protein excess neither affected rumen passage kinetics nor Mg absorption and retention. Absorption of Mg was correlated with rumen liquid volume which both decreased with decreasing daily NDF intake (NDFi, 11.8 ± 2.4 l/kg NDFi). Consequently, daily Mg absorption decreased by 1.32 ± 0.28 g/kg decreasing NDFi. To conclude, in addition to the known antagonistic effect of dietary K, the present data indicate that Mg absorption was dependent from NDFi which modified rumen liquid volume, but was independent of dietary protein excess likely associated to low NDF herbages.     

Effects of acute and chronic heat stress on feed sorting behaviour of lactating dairy cows

Nutritional strategies to mitigate the negative effects of heat stress on animal welfare and productivity often involve changes in ration formulation. However, cattle commonly sort their ration in favour of certain components, and it is not clear how feed sorting responds to heat stress. This study investigated the association between heat stress and feed sorting behaviour. Lactating Holstein dairy cows (n = 32; parity = 2.8±1.2; mean±SD) were housed in a free stall barn and milked 3×/day. Cows were fed individually using the Calan Broadbent Feeding System and offered ad libitum access to a total mixed ration (containing on a dry matter basis: 3.3% ryegrass hay, 16.5% ryegrass baleage, 24.7% corn silage, 11.1% brewers grains, 19.7% ground corn, 19.8% concentrate and 4.9% protein/mineral supplement), provided 1×/day. Beginning at 186±60 days in milk, cows were exposed to either: heat stress conditions (HT; n = 15) (average temperature–humidity index: 77.6), or evaporative cooling (CL; n = 17), consisting of misters and fans over the freestall and feed bunks. Data were collected during a 4-day baseline period, and two 4-day experimental periods: starting at 10 days after implementing treatments (defined as acute heat stress for HT cows), and at 62 days after implementing treatments (defined as chronic heat stress for HT cows). Daily feed intake and physiological responses to heat stress (body temperature, respiration rate) were recorded. Samples of fresh and refused feed were collected daily from individual cows for particle size analysis. The particle size separator had three screens (19, 8 and 1.18 mm) and a bottom pan, resulting in 4 fractions (long, medium, short and fine particles). Feed sorting was calculated as the actual intake of each particle size fraction expressed as a percentage of the predicted intake of that fraction. During both heat stress periods, HT cows sorted for long particles more than CL cows (105.0% v. 100.6%; SE = 1.1). During acute heat stress, HT cows sorted to a greater extent than CL cows against medium and short particles, whereas sorting of these fractions did not differ during chronic heat stress. Body temperature and respiration rate were associated across treatments with the extent of sorting for long particles and against short particles during acute heat stress. These results suggest that feed sorting is particularly influenced during acute heat stress, and that sorting for longer particles may increase in heat stress.     

Restoration of in situ fiber degradation and the role of fibrolytic microbes and ruminal pH in cows fed grain-rich diets transiently or continuously

In this study, we used two different grain-rich feeding models (continuous or transient) to determine their effects on in situ fiber degradation and abundances of important rumen fibrolytic microbes in the rumen. The role of the magnitude of ruminal pH drop during grain feeding in the fiber degradation was also determined. The study was performed in eight rumen-fistulated dry cows. They were fed forage-only diet (baseline), and then challenged with a 60% concentrate diet for 4 weeks, either continuously (n=4 cows) or transiently (n=4 cows). The cows of transient feeding had 1 week off concentrate in between. Ruminal degradation of grass silage and fiber-rich hay was determined by the in situ technique, and microbial abundances attached to incubated samples were analyzed by quantitative PCR. The in situ trials were performed at the baseline and in the 1^st and the last week of concentrate feeding in the continuous model. The in situ trials were done in cows of the transient model at the baseline and in the 1^st week of the re-challenge with concentrate. In situ degradation of NDF and ADF of the forage samples, and microbial abundances were determined at 0, 4, 8, 24 and 48 h of the incubation. Ruminal pH and temperature during the incubation were recorded using indwelling pH sensors. Compared with the respective baseline, both grain-rich feeding models lowered ruminal pH and increased the duration of pH below 5.5 and 5.8. Results of the grass silage incubation showed that in the continuous model the extent of NDF and ADF degradation was lower in the 1^st, but not in the last week compared with the baseline. For the transient model, degradation of NDF of the silage was lower during the re-challenge compared with the baseline. Degradation of NDF and ADF of the hay was suppressed by both feeding models compared with the respective baseline. Changes in fiber degradation of either grass silage or hay were not related to the magnitude of ruminal pH depression during grain-rich feeding. In both feeding models total fungal numbers and relative abundance of Butyrivibrio fibrisolvens attached to the incubated forages were decreased by the challenge. Overall, Fibrobacter succinogenes was more sensitive to the grain challenge compared with Ruminococcus albus and Ruminococcus flavefaciens. The study provided evidence for a restored ruminal fiber degradation after prolonged time of grain-rich feeding, however depending on physical and chemical characteristics of forages.     

Effect of stage of maturity of grass at harvest on intake,chewing activity and distribution of particle size in faeces from pregnant ewes

This study was conducted to investigate the effect of stage of maturity at harvest on the intake of grass silage, eating and ruminating activity and the distribution of faecal particle size in ewes during late pregnancy. A total of 18 Swedish Finull × Dorset 85 ± 8 kg (mean ± s.d.) ewes bearing twins were randomly assigned to three dietary treatments 6 weeks before lambing. The treatments included ad libitum feeding with early harvested (EH), medium harvested (MH) or late harvested (LH) primary-growth grass silage with 45%, 58% and 63% NDF on a dry matter (DM) basis, respectively. Intake and chewing activity were recorded and faeces were sampled over 4 continuous days for each individual ewe. The faeces samples were washed in nylon bags, freeze dried and sieved with pore sizes from 2.4 mm to 0.1 mm; particles less than 0.1 mm in size were also collected. Subsamples of each sieving fraction were scanned and the dimensions of the individual particles in each sieving fraction were measured by image analysis. In addition, the number of particles longer than 7 mm was counted from the particles retained on a sieve with a pore size of 2.4 mm using a simple wet sieving technique. The time spent eating and ruminating per kg of DM intake was affected by the stage of maturity at harvest; it was shorter in ewes fed EH compared with ewes fed MH and LH (P < 0.05). In comparison with feeding LH, feeding EH resulted in the retention of a larger proportion of particles in the lower and upper sieve fractions (<0.2 mm and >1 mm, respectively, P < 0.01), a smaller mean particle size (P < 0.05) and a smaller mean particle width in faeces (P < 0.01). The results from the simple wet sieving technique confirmed the results from dry sieving and image analysis, showing a higher number of large particles in faeces from ewes fed the EH compared with the ewes fed the MH and LH (P < 0.001). In conclusion, the distribution of faecal particle size might be considered as a footprint of the characteristics of forage fibre eaten by ewes.