Abstract: The effects of an exogenous enzyme preparation, the application method and feed type on ruminal fermentation and microbial protein synthesis were investigated using the rumen simulation technique (Rusitec). Steam-rolled barley grain and chopped alfalfa hay were sprayed with water (control, C), an enzyme preparation with a predominant xylanase activity (EF), or autoclaved enzyme (AEF) 24 h prior to feeding, or the enzyme was supplied in the buffer infused into the Rusitec (EI). Microbial N incorporation was measured using (15NH4)2SO4 in the buffer. Spent feed bags were pummelled mechanically in buffer to segregate the feed particle-associated (FPA) and feed particle-bound (FPB) bacterial fractions. Enzymes applied to feed reduced neutral-detergent fibre content, and increased the concentration of reducing sugars in barley grain, but not alfalfa hay. Ruminal cellulolytic bacteria were more numerous with EF than with C. Disappearance of DM from barley grain was higher with EF than with C, but alfalfa was unaffected by EF. Treatment EF increased incorporation of 15N into FPA and FPB fractions at 24 and 48 h. In contrast, AEF reduced the 24 h values, relative to C; AEF and C were similar at 48 h. Infused enzyme (EI) did not affect 15N incorporation. Xylanase activity in effluent was increased by EF and EI, compared to C, but not by AEF. Xylanase activity in FPA was higher at 48 h than at 24 h with all treatments; it was higher with EF than C at 24 and 48 h, but was not altered by AEF or EI. Applying enzymes onto feeds before feeding was more effective than dosing directly into the artificial rumen for increasing ruminal fibrolytic activity.

Abstract: C.S. MCSWEENEY, B. PALMER, R. BUNCH AND D.O. KRAUSE. 2001. Aims: To determine the effect of condensed tannins in Calliandra calothyrsus (calliandra) on rumen microbial function. Methods and Results: Microbial populations, ruminal protein synthesis and fermentation end-products were measured in sheep fed roughage hay supplemented with calliandra (30%), with and without inclusions of polyethylene glycol (PEG) to counteract the effect of tannin. Molecular and conventional enumeration techniques were used to quantify rumen bacteria, fungi and protozoa, and protein synthesis was predicted from estimates of urinary purine excretion. The total number of cellulolytic bacteria, including populations of Fibrobacter succinogenes and Ruminococcus spp., was significantly lower in sheep supplemented with calliandra and these populations increased when animals were treated with PEG. By contrast, protozoa and fungi and the microbial group containing Bacteroides-Porphyromonas-Prevotella bacteria appeared to be less affected. The efficiency of microbial protein synthesis in the rumen was not altered significantly. Conclusions: Calliandra caused significant shifts in rumen microbial populations without changing the efficiency of protein synthesis. Significance and Impact of the Study: The effect of calliandra tannins on rumen digestion may result more from complexing with nutrients than direct inhibition of micro-organisms.

Abstract: Both the anti-nutritional and beneficial effects of secondary compounds in a range of temperate and tropical forages have been reviewed. Major secondary compounds in temperate and tropical forage plants occur in the phenolic fraction and include condensed and hydrolysable tannins, phenolic monomers and lignin. Condensed tannins (CT) bind to plant protein by pH-reversible hydrogen bonding. In temperate legume forages this reduces rumen protein degradation and can increase the absorption of essential amino acids (EAA) from the small intestine, with reactivity depending on CT concentration, molecular weight and chemical structure. Low concentrations of CT in Lotus corniculatus (20-40g/kg DM) increased EAA absorption by 62% and increased wool growth (15%) and ovulation rate (25%) in grazing sheep and increased milk production in ewes and dairy cows, all without changing voluntary feed intake (VFI). High concentrations of CT in Lotus pedunculatus (80-100 g/kg DM) depressed VFI and depressed rates of body and wool growth in grazing sheep. Sulla, containing 80-120 g CT/kg DM, was particularly effective for counteracting the effects of parasitism and for promoting high rates of body growth in parasitised lambs. CT is present in tropical species such as Leucaena and Acacia at higher concentrations (60-200 g/kg DM) than in temperate species. Action of CT reduced rumen protein degradation in sheep fed tropical forages, but as yet there is no convincing evidence that this leads to increases in EAA absorption from the small intestine or that CT increases animal production. Further research is needed in these areas with tropical forages, particularly on the relationship between CT structure and its reactivity with proteins. Increasing CT concentration did not depress rumen microbial protein synthesis in sheep fed either temperate or tropical forages, until CT concentration exceeded 130 g/kg DM. Effect of CT upon undegraded, dietary protein release in the small intestine and upon endogenous protein secretion is defined as a future research area.

Abstract: Forage proteins are degraded rapidly by rumen microorganisms and therefore supply relatively small quantities of undegraded intake protein (UIP). Growing cattle with high metabolizable protein requirements and lactating beef and dairy cows respond to UIP supplementation when fed high-forage diets, even though degradable intake protein (DIP) is adequate. This observation suggests that an accurate estimate of forage UIP is needed to establish optimal supplementation conditions. Microbial protein must be quantitated in duodenal or in situ residue samples to accurately measure forage UIP. Purines commonly are used as a microbial protein marker. Recent reports suggested that the original purine procedure generates interfering compounds that reduce estimates of microbial protein. Reanalysis of samples with a modified purine procedure yielded three to four times more purines in both duodenal samples and NDF residue incubated in situ. An alternative in situ procedure removes the microorganisms by refluxing with neutral detergent after ruminal incubation. This alternative correlates highly to the purine-corrected in situ procedure, and it is less

Abstract: Effects of dietary factors including kernel thickness of processed barley grain, ratio of forage to concentrate, and forage particle length on chemical composition of bacteria, bacterial colonization of feed particles and distribution in the rumen, and duodenal flow of bacteria in dairy cows were evaluated. The experiment was designed as a double 4 x 4 quasi-Latin square with a 2 x 2 x 2 factorial arrangement of treatments using eight lactating cows with ruminal and duodenal cannulas. Barley grain was steam-rolled to two thicknesses: coarse (1.60 mm) or flat (1.36 mm); ratio of forage:concentrate was low (35:65) or high (55:45) (DM basis); and forage particle length was long (7.59 mm) or short (6.08 mm). Cows were offered ad libitum access to a total mixed diet. Chemical composition was different (P < 0.01) between liquid-associated bacteria (LAB) and solid-associated bacteria (SAB). Reduced barley thickness increased (P < 0.05) N content and 15N enrichment, but a high ratio of forage:concentrate decreased (P < 0.01) 15N enrichment of both the LAB and SAB. Significant differences between AA composition of the LAB and SAB were observed for 15 out of 17 AA studied. Bacterial colonization was linearly increased (P < 0.01) from about 5 to 70% as particle length of rumen contents was reduced from 3.35 mm to 0.15 mm (sieve size). The degree of colonization on each fraction of the rumen particulate matter was only affected (P < 0.10) by the ratio of forage:concentrate, with consistently higher (P < 0.10) bacterial colonization noted for high than for low forage:concentrate diets. Of the total bacterial mass within the rumen, less than 20% was associated with the liquid and over 70% was associated with the small particles that passed through the 0.6-mm sieve. Although the bacterial pool in the rumen was lower (P < 0.04) when flatly rolled barley rather than coarsely rolled barley was fed, bacterial flow to the duodenum was greater (P < 0.10) with increasing ratio offorage:concentrate. The present results confirm the differences of chemical composition and biomass for LAB and SAB. Manipulation of dietary factors such as ratio of forage:concentrate have the potential to alter bacterial colonization of rumen particles and the relative proportion of LAB to SAB, which were positively correlated to bacterial flow to the duodenum.

Abstract: The objective of the current study was to evaluate the effect of unrepresentative sampling of digesta particulate matter entering the omasal canal on the accuracy of fiber flow measurements. The experimental design comprised one period, one diet, and three cows as experimental units. Within each cow, the physical and chemical composition of digesta particulate matter was assessed at seven sites within the digestive tract. Three Finnish Ayshire dairy cows, equipped with ruminal and simple-T duodenal cannulas, in extended lactation were offered grass silage twice daily on an ad libitum basis. Digesta samples were collected from the rumen (dorsal and ventral sac), reticulum, omasal canal, omasum, duodenum, and rectum to determine particle size distribution in digesta, chemical composition of various particle size fractions, and distribution of two flow markers (Cr-labeled straw and indigestible NDF [INDF]) among particle size fractions. Digesta samples were wet-sieved using sieves of 2.50, 1.25, 0.630, 0.315, 0.160, and 0.080 mm. Particulate matter was analyzed for OM, NDF, and Cr concentrations, and INDF concentration was determined based on 12-d ruminal incubation. The particle size of digesta entering the omasal canal was larger compared with the omasum or the duodenum, suggesting that omasal canal samples were not representative of particle size distribution truly escaping the rumen. The concentration of potentially digestible NDF (PDNDF) decreased with decreasing particle size. The PDNDF concentration of particulate matter retained on all sieves was greatest in the rumen and gradually decreased along the digestive tract. From the reticulorumen to the omasum, the decrease was associated with decreased particle size, reflecting selective passage of particulate matter. In contrast, from the omasum to the duodenum and rectum, the PDNDF concentration decreased within each particle size fraction without effect on particle size, indicating a nonselective passage of particulate matter between these sites. Variation between particle size fractions was slightly greater for Cr concentration than for INDF concentration, indicating that unrepresentative sampling of particulate matter had a greater effect on Cr concentration compared with that of INDF. Owing to unrepresentative sampling, NDF entering the omasal canal was overestimated by 5% using INDF and underestimated by 7% using Cr as a particle phase marker. Of total NDF digestibility, proportionally 0.90, 0.07, and 0.03 occurred in the reticulorumen, omasum, and intestines, respectively. The current results indicate that, despite unrepresentative sampling of digesta particulate matter entering the omasal canal, the errors in determined NDF flow were small. The omasum may have a greater role in postruminal NDF digestion than the intestines.

Abstract: ruminally cannulated cows were fed a forage diet (93% hay + 7% straw) and a mixed diet (33 % hay + 7% straw + 40% barley) in a 2 ¥ 2 crossover experimental design. In sacco degra- dation of forage, fibrolytic activities (polysaccharidases and glycosidases) of the solid-associated bacteria (SAB), and distribution of the 3 main cellulolytic bacterial species (Fibrobacter succinogenes, Ruminococcus albus, Ruminococcus flavefaciens) were determined for both diets. Barley supple- mentation decreased the hay degradation rate and mainly the polysaccharidase activities of the SAB (30% on average). The sum of rRNA of the 3 cellulolytic bacterial species represented on average 17% of the total bacterial signal and R. albus was the dominant cellulolytic bacterial species of the 3 stud- ied. Barley supplementation did not modify the proportion of the 3 cellulolytic bacteria attached to plant particles. The negative effect of barley on the ruminal hay degradation rate is due to a decrease in fibrolytic activity of the SAB, and not to a modification of the balance of the three cellulolytic bac- terial species examined.

Abstract: A series of experiments was conducted to test the hypothesis that ‘synchronising’ the availability of carbohydrate and nitrogen (N) in the rumen of sheep, either by the feeding of carbohydrate-based supplements in relation to pasture intake or by provision of pasture with a higher ratio of these 2 nutrients, would benefit microbial protein synthesis (MPS). The response variables were rumen pH and ammonia (NH3), and urine allantoin was measured as an indicator of MPS. The animals used were adult castrate male sheep fitted with a rumen cannula. In experiment 1, rumen NH3 and pH were found to be at a maximum and minimum, respectively, 1–2 h after feeding kikuyu (Pennisetum clandestinum) grass and 3–4 h after feeding biennial ryegrass (Lolium multiflorum) with rumen N degradability (at 18 h in sacco) being 55 and 75%, respectively. In experiment 2, the infusion of sucrose at 4 h post-ingestion of ryegrass pasture (the anticipated time of peak rumen NH3 concentration) lowered peak NH3 concentration by 40 mg/dL (from 187 to 147 mg/dL) and raised urine allantoin excretion from 6.0 to 9.0 mmol/sheep.day compared with infusion of sucrose at –1, +1 or +7 h. When sucrose was infused, 1 h after being fed kikuyu (synchronous) at the expected time of peak rumen NH3 concentration, the NH3 peak (from 152 to 110 mg/dL) and rumen pH were reduced and urinary allantoin excretion raised (from 6.6 to 7.6 mmol/sheep.day) compared with sheep given sucrose 7 h after ingestion of kikuyu grass (asynchronous). In experiment 3, the feeding of fine-rolled barley grain at, or 2 h before, feeding kikuyu grass, to ‘synchronise’ the availability of rumen-fermentable carbohydrates (RFC) in the grain with the availability of N released from the grass, substantially reduced rumen NH3 and pH, and increased urine allantoin levels (from 10.1 to 11.8 mmol/sheep.day) compared with feeding barley at 4 or 6 h before feeding grass. In experiment 4, sheep were fed kikuyu or ryegrass cut in the early morning (AM) or late afternoon (PM), with the nutrient content varying only in water-soluble carbohydrates (64 v. 114 g/kg DM for ryegrass and 41 v. 80 g/kg DM for kikuyu, respectively) and percentage of dry matter. Sheep fed PM-ryegrass had a significantly lower rumen pH compared with sheep fed AM-ryegrass but there was no difference in rumen NH3 concentrations. Urinary allantoin secretion was substantially higher in the PM group than the AM group (8.7 v. 3.9 mmol/sheep.day). When sheep were fed PM-kikuyu the pH fell to lower values than for the AM-kikuyu (6.4 v. 6.8) and there was again no effect on rumen NH3 levels. Urine allantoin levels were higher (P = 0.006) (6.2 v. 5.6 mmol/ sheep.day) in PM-fed sheep. The results of these studies support the hypothesis that there are benefits in synchronising the availability of RFC with N in the rumen on MPS but this is not always associated with significant changes in rumen pH and NH3. The implications of these results are discussed in relation to the situation in which synchronisation of RFC and N may potentially benefit dairy cattle.

Abstract: The objective of this study was to determine the effects on animal performance and rumen digestion characteristics of supplementing cattle grazing autumn grass offered at two levels of allowance, with concentrates formulated from different carbohydrate sources. Seventy-two steers (494 (s.e. 38·4) kg) were assigned to one of six treatments which were arranged in a two grass allowances (5·5 or 11 kg grass dry matter (DM) per head per day) by three concentrate types (starch, starch + fibre or fibre-based diets) factorial design. The experiment commenced on 22 August and had a mean duration of 84 days. Concentrates were formulated to have similar metabolizable energy and crude protein and animals were offered 5 kg per head daily. Grass allowances were offered daily and concentrates were offered individually. Six rumen fistulated steers were used concurrently in a Latin square design to determine the effect of the diets on rumen digestion characteristics. Grass and concentrates degradabilities were measured using the nylon bag technique. The concentrate DM was degraded at 0·189, 0·102 and 0·094 per h for the starch, starch + fibre and fibre-based diets, respectively. Concentrate carbohydrate source and grass allowance did not affect (P > 0·05) degradation rate of grass DM or nitrogen in the rumen, rumen fluid pH, rumen fluid concentration of volatile fatty acids, total lactic acid or ammonia concentration or plasma concentration of glucose, urea or beta-hydroxybutyrate. Animals offered the low grass allowance had larger (P < 0·05) diurnal variation in the rumen fluid acetate: propionate ratio than those offered the high grass allowance. Increasing grass allowance increased (P < 0·01) carcass gain whereas concentrate type had no effect. There was no interaction between concentrate carbohydrate source and grass allowance on any of the production variables measured. It is concluded that the rumen environment in cattle grazing autumn grass buffered the effects of concentrate carbohydrate degradability such that similar productivity was achieved with each concentrate.

Abstract: Feeding leguminous fodder that is high in protein can improve rumen fermentation parameters leading to increased digestibility and intake of low quality fibrous feeds, and hence improved animal production. The effect on dry matter (DM) degradation and rumen fermentation parameters of feeding urea or tree legume foliage (Erythrina poeppigiana or Gliricidia sepium), as protein supplements to a basal diet of Hyparrhenia rufa hay, rice bran and molasses, was studied with rumen fistulated steers (Zebu × Creole crossbreeds). In a second experiment, Jersey × Creole crossbreeds were used to determine the effect of the above three treatments on milk production. Both experiments were conducted at CATIE's experimental farm, Turrialba, Costa Rica (9°58′ N; 83°31′ W; 639 m above sea level; mean annual precipitation of 2600 mm; mean daily temperature 22 °C). The disappearance of hay DM in the rumen increased with time to 144 hours and potential degradation of hay DM was significantly higher (P 0.05) when cows received either E. poeppigiana or G. sepium foliages as protein supplements (7.3 and 7.4 kg milk/cow/day, respectively) but they were superior (P < 0.05) to urea supplementation (6.7 kg milk/cow/day).

Abstract: Experimental data on phytate phosphorus utilisation by ruminants are scarce. The aim of this study was to estimate the phytase activity of rumen micro-organisms when phytate phospho- rus supply is high. A semi-continuous culture system fermentor (RUSITEC) was used. The inoculum was obtained from eight goats fed on either high or low forage level diets. Experimental buffers only differed by the nature of phosphorus monosodium phosphate vs. corn sodium phytate. The nylon bags containing 15 g DM of substrate were removed after a 48-hour incubation period. The sys- tem was maintained for 15 days: 5 days for adaptation, in order to obtain a steady state, and 10 days for sampling and recording. No significant differences were observed for DM digestibility, gas pro- duction, pH, N-NH 3 , and SCFA for the different treatments. Bacterial efficiency of phytate phosphorus utilisation was significantly higher ( p < 0.001) with organic P, but remained lower than the data usually reported in the literature. These results may be explained by the relative saturation of bacte- rial phytase activity when the buffer contains a high level of phytate phosphorus.

Abstract: The effect of varying hay/barley-proportions in the feed ration on biotin metabolism of rumen microbes was studied by means of the rumen simulation technique RUSITEC. The stepwise replacement of hay by barley decreased dietary biotin and the net output of biotin by the microbial metabolism. It is concluded that rumen microbes utilise more and/or synthesize less biotin with increasing proportions of dietary barley. These results indicate that a critical reconsideration of current views with regard to the supply and requirement of the high yielding dairy cow for biotin is necessary.

Abstract: Three sheep were fed a pelleted high-roughage diet either once, 6, or 24 times per day in a 3 x 3 Latin square trial. During each 21-d period, 14 d were allowed for adaptation followed by a 7-d collection period, in which samples for microbial counts were taken on d 1 and 5 and several rumen parameters were measured on d 2 and 6. Bacterial concentrations were not different between feeding frequencies on the first sampling day but were higher (P < 0.05) on the second sampling day when the sheep were fed 24 times a day. Fungal concentrations were not different among feeding frequencies on either sampling day. No effects of feeding frequency were observed for the concentration of cellulolytic bacteria or fungi. On d 2, ruminal volume was larger (P < 0.05) with six feedings than with one feeding and fluid volume turnover was greater (P < 0.05) when sheep were fed 24 times per day. Rumen pH values were higher (P < 0.01) on both d 1 and 5 when the sheep were fed once a day and the percentage of rumen dry matter was highest (P < 0.02) with 24 feedings. These findings would suggest that if the same amount of a given diet is fed daily, the number of feedings does not markedly affect microbial concentrations, rumen volume, or liquid turnover time.

Abstract: Incubations were carried out with a batch culture system to study the effects of different N sources on the fermentation by ruminal micro-organisms from Merino sheep of two fibre substrates derived from feedstuffs that differed in their fermentation rate. The substrates were neutral-detergent fibre (NDF) from barley straw and sugarbeet pulp. N sources were ammonia (NH4Cl) and peptides (Trypticase). Three treatments were made by replacing ammonia-N with peptide-N at levels of 0 (AMMO), 33 (PEPLOW) and 66 % (PEPHIGH) of total N. There were no differences (P>0.05) between treatments in NDF degradation for both the barley straw and the sugarbeet pulp. Peptides increased (P 0.05) by the presence of peptides compared with the AMMO treatment, but values were greater for the PEPHIGH compared with the PEPLOW N source, reaching statistical significance (P 0.05) by N source. These results indicate that N forms other than ammonia are needed for maximal growth of fibre-digesting ruminal micro-organisms.

Abstract: The hypothesis that South African game animals that use browse containing condensed tannins (CT) could have rumen microbial populations better able to ferment tanniniferous forages was explored Rumen fluid (RF) from a range of browsing ruminants was used to ferment tanniniferous forages in vitro and the results were compared with the values obtained with rumen fluid from sheep Leaves of the shrub legumes Leucaena leucocephala, L pallida, L trichandra, Calliandra calothyrsus,Gliricidia sepium, and Acacia boliviana were used as substrates In vitro digestibility of dry matter (IVDMD) and nitrogen (IVND) was measured in the presence or absence of polyethylene glycol (PEG) 4000 to assess the adverse effects of the CT in the feeds Rumen fluid from giraffe, greater kudu, eland, duiker, impala, nyala, goat, and gnu was compared with that from sheep Ability to metabolise or block the adverse effects of CT was expected to result in only a small or no response to PEG in terms of IVDMD and IVND with rumen fluid from browsers, and therefore to result in a significant animal feed and animal PEG interaction There were no significant (P > 005) animal feed interactions For every animal species there was a response to PEG in terms of IVND, and this response was linearly related to the CT in the feed as measured by PEG binding using 14C-labelled PEG Rumen fluid from browsers had a lower pH (578 0060 v 668 0052) and a higher dry matter content (1473 102 mg v 415 43 mg/100 mL RF) than that of grazers (sheep and gnu) However, when measured at 2 pH levels (67 and 58), IVDMD and IVND of the feeds were not improved at the lower pH with rumen fluid from the browsers Rather, digestibility was depressed at the lower pH with rumen fluid from all animal species We conclude from these in vitro studies that rumen microbial populations from browsers are not able to ferment tanniniferous forages better than rumen microbial populations from grazers It seems likely that these browsers do not have rumen bacteria capable of degrading the CT in these shrub legumes Production on such tanniniferous feeds may well depend on tannin binding with proline-rich saliva rather than on metabolism of or tolerance to CT by rumen bacteria