Rumen Of Sheep Research Articles

Identification and distribution of cellobiose 2-epimerase genes by a PCR-based metagenomic approach.

Cellobiose 2-epimerase (CE) catalyzes the reversible epimerization of cellobiose to 4-O-β-D-glucopyranosyl-D-mannose. By using a PCR-based metagenomic approach, 71 ce-like gene fragments were obtained from wide-ranging environmental samples such as sheep rumen, soils, sugar beet extracts, and anaerobic sewage sludge. The frequency of isolation of the fragments similar to known sequences varied depending on the nature of the samples used. The ce-like genes appeared to be widely distributed in environmental bacteria belonging to the phyla Bacteroidetes, Chloroflexi, Dictyoglomi, Firmicutes, Proteobacteria, Spirochaetes, and Verrucomicrobia. The phylogenetic analysis suggested that the cluster of CE and CE-like proteins was functionally and evolutionarily separated from that of N-acetyl-D-glucosamine 2-epimerase (AGE) and AGE-like proteins. Two ce-like genes containing full-length ORFs, designated md1 and md2, were obtained by PCR and expressed in Escherichia coli. The recombinant mD1 and mD2 exhibited low K m values and high catalytic efficiencies (k cat/K m) for mannobiose compared with cellobiose, suggesting that they should be named mannobiose 2-epimerase, which is involved in a new mannan catabolic pathway we proposed.

Technical note: Protozoa-specific antibodies raised in sheep plasma bind to their target protozoa in the rumen.

Binding of IgG antibodies to Entodinium spp. in the rumen of sheep (Ovis aries) was investigated by adding IgG, purified from plasma, directly into the rumen. Plasma IgG was sourced from sheep that had or had not been immunized with a vaccine containing whole fixed Entodinium spp. cells. Ruminal fluid was sampled approximately 2 h after each antibody dosing. Binding of protozoa by a specific antibody was detected using an indirect fluorescent antibody test. An antibody titer in the ruminal fluid was determined by ELISA, and the concentration of ruminal fluid ammonia-N and ruminal pH were also determined. Entodinium spp. and total protozoa from IgG-infused sheep were enumerated by microscopic counts. Two-hourly additions of IgG maintained a low antibody titer in the rumen for 12 h and the binding of the antibody to the rumen protozoa was demonstrated. Increased ammonia-N concentrations and altered ruminal fluid pH patterns indicated that additional fermentation of protein was occurring in the rumen after addition of IgG. No reduction in numbers of Entodinium spp. was observed (P>0.05). Although binding of antibodies to protozoa has been demonstrated in the rumen, it is unclear how much cell death occurred. On the balance of probability, it would appear that the antibody was degraded or partially degraded, and the impact of this on protozoal populations and the measurement of a specific titer is also unclear.

Effect of stoned olive pomace on rumen microbial communities and polyunsaturated fatty acid biohydrogenation: an in vitro study.

BackgroundStoned olive pomace (SOP), which represents approximately 50% of the conversion process of olives to olive oil, is largely not utilised and creates costs for its disposal and has negative environmental impacts. In vitro trial experiments were employed to study the effect of feeds integrated with this bio-waste, which is rich in polyphenols, on rumen biohydrogenation, using sheep rumen liquor as inoculum.ResultsFatty acid (FA) analysis and a polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE) approach aimed at characterising the microbial community indicated that including SOP in feeds at the level of 50 g/kg and 90 g/kg induced changes in the FA profile and microbial populations. The simultaneous decrease of Butyrivibrio proteoclasticus and accumulation of vaccenic acid was observed. A depression in the populations of Neisseria weaveri, Ruminobacter amylophilus and other unclassified bacteria related to members of the Lachnospiraceae and Pasteurellaceae families was detected, suggesting that these microbial groups may be involved in rumen biohydrogenation.ConclusionsSupplementation of feeds with SOP alters the rumen bacterial community, including bacteria responsible for the hydrogenation of vaccenic acid to stearic acid, thereby modifying the FA profile of the rumen liquor. Hence, a use of SOP aimed to produce meat or dairy products enriched in functional lipids can be hypothesised.

Genetic diversity and expression profiles of cysteine phytases in the sheep rumen during a feeding cycle.

Cysteine phytase is the main phytate-degrading enzyme of ruminant animals. To explore the genetic diversity and dynamic expression profile of cysteine phytase in sheep rumen during a feeding cycle, four transcript (0, 4, 9 and 16h after feeding) and one DNA (9h after feeding) clone libraries were constructed, respectively. A total of 46 distinct gene fragments were identified, and most of these sequences had low identities (<60%) with known phytases. Great divergence was found in the constitution and abundance of genes at the genome and transcriptional levels, and the transcript data are more reliable to reflect the information of functional genes. Phylogenetic analysis indicated that the genes from uncultured bacteria instead of Firmicutes played the major phytate-degrading role. Further comparative analysis revealed the dynamic constitution of cysteine phytase genes in rumen at different time points. Ruminal phytases, that are cysteine phytases, are novel in sequences and functions. Great divergence in the constitution and abundance of cysteine phytase genes at the genome and transcriptional levels suggested that transcript data are more reliable to reflect the information of functional genes. Phylogenetic and rarefaction analyses indicated that the cysteine phytase genes from uncultured bacteria instead of Firmicutes play the major phytate-degrading role in rumen, and their constitution is dynamic at different time points. This study provides a new insight into ruminal cysteine phytase genes and undermines their expression profiles over a whole feeding cycle.

Low ambient temperature elevates plasma triiodothyronine concentrations while reducing digesta mean retention time and methane yield in sheep.

Ruminant methane yield (MY) is positively correlated with mean retention time (MRT) of digesta. The hormone triiodothyronine (T3 ), which is negatively correlated with ambient temperature, is known to influence MRT. It was hypothesised that exposing sheep to low ambient temperatures would increase plasma T3 concentration and decrease MRT of digesta within the rumen of sheep, resulting in a reduction of MY. To test this hypothesis, six Merino sheep were exposed to two different ambient temperatures (cold treatment, 9 ± 1 °C; warm control 26 ± 1 °C). The effects on MY, digesta MRT, plasma T3 concentration, CO2 production, DM intake, DM digestibility, change in body weight (BW), rumen volatile fatty acid (VFA) concentrations, estimated microbial protein output, protozoa abundance, wool growth, water intake, urine output and rectal temperature were studied. Cold treatment resulted in a reduction in MY (p < 0.01); digesta MRT in rumen (p < 0.01), hindgut (p = 0.01) and total digestive tract (p < 0.01); protozoa abundance (p < 0.05); and water intake (p < 0.001). Exposure to cold temperature increased plasma T3 concentration (p < 0.05), CO2 production (p = 0.01), total VFA concentrations (p = 0.03) and estimated microbial output from the rumen (p = 0.03). The rate of wool growth increased (p < 0.01) due to cold treatment, but DM intake, DM digestibility and BW change were not affected. The results suggest that exposure of sheep to cold ambient temperatures reduces digesta retention time in the gastrointestinal tract, leading to a reduction in enteric methane yield. Further research is warranted to determine whether T3 could be used as an indirect selection tool for genetic selection of low enteric methane-producing ruminants.

The fate of glycerol entering the rumen of dairy cows and sheep.

This study investigated the fate of glycerol entering the rumen, in particular whether glycerol could be absorbed across the rumen epithelium. Three non-lactating rumen-fistulated cows were used to calculate the overall disappearance rate of glycerol in vivo and evaluate the rate of ruminal glycerol absorption. Rumen epithelial tissues isolated from sheep were used to characterise glycerol transport properties. The rate of rumen microbial degradation of glycerol was then studied in an in vitro system under anaerobic and thermo-regulated conditions. The results showed that glycerol can be absorbed from the rumen in significant amounts. The fractional rate of absorption of glycerol was not affected by variations in glycerol concentration in the buffer solution in the in vivo study. The glycerol absorption apparently occurred largely by passive diffusion and was probably not facilitated by carriers. Glycerol also disappeared via microbial digestion and outflow from the rumen through the omasal orifice.

In vivo measurement of the absorption of strontium in the rumen and small intestine of sheep as an index of calcium absorption capacity.

In the present study, a method was developed for determining the alimentary tract Ca absorption capacity of ruminant animals by measuring the absorption rate of Sr after the administration of an oral dose of strontium chloride acting as a tracer analogue of Ca. A close correlation between the absorption rates of the two tracers was observed upon simultaneous administration of an oral dose of stable Sr and radioactive calcium (r 0·98). The Ca absorption capacity of the rumen and small intestine was determined separately by either directing the solution into the rumen or by diverting it into the post-ruminal tract by vasopressin-induced closure of the ruminoreticular groove. The animals were treated with 1α-hydroxyvitamin D3 administered via subcutaneously implanted mini-osmotic pumps. The effect of elevated plasma 1,25-dihydroxycholecalciferol concentrations on the Ca absorption capacity of the alimentary tract was then determined. An increased rate of Sr absorption was observed in both the rumen and small intestine of sheep after treatment, although it is unclear whether the rumen possesses the same vitamin D-dependent Ca absorption pathway as the small intestine.

Utilization of microbial selenium collected from the rumen of sheep

1. Selenium species in diet containing carnosic acid, fish and rapeseed oils affect fatty acid profiles in lamb muscles A. Rozbicka-Wieczorek, M. Czauderna, E. Więsyk, A. Radzik-Rant Journal of Animal and Feed Sciences CrossRef

A study on gastrointestinal tract characteristics of ram lambs at the same weights from six Turkish sheep breeds

A study was conducted, first, to compare the gastrointestinal tract (GIT), reticulo-rumens and intestinal capacities (relative to body weight) of ram lambs from six Turkish sheep breeds: Turkish Merino (TM, Anatolia Merino), Akkaraman (AK), Kıvırcık (KV), Awassi (AW), Karayaka (KY) and Morkaraman (MK), and second, to examine the influence of sheep breed on the pH of the digesta in the various segments (rumen, jejunum and caecum) of the GIT. Six ram lambs from each breed were slaughtered at 40.2 ± 0.80 kg liveweight. Lambs from the AK breed had a heavier GIT weight than the AW, KY or MK breeds. The weight of the full stomach was higher in AK lambs compared with the other lambs, except for KV lambs, while the relative weight of the empty stomachs was heavier in MK ram lambs than in TM, AW and KY lambs. The AK lambs had the highest relative empty intestinal weight. The relative weight of empty intestines was heavier in KV lambs than in KY lambs. The pH of the rumen content was higher in TM and AK lambs compared with KY and MK lambs, while the pH of the caecum content of KV lambs was higher than those of KY and MK lambs. In conclusion, significant differences were found among the six breeds in terms of some variables that were studied, although a higher proportional GIT weight in the fat-tail breeds such as the MK, AK and AW was not reflected in all components or in the pH of the GIT content.Keywords: Native sheep breed, digestive system, rumen, small and large intestine, pH value

Effects of ethyl-3-nitrooxy propionate and 3-nitrooxypropanol on ruminal fermentation, microbial abundance, and methane emissions in sheep

The aim of this work was to investigate the effect of feeding ethyl-3-nitrooxy propionate (E3NP) and 3-nitrooxypropanol (3NP), 2 recently developed compounds with potential antimethanogenic activity, in vitro and in vivo in nonlactating sheep on ruminal methane production, fermentation pattern, the abundance of major microbial groups, and feed degradability. Three experiments were conducted, 1 in vitro and 2 in vivo. The in vitro batch culture trial (experiment 1) tested 2 doses of E3NP and 3NP (40 and 80μL/L), which showed a substantial reduction of methane production (up to 95%) without affecting concentration of volatile fatty acids (VFA). The 2 in vivo trials were conducted over 16 d (experiment 2) and 30 d (experiment 3) to study their effects in sheep. In experiment 2, 6 adult nonpregnant sheep, with permanent rumen cannula and fed alfalfa hay and oats (60:40), were treated with E3NP at 2 doses (50 and 500mg/animal per day). After 7, 14, and 15 d of treatment, methane emissions were recorded in respiration chambers and rumen fluid samples were collected for VFA analysis and quantification of bacterial, protozoal, and archaeal numbers by real-time PCR. Methane production decreased by 29% compared with the control with the higher dose of E3NP on d 14 to 15. A decrease in the acetate:propionate ratio was observed without detrimental effects on dry matter intake. In experiment 3, 9 adult nonpregnant sheep, with permanent rumen cannula and fed with alfalfa hay and oats (60:40), were treated with E3NP or 3NP at one dose (100mg/animal per day) over 30 d. On d 14 and d 29 to 30, methane emissions were recorded in respiration chambers. Rumen fluid samples were collected on d 29 and 30 for VFA analysis and quantification of bacterial, protozoal, and archaeal numbers by real-time PCR. In addition, on d 22 and 23, samples of oats and alfalfa hay were incubated in the rumen of sheep to determine dry matter ruminal degradation over 24 and 48h, respectively; no effect was observed (78.6, 78.3, and 78.8% of alfalfa and 74.2, 74.0, and 70.6% of oats in control, E3NP, and 3NP groups, respectively). A reduction in methane production was observed for both additives at d 14 and d 29 to 30. In both treatments, the acetate:propionate ratio was significantly decreased. Likewise, total concentrations of the analyzed microbial groups in the rumen showed no difference among treatments and doses for both experiments. Both tested compounds showed promise as methane inhibitors in the rumen, with no detrimental effects on fermentation or intake, which would need to be confirmed in lactating animals.

Determination of the 137Cs and 90Sr radioisotope activity concentrations found in digestive organs of sheep fed with different feeds

In this study, 12 weanling lambs were fed for 6 months at the Vetenary Control and Research Institute of Elazığ in Turkey. Sheep were divided into two groups according to feeding type. Each group of sheep was fed a different amount of feed. The activity concentrations of 137Cs and 90Sr were determined in rumen, small intestine and large intestine tissue samples of sheep fed various feeds and quantities.Transfer coefficients and concentration ratios were estimated. The highest average 137Cs and 90Sr activity concentrations were 82 ± 22 Bq kg−1 fw in the straw group small intestine sample and 74 ± 9.0 Bq kg−1 fw in the fresh lucerne group rumen sample, respectively. Radioactivity levels for 137Cs and 90Sr are under the maximum permitted radioactive pollution level for all the samples. The transfer coefficient of 137Cs and 90Sr in the samples of sheep ranged from 1.0 × 10−1 to 2.1 × 100 d kg−1 and 1.7 × 10−1 to 2.3 × 100 d kg−1, respectively. Calculated transfer coefficients of 90Sr and 137Cs in the samples of sheep were higher than the expected values. Geometric means of Ff and CR for 137Cs were maximum in the fresh lucerne group large intestine sample. Geometric means of Ff and CR for 90Sr were maximum in the fresh lucerne group small intestine sample.

A C-Terminal Proline-Rich Sequence Simultaneously Broadens the Optimal Temperature and pH Ranges and Improves the Catalytic Efficiency of Glycosyl Hydrolase Family 10 Ruminal Xylanases

Efficient degradation of plant polysaccharides in rumen requires xylanolytic enzymes with a high catalytic capacity. In this study, a full-length xylanase gene (xynA) was retrieved from the sheep rumen. The deduced XynA sequence contains a putative signal peptide, a catalytic motif of glycoside hydrolase family 10 (GH10), and an extra C-terminal proline-rich sequence without a homolog. To determine its function, both mature XynA and its C terminus-truncated mutant, XynA-Tr, were expressed in Escherichia coli. The C-terminal oligopeptide had significant effects on the function and structure of XynA. Compared with XynA-Tr, XynA exhibited improved specific activity (12-fold) and catalytic efficiency (14-fold), a higher temperature optimum (50°C versus 45°C), and broader ranges of temperature and pH optima (pH 5.0 to 7.5 and 40 to 60°C versus pH 5.5 to 6.5 and 40 to 50°C). Moreover, XynA released more xylose than XynA-Tr when using beech wood xylan and wheat arabinoxylan as the substrate. The underlying mechanisms responsible for these changes were analyzed by substrate binding assay, circular dichroism (CD) spectroscopy, isothermal titration calorimetry (ITC), and xylooligosaccharide hydrolysis. XynA had no ability to bind to any of the tested soluble and insoluble polysaccharides. However, it contained more α helices and had a greater affinity and catalytic efficiency toward xylooligosaccharides, which benefited complete substrate degradation. Similar results were obtained when the C-terminal sequence was fused to another GH10 xylanase from sheep rumen. This study reveals an engineering strategy to improve the catalytic performance of enzymes.

Interaction of rumen bacteria as assumed by colonization patterns on untreated and alkali‐treated rice straw

Colonization patterns of representative rumen bacteria were compared between untreated rice straw (UTS) and sodium hydroxide-treated rice straw (SHTS). UTS and SHTS were incubated in the rumen of sheep for 10 min, 1, 2, 6, 12, 24, 48 and 96 h using the nylon bag method. The population sizes of 13 representative bacterial species or groups were quantified by real-time PCR. The total bacterial population size (abundance) was similar in both UTS and SHTS. Fibrobacter succinogenes showed a higher population size compared to other fibrolytic species and was detected at a higher level in SHTS (3.7%) than in UTS (2.6%). Ruminococcus albus and Ruminococcus flavefaciens were also detected at higher levels in SHTS (0.15% and 0.29%) than in UTS (0.03% and 0.18%). Population sizes of non-fibrolytic species, such as Selenomonas ruminantium, Anaerovibrio lipolytica and Succinivibrio dextrinosolvens were higher in UTS than in SHTS. Coefficient of determination (r(2) ) on population changes between bacterial species or groups were higher in UTS than in SHTS, suggesting the necessity of stronger bacterial interactions for UTS digestion. Therefore, not only colonization of fibrolytic species, but also synergistic interactions between different bacterial species may be key to the ruminal digestion of rice straw.

Study of effectiveness of bacteria isolated from rumen microbes with acetogen medium as methanogenesis inhibitor

Ruminal methanogenesis has disadvantageous effects on ruminant animals and environment of atmosphere. Inhibition of methane produced through reduction of carbondioxide has been conducted by bacteriological approach. The approach involved the isolation of bacteria from rumens of sheep (IBD) and buffalo (IBK) using medium for CO-utilizing acetogens. The isolate of bacterium was multiplied with the usual culture medium and then used as inoculum to degrade a substrate of King grass under constant temperature (390C) for 48 hours. Fresh rumen fluid of sheep (CRDS) was used as comparing inoculum. Measurementswere carbondioxide and methane gasses, pH, volatile fatty acid (VFA) and NH3-N contents, bacterial count, and dry matter digestibility ( in vitro DMD). The data measured were analyzed by using completely randomized design. The results showed that morphological cell of IBD was oval pleomorphic with Gram negative type, and cell of IBK was rod with Gram negative type. Percentage of CH4 produced by inoculum of IBD was lower than CRDS but was not significantly different (29.47 vs. 33.07%), while the percentage of methane produced by inoculum of IBK was very significantly lower than CRDS (24.29 vs. 33.07%) (P<0.01). Acetate/propionate ratio as a result of substrate fermentation by inoculum of IBD (3.55) and IBK (3.79) were very significantly higher than that of CRDS (2.43) (P<0.01) . It is concluded from this experiment that isolates used were effective to inhibit the methanogenesis and the species contained in the isolates were indicated to be homoacetogenic bacteria. Key words: Bacterial isolate, acetogen medium, methanogenesis, inhibitor

Technical note: Comparison of automated ribosomal intergenic spacer analysis and denaturing gradient gel electrophoresis to assess bacterial diversity in the rumen of sheep1

The aim of this study was to compare automated ribosomal intergenic spacer analysis (ARISA) and denaturing gradient gel electrophoresis (DGGE) techniques to assess bacterial diversity in the rumen of sheep. Sheep were fed 2 diets with 70% of either alfalfa hay or grass hay, and the solid (SOL) and liquid (LIQ) phases of the rumen were sampled immediately before feeding (0 h) and at 4 and 8 h postfeeding. Both techniques detected similar differences between forages, with alfalfa hay promoting greater (P < 0.05) bacterial diversity than grass hay. In contrast, whereas ARISA analysis showed a decrease (P < 0.05) of bacterial diversity in SOL at 4 h postfeeding compared with 0 and 8 h samplings, no variations (P > 0.05) over the postfeeding period were detected by DGGE. The ARISA technique showed lower (P < 0.05) bacterial diversity in SOL than in LIQ samples at 4 h postfeeding, but no differences (P > 0.05) in bacterial diversity between both rumen phases were detected by DGGE. Under the conditions of this study, the DGGE was not sensitive enough to detect some changes in ruminal bacterial communities, and therefore ARISA was considered more accurate for assessing bacterial diversity of ruminal samples. The results highlight the influence of the fingerprinting technique used to draw conclusions on factors affecting ruminal bacterial diversity.

The symbiotic intestinal ciliates and the evolution of their hosts

The evolution of sophisticated differentiations of the gastro-intestinal tract enabled herbivorous mammals to digest dietary cellulose and hemicellulose with the aid of a complex anaerobic microbiota. Distinctive symbiotic ciliates, which are unique to this habitat, are the largest representatives of this microbial community. Analyses of a total of 484 different 18S rRNA genes show that extremely complex, but related ciliate communities can occur in the rumen of cattle, sheep, goats and red deer (301 sequences). The communities in the hindgut of equids (Equus caballus, Equus quagga), and elephants (Elephas maximus, Loxodonta africanus; 162 sequences), which are clearly distinct from the ruminant ciliate biota, exhibit a much higher diversity than anticipated on the basis of their morphology. All these ciliates from the gastro-intestinal tract constitute a monophyletic group, which consists of two major taxa, i.e. Vestibuliferida and Entodiniomorphida. The ciliates from the evolutionarily older hindgut fermenters exhibit a clustering that is specific for higher taxa of their hosts, as extant species of horse and zebra on the one hand, and Africa and Indian elephant on the other hand, share related ciliates. The evolutionary younger ruminants altogether share the various entodiniomorphs and the vestibuliferids from ruminants.

Effect of dietary potassium supplementation on the calcium absorption capacity in the rumen and abomasum and fractional excretion of urinary minerals in sheep

The aim of this study was to determine the effect of potassium (K) supplementation on the calcium (Ca) absorption capacity from the rumen and abomasum of sheep. The Ca absorption capacity from the rumen and abomasum of sheep was measured using stable strontium (Sr) as a Ca-analogue tracer method. The sheep, cannulated at either the rumen or abomasum, were randomly allocated to one of two groups (control or K-supplemented) and fed in individual metabolism pens twice daily with a diet comprising oaten hay, lucerne chaff and barley fortified with or without potassium carbonate (K2CO3). The K content of the diet of the K-supplemented animals was 3.1% of dry matter compared with 1.4% for the control animals. The animals were fed their respective diets for a period of 2 weeks. The fractional absorption capacity (FC) of Ca was estimated before, during and after the treatment period. Supplementation with K decreased the FC of Ca in both the rumen and abomasum during treatment and increased the FC of Ca in the abomasum post-treatment. Supplementation with K also increased the fractional excretion of K in the urine, but decreased the fractional excretion of Ca and magnesium (Mg) (P &lt; 0.05), showing that K supplementation significantly affected Ca and Mg metabolism. Results suggest that renal conservation of Ca and Mg is an important mechanism controlling the Ca and Mg pool for vital functions of the body. In times of high demand for these minerals during lactation and pregnancy, high K in the diets may predispose sheep to hypomagnesaemia and hypocalcaemia, which in turn will have a negative impact on productivity and economic returns.

The production of acetate, propionate and butyrate in the rumen of sheep: fitting models to 14C- or 13C-labelled tracer data to determine synthesis rates and interconversions

A procedure is described for solving an open, fully exchanging, three-compartment model representing ruminal volatile fatty acids (VFA) kinetics in sheep. This model was solved using results from a published study in which labelled VFA, viz. 1-14C-acetate (Ac), 1-14C-propionate (Pr) and 1-14C-butyrate (Bu), were infused individually at a constant rate for 240 min into the rumen of sheep on different occasions. During the tracer infusions, the sheep were given 75 g of lucerne (Medicago sativa) every hour. The patterns of increasing specific radioactivity (SA) during the infusions were described by differential equations that were solved using the computer software, WinSAAM. This linear kinetic analysis gave estimates of the rates of synthesis and absorption of Ac, Pr and Bu and the carbon interconversions between each acid. The sizes of the Ac, Pr and Bu compartments (10.5, 3.9 and 2.1 g C respectively), were also estimated, which is not possible with commonly used algebraic analyses. The model output showed that tracer : tracee equilibrium (plateau SA) had not been reached in the Ac, Pr and Bu compartments during the 240 min of tracer infusion and the algebraic method of analysis used in the original study was therefore compromised. The procedures described here eliminated this source of error; in addition, confidence in the model solution was improved because all data representing the build-up to plateau SA were used, rather than just estimates of ‘plateau’ SA. After accounting for VFA interconversions, the rates of VFA absorption (or incorporation into other materials such as microbial polymers or methane) were 85, 48 and 49 g C/day, or 78%, 91% and 73% of the total production of each VFA, respectively. The variability in the observed SA responses to the tracer infusions was relatively large and this is discussed. Model solutions using the linear kinetic analysis and the previously widely used algebraic analyses are compared. The effect of positional labelling in tracer VFA is also discussed in the light of evidence that the rate of absorption of Pr may be overestimated when [1-14C]-Pr rather than [2-14C]-Pr or uniformly labelled [14C]-Pr is used as the Pr tracer.

Carbon precursors of methane synthesis in the rumen of sheep dosed with ionophores

Rates of methane (CH4) production and the sources of carbon (C) for its synthesis were studied in four mature ewes when dosed with a CH4-mitigating ionophore ICI-111075, or monensin, or when untreated. The sheep were given 700 g/day of chaffed lucerne hay in equal portions every hour, before and during experiments in which 14C-labelled NaHCO3–, acetate, propionate, lactate and formate were infused intraruminally over 12 h and the specific radioactivity of C (SR) in each of these substrates was determined. During these infusions, the SR of material in the primary pool (the tracer infusion site) and in secondary metabolites of this material (secondary pools) approached asymptotic or ‘plateau’ values. The rate of infusion of 14CH4 (kBq/day) divided by the plateau SR value (kBq/g C) gave estimates of the rate of irreversible loss of CH4 (g C/day). These calculations indicated that CH4 production rate was reduced by 72% when sheep were dosed with ICI-111075 and by 58% when dosed with monensin. With monensin, the reduction in CH4 production was not associated with hydrogen (H2) accumulation in the rumen headspace gases whereas with ICI-111075, the decrease in CH4 production was associated with marked H2 accumulation in the headspace gases. When plateau SR were attained during the tracer infusions, the percentage ratio, (SR in any secondary pool: SR in the primary pool) gave an estimate of the fraction of C in that secondary pool derived from material of the primary pool. Calculated in this way, the percentage of CH4-C derived from rumen fluid carbon dioxide (CO2) averaged 59% in untreated sheep, and when sheep were dosed with ICI-111075, the corresponding percentage averaged 12%. These findings indicate there are sources of C for rumen CH4 synthesis other than rumen fluid CO2. However, there was no evidence that C from acetate, propionate, lactate or formate in rumen fluid were direct sources of the unidentified CH4-C. One plausible explanation for these findings is that CH4 is synthesised within naturally occurring microbial biofilms (attached to feed particles or the rumen wall) from CO2 that is produced locally by fermentation of unlabelled substrates within the biofilms. It is postulated that such pools of CO2 would be kinetically distinct and, during the infusion of 14C-labelled substrates, would exhibit a lower SR than the CO2 in the surrounding rumen fluid.

Epithelia of the ovine and bovine forestomach express basolateral maxi-anion channels permeable to the anions of short-chain fatty acids

It has long been established that the absorption of short-chain fatty acids (SCFA) across epithelia stimulates sodium proton exchange. The apically released protons are not available as countercations for the basolateral efflux of SCFA anions and a suitable transport model is lacking. Patch clamp and microelectrode techniques were used to characterize an anion conductance expressed by cultured cells of the sheep and bovine rumen and the sheep omasum and to localize the conductance in the intact tissue. Cells were filled with a Na-gluconate solution and superfused with sodium salts of acetate, propionate, butyrate, or lactate. Reversal potential rose and whole cell current at +100mV decreased with the size of the anion. Anion-induced currents could be blocked by diisothiocyanato-stilbene-2,2'-disulfonic acid (DIDS), NPPB (200 μmoll(-1)), or pCMB (1mmoll(-1)). In patches of bovine ruminal cells, single channels were observed with a conductance for chloride (327 ± 11 pS), acetate (115 ± 8 pS), propionate (102 ± 10 pS), butyrate (81 ± 2 pS), and gluconate (44 ± 3 pS). Channels expressed by sheep rumen and omasum were similar. Microelectrode experiments suggest basolateral localization. In conclusion, forestomach epithelia express basolateral maxi-anion channels with a permeability sequence of chloride > acetate > propionate > butyrate. SCFA absorption may resemble functionally coupled transport of NaCl, with the Na(+)/K(+)-ATPase driving the basolateral efflux of the anion through a channel. Since protons are apically extruded, the model accurately predicts that influx of buffers with saliva is essential for the pH homeostasis of the ruminant forestomach.