Enteric methane emission from Jersey cows during the spring transition from indoor feeding to grazing.

Organic dairy cows in Denmark are often kept indoors during the winter and outside at least part time in the summer. Consequently, their diet changes by the season. We hypothesized that grazing might affect enteric CH4 emissions due to changes in the nutrition, maintenance, and activity of the cows, and they might differentially respond to these factors. This study assessed the repeatability of enteric CH4 emission measurements for Jersey cattle in a commercial organic dairy herd in Denmark. It also evaluated the effects of a gradual transition from indoor winter feeding to outdoor spring grazing. Further, it assessed the individual-level correlations between measurements during the consecutive feeding periods (phenotype × environment, P × E) as neither pedigrees nor genotypes were available to estimate a genotype by environment effect. Ninety-six mixed-parity lactating Jersey cows were monitored for 30 d before grazing and for 24 d while grazing. The cows spent 8 to 11 h grazing each day and had free access to an in-barn automatic milking system (AMS). For each visit to the AMS, milk yield was recorded and logged along with date and time. Monitoring equipment installed in the AMS feed bins continuously measured enteric CH4 and CO2 concentrations (ppm) using a noninvasive "sniffer" method. Raw enteric CH4 and CO2 concentrations and their ratio (CH4:CO2) were derived from average concentrations measured during milking and per day for each cow. We used mixed models equations to estimate variance components and adjust for the fixed and random effects influencing the analyzed gas concentrations. Univariate models were used to precorrect the gas measurements for diurnal variation and to estimate the direct effect of grazing on the analyzed concentrations. A bivariate model was used to assess the correlation between the 2 periods (in-barn vs. grazing) for each gas concentration. Grazing had a weak P × E interaction for daily average CH4 and CO2 gas concentrations. Bivariate repeatability estimates for average CH4 and CO2 concentrations and CH4:CO2 were 0.77 to 0.78, 0.73 to 0.80, and 0.26, respectively. Repeatability for CH4:CO2 was low (0.26) but indicated some between-animal variation. In conclusion, grazing does not create significant shifts compared with indoor feeding in how animals rank for average CH4 and CO2 concentrations and CH4:CO2. We found no evidence that separate evaluation is needed to quantify enteric CH4 and CO2 emissions from Jersey cows during in-barn and grazing periods.

Defining risk for low reticuloruminal pH during the diet transition period in a commercial feedlot in western Canada.

The objective of the current study was to measure reticuloruminal pH in cattle in a commercial feedlot setting to determine the incidence and extent of low reticuloruminal pH for steers and heifers as they transition to a high-concentrate finishing diet. Reticuloruminal pH was measured in 16 "mixed breed" steers (4 steers/pen with 4 pens) and 16 "mixed breed" heifers (4 heifers/pen with 4 pens) housed in commercial feedlot pens, with 227 ± 13 and 249 ± 6 cattle/pen cohort steers and heifers, respectively, for the diet transition period. Cattle were transitioned from a diet of 53.5% forage and 46.5% concentrate to a diet of 9.5% forage and 90.5% concentrate on a DM basis using a 40-d transition with 5 dietary steps with the diets containing 41.4, 44.8, 49.8, 52.5, 55.1, and 64.0% nonfibrous carbohydrate. In addition, wheat replaced barley as the grain source during the dietary transition. Reticuloruminal pH was measured using orally administered pH measurement devices that were retrieved at slaughter. Data were analyzed using a mixed model including the fixed effects of sex, diet, and the 2-way interaction to evaluate the effect of diet and sex and with the fixed effects of sex, diet, and day relative to each dietary change along with the 2- and 3-way interactions to evaluate temporal responses as a result of diet change. A repeated measures statement was included for the effect of day. Both the mean and minimum reticuloruminal pH values decreased as the proportion of concentrate in the diet increased ( < 0.001). The area and duration that pH was <5.6 increased with greater inclusion of concentrate in the diet ( < 0.001). The number of cattle experiencing low reticuloruminal pH, defined as pH < 5.6 for >180 min, increased with increasing concentrate, and by the end of the 40-d dietary transition, 83% of the cattle had experienced at least 1 bout of low reticuloruminal pH, with most experiencing between 1 and 3 bouts/diet. These data are interpreted to suggest that cattle are at high risk for experiencing low reticuloruminal pH during the dietary transition but that the extent of low reticuloruminal pH is mild. Moreover, the data suggest that the risk for low reticuloruminal pH increases with increasing proportion of concentrate in the diet. The results also suggest that susceptibility to low reticuloruminal pH may differ between steers and heifers.

Structural, histochemical and immunocytochemical study of the forestomach mucosa in domestic ruminants.

The forestomach plays an important role in the digestion physiology of ruminants. The aim of this study is to clarify the morpho-functional role of the mucosa in each of the three compartments of the forestomach in three domestic ruminants species, viz cattle, buffalo and sheep, by means of structural, histochemical and immunocytochemical methods, including transmission electron microscopy, light microscopy and scanning electron microscopy. These methods were chosen to demonstrate the indirect evidence for the presence of nitric oxide (NO) employing NADPHd and nitric oxide synthase I (NOS I). The various cell layers of the forestomach epithelium are described and illustrated in detail. An intense NADPHd staining was observed in the granulosa, spinosa and basal layers of the epithelium, in particular in the cytoplasm over the nucleus. NOS I immunoreactivity was found in all specimens of the forestomach mucosa. The results of this study might reflect a possible role of NO in delaying the onset of cellular apoptosis in the forestomach mucosa of the domestic ruminants, by playing a role in the production of cell energy.

Expression of mRNA for proglucagon and glucagon-like peptide-2 (GLP-2) receptor in the ruminant gastrointestinal tract and the influence of energy intake.

Glucagon-like peptide-2 (GLP-2) is a potent trophic gut hormone, yet its function in ruminants is relatively unknown. Experiment 1 was conducted as a pilot study to establish the presence of GLP-2 in ruminants and to ascertain whether it was responsive to increased nutrition, as in non-ruminants. Concentrations of intact GLP-2 in the blood and gut epithelial mRNA expression of proglucagon (GCG) and the GLP-2 receptor (GLP2R) were measured in 4 ruminally, duodenally, and ileally cannulated steers. Steers were fed to meet 0.75 x NE(M) for 21 d, and then increased to 1.75 x NE(M) requirement for another 29 d. Blood samples and ruminal, duodenal, and ileal epithelium biopsies were collected at low intake (Days -6 and -3), acute high intake (Days 1 and 3), and chronic high intake (Days 7 and 29) periods. Experiment 2 investigated the mRNA expression pattern of GCG and GLP2R in epithelial tissue obtained from the forestomachs (rumen, omasum, and abomasum) and intestines (duodenum, jejunum, ileum, and colon) of 18 forage-fed Angus steers (260 kg BW). In Experiments 1 and 2, real-time polymerase chain reaction showed that expression of GCG and GLP2R mRNA was detectable in forestomach tissues, but expression was greater (P < 0.001) in small intestinal and colon tissue. High energy intake tended (P = 0.07) to increase plasma GLP-2 during the acute period and was paralleled by a 78% increase (P = 0.07) in ileal GCG mRNA expression. After this initial adaptation, duodenal GCG mRNA expression increased (P = 0.08) during the chronic high intake period. Duodenal GLP2R mRNA expression was not affected by energy intake, but ileal GLP2R expression was increased after 29 d of high energy intake compared to both the low and acute high intake periods (P = 0.001 and P = 0.01, respectively). These data demonstrate that cattle express GCG and GLP2R mRNA primarily in small intestinal and colon tissues. Increased nutrient intake increases ileal GCG mRNA and plasma GLP-2, suggesting that GLP-2 may play a role in the trophic response of the ruminant gastrointestinal tract to increased feed intake.

Different responses in postprandial plasma ghrelin and GH levels induced by concentrate or timothy hay feeding in wethers.

Ghrelin and growth hormone (GH) play a key role in regulating energy balance, metabolic hormone secretion and food intake. Ghrelin and GH responses to dietary compositions have not yet been fully clarified, although there may be significant relationships between dietary compositions and ghrelin and GH responses. In the present study, therefore, we assessed whether dietary compositions influence postprandial plasma ghrelin and GH levels in wethers. Four wethers were respectively fed concentrate (C) or timothy hay (R) for 14 days. The levels of total digestive nutrients (TDN) and crude protein (CP) were adjusted to be at the same level. The basal ghrelin in both groups was rapidly and significantly decreased after feeding. Although the decline of ghrelin levels in C was greater and shorter than that in R, no significant difference was observed in the area under the curve (AUC) or in the incremental area. The plasma GH levels were also rapidly and significantly decreased after feeding in both groups and a significant difference was observed between the two groups for AUC of GH. Interestingly, the circadian changes in the plasma ghrelin levels were close to those in the GH levels in C, but this was not the case in R. These data suggest that dietary compositions influence postprandial plasma ghrelin and GH levels, and that these differences may be caused by several factors, including nutrients and ruminal fermentation.

Determination of ellagic acid in oak leaves and in sheep ruminal fluid by ion-pair RP-HPLC.

An isocratic ion-pair high-performance liquid chromatography (IP-RP-HPLC) method with UV detection was developed to identify and quantify ellagic acid (EA). This phenolic compound is widely distributed in the plants and is often present in the diet of ruminants. The method was validated and validation parameters were: linearity range 5-100 mg/L; correlation coefficient, 0.9995; mean recoveries (99.94 and 101.07%) and detection limit 1.4 mg/L. Method was applied for the determination of ellagic acid in oak leaves and in ruminal fluid from to a vitro ruminal system. The proposed method proved to be rapid and accurate and can be successfully used in ruminant nutrition studies.

Reticular groove and reticulum are innervated by myenteric neurons with different neurochemical codes.

The reticulum and the reticular groove are functional distinct compartments within the ovine forestomach. While the reticulum takes part in various motor functions, such as mixing, retaining, and rejecting the forestomach ingesta, the reticular groove serves mainly as a bypass between the esophagus and the abomasum. To accomplish these different tasks, the compartments develop specific motility patterns that are controlled by intrinsic neural circuits. In this study the intrinsic innervation by myenteric neurons was analyzed by quadruple immunohistochemistry against cholineacetyl transferase (ChAT), nitric oxide synthase (NOS), substance P (SP), and vasoactive intestinal peptide (VIP). Four neurochemically different subpopulations of myenteric neurons were found in the reticulum and the floor of the reticular groove: ChAT/-, ChAT/SP, NOS/-, and NOS/VIP. The neuronal proportions were calculated relative to all myenteric neurons. Neurons of the reticulum were mostly immunoreactive for ChAT (89% +/- 3%), whereas neurons adjacent to the reticular groove predominantly expressed a nitrergic phenotype (62% +/- 4%). ChAT-positive neurons were also immunoreactive for SP (ChAT/SP: 64% +/- 3% reticulum; 25% +/- 1% reticular groove) or were purely cholinergic (ChAT/-: 25% +/- 4% reticulum; 13% +/- 3% reticular groove). NOS-positive neurons colocalized VIP (NOS/VIP: 10% +/- 3% reticulum; 46% +/- 1% reticular groove) or none of the other neurotransmitters (NOS/-: 1% +/- 1% reticulum; 17% +/- 3% reticular groove). Analysis of the soma sizes revealed that in both compartments the nitrergic neurons were significantly larger than the cholinergic neurons. It is suggested that the specific neurochemical code in combination with a specific morphology leads to a precise regulation of the specialized tasks of the reticulum and reticular groove by subpopulations of myenteric neurons.

Analysis of first gastric compartment fluid collected via percutaneous paracentesis from healthy llamas.

OBJECTIVE: To evaluate the safety and efficacy of percutaneous paracentesis for fluid collection from the first gastric compartment of healthy llamas and to describe characteristics of that fluid. DESIGN: Prospective study. ANIMALS: 10 healthy adult llamas. PROCEDURE: Physical examinations were performed prior to sample collection and for 14 days afterwards. A CBC was performed prior to sample collection and 5 days later. A 16-gauge, 7.5-cm stainless steel needle, positioned approximately 20 cm caudal to the costochondral junction of the last rib, was pointed in a dorsocraniomedial direction and pushed through the abdominal wall into the lumen of the first gastric compartment. Fluid was aspirated and analyzed immediately for color, odor, consistency, pH, methylene blue reduction (MBR) time, protozoa, and bacteria. RESULTS: Fluid samples were obtained from 9 of 10 llamas. Mean volume was 4.1 ml, mean pH was 6.67, and mean MBR time was 173 seconds. Odor was slightly acidic, color was light brown-green to light yellow-green, and consistency was moderate. Small protozoa with variable iodine staining and gram-negative bacteria were commonly detected. With few exceptions, results of physical examinations and CBC remained within reference ranges. CLINICAL IMPLICATIONS: Fluid samples from the first gastric compartment can be successfully obtained by percutaneous paracentesis. Fluid characteristics were similar to those of fluid collected via orogastric tube in llamas and cattle.

Comparative study of forestomach digestion in llamas and sheep.

To compare digestion in the forestomach of llamas and sheep, the animals were fed four different diets: hay alone (H), low in nitrogen; the same hay with soybean meal (HS), with barley (HB) and with both soybean meal and barley (HSB). The sheep intakes were restricted to obtain about the same intake level in the two species. On average, the digestibilities of DM, OM and NDF were significantly higher in llamas: respectively, +2.7, 3.6 and 5.3 for the four diets. Added barley impaired hay digestion in the sheep, but very little in the llamas. The llamas retained nitrogen better than the sheep owing to very low urine losses. For hay alone, the retention time of digesta in the forestomach was higher in the llamas than in the sheep. In contrast, there was no difference between species for the other diets. The pH and ammonia levels were higher in llamas. In contrast, the SCFA levels were lower. In all cases the in-situ rate of digestion was greater in llamas. The low intake of llamas generally observed in the literature does not account for their better digestion. The stability of the two first compartment pH levels and an excellent cellulolytic activity are determining factors in the better digestion efficiency of plant cell walls in the llamas. However, higher NH3 levels were observed in llamas, although the urinary N excretion was lower.

[Comparative ontogenic analysis of the epithelium of the non-glandular stomach compartments of merino sheep]. / Estudio ontogenico comparativo de la lamina epitelial de los compartimentos gastricos no glandulares en ovinos merinos.

A total of 74 embryos and fetuses were used in a comparative analysis of the epithelium of the non-glandular stomach compartments of merino sheep during development. The mechanical protection showed by the tegumentary epithelium in the superficial layers of the rumen, reticulum and omasum is supported by a buffer system of neutral mucopolysaccharides secreted by the deeper strata. Neutral mucopolysaccharides first appeared in epithelial cells at 46 days of fetal life. Acid mucopolysaccharides, mucins, and mucoid compounds were not detected. Growth curves and formulas were constructed for the epithelial layers.

Forestomach acidosis in six New World camelids.

Forestomach acidosis was diagnosed in 2 llamas and 4 alpacas. All were young, group-housed, sexually intact males. Clinical signs included forestomach atony, lethargy, ataxia, diarrhea, and tachycardia. Forestomach distention was observed in only 1 llama. Clinicopathologic abnormalities included low forestomach fluid pH, hyperchloremia, hypokalemia, and metabolic acidosis. Although camelids differ from domestic ruminants in typical management practices and behavioral, anatomic, and physiologic characteristics, they are, nonetheless, susceptible to forestomach acidosis. Gastric fluid analysis was essential for an accurate diagnosis. Four of 6 camelids recovered after PO and IV treatment with alkalinizing agents and fluids, antibiotics, and thiamine.

Immunohistochemical localization of chondroitin sulfate in the forestomach of the sheep.

The localization of chondroitin sulfate glycosaminoglycans in the forestomach mucosa was examined by use of monoclonal mouse IgMs against chondroitin sulfate (CS-56 and 2H6). Under the light microscope, immunoreactive sites were observed in subepithelial regions and around blood vessels of the rumen, reticulum and omasum. In the reticular papillae and omasal papillae, large areas of core regions also reacted with the monoclonal antibodies. The cell surfaces of fibroblasts in the core region of the epithelium were intensely stained. Under the electron microscope, heavy staining of collagen fibrils beneath the basement membrane of the mucosal epithelium was observed. In addition, products of immunoreactions were localized at the abluminal surfaces of endothelial cells, in pinocytotic vesicles and on pits of the endothelium, as well as surrounding collagen fibrils. At the core regions of reticular papillae and omasal papillae, cell surfaces, interstitial area of collagen fibrils and elastin fibers were stained. Some of the rough endoplasmic reticulum in fibroblasts was also stained. These results suggest that chondroitin sulfate in the forestomach mucosa might serve as a charge barrier in the transport of charged molecules.

Light microscopic studies of the stomach of the lesser mouse deer (Tragulus javanicus).

The stomach of the lesser mouse deer was studied at the light microscopic level using histological and immunohistochemical methods. The stomach was clearly differentiated into rumen, reticulum including reticular groove, a small transition zone and abomasum. The mucosal surface of the rumen, reticulum and transition zone was lined with a stratified squamous epithelium and that of the abomasum with a simple columnar type. The epithelial keratinization was weak in the rumen, floor of the reticular groove and transition zone, while it was strong in the reticulum, especially on the tip of the reticulum papillae. Large sinusoidal capillaries were often present in the ruminal papillae. In the ruminal mucosa, a thin layer of alpha-smooth muscle actin immunoreactive cells was demonstrated by immunohistochemistry. The muscularis mucosae of the reticulum was continuous and well-developed. The transition zone appeared as a nonglandular area having many low mucosal folds and two layers of tunica muscularis. The abomasal mucosa consisted of cardiac, proper gastric and pyloric glands. Cells immunoreactive for bovine pepsinogen and bovine prochymosin antisera were demonstrated in the abomasum. It is suggested that the characteristic features observed might be adaptations to a relatively rapid passage and rapid absorption of the fermentation products. There is some evidence that the transition zone is not a part of either the floor of the reticular groove or the abomasum, suggesting a possible reevaluation of the term used for the reticulo-abomasal orifice in the mouse deer.

Ion chromatographic methods for the detection of starch hydrolysis products in ruminal digesta.

Dionex high-performance ion chromatographic methods were evaluated for separation and quantitation of plant sugars and starch digestion products in the ruminal digesta of cattle. Mono- and disaccharides were eluted from a Dionex CarboPac PA1 column with sodium hydroxide used isocratically or as a pH gradient. Maltooligosaccharides which had a degree of polymerization (DP) less than 30 glucose residues were eluted in 60 min by a sodium hydroxide eluent containing a sodium acetate gradient. Carbohydrates were detected amperometrically. Responses were linear (r2 greater than 0.99) for glucose, disaccharides and maltooligosaccharides (DP less than 8). Precipitation and solid-phase extraction methods were evaluated for clean-up of samples of feedstuffs, ruminal contents, and bacterial culture fluids. Perchloric acid precipitation hydrolyzed sucrose but did not affect recoveries of cellobiose, isomaltose or maltose. Ethanol in concentrations of 79 and 86% precipitated maltooligosaccharides having chain lengths larger than 14 and 9 glucose residues, respectively. Maltooligosaccharide recoveries from solid-phase extraction columns varied with maltooligosaccharide size and column packing. Recoveries were greater than 94% for short chains (DP less than 6) eluted from phenyl-substituted columns and variable for all oligosaccharides eluted from C18 columns. Applications of these methods are presented and include: (1) detection of sugars in ruminant feed, (2) monitoring changes in ruminal sugars after feeding and (3) monitoring changes in extracellular sugars and oligosaccharides in the culture fluids of the ruminal bacterium, Bacteroides ruminicola.