Ovine Gastrointestinal Tract Research Articles

Survival of the nematophagous fungus Duddingtonia flagrans to in vitro segments of sheep gastrointestinal tract

The nematophagous fungus Duddingtonia flagrans is used in integrated management of gastrointestinal nematodes in ruminants. The chlamydospores of the fungus, orally administered, pass through the segments of the ruminant digestive tract and, in the feces, capture the nematodes preventing their migration to grasslands. The drastic conditions of the gastrointestinal segments can negatively affect the fungus' biocontrol activity. The aim of this study was to assess the effect of in vitro conditions of the sheep's main gastrointestinal segments on the concentration, viability and nematode predatory ability of D. flagrans chlamydospores. The segments evaluated separately in vitro were the oral cavity, rumen, abomasum, and small intestine. The results showed that chlamydospores concentration was not affected by exposure to the different segments. The viability of the chlamydospores after exposure to the oral cavity (2.53 × 106 CFU/mL) and small intestine (1.24 × 105 CFU/mL) was significantly lower than its control treatment, with values of 6.67 × 106 CFU/mL and 2.31 × 105 CFU/mL respectively. Nematode predatory ability after rumen exposure was reduced by 7% compared to the control treatment, by 25% after abomasum exposure and by 17% after small intestine. This study revealed the individual in vitro effect of each segment of ovine gastrointestinal tract on the integrity of this strain of the fungus D. flagrans affecting its viability and nematode predatory ability under the evaluated conditions. Delivery systems could be designed to protect chlamydospores considering the impact of each gastrointestinal segment.

Messenger RNA expression and localization of xenin in the gastrointestinal tract in sheep

The present study aimed to determine the primary sequence of ovine xenin and clarify the mRNA expression and peptide localization of xenin in the gastrointestinal tract in sheep. The colocalization of xenin and glucose-dependent insulinotropic polypeptide was also compared in the antrum and duodenum. Analysis of the nucleotide sequence of ovine xenin revealed a high degree (97.9%) of sequence homology of the sequence between sheep and cattle, and the amino acids sequence determined for ovine xenin coincided (100%) with that of other mammalian species. Real-time quantitative PCR for ovine xenin did not show regional difference in the mRNA expression ratio of xenin. In contrast to the real-time quantitative PCR results, anti-xenin positive cells were abundantly localized in the abomasal antrum (P < 0.01) and at a lesser amount in the duodenum, but no antixenin positive cells were observed in the other regions. Anti-xenin single-positive cells were in a majority in the abomasal antrum, whereas anti-xenin single-positive cells, and anti-GIP single-positive cells, and double-positive cells were even colocalized in the duodenum. These results suggest that abomasal antrum is a major source of xenin in the ovine gastrointestinal tract.

Eosinophil interactions with Haemonchus contortus larvae in the ovine gastrointestinal tract

Sheep were immunized by weekly oral infections with Haemonchus contortus for 9 weeks followed by anthelmintic treatment. They were challenged either 9 or 22 weeks later with PBS (sham controls) or one million exsheathed L3 surgically injected in the abomasum, and killed 24 h or 48 h later. Sheep challenged 9 weeks after immunization displayed varying degrees of tissue eosinophilia that showed a significant inverse relationship with the number of intra-epithelial mast cells (globule leucocytes). Close association of eosinophils with tissue larvae was observed mainly in the gastric pits (24 h) or on the mucosal surface (48 h). All L3-challenged sheep in this group had detectable globule leucocytes and tissue IL-4 mRNA, as measured by Southern blot RT-PCR. In contrast, sheep challenged 22 weeks after immunization had no detectable globule leucocytes or IL-4 mRNA and although they exhibited consistent tissue eosinophilia, eosinophils were not closely associated with tissue larvae. Scanning and transmission electron microscopy of sheep sensitized and rested for 9 weeks before challenge showed that L3 surrounded by eosinophils were at varying stages of damage and structural collapse. These studies strongly indicate that eosinophils can damage and probably kill gastrointestinal nematode larvae in vivo. In addition, they also suggest that effective killing by tissue eosinophils may depend on other microenvironmental factors such as intra-epithelial mast cells and IL-4.

Seasonal dynamics of antibiotic-resistantEnterobacteriaceae in the gastrointestinal tract of domestic sheep

Considerable variation in counts of antibiotic-resistant enterobacteria in the ovine gastrointestinal tract was observed. The occurrence of ruminal and fecal isolates resistant to ampicillin (Ap), kanamycin (Km) and tetracycline (Tc) culminated in summer months, followed by rapid decline in subsequent months. Using PCR the tem1bla (Apr), aphA1 (Kmr) and tetB (Tcr) genes were found to be predominant. Under in vitro conditions all resistance genes were transferable into laboratory Escherichia coli strain with relatively high frequency (10(-3) transconjugants per recipient).

Monocarboxylate transporter 1 gene expression in the ovine gastrointestinal tract

In this study, we investigated the tissue distribution and expression of monocarboxylate transporter 1 (MCT1) along the gastrointestinal tract of sheep. Western blot analysis suggested the presence of MCT1 as a 43-kDa protein in immunoblots of membranes from the various tissues examined. The results of Western blotting were further confirmed by immunohistochemical studies, which revealed intense immunoreactivity for the MCT1 protein in the forestomach (rumen, reticulum and omasum) and large intestine (caecum, proximal and distal colon). Moderate reactivity, however, was detected in the abomasum, while no immunoreactivity could be seen in any regions of the small intestine examined. Furthermore, MCT1 was expressed at the mRNA level as determined by reverse transcriptase polymerase chain reaction (RT-PCR), which showed a band of the expected size (300 bp) in all tissues examined. From these results we concluded that MCT1 protein is highly expressed and distributed in the stomach and large intestine of sheep suggesting that MCT1 may play a significant role in the transport of short chain fatty acids and their metabolites in the gastrointestinal tract of ruminants.

Serotypes and virulence gene profiles of shiga toxin-producing Escherichia coli strains isolated from feces of pasture-fed and lot-fed sheep.

Shiga toxin-producing Escherichia coli (STEC) strains possessing genes for enterohemolysin (ehxA) and/or intimin (eae), referred to here as complex STEC (cSTEC), are more commonly recovered from the feces of humans with hemolytic uremic syndrome and hemorrhagic colitis than STEC strains that do not possess these accessory virulence genes. Ruminants, particularly cattle and sheep, are recognized reservoirs of STEC populations that may contaminate foods destined for human consumption. We isolated cSTEC strains from the feces of longitudinally sampled pasture-fed sheep, lot-fed sheep maintained on diets comprising various combinations of silage and grain, and sheep simultaneously grazing pastures with cattle to explore the diversity of cSTEC serotypes capable of colonizing healthy sheep. A total of 67 cSTEC serotypes were isolated, of which 21 (31.3%), mainly isolated from lambs, have not been reported. Of the total isolations, 58 (86.6%) were different from cSTEC serotypes isolated from a recent study of longitudinally sampled healthy Australian cattle (M. Hornitzky, B. A. Vanselow, K. Walker, K. A. Bettelheim, B. Corney, P. Gill, G. Bailey, and S. P. Djordjevic, Appl. Environ. Microbiol. 68:6439-6445, 2002). Our data suggest that cSTEC serotypes O5:H(-), O75:H8, O91:H(-), O123:H(-), and O128:H2 are well adapted to colonizing the ovine gastrointestinal tract, since they were the most prevalent serotypes isolated from both pasture-fed and lot-fed sheep. Collectively, our data show that Australian sheep are colonized by diverse cSTEC serotypes that are rarely isolated from healthy Australian cattle.

Ontogenesis of peptide transport and morphological changes in the ovine gastrointestinal tract

The purpose of this study was to examine developmental and dietary regulation of the potential for peptide transport via PepT1 and morphological changes in the gastrointestinal tract of lambs. A 2×4 factorial arrangement of treatments with four blocks was created based upon gender, birth type (single or twin), birth weight, birth date, and breeding with 32 cross-bred lambs. Lambs were randomly allotted at birth to receive or not to receive a creep diet and all lambs were allowed to nurse. Sampling times of 2, 4, 6, or 8 weeks were randomly allotted to lambs. Samples for RNA extraction and histological evaluation were taken from the dorsal rumen, ventral rumen, omasum, duodenum, jejunum, and ileum. The DMI was similar for ewes nursing lambs that had or did not have access to creep feed (2,630 and 2, 574 g h −1 per day, respectively). Lambs with access to creep feed consumed DM at the rate of 2, 22, 129, 219, and 227 g h −1 per day, respectively when they were approximately 12, 13, 20, 38, and 52 days of age. Cumulative weight gain was similar for both groups of lambs and increased linearly ( P<0.001) with age. Villi were about 7% shorter ( P<0.09) in lambs receiving creep feed. Papillary height and width increased linearly ( P<0.01) with age. In the stomach, total and keratinized epithelial cells decreased ( P<0.03 and <0.01, respectively) with age and were fewer ( P<0.01) in lambs receiving creep feed. Even with the modest intakes observed in this study, creep feeding appeared to slightly alter the mucosal structure of the small intestine and was advantageous in that it stimulated papillae growth, thus predisposing the rumen for the introduction of feed into the diet. A 2.8 kb oPepT1 mRNA was present in all tissues studied by 2 weeks and age did not significantly influence the abundance of oPepT1 mRNA in the small intestine or stomach. Abundance of oPepT1 mRNA was greatest in the jejunum ( P<0.01) of the small intestine and in the dorsal rumen ( P<0.01) of the stomach. Lambs not receiving the creep diet had a greater ( P<0.02) abundance of oPepT1 mRNA in the rumen, particularly the dorsal rumen. Because no dry feed and little or no milk entered the rumen when no creep was fed, it is possible that a stimulus for development from the non-luminal direction, possibly blood-borne, may be involved in the ontogenesis of oPepT1. That PepT1 mRNA was present indicates that peptide transport occurs in the young lamb and the rumen and omasum appear to be involved in this process.

Dietary magnesium increases calcium absorption of ovine small intestine in vivo and in vitro.

The purpose of this study was to investigate whether or not an increase in dietary Mg intake increases Ca absorption in the ovine gastrointestinal tract. In an in vivo experiment, an increase in the infused MgCl2 level (0.0, 25.0 and 75.0 mg Mg x kg BW(-1) x day(-1) with 75.0 mg Ca x kg BW(-1) x day(-1) as CaCl2) into the rumen for ten days significantly decreased fecal excretion but increased urinary excretion (P < 0.05) of Ca in five castrated male sheep. Apparent Ca absorption tended to increase (P = 0.067) whilst the retention and plasma concentration of Ca were not changed. In an in vitro experiment with isolated segments from the rumen, upper jejunum, cecum and upper colon under the presence of an electrochemical gradient, the mucosal to serosal Ca flux rate was significantly greater in the presence of 60.0 mM as compared with 1.2 mM MgCl2 (P < 0.05). From these results, we conclude that the mucosal Mg has the ability to increase the Ca absorption in the gastrointestinal tract in sheep when the dietary Mg level is raised.

Antimicrobial Peptide Expression Is Developmentally Regulated in the Ovine Gastrointestinal Tract ,

Antimicrobial peptides are abundant components of the innate immune system present in species throughout the plant and animal kingdoms. In mammals, these immune peptides have been localized to epithelial tissues of the pig, mouse, rat, cow and human gastrointestinal tracts. We have identified in sheep two members of the β-defensin antimicrobial peptide gene family that are expressed in a unique pattern throughout the gastrointestinal tract. Sheep β-defensin 1 mRNA is the most prevalent from tongue to colon with the exception of the distal ileum, where β-defensin 2 mRNA predominates. Sheep β-defensin expression varies significantly between animals and is developmentally regulated both pre- and postnatally. These changes in antimicrobial peptide expression may correlate with anatomical differentiation as well as physiologic adaptations to extra-uterine life.

Effect of L364 718 on interdigestive pancreatic exocrine secretion and gastroduodenal motility in conscious sheep

The present study examined roles of endogenous cholecystokinin (CCK) and CCK-A receptors in the regulation of pancreatic exocrine secretion and gastroduodenal motility in conscious sheep during interdigestive period. Interdigestive exocrine secretion of ovine pancreas changed cyclically corresponding with cycle of duodenal migrating myoelectric complexes (MMC). During second phase of the duodenal MMC, intravenous injection of L364 718 at 2.45 μmol kg −1 inhibited exogenous CCK-8-induced pancreatic exocrine secretion. Intravenous infusion of the antagonist at 2.45 μmol kg −1/5 min for 5 min also inhibited significantly the pancreatic enzyme secretion without CCK-stimulation to half of that in the control, but not the fluid and bicarbonate secretion. Atropine infusion (i.v.) at 72.0 nmol kg −1/5 min significantly inhibited not only enzyme but also fluid and bicarbonate secretion. Corresponding to the inhibition of the exocrine secretion, L364 718 induced premature phase III in duodenal electromyogram (EMG) in three of the five sheep. Omasal EMG was inhibited slightly but significantly by L364 718, however, neither regular ruminal contractions nor abomasal EMG were altered by L364 718. In contrast, the atropine infusion inhibited only amplitude of ruminal contractions. These results suggest that endogenous CCK contributes to the regulation of interdigestive pancreatic exocrine secretion, omasal contractions and duodenal MMC in the ovine gastrointestinal tract via CCK-A receptors.

THE DISTRIBUTION OF VASOACTIVE INTESTINAL POLYPEPTIDE‐LIKE, SUBSTANCE P‐LIKE AND BOMBESIN‐LIKE IMMUNOREACTIVITY IN THE DIGESTIVE SYSTEM OF THE SHEEP

Peptides which are possibly related to the non-cholinergic, non-adrenergic division of the autonomic nervous system have been identified by immunofluorescence in the digestive system of mature sheep. Vasoactive intestinal polypeptide-, substance P-, and bombesin-like immunoreactivity were localized in neural elements throughout the ovine gastrointestinal tract (g.i.t.). Vasoactive intestinal polypeptide-like immunoreactivity (VIP-l.i.) was demonstrable in the submaxillary, parotid and the sublingual salivary glands close to small blood vessels and the acini. VIP-l.i. was also demonstrable in the upper oesophagus in connective tissue near small blood vessels. In the forestomachs, abomasum, and small and large intestines reactive fibres were present in the mucosa, submucosa, smooth muscle layers and the plexuses. The plexuses also contained reactive nerve cell bodies. VIP-reactive fibres were found in the pancreas, the gall bladder and the common bile and pancreatic duct but were not found in the intestinal mesentery, portal vein, and liver tissue. Substance P-like immunoreactivity (SP-l.i.) was demonstrable in nerve fibres in all the layers of the g.i.t. and in nerve cell bodies in the gut plexuses. The pancreas and the gall bladder also contained a few scattered fibres. Additionally, SP-l.i. was present in open-type endocrine cells throughout the mucosa of the small and large intestines but no SP-l.i. was found in the salivary glands or the oesophagus. Bombesin-like immunoreactivity (B-l.i.) was associated with nerve fibres and was demonstrable in the mucosa and myenteric plexuses throughout the g.i.t. B-l.i. in the smooth muscle appeared to be restricted to nerve fibres in the forestomachs, the abomasum, and the upper small intestine. No B-l.i. was found in the salivary glands, oesophagus, liver tissue, pancreas, gall bladder or intestinal mesentery.

Digestion and Absorption of Protein Along Ovine Gastrointestinal Tract

Digestion and absorption of protein were determined in ovine gastrointestinal tract with cerium-141 as an unabsorbed reference substance. Nitrogen flows changed little in rumen and reticulum, but in the proximal small intestine flows increased because of secretion of .9g nitrogen per day per kg body weight. This secretion included trypsin, chymotrypsin, elastase, and carboxypeptidases A and B; maximal activity was in proximal segments of the small intestine and decreased with distance from the pylorus. Activity of chymotrypsin decreased more rapidly than that of trypsin.Amino acid flows reflected the influx of protein in the duodenum; absorption was approximately 55% in the terminal ileum. No major changes of proportions of individual amino acids were observed. Overall nitrogen absorption was 72.6% of which 6% was in the large intestine.The major soluble protein fraction in the gastrointestinal tract consisted of peptides with molecular weight 7,000 to 14,000 daltons. Soluble high molecular weight protein was observed only in rumen and duodenum. Low molecular weight peptides and amino acids accumulated only in the proximal small intestine. Solubilization of protein and breakdown of peptides of 7,000 to 14,000 molecular weight appear to be rate limiting for protein absorption in sheep.

Movement and Absorption of Major Minerals and Water in Ovine Gastrointestinal Tract

Movement of calcium, chloride, magnesium, potassium, sodium, and water in the intestine of sheep was determined with cerium-141 as an unabsorbed reference substance. Net secretion of phosphorus and sodium, but not of potassium and calcium, was observed to the rumen. Water disappeared from the omasum and was secreted in the abomasum. Chloride secretion occurred in the omasum and more in the abomasum.Extensive secretion in the duodenum increased flows of sodium and potassium 8 to 9 times; of calcium, chloride, and phosphorus 3 to 6 times; of magnesium 1.3 times; and of water 12 times. Reabsorption was rapid in the small intestine, the major site of disappearance of all ions. Rates of disappearance decreased with increasing distance from the pylorus. Little further net disappearance of calcium, phosphorus, magnesium, or potassium was found in the large intestine, whereas water, sodium, and chloride disappearance continued. Overall absorption of sodium, potassium, and chloride was approximately 90%, of phosphorus 63%, of calcium 38%, and of magnesium 71%.This study highlights the importance of endogenous secretions in mineral absorption in the sheep.