Gastrointestinal Tract Of Ruminants Research Articles

Digestibility of Duddingtonia flagrans chlamydospores in ruminants: in vitro and in vivo studies

BackgroundThe use of Duddingtonia flagrans as a tool for the biological control of gastrointestinal nematodes (GIN) is a promising alternative to anthelmintics. The chlamydospores of D. flagrans are orally dosed and their thick cell wall gives them the capacity to resist digestion and pass through the gastrointestinal tract (GIT). Chlamydospores reaching the faeces are able to germinate and trap nematode larvae. The efficacy of this control method is based on reducing the numbers of infective larvae leaving the faeces. Techniques have recently been developed for quantifying the numbers of chlamydospores in faeces. As the number of non-digested spores could be relevant in the design and optimization of dosing programmes for the control of GIN infective larvae, the aim of the present study was to estimate the loss of D. flagrans chlamydospores during their passage through the ruminant gastrointestinal tract using in vitro and in vivo techniques.ResultsAfter in vitro rumen digestion, chlamydospore recovery was not different from the quantity originally incubated (undigested spores) (P > 0.05). In vitro rumen+abomasum digestion caused nearly 36% loss of the chlamydospores originally incubated (P < 0.05). Germination of chlamydospores classified as viable was 24.3%. Chlamydospores classified as non-viable did not germinate. Rumen digestion resulted in more spore germination (R1 = 35.7% and R2 = 53.3%) compared to no digestion (time 0 h = 8.7%). Subsequent abomasal digestion reduced germination (R1+A = 25%) or stopped it (R2+A = 0%). In vivo apparent chlamydospore digestibility in sheep showed a loss of 89.7% of the chlamydospores (P < 0.05).ConclusionsThe loss of chlamydospores was evident under in vitro and in vivo conditions. Negligible amounts of spores were lost during the in vitro rumen digestion. However, in vitro rumen+abomasum digestion resulted in a chlamydospore loss of approximately 36%. In vivo passage through the sheep GIT resulted in a total loss of 89.7% of the orally administered spores.

Tissue Expression of Fasting‐Induced Adipocyte Factor in Cattle

Fasting‐induced adipocyte factor (FIAF; also known as angiopoietin‐like 4) is a plasma protein that stimulates the oxidation of fatty acids and inhibits fat accumulation. The gastrointestinal tract appears to play an important role in regulating plasma FIAF concentration in some situations. Bovine FIAF shares 87% homology with the mouse protein. Our aim was to determine tissue expression of FAIF in the bovine. Rumen, omasum, abomasum, duodenum, jejunum, ileum, colon, pancreas, liver and adipose tissue samples were collected postmortem from 2 steers. FIAF mRNA was quantified by qPCR, and was most abundant in liver (420 arbitrary units; AU) and adipose tissue (320 AU). We detected FIAF mRNA throughout the gastrointestinal tract, with values ranging from 10 AU in the abomasum to 28 AU in the abomasum. By Western blot, FIAF protein was most abundant in liver and adipose tissue. However, FIAF protein was barely detectable in the rumen or duodenum, but was highly abundant in the omasum. Finally, cross‐sections of the rumen wall were used for immunohistochemistry staining, and strong signals for FIAF were detected in the epithelial layer. These findings suggest host/microbe interactions may influence FIAF expression in the ruminant gastrointestinal tract.

Absorption, Tissue Distribution, And Elimination of Residues after 2,4,6-Trinitro[14C]toluene Administration to Sheep

The compound 2,4,6-trinitrotoluene (TNT) is a persistent contaminant of some industrial and military sites. Biological bioremediation techniques typically rely on the immobilization of TNT reduction products rather than on TNT mineralization. We hypothesized that sheep ruminal microbes would be suitable for TNT destruction after phytoremediation of TNT-contaminated soils by cool-season grasses. Therefore we investigated the fate of [14C]TNT in ruminating sheep to determine the utility of ruminant animals as a portion of the bioremediation process. Three wether sheep were dosed with 35.5 mg each of dietary unlabeled TNT for 21 consecutive days. On day 22 sheep (41.9 +/- 3.0 kg) were orally dosed with 35.5 mg of [14C]TNT (129 microCi; 99.1% radiochemical purity). Blood, urine, and feces were collected at regular intervals for 72 h. At slaughter, tissues were quantitatively collected. Tissues and blood were analyzed for total radioactive residues (TRR); excreta were analyzed for TRR, bound residues, and TNT metabolites. Plasma radioactivity peaked within 1 h of dosing and was essentially depleted within 18 h. Approximately 76% of the radiocarbon was excreted in feces, 17% in urine, with 5% being retained in the gastrointestinal tract and 1% retained in tissues. Parent TNT, dinitroamino metabolites, and diaminonitro metabolites were not detected in excreta. Ruminal and fecal radioactivity was essentially nonextractable using ethyl acetate, acetone, and methanol; covalent binding of fecal radioactive residues was evenly distributed among extractable organic molecules (i.e., soluble organic matter, soluble carbohydrate, protein, lipid, and nucleic acid fractions) and undigested fibers (cellulose, hemicellulose, and lignin). This study demonstrated that TNT reduction within the ruminant gastrointestinal tract leads to substantial immobilization of residues to organic matter, a fate similar to TNT in other strongly reducing environments.

Preliminary X-ray characterization of a novel type of anchoring cohesin from the cellulosome of Ruminococcus flavefaciens.

Ruminococcus flavefaciens is an anaerobic bacterium that resides in the gastrointestinal tract of ruminants. It produces a highly organized multi-enzyme cellulosome complex that plays a key role in the degradation of plant cell walls. ScaE is one of the critical structural components of its cellulosome that serves to anchor the complex to the cell wall. The seleno-L-methionine-labelled derivative of the ScaE cohesin module has been cloned, expressed, purified and crystallized. The crystals belong to space group C2, with unit-cell parameters a = 155.6, b = 69.3, c = 93.0 A, beta = 123.4 degrees, and contain four molecules in the asymmetric unit. Diffraction data were phased to 1.95 A using the anomalous signal from the Se atoms.

Expression, cellular localization, and functional role of monocarboxylate transporter 4 (MCT4) in the gastrointestinal tract of ruminants

This is the first study to determine the precise cellular localization of monocarboxylate transporter 4 (MCT4), along with its co-existence with its chaperone, CD147 in the ruminant gastrointestinal tract. Quantitative Western blot analysis demonstrated that the abundance of MCT4 protein was in the order of forestomach > large intestine > abomasum ≥ small intestine. Immunohistochemistry and immunofluorescence confocal laser microscopy showed that MCT4 in the forestomach was confined to the cell membranes of strata corneum and granulosum, while diffuse cytoplasmic staining for MCT4 was visualized in strata spinosum and basale. In the epithelium cells lining the abomasum, MCT4 immunoreactive positivities were predominantly localized on the basolateral membranes. In the small intestine, MCT4 was localized at the brush borders and the basolateral membranes of the epithelial cells lining the villi, however it was mostly found on the apical membranes of the crypt cells. In the large intestine, the immunoreactivity for MCT4 differed between the surface epithelium and the crypts; in the surface epithelium, MCT4 was mainly localized at the apical membranes, whereas in the crypts it was predominantly expressed on the basolateral membranes of the lining epithelial cells. MCT4 was remarkably co-existed with CD147 along the bovine gastrointestinal tract. Our results suggest that MCT4 can play an important role in the transport of SCFA. The study also explored the potential functional collaboration between MCT1 and MCT4 and provided new insights into the mechanisms that mediate the transport of SCFA and other monocarboxylates in the different segments of the ruminant gastrointestinal tract.

Screening of Bacteria from the Cattle Gastrointestinal Tract for Inhibitory Activity against Enterohemorrhagic Escherichia coli O157:H7, O111:H−, and O26:H11

A quick and reproducible microgel plate assay was adapted to screen bacteria from cattle gastrointestinal tracts for production of compounds inhibitory to the growth of three enterohemorrhagic Escherichia coli (EHEC) serotypes: O157:H7, O111:H−, and 026:H11. The inhibitory activity of 309 bacteria, isolated on several agar media, was assessed by a microgel assay performed in 96-well microtiter plates. Fifty-three isolates secreted inhibitory compounds with a molecular weight of less than 1,000. In 12 isolates, the inhibitory activity was attributable to compounds other than lactic or acetic acid. These compounds were highly heat tolerant, with varying sensitivity to digestion by proteolytic enzymes. The inhibitory isolates were identified as lactic acid–producing bacteria on the basis of a combination of analyses, including 16S-rDNA restriction fragment length polymorphisms, 16S-rDNA gene sequences, and fermentation end products. The lactic acid bacteria of ruminants may contain antibacterial compounds not yet described. Naturally occurring populations of lactic acid bacteria may have potential as probiotics, to reduce the carriage of EHEC in the gastrointestinal tract of ruminants.

In Vivo Transduction of an Stx-Encoding Phage in Ruminants

We assessed the ability of a kanamycin-marked Stx phage to move into a commensal, ovine Escherichia coli strain in the ruminant gastrointestinal tract. Transduction was detected in 19/24 sheep tested, resulting in the recovery of 47 transductants. Subtherapeutic doses of the quinolone antibiotic enrofloxacin did not increase the rate of transduction.

Biotic and Abiotic Factors Influencing In Vitro Growth of Escherichia coli O157:H7 in Ruminant Digestive Contents

The gastrointestinal tract (GIT) of ruminants is the main reservoir of enterohemorrhagic Escherichia coli, which is responsible for food-borne infections in humans that can lead to severe kidney disease. Characterization of biotic and abiotic factors that influence the carriage of these pathogens by the ruminant would help in the development of ecological strategies to reduce their survival in the GIT and to decrease the risk of contamination of animal products. We found that growth of E. coli O157:H7 in rumen fluid was inhibited by the autochthonous microflora. Growth was also reduced when rumen fluid came from sheep fed a mixed diet composed of 50% wheat and 50% hay, as opposed to a 100% hay diet. In fecal suspensions, E. coli O157:H7 growth was not suppressed by the autochthonous flora. However, a probiotic strain of Lactobacillus acidophilus inhibited E. coli O157:H7 growth in fecal suspensions. The inhibitory effect was dose dependent. These lactic acid bacteria could be a relevant tool for controlling O157:H7 development in the terminal part of the ruminant GIT, which has been shown to be the main site of colonization by these pathogenic bacteria.

Intestinal Protein Supply Alters Amino Acid, but Not Glucose, Metabolism by the Sheep Gastrointestinal Tract

This study was intended to establish the extent which amino acids (AAs) and glucose are net metabolized by the gastrointestinal tract (GIT) of ruminant sheep when intestinal protein supply is varied. Wether sheep (n = 4, 33 +/- 2.0 kg) were fitted with catheters for measurement of net absorption by the mesenteric (MDV) and portal-drained (PDV) viscera and a catheter inserted into the duodenum for casein infusions. Sheep received a fixed amount of a basal diet that provided adequate metabolizable energy (10.9 MJ/d) but inadequate metabolizable protein (75 g/d) to support 300-g gain per day. Four levels of casein infusion [0 (water), 35, 70, and 105 g/d], each infused for 5.5 d, were assigned to sheep according to a 4 x 4 Latin square design. [methyl-(2)H(3)]leucine was infused (8 h) into the duodenum while [1-(13)C]leucine plus [6-(2)H(2)]glucose were infused (8 h) into a jugular vein. With the exception of glutamate and glutamine, net absorption of AAs increased linearly (P < 0.05, R(2) = 0.46-1.79 for MDV; P < 0.05, R(2) = 0.6-1.58 for PDV) with casein infusion rate. Net absorption by the PDV accounted for <100% of the additional supplies of leucine, valine, and isoleucine (0.6-0.66, P < 0.05) from casein infusion, whereas net absorption by the MDV accounted for 100% of the additional essential AA supply. Glucose absorption (negative) and utilization of arterial glucose supply by the GIT remained unchanged. There was a positive linear (P < 0.05) relation between transfer of plasma urea to the GIT and arterial urea concentration (MDV, P < 0.05, r = 0.90; PDV, P < 0.05, r = 0.93). The ruminant GIT appears to metabolize increasing amounts of the branched-chain AAs and certain nonessential AAs when the intestinal supply of protein is increased.

Uso da cutina na estimativa das digestões total e parcial de alguns componentes de rações contendo diferentes fontes de nitrogênio, em bovinos

Seis novilhos Holandês x Zebu canulados no rúmen e no abomaso foram alimentados com rações contendo dois níveis de casca de soja e três fontes protéicas para determinação das digestões total e parcial no rúmen e nos intestinos. A cutina foi utilizada como indicador interno para se quantificarem a MS da digesta abomasal e a matéria seca fecal. Após 15 dias de adaptação dos animais às rações e às condições experimentais, foram coletadas 12 amostras por animal e por período das digestas do abomaso e das fezes. Foram determinados os teores de cutina, MS, MO, PB, FDN, FDA e CNF nas amostras compostas por animal e por período experimental. Não foram observados efeitos das fontes protéicas sobre as quantidades de MS, MO, FDN, FDA e PB no rúmen, no abomaso e no intestino. A amiréia proporcionou maiores ingestões de CNF e menores coeficientes de digestibilidades da MS, FDN, FDA e de CNF que as demais fontes protéicas. À exceção da PB, o teor de casca de soja afetou positivamente a digestão desses componentes no trato digestivo total e no rúmen. Os resultados indicaram potencial da cutina como indicador da taxa de passagem dos componentes nutritivos ao longo do trato digestivo.

Expression of mRNA for sodium‐glucose transporter 1 and fatty acid translocase in the ruminant gastrointestinal tract before and after weaning

ABSTRACTThe mRNA expression of sodium‐glucose transporter 1 (SGLT1) and fatty acid translocase (CD36) in the gastrointestinal tract of Holstein cattle and Saanen goats before and after weaning was investigated. Before weaning, the expression of both SGLT1 and CD36 was highest in the jejunum, relative to the other parts of the gastrointestinal tract in both species. The expression of SGLT1 and CD36 in the duodenum was second highest in the goats. After weaning, SGLT1 and CD36 expression in the small intestine significantly decreased in both species. The expression of both types of transporters was also detected in the forestomach. From these results, it was concluded that the jejunum is probably the major absorption site for glucose and long‐chain fatty acids before weaning, and that the expression of both types of transporters decreases after weaning in cattle and goats.

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.

Characterization of Streptococcus bovis from the rumen of the dromedary camel and Rusa deer.

Isolation and characterization of Streptococcus bovis from the dromedary camel and Rusa deer. Bacteria were isolated from the rumen contents of four camels and two deer fed lucerne hay by culturing on the semi-selective medium MRS agar. Based on Gram morphology and RFLP analysis seven isolates, MPR1, MPR2, MPR3, MPR4, MPR5, RD09 and RD11 were selected and putatively identified as Streptococcus. The identity of these isolates was later confirmed by comparative DNA sequence analysis of the 16S rRNA gene with the homologous sequence from S. bovis strains, JB1, C14b1, NCFB2476, SbR1, SbR7 and Sb5, from cattle and sheep, and the Streptococcus equinus strain NCD01037T. The percentage similarity amongst all strains was >99%, confirming the identification of the camel isolates as S. bovis. The strains were further characterized by their ability to utilize a range of carbohydrates, the production of volatile fatty acids (VFA) and lactate and the determination of the doubling time in basal medium 10 supplemented with glucose. All the isolates produced l-lactate as a major fermentation end product, while four of five camel isolates produced VFA. The range of carbohydrates utilized by all the strains tested, including those from cattle and sheep were identical, except that all camel isolates and the deer isolate RD11 were additionally able to utilize arabinose. Streptococcus bovis was successfully isolated from the rumen of camels and deer, and shown by molecular and biochemical characterization to be almost identical to S. bovis isolates from cattle and sheep. Streptococcus bovis is considered a key lactic acid producing bacterium from the gastrointestinal tract of ruminants, and has been implicated as a causative agent of lactic acidosis. This study is the first report of the isolation and characterization of S. bovis from the dromedary camel and Rusa deer, and suggests a major contributive role of this bacterium to fermentative acidosis.

Thermoregulation of the Escherichia coli O157:H7 pO157 ecf operon and lipid A myristoyl transferase activity involves intrinsically curved DNA.

Escherichia coli O157:H7 survives in diverse environments from the ruminant gastrointestinal tract to cool nutrient-dilute water. We hypothesized that the gene regulation required for this flexibility includes intrinsically curved DNA that responds to environmental changes. Three intrinsically curved DNAs were cloned from the E. coli O157:H7 virulence plasmid (pO157), sequenced and designated Bent 1 through Bent 3 (BNT1, BNT2 and BNT3). Compared to BNT1 and BNT3, BNT2 had characteristics typical of intrinsically curved DNA including electrophoretic gel retardation at 4 degrees C, six partially phased adenine:thymine tracts and transcriptional activation. BNT2::lacZ operon fusions showed that BNT2 activated transcription at 24 degrees C compared to 37 degrees C and was partially repressed by a bacterial nucleoid-associated protein H-NS. BNT2 regulated the E. coli attaching and effacing gene-positive conserved fragments 1-4 (ecf1-4) that are conserved in Shiga toxin-producing E. coli associated with human disease. Experimental analyses showed that ecf1-4 formed an operon. ecf1, 2 and 3 encoded putative proteins associated with bacterial surface polysaccharide biosynthesis and invasion and ecf4 complemented a chromosomal deletion of lpxM encoding lipid A myristoyl transferase. Mass spectrometric analysis of lipid A from ecf and lpxM single and double mutants showed that myristoylation was altered at lower temperature.

Intake and digestibility of untreated and urea treated rice straw base diet fed to sheep

Rice straw as one of agricultural by-products has low quality due to low content of essensial nutrients like protein, energy, minerals and vitamin as well as poor palatability and digestibility. Therefore, the quality of rice straw needs to be improved in order to increase its utilization by gastrointestinal tract of ruminants. The purpose of this study is to compare untreated and urea treated rice straw as basal diets for sheep. Twelve mature Merino wethers (average body weight 53.62 + 3.44 kg) were separated into 4 groups based on their live weight with each groups assigned three diets, that are: diet 1 untreated rice straw with high forage legume content, diet 2 urea ensiled rice straw and diet 3 rice straw sprayed with urea solution at feeding time. Diets were allocated based on a randomized complete block design. Urea ensiled rice straw was prepared by spraying chopped straw with urea solution to yield straw containing 4% urea and 40% moisture, then kept in air tight polythylene bags for 6 weeks. The untreated, ensiled and urea supplemented rice straw were mixed with other feed ingredients to provide isoenergetic and isonitrogenous diets. Diets were formulated to meet maintenance requirement according to NRC. Sheep were adapted to experimental diets for 15 days, and after adaptation period, a metabolism trial was conducted. Results reveal that dry matter intake permetabolic body weight (DMI/W0.75), DE (digestible energi) intake and apparent digestibility of NDF (neutral detergent fibre) were not significantly different between diet 1 and diet 2. Apparent digestibility of DM (dry matter), OM (organic matter), and ADF (acid detergent fibre), as well as N retention were not significantly different between three diets. Positive result in N retention was only observed in diet 2, while others were negative. It may be concluded from this study that untreated rice straw basal diet supplemented with forage legume offer an alternative method other than urea ensiled for improving the nutritional value of rice straw as a ruminant feed on small farmer. Key words : Rice straw, urea treated, digestibility, and sheep

Aflatoxin binders I: in vitro binding assay for aflatoxin B1 by several potential sequestering agents.

Nine potential proprietary sequestering agents consisting of 4 activated charcoals, 3 sodium bentonites, a calcium bentonite, and an esterified glucomannan were compared in a novel in vitro assay for aflatoxin B1 (AFB1) binding. Agents were evaluated in 10% methanol prepared as 1% stirred suspensions at pH 3, 7, 10 and pH-unadjusted, with or without AFB1 at 5 microg/ml. All nine agents bound more than 95% of the 5 microg of AFB1 in solution, regardless of pH. The sodium bentonites bound 98, 95, and 98% of the AFB1. The four activated charcoals bound over 99%, the calcium bentonite bound 98%, and the esterified glucomannan bound 97% of the AFB1 in solution. The results suggested that the sequestering agents tested here had sufficient potential to bind AFB1 at pH values commonly found in the gastrointestinal tracts of ruminants and other animals.

Validity of specifically applied rare earth elements and compartmental models for estimating flux of undigested plant tissue residues through the gastrointestinal tract of ruminants

Validity of specifically applied rare earth elements and compartmental models for estimating flux of undigested plant tissue residues through the gastrointestinal tract of ruminants

Effect of diet on Shiga toxin-producing Escherichia coli (STEC) growth and survival in rumen and abomasum fluids.

The gastrointestinal tract of ruminants is the main reservoir for Shiga toxin-producing Escherichia coli (STEC) strains, potentially pathogenic for humans. We used for the first timerumen fluid in which no exogenous carbon source or other supplement was added to compare acid resistance and growth of STEC in physiological physico-chemical conditions. We showed that acidic conditions resulting from the combination of high volatile fatty acid concentration and moderately acidic pH did not alter the survival of STEC, and that human non-O157:H7 STEC isolates were able to persist in the rumen contents in spite of acid stress, low oxygen availability and nutrient deprivation, in the same manner as bovine STEC isolates do. Furthermore, our results support the hypothesis that a grain-rich diet may induce mechanisms of STEC acid resistance in the rumen that allow STEC survival in the abomasum.

Validity of specifically applied rare earth elements and compartmental models for estimating flux of undigested plant tissue residues through the gastrointestinal tract of ruminants.

The validity of using rare earth elements as flow markers of undigested residues was evaluated by comparing mean gastrointestinal residence time (GMRT) of rare earths specifically applied to cottonseed hulls (CSH) to that of the indigestible fiber of CSH. Feces were collected from five lambs fed a mineral supplemented diet of CSH containing 52 g CP/kg DM and five lambs fed a CSH plus cottonseed meal diet (CSH+CSM) containing 123 g CP/kg DM. Rare earth elements (La, Yb, and Tb) specifically bound to CSH were included in the diet for a 5-d period and then deleted from the diet for a 3-d period. Following the last fecal collection, lambs were slaughtered for collection of digesta from segments of the gastrointestinal tract. Potentially indigestible NDF (PIF) was determined in diets and digesta from each segment of the gastrointestinal tract. Mean turnover rate, time delay, and GMRT for each rare earth element was estimated by fitting an age-dependent compartment model to profiles of markers appearing in the feces (compartmental model-marker method, CMM). The GMRT also was computed by the indigestible entity pool dilution method (IEPD) as grams of PIF in sampled segment/mean intake rate of PIF proceeding slaughter, g/h. The GMRT computed by the CMM and the IEPD methods did not significantly (P < 0.05) differ (99.6 vs 94.8 h and 58.9 vs 59.5 h for CMM vs IEPD and CSH and CSH+CSM diets, respectively). Regression of GMRT estimated for rare earths vs PIF yielded a highly significant regression (P = 0.001) with a regression coefficient of 0.94 +/- 0.016. It was concluded that rare earth elements applied to specific feeds are valid flow markers for the undigested residues derived from such marked feeds.

Genomic and Proteomic Analysis of Microbial Function in the Gastrointestinal Tract of Ruminants - Review -

Genomic and Proteomic Analysis of Microbial Function in the Gastrointestinal Tract of Ruminants - Review -