Functional interactions of manno-oligosaccharides with dietary threonine in chicken gastrointestinal tract. I. Growth performance and mucin dynamics.

1. An experiment was conducted to evaluate the interactive effects of manno-oligosaccharides (MOS; Bio-MOS®) and dietary threonine on the growth performance in relation to intestinal mucin dynamics in broiler chickens from 1 to 21 and from 22 to 35 d of age. Two concentrations of MOS (0 or 2 g/kg for d 1 to 21; and 0 or 1 g/kg for d 22 to 35) and three concentrations of threonine (0.0, 1.0 and 1.2 of National Research Council (NRC), 1994, recommendations) were included in the experimental diets for each age group. 2. Body weight gain was significantly lower in threonine-deficient birds compared with those fed on adequate or excess threonine diets. Positive interaction between MOS and threonine supplementation on body weight gain was apparent in all phases of growth due mainly to the significantly poorer performance of birds given excess threonine in the absence of MOS. 3. The duodenal and ileal adherent mucous thickness were reduced at 14 and 28 d in threonine-deficient birds. Nevertheless, MOS significantly increase duodenal adherent mucous thickness at 14 d and ileal mucous thickness at 14 and 28 d. At 14 d, a significant MOS and threonine interaction on the jejunal adherent mucous thickness was also noted in that there was no difference between adequate and excess threonine groups in the absence of MOS, but a significant increase with excess threonine and MOS supplementation. 4. Dietary threonine greatly influenced mucin synthesis at the translational stage with no effect on jejunal MUC2 gene expression. Conversely, MOS modulated the transcriptional stage of intestinal mucin synthesis by consistently up-regulating jejunal MUC2 gene expression which was independent of dietary threonine concentration. There were no significant interactions between threonine and MOS on all the goblet cell densities. However, there was a MOS and threonine interaction on the staining intensities of jejunal sulphomucins due mainly to the significantly lower staining intensities in birds fed excess threonine in the absence of MOS. 5. The ameliorative effect of MOS on the growth-suppressive effects of excess threonine is likely to be linked to its modulating effects on the intestinal mucin dynamics.

Functional interactions of manno-oligosaccharides with dietary threonine in chicken gastrointestinal tract. III. Feed passage rate.

1. A 3 × 2 factorial experimental design was used to investigate the interaction between threonine concentration (0.70, 1.0 and 1.3 of National Research Council (NRC), 1994, recommendations) and manno-oligosaccharides (0 and 2 g/kg) on feed passage rate in relation to intestinal microbial activities and crude mucin turnover. 2. There was no interaction between the effects of manno-oligosaccharides (MOS) and dietary threonine on total tract transit time. However, an interaction between MOS and threonine was apparent where increasing threonine in the absence of MOS led to a reduction in the mean retention time, but a trend in the opposite direction in the presence of MOS. The ileal mean retention time at deficient and adequate concentrations of threonine was also significantly shorter in the presence of MOS. 3. In the jejunum, dietary MOS interacted with threonine to increase the villus-to-crypt ratio with deficient and adequate concentrations of threonine but not with an excess. In the ileum, MOS had no effect on the villus-to-crypt ratio at the deficient and adequate concentrations of threonine but significantly increased the ileal villus-to-crypt ratio with an excess. 4. There were significant interactions between MOS and dietary threonine in their effects on ileal flow of crude mucin, with MOS supplementation increasing mucin concentration and output when threonine was adequate but not when deficient or in excess. 5. Neither MOS nor threonine affected volatile fatty acids and intestinal musculature. No effects of gut microflora or voluntary feed intake on feed passage rate was attributable to dietary threonine or MOS supplementation.

Functional interactions of manno-oligosaccharides with dietary threonine in chicken gastrointestinal tract. II. Mucosal development, mucin dynamics and nutrient utilisation.

1. A 3 × 2 factorial experimental design was used to investigate the interaction between threonine concentration (0.7, 1.0 and 1.3 of National Research Council (NRC), 1994, recommendations) and manno-oligosaccharides (MOS) supplemented at 0 and 2 g/kg on growth performance in relation to intestinal flow of crude mucins, mucosal development and nutrient utilisation. 2. There was no interaction between MOS and dietary threonine in any performance variable analysed, except for body weight gain during the period to 14 d of age, where body weight gain was significantly lower in birds fed excess threonine in the absence of MOS. Dietary MOS was also observed to significantly increase the body weight gain at deficient and adequate concentrations of threonine. 3. Dietary treatments had no significant effect on either the ileal external muscularis thickness or crypt depth. However, there was a MOS and threonine interaction in the ileal villus to crypt ratio and ileal crude mucin output with both being increased only at the adequate threonine concentration. 4. Dietary MOS tended to interact with threonine to increase the ileal uptake of D-glucose and L-threonine, but the effect was only apparent in birds fed on the deficient or excess threonine diet. There was no significant interaction between MOS and threonine on ileal digestibility of amino acids. Supplementation of MOS or increased dietary threonine significantly increased the apparent and standardised ileal digestibility of threonine. 5. Results from the current study indicate the possible link between the modulating effects of these supplements on intestinal mucosal development and mucin dynamics. This, in turn, may suggest a relatively higher proportion of mature enterocytes and absorptive area, which would be expected to improve the capacity for digestion and mucosal nutrient absorption.

Characterisation and response of intestinal microflora and mucins to manno-oligosaccharide and antibiotic supplementation in broiler chickens.

1. An experiment was conducted to characterise and evaluate, in comparison to zinc bacitracin (ZnB), the response of intestinal microflora and mucins to manno-oligosacchares (MOS, Bio-MOS(R), Alltech Biotechnology, Nicholasville, Kentucky, USA). 2. Supplementation of MOS and ZnB selectively increased the intensity of sulphomucins. As revealed by the plate culture method, MOS accelerated the maturation of gut microflora by promoting the growth of lactobacilli in the ileal mucosa and vice versa on ileal and caecal clostridia. Unlike MOS, ZnB suppressed the growth of intestinal bacteria, especially those of lactobacilli and clostridia. Use of T-RFLP further revealed that MOS increased the diversity of lactobacilli in the ileum and ileal mucosa but the opposite was observed for ZnB. It also appears that MOS and ZnB possessed a common property in differentially favouring the growth of certain Lactobacillus species. There was also evidence to show that both MOS and ZnB also increased the homogeneity of the gut microflora, possibly through the regulation of the overall gut bacterial communities. 3. Improvement in intestinal microbial homogeneity and mucin synthesis, coupled with the differential selections for certain specific beneficial bacterial species, may ultimately be proven to be the target mechanisms in the search for more effective alternatives to antibiotics.

Comparison of alternatives to in-feed antimicrobials for the prevention of clinical necrotic enteritis.

AIMS: The capacity for Lactobacillus johnsonii and an organic acid (OA) blend to prevent Clostridium perfringens-induced clinical necrotic enteritis (NE) in chickens was studied. METHODS AND RESULTS: Cobb 500 birds were allocated into six groups (n = 25 birds/pen, eight pens/treatment); Unchallenged, Challenged, Antimicrobial (zinc bacitracin (ZnB)/monensin), OA, probiotic Lact. johnsonii and probiotic sham (Phosphate-buffered saline). All birds were challenged with Eimeria spp. and Cl. perfringens except for unchallenged controls. Birds fed antimicrobials were protected from NE development as indicated by maintenance of body weight, low mortality and clostridium levels, and decreased intestinal macroscopic lesion scores compared to challenged controls (P < 0.05). Lactobacillus johnsonii-fed birds had reduced lesion scores, whilst OA-fed birds had decreased Cl. perfringens levels. Both Lact. johnsonii and OA-fed birds had improved feed efficiency between days 0 and 28 compared to challenged controls; however, mortality and body weights were not improved by either treatment. Microbial profiling indicated that the challenge procedure significantly altered the jejunal microbiota. The microbiota of antimicrobial-fed birds was significantly different from all other groups. CONCLUSIONS: Whilst Lact. johnsonii and OA altered specific intestinal parameters, significant protection against NE was not observed. SIGNIFICANCE AND IMPACT OF THE STUDY: Lactobacillus johnsonii and OA did not prevent NE; however, some improvements were evident. Other related treatments, or combinations of these two treatments, may provide greater protection.

Effect of potassium diformate on growth performance and gut microbiota in broiler chickens challenged with necrotic enteritis.

1. The effect of potassium diformate (KDF) on mortality, growth performance, microbial populations, pH and short chain fatty acid concentrations in the intestinal tract of broiler chickens challenged with necrotic enteritis was investigated. 2. An experiment with 1050 Cobb male broiler chickens was conducted from 1 to 35 d of age. There were 7 treatment groups: (1) unchallenged negative control, (2) unchallenged KDF (4.50 g/kg feed), (3) challenged negative control, (4) challenged positive control (100 ppm monensin and 45 ppm Zn-bacitracin), and (5) to (7) challenged KDF (inclusion rate 2.25, 4.50 and 6.75 g/kg feed, respectively). 3. The necrotic enteritis challenge caused significant growth depression except in the challenged positive control group. Inclusion of KDF in feed had no significant effect on weight gain or feed conversion ratio, neither under challenged nor unchallenged conditions. Necrotic-enteritis-related mortalities were reduced in response to the positive control feed and KDF at 2.25 and 4.50 g/kg rates, but with no effect at the 6.75 g/kg rate. 4. There was no effect of KDF on Clostridium perfringens numbers in jejunum during the necrotic enteritis challenge (d 15). Post challenge (d 35), 4.5 g/kg KDF reduced the number of C. perfringens and Enterobacteria in jejunum compared with the negative control group. 5. Intestinal pH was not affected by adding KDF to the feed. The challenge reduced jejunum pH compared with the unchallenged treatment groups and challenged positive control group. 6. Chickens fed the KDF diets had detectable concentrations of formic acid in the jejunum. There was a tendency towards higher concentrations of acetic acid and lactic acid in the small intestine of unchallenged birds, whereas challenged birds had higher concentrations of butyric acid in the caeca. 7. It is concluded that KDF holds promise as an agent to control necrotic enteritis in broiler chickens and it may be possible to enhance the efficacy of KDF by manipulating dietary properties that may influence the dissociation kinetics of KDF in the gut.

Intestinal function and gut microflora of broiler chickens as influenced by cereal grains and microbial enzyme supplementation.

A study was conducted to investigate the effect of the key cereal grains and a microbial enzyme supplement on broiler chicken performance, gut microflora and intestinal function. Ingestion of the barley-based diet was associated with low 28-day body weight, decreased feed intake and high FCR. The supplemental enzyme increased feed intake and weight gain of the chickens on a wheat-based diet. The pH of the gizzard and caecal contents varied with the grain type. Enzyme supplementation reduced ileal viscosity, particularly in birds that received the diet based on wheat. The birds on the barley-based diet had lower ileal digestibility of dry matter, protein and energy than those given maize and sorghum-based diets. The ileal digestibility of starch was increased by enzyme supplementation. Enzyme supplementation increased the number of total anaerobic bacteria in the gizzard of birds fed on sorghum and increased lactobacilli in the gizzard of those fed both sorghum and wheat. The birds fed the sorghum-based diet had the lowest counts of caecal total anaerobic bacteria and lactobacilli. Jejunal villus height and villus:crypt ratio of birds fed the barley-based diet were the lowest when compared with those fed the other diets. Enzyme application induced an increase in villus height and villus:crypt ratio of birds on wheat, crypt depth on barley and a reduction in crypt depth of chickens on the sorghum-based diets. The highest activity of maltase and the lowest activity of sucrase were observed in tissue from birds fed on maize and sorghum-based diets respectively. The differences in the performance of broilers on cereal grains could be explained by changes in intestinal morphology, enzyme activities and gut microflora as well as nutrient digestibility. The improved performance by supplemental enzyme in wheat-fed chickens was associated with beneficial changes in intestinal morphology and digesta viscosity.

Effects of dietary mannanoligosaccharide on growth performance, nutrient digestibility and gut development of broilers given different cereal-based diets.

A feeding trial was conducted to evaluate the effects of mannanoligosaccharide (MOS) on the growth performance, nutrient digestibility and gut development of broilers given a corn or a wheat-based diet over a 21-day experimental period. Dietary MOS improved the growth performance of birds given the wheat-based diet compared to that of birds given the corn-based diet during 7-21 days of age. In line with this, the ileal digestibility of starch was increased by MOS at 21 days of age. The addition of MOS modulated the development of gut microflora. From day 7 to day 21, the numbers of mucosa-associated coliforms along the small intestine were decreased; whereas the numbers of mucosa-associated lactobacilli were increased by MOS, regardless of the cereal type in the diets. Dietary MOS also reduced the counts of coliforms and Clostridium perfringens in the caeca of birds by 21 days of age. Villus height at the jejunum was not affected by MOS but the crypt depth and the muscularis thickness were reduced. The specific activities of maltase and alkaline phosphatase were increased in birds given the MOS-supplemented diet; whereas the development of leucine aminopeptidase was delayed by MOS. All these changes in the mucosal morphology and function were dependent on the type of cereal and/or the age of the birds.

Effects of mannanoligosaccharide and fructooligosaccharide on the response of broilers to pathogenic Escherichia coli challenge.

1. The effects of mannanoligosaccharide (MOS, Bio-Mos, Alltech Inc.) on the growth performance and digestive system, particularly gut microflora, were tested and compared with fructooligosaccharide (FOS, Raftilose P95, Orafti) using 1-d-old birds in an Escherichia coli challenge model. The experiment lasted for 3 weeks and zinc bacitracin (ZnB) was used as a positive control. 2. Dietary MOS had positive effects on body weight gain (BWG) or/and feed conversion efficiency (FCE) of the challenged birds compared to the negative control at the end of weeks 1 and 3. Similar results were obtained for ZnB treatment. In contrast, FOS supplementation improved only the BWG of the challenged birds at 21 d of age. Within the unchallenged birds, a large improvement in BWG was noticed for FOS treatment at the end of the experiment, with the BWG of birds on ZnB and MOS treatments being intermediate. The FCE of the unchallenged birds was not affected by the dietary additives. 3. The addition of MOS reduced the number of mucosa-associated coliforms in the jejunum of the challenged birds on d 7. On d 21, FOS tended to increase the number of jejunal mucosa-associated lactobacilli in both the challenged and unchallenged birds. The number of Clostridium perfringens in the gut lumen was reduced by only ZnB. 4. Dietary MOS reduced the jejunal crypt depth of birds on d 7, regardless of the challenge. The FOS supplement did not affect the gut morphology, however, the concentration of lactic acid in the ileum was increased and, depending on the challenge, the intestinal pH was decreased by FOS at different ages. 5. In conclusion, the effects of MOS or FOS on the composition and activities of gut microflora and mucosal morphology of birds were related to E. coli challenge as well as the age of birds, which may be involved in the observed different growth-improving effects of the tested dietary additives.

Effects of mannanoligosaccharide in broiler chicken diets on growth performance, energy utilisation, nutrient digestibility and intestinal microflora.

1. A study was undertaken to evaluate the effects of mannanoligosaccharide (MOS, Bio-MOS, Alltech Inc.) on the growth performance, energy utilisation, nutrient digestibility and intestinal microflora of birds given a sorghum-wheat based diet. Two MOS levels (1 and 2 g/kg) were included in the diet. 2. Inclusion of MOS at both levels in the diet improved the apparent metabolisable energy (AME) values of the diet. However, these effects were not as pronounced as those of zinc bacitracin (ZnB) treatment. Dietary ZnB also significantly improved the net energy value of the diet. No significant differences between the different levels of MOS were noticed in the growth performance, AME and net energy values of the diet. Compared to the negative control, inclusion of 2 g/kg MOS tended to improve feed conversion efficiency (FCE) in the starter phase. 3. Dietary MOS did not affect the apparent total tract digestibility of nutrients compared to the negative control. In contrast, ZnB significantly improved the protein digestibility and tended to increase the starch digestibility. The addition of MOS reduced the concentration of arabinose in the soluble non-starch polysaccharides (NSP) fraction in the excreta of birds; whereas, the concentrations of individual sugars in the insoluble NSP and free sugar fractions were increased by ZnB. 4. A decrease in the populations of lactobacilli and coliforms in the ileal and caecal lumen was observed for MOS and ZnB treatments. Correspondingly, pH and microbial fermentation in the gut was altered. The addition of MOS tended to reduce the coliform load at the gut mucosa. 5. Results from the current study suggest that MOS can improve the apparent energy utilisation of the diet and tend to improve FCE of birds in the first three posthatch weeks, which may be partly related to the modulatory effects of MOS on the gut microflora.

Effect of feed particle size and feed processing on morphological characteristics in the small and large intestine of pigs and on adhesion of Salmonella enterica serovar Typhimurium DT12 in the ileum in vitro.

A 2 x 2 factorial experiment with pigs was undertaken to investigate the effect of particle size (fine and coarse) and feed processing (pelleted and nonpelleted) on morphological characteristics in the small intestine, cecum, and colon of pigs and on the adhesion of Salmonella enterica serovar Typhimurium DT12 to the ileum in vitro. Ninety-six pigs (average BW = 33 +/- 7 kg) were fed the experimental diets. After 4 wk, 24 pigs were selected (six pigs per diet) and euthanized, and tissue samples were taken from the mid and distal small intestine, cecum, and distal colon. The effects of particle size and feed processing on villus height and crypt depth in the small intestine were minor. Feeding coarse diets increased (P = 0.05) the crypt depth in the colon. The crypt depth was 420 +/- 12 and 449 +/- 12 microm in pigs fed finely and coarsely ground feed, respectively. Pigs fed pelleted diets had a larger (P = 0.01) staining area for neutral mucins, as well as for acidic and sulfomucins on the villi of the distal small intestine than pigs fed nonpelleted diets. The area was 41, 46, and 33% larger for neutral, acidic, and sulfomucins, respectively. The mucin-staining areas of the crypts in the cecum and the colon were not affected by the experimental diets. Examination of lectin binding characteristics of the distal small intestine and the cecum did not reveal any differences between the experimental diets. Using a pig intestine organ culture model, Salmonella adhered less (P < 0.05) to the ileal tissue of pigs fed the nonpelleted diets than to those fed pelleted diets; the adherence was 60% less in these pigs. Results of this study suggest that pigs fed pelleted diets secrete mucins that are capable of binding Salmonella enterica serovar Typhimurium DT12 and thereby allowing for colonization. Therefore, pigs fed a nonpelleted diet are better protected against Salmonella infections than pigs fed a pelleted diet.

Effects of nondigestible oligosaccharides on Salmonella enterica serovar Typhimurium and nonpathogenic Escherichia coli in the pig small intestine in vitro.

An in vitro intestinal tissue model was developed for the investigation of bacterial association in the pig small intestine under different dietary regimes. In preliminary experiments, jejunal and ileal tissue was taken from Danish Landrace pigs fed standard diet and inoculated with either Salmonella or nonpathogenic Escherichia coli strains. Higher numbers of salmonellae associated with the ileal tissues, but the numbers did not reach significance. Hence, jejunal sections were inoculated with nonpathogenic E. coli and ileal sections were inoculated with salmonellae in the presence of mannose or commercial nondigestible oligosaccharides (NDO) at 2.5%. There was a significant decrease in E. coli associated with the jejunum in the presence of mannose (P < 0.05). Furthermore, in pigs fed a diet supplemented with commercial NDO at 4% there was a significant reduction in the numbers of E. coli in jejunal organ cultures of pigs fed the FOS diet (P < 0.05). There was a reduction, though not a significant one, in the association of Salmonella sp. to the ileal sections of pigs fed the commercial FOS diet. The feeding of commercial GOS or its addition to organ cultures did not affect E. coli or Salmonella numbers.