Effects of a feed additive blend on broilers challenged with heat stress.

We evaluated a blend of medium-chain fatty acids (MCFA), organic acids, and a polyphenol antioxidant on gut integrity. Eighty Ross Broilers were exposed to 20-22°C (control - normothermic) or to 35-39.5°C (heat stress) for eight hours a day for a period of 1 or 5 days. Birds were fed a standard diet, or a diet supplemented with the test blend. Thereafter, birds were euthanized, and intestinal sections were excised for morphological, morphometric and gene expression analyses. Blood samples were collected for glucose-6-phosphate dehydrogenase (G6PD), glutathione peroxidase (GSH-Px) activity and trolox equivalent antioxidant capacity (TEAC) determination. Heart and liver tissues were used to quantify the expression of heat shock proteins 60 and 70 (HSP60 and HSP70, respectively) and inhibitor of kappa light chain gene enhancer in B cells alpha (IKBA). The jejunum was the most sensitive intestinal section, where heat stress modulated the expression of HSP70, of the inflammatory markers IKBA, interleukin 8 (IL-8), interferon gamma (IFNγ), and toll-like receptor 4 (TLR4). Moreover, expression of tight junctions (CLDN1, ZO1 and ZO2) and nutrient transporters (PEPT1 and EAAT3) was modulated especially in the jejunum. In conclusion, the feed additive blend protected intestines during heat stress from the decrease in villus height and crypt depth, and from the increase in villus width. Especially in the jejunum, heat stress played an important role by modulating oxidative stress and inflammation, impairing gut integrity and nutrient transport, and such deleterious effects were alleviated by the feed additive blend. RESEARCH HIGHLIGHTS Jejunum is the most sensitive intestinal segment during heat stress. Heat stress affects the expression of tight junctions and nutrient transporters. Feed management helps to alleviate the disturbances caused by heat stress. A blend of MCFA, organic acids and a polyphenol protects broilers under heat stress.

Quantitative histo-morphometric analysis of heat-stress-related damage in the small intestines of broiler chickens.

The aim of the current research was to present a methodological approach allowing reproducible morphometric and morphological (Chiu/Park scale) analyses of the alterations in the intestines of broilers exposed to heat stress. Ross broilers were exposed over four consecutive days to a high-temperature regime in controlled climate rooms, with a day temperature of 39°C (±1°C) and a night temperature of 25°C (±1°C), respectively. A control group was kept at an ambient temperature of 25°C (±1°C) during the entire experimental period. At the end of the exposure period, the birds were sacrificed and specimens were taken of the duodenum, jejunum and ileum for histology. Blood was collected for oxidative stress analysis. Histo-morphological and morphometric analyses of the intestines indicated that the duodenum and jejunum showed more damage than the ileum. The major alterations in the control intestines were limited to the villus tips, while heat stress led to villus denudation and crypt damage. When compared with morphologically normal villi, heat-stress-associated alterations were also observed in villus height (decreased), villus breadth at base (increased) and epithelial cell area (decreased). Birds exposed to heat stress presented with an increase in glutathione peroxidase activity and a decreased antioxidant capacity. It can be concluded that the chosen model allows a reproducible quantification of heat stress effects, which is suitable for the evaluation of dietary intervention strategies to combat heat stress conditions.

Phytase in non-ruminant animal nutrition: a critical review on phytase activities in the gastrointestinal tract and influencing factors.

This review focuses on phytase functionality in the digestive tract of farmed non-ruminant animals and the factors influencing in vivo phytase enzyme activity. In pigs, feed phytase is mainly active in the stomach and upper part of the small intestine, and added phytase activity is not recovered in the ileum. In poultry, feed phytase activities are mainly found in the upper part of the digestive tract, including the crop, proventriculus and gizzard. For fish with a stomach, phytase activities are mainly in the stomach. Many factors can influence the efficiency of feed phytase in the gastrointestinal tract, and they can be divided into three main groups: (i) phytase related; (ii) dietary related and (iii) animal related. Phytase-related factors include type of phytase (e.g. 3- or 6-phytase; bacterial or fungal phytase origin), the pH optimum and the resistance of phytase to endogenous protease. Dietary-related factors are mainly associated with dietary phytate content, feed ingredient composition and feed processing, and total P, Ca and Na content. Animal-related factors include species, gender and age of animals. To eliminate the antinutritional effects of phytate (IP6), it needs to be hydrolyzed as quickly as possible by phytase in the upper part of the digestive tract. A phytase that works over a wide range of pH values and is active in the stomach and upper intestine (along with several other characteristics and in addition to being refractory to endogenous enzymes) would be ideal.

Validation of a dual in vivo-in vitro assay for predicting the digestibility of nutrients in humans.

BACKGROUND: The validation of a dual in vivo-in vitro digestibility assay ('dual digestibility assay') for separately predicting the upper-tract, hindgut and total tract digestibility of nutrients in humans, as estimated using organic matter digestibility (OMD), is described. Human upper-tract OMD was predicted using an animal (rat) model with digesta from the terminal ileum collected from rats fed one of four complete human diets (wheat bran diet, pectin diet, mixed low-fibre diet, mixed high-fibre diet). Large intestinal OMD was predicted using an in vitro hindgut fermentation assay employing a human faecal inoculum and with the rat ileal digesta as the substrate. RESULTS: A comparison of total tract OMD of the four diets from a human balance study (OMDhuman ) with that predicted using the dual digestibility assay (OMDdual ) showed no significant differences (P > 0.05). OMDhuman and OMDdual were highly correlated (r = 0.953, P = 0.047). CONCLUSION: The dual digestibility assay accurately predicts the uptake of dietary nutrients (as grams of organic matter) in humans over the total tract. The assay is able to separately quantify the digestibility of nutrients in the upper and lower digestive tracts. The validation of the dual digestibility assay needs to be extended to a wider range of human diets.

NMR-based metabonomics detection of differences in the metabolism of hydrolysed versus intact protein of similar amino acid profile.

BACKGROUND: Proton nuclear magnetic resonance (NMR)-based metabonomics has only recently been applied to nutritional research. The limitation of any analytical technique is its sensitivity in detecting the smallest variation. Alterations in nutrition often produce only subtle metabolic modulations. The objective of this study was to determine if NMR-based metabonomics could detect variations in the metabolic profile of urine from pigs digesting either native casein (NC) or the same casein that had been enzymatically hydrolysed (EHC). NMR permits simultaneous detection of a large number of metabolites, thus allowing detection of unanticipated metabolic fluctuations that may otherwise have gone undetected with the use of only targeted analysis. RESULTS: Partial least squares discriminant analysis identified significantly (P < 0.05) higher urinary excretions of leucine, valine, taurine and glycine by pigs on the EHC-based diet. CONCLUSION: NMR-based metabonomics is a sensitive method that can uncover unanticipated metabolic changes brought about by physicochemical changes to the feedstock (i.e. hydrolysis). The data show a lower efficiency of retention by the kidney of some amino acids following ingestion of a hydrolysed protein.

Predicted apparent digestion of energy-yielding nutrients differs between the upper and lower digestive tracts in rats and humans.

The apparent digestibility of energy-yielding nutrients (carbohydrate, protein, and fat) was predicted in the human upper digestive tract and large bowel separately for 4 diverse diets containing either a single dietary fiber source [wheat bran and pectin (PE) diets] or mixed fiber sources [low-fiber (LF) and high-fiber (HF) diets). A human balance study was undertaken to determine fecal energy and nutrient excretion and a rat model was used to predict human ileal energy and nutrient excretion. Total tract energy digestibility ranged from 92 (HF diet) to 96% (PE diet and LF diet), while at the ileal level it ranged from 79 to 86% for the HF diet to the LF diet. The predicted upper-tract digestion of starch, sugars, and fat was high, with ileal digestibilities exceeding 90% for all diets. Nonstarch polysaccharides were poorly digested in the upper tract for all diets except in the PE diet. The daily quantity of protein excreted at the ileal level was between 2 (HF diet) and 5 (PE diet) times higher than that at the fecal level. The large differences between fecal and ileal nutrient loss highlight that fecal digestibility data alone provide incomplete information on nutrient loss. There is a need to be able to routinely determine the uptake of energy in the upper and lower digestive tracts separately.

The influence of whey protein and glycomacropeptide on satiety in adult humans.

Protein is often considered the most satiating macronutrient. The objective was to determine the short-term effect of mixtures of whey protein and glycomacropeptide (GMP) versus a carbohydrate control on satiety in healthy adult humans. The study was a randomised crossover Latin Square design. On 4 separate days, fifty healthy subjects (19 males and 31 females) received a subject-specific breakfast (08:00 h), a preload drink (12:00 h) and lunch (12:30 h). The preload drink was presented as a milkshake with either maltodextrin carbohydrate (control), whey protein isolate (WPI) with no GMP, WPI with naturally present 21% GMP or WPI with naturally present 21% GMP plus added GMP. Satiety was assessed using visual analogue scales (VAS) and by determining ad libitum food intake during a cafeteria style meal offered 30 min after the preload. The VAS indicated that the lower GMP treatment induced a greater feeling of fullness immediately after consumption of the preload compared with the other treatments. Energy and macronutrient intake at lunch did not differ significantly (p>0.05) between treatments although subjects chose to eat foods higher in carbohydrate and lower in protein after the protein preloads. Women consumed the least amount of protein after the protein preloads whereas no difference was found in men. There was some evidence that whey proteins and their components enhance satiety over a short-term period compared to carbohydrate but there was no consistent effect of either whey protein alone or glycomacropeptide.

Productive performance of commercial growing and finishing pigs supplemented with a Buttiauxella phytase as a total replacement of inorganic phosphate.

The objective of this study was to test if a novel phytase from Buttiauxella sp. can replace all added inorganic phosphate in a diet with reduced Ca and metabolizable energy (ME) fed to commercial pigs from 12 kg body weight (BW) until slaughter, whilst maintaining performance and carcass quality parameters. Four dietary treatments were tested in a completely randomized design with 9 replicate pens, each containing 31 mixed sex Newsham Choice pigs. Diets included a positive control (PC) based on corn, soybean meal, wheat middling and bakery meal, meeting all nutrient requirement of pigs; a negative control (NC) excluded inorganic phosphate and with reduced Ca (-0.13%) and ME (-0.15 MJ/kg); and NC supplemented with Buttiauxella phytase at 500 or 1,000 FTU/kg feed. Diets were fed ad libitum in mash form in 5 phases: starter (12 to 25 kg BW), grower 1 (25 to 50 kg BW) and 2 (50 to 75 kg BW), and finisher 1 (75 to 100 kg BW) and 2 (100 kg BW to slaughter). The NC group showed lower (P < 0.05) average daily feed intake (ADFI) and average daily gain (ADG) in starter and grower phases, lower gain to feed ratio (G:F) in starter and grower 1 compared with PC. Pigs receiving the high dose of phytase of 1,000 FTU/kg had improved performance vs. the 500 FTU/kg phytase treatment in starter and grower 1 phase compared with the PC in grower 1 phase. Increasing phytase dose resulted in a linear increase in ADG (12 to 120 kg BW) and G:F (50 to 75 kg BW). A comparison of treatment groups over the full production period from 12 kg BW until slaughter showed that both 500 and 1,000 FTU/kg phytase treatments were able to maintain growth performance and carcass characteristics compared with PC. The application of Buttiauxella phytase could therefore be used as an effective strategy to replace all inorganic phosphate in diets of pigs fed corn, soybean meal, wheat middling and bakery meal based diets from 12 kg BW. An economic analysis showed greater return from both phytase treatments vs. the PC and favored the higher phytase dose at 1,000 FTU/kg vs. the traditional dose of 500 FTU/kg. The latter was mainly related to the improved performance of the higher dose in younger pigs to 75 kg BW.

Accuracy of the Atwater factors and related food energy conversion factors with low-fat, high-fiber diets when energy intake is reduced spontaneously.

BACKGROUND: Systems to calculate metabolizable energy (ME) in foods and diets are often based on Atwater factors. The accuracy of these factors with low-fat diets high in fiber is unknown when food intake is reduced spontaneously. OBJECTIVE: The objective was to evaluate the accuracy of Atwater factors and other systems for calculating ME available from low-fat, high-fiber diets when food intake was reduced spontaneously. DESIGN: The ME contents of a high-fat, low-fiber diet and 2 low-fat diets, one high in fruit and vegetable fiber and the other high in cereal fiber, were determined in a randomized parallel study in humans (n = 27) and compared with various factorial and empirical models for calculating ME. RESULTS: Food intakes decreased with both the high fruit and vegetable fiber and cereal fiber diets. The difference between ME calculated by using Atwater and similar factors and determined ME values was up to 4% for the refined diet and up to 11% for the low-fat, high-fiber diets. Various factorial and empirical systems for calculating food energy failed to reflect the results of the direct determinations. CONCLUSION: Atwater factors were inaccurate with low-fat, high-fiber diets. Although modified Atwater factors may be accurate under standardized conditions of zero-nitrogen and zero-energy balance, they overestimate energy availability from high-fiber fruit and vegetable and cereal diets when food intake is reduced spontaneously in addition to when intake is reduced voluntarily.

Optimisation of inoculum concentration and incubation duration for an in vitro hindgut dry matter digestibility assay.

The aim was to optimise inoculum concentration and incubation duration for a published in vitro hindgut digestibility assay using ileal digesta (sampled from the chicken or rat) pertaining to a mixed human diet as the substrate. The study also sought to investigate the digestibility of the inoculum itself and the importance of correcting for this in the in vitro hindgut digestion assay. For two assays, hindgut dry matter digestibility (DMD) generally increased with inoculum concentration. A sharp increase in DMD observed at high inoculum concentrations may have been related to problems with filtering the inoculum. An inoculum concentration of 160 g/L was considered optimal based on close agreement of observed values with previously published in vivo hindgut dry matter digestibility for similar diets. One of the methods was chosen for optimisation of the duration of incubation. Ileal substrate organic matter digestibility (OMD) increased with increasing time of incubation for all diets. An incubation duration of 18 h using a mean inoculum digestibility value for calculation purposes was considered optimal based on observed in vivo hindgut DMD values in humans, but there was little difference in estimated in vitro hindgut DMD between 18 and 24h incubation durations. Although considerably lower than the OM digestibility of the substrate (no less than 51% after 48 h), the OM digestibility of the inoculum (13% after 48 h) itself was of significance in calculating estimated digestibility. The optimised assay gave realistic hindgut OMD values ranging from 55% to 79% (Wheat Bran Diet and Pectin Diet, respectively) using an 18-h incubation duration.

Post-natal development of the porcine microbiota composition and activities.

The current study describes the development of the porcine microbiota and its metabolic activities during the neonatal and weaning period. Using 16S rRNA-based approaches, we first analysed the ileal and colonic microbiota of neonatal piglets at days 2, 5 and 12 after birth. To further investigate the effect of weaning at 3 weeks of age, 19-day-old piglets (n = 64) were randomly allocated into two groups. Half of the piglets remained with their sows throughout the study, while the remaining piglets were weaned. As revealed by sequence analysis of 16S rRNA gene amplicons, the samples of 2-day-old piglets harboured a consortium of bacteria related to Escherichia coli, Shigella flexneri, Lactobacillus sobrius, Lactobacillus reuteri and Lactobacillus acidophilus. Moreover, species-specific real-time polymerase chain reaction assays unveiled that L. sobrius and L. reuteri predominated in the ileal samples of the neonatal and unweaned piglets with population levels up to 7 x 10(8) cells per gram of lumen content. Following weaning, however, these two lactobacilli were detected at significantly lower levels (< 10(3)) in the ileal samples. Furthermore, a shift in composition and metabolic activities of the predominant microbiota, and emergence of clostridia and E. coli, were encountered in the intestinal samples of the piglets after the early post-weaning period.

Specific response of a novel and abundant Lactobacillus amylovorus-like phylotype to dietary prebiotics in the guts of weaning piglets.

Using 16S rRNA gene-based approaches, we analyzed the responses of ileal and colonic bacterial communities of weaning piglets to dietary addition of four fermentable carbohydrates (inulin, lactulose, wheat starch, and sugar beet pulp). An enriched diet and a control diet lacking these fermentable carbohydrates were fed to piglets for 4 days (n = 48), and 10 days (n = 48), and the lumen-associated microbiota were compared using denaturing gradient gel electrophoresis (DGGE) analysis of amplified 16S rRNA genes. Bacterial diversities in the ileal and colonic samples were measured by assessing the number of DGGE bands and the Shannon index of diversity. A higher number of DGGE bands in the colon (24.2 +/- 5.5) than in the ileum (9.7 +/- 4.2) was observed in all samples. In addition, significantly higher diversity, as measured by DGGE fingerprint analysis, was detected in the colonic microbial community of weaning piglets fed the fermentable-carbohydrate-enriched diet for 10 days than in the control. Selected samples from the ileal and colonic lumens were also investigated using fluorescent in situ hybridization (FISH) and cloning and sequencing of the 16S rRNA gene. This revealed a prevalence of Lactobacillus reuteri in the ileum and Lactobacillus amylovorus-like populations in the ileum and the colon in the piglets fed with fermentable carbohydrates. Newly developed oligonucleotide probes targeting these phylotypes allowed their rapid detection and quantification in the ileum and colon by FISH. The results indicate that addition of fermentable carbohydrates supports the growth of specific lactobacilli in the ilea and colons of weaning piglets.