Effect of dietary carbohydrates on bacterial cholyltaurine hydrolase in poultry intestinal homogenates.

The bile salt hydrolase activity in intestinal homogenates reflects composite activities of the gastrointestinal microbial consortia. We have proposed that specific transformations of conjugated bile acids by the intestinal microflora result in the production of metabolites which depress the growth of poultry. The influence of dietary carbohydrates on the physical and kinetic properties of cholyltaurine hydrolase activity, one such bile acid-transforming enzyme in gastrointestinal homogenates of young chickens, was characterized by using a sensitive radiochemical assay. Cholyltaurine hydrolase activity in crude extracts of ileal homogenates was increased twofold by 0.25% Triton X-100 and a freeze-thaw cycle. The pH optimum for cholyltaurine hydrolase from ileal homogenates was very broad and reflected the pH range of poultry intestinal contents (i.e., 5.8 to 6.4). The carbohydrate component of the diet did not affect the apparent temperature optimum (41 degrees C) or stability profile, nor did it affect the apparent Km for taurocholic acid hydrolysis (approximately 0.43 mM). The enzymes in intestinal homogenates were active on all taurine-conjugated bile acids tested. The carbohydrate component of the diet did, however, affect the specific activity of cholyltaurine hydrolase in ileal homogenates from chickens. The levels of cholyltaurine hydrolase activity (rye greater than sucrose greater than corn) in homogenates from birds fed the different diets were directly related to the amount of growth depression (rye greater than sucrose greater than corn) associated with feeding these dietary carbohydrates. These data suggest that intestinal levels of cholyltaurine hydrolase are correlated with the amount of carbohydrate-induced growth depression in poultry.     

Subtherapeutic levels of antibiotics in poultry feeds and their effects on weight gain, feed efficiency, and bacterial cholyltaurine hydrolase activity.

A radiochemical method was developed to estimate cholyltaurine hydrolase potentials and rates of cholyltaurine hydrolysis in chicken intestinal homogenates. This method was used to monitor the effects of antibiotic feed additives on cholyltaurine hydrolase activity. Avoparcin, bacitracin methylenedisalisylic acid, efrotomycin, lincomycin, penicillin G procaine, and virginiamycin improved rate of weight gain and feed conversion of chicks and decreased cholyltaurine hydrolase activity in ileal homogenates relative to those of nonmedicated control birds. The results provided the first evidence that feeding selected antibiotics at subtherapeutic levels can affect bile acid-transforming enzymes in small-intestinal homogenates. The inverse relationship between growth performance and cholyltaurine hydrolase activity raises the possibility that specific inhibitors of this enzyme may promote weight gain and feed conversion in livestock and thereby reduce or eliminate the need for antibiotic feed additives.     

Subtherapeutic levels of antibiotics in poultry feeds and their effects on weight gain, feed efficiency, and bacterial cholyltaurine hydrolase activity

A radiochemical method was developed to estimate cholyltaurine hydrolase potentials and rates of cholyltaurine hydrolysis in chicken intestinal homogenates. This method was used to monitor the effects of antibiotic feed additives on cholyltaurine hydrolase activity. Avoparcin, bacitracin methylenedisalisylic acid, efrotomycin, lincomycin, penicillin G procaine, and virginiamycin improved rate of weight gain and feed conversion of chicks and decreased cholyltaurine hydrolase activity in ileal homogenates relative to those of nonmedicated control birds. The results provided the first evidence that feeding selected antibiotics at subtherapeutic levels can affect bile acid-transforming enzymes in small-intestinal homogenates. The inverse relationship between growth performance and cholyltaurine hydrolase activity raises the possibility that specific inhibitors of this enzyme may promote weight gain and feed conversion in livestock and thereby reduce or eliminate the need for antibiotic feed additives.     

Lactobacilli and bile salt hydrolase in the murine intestinal tract.

Mice that have a complex intestinal microflora but that do not harbor lactobacilli were used to determine the contribution of lactobacilli to the total bile salt hydrolase activity in the murine intestinal tract. Bile salt hydrolase activity in the ileal contents of these mice was reduced 86% in the absence of lactobacilli and by greater than 98% in the absence of lactobacilli and enterococci compared with samples from conventional mice. Bile salt hydrolase activities were lower in ileal and cecal contents from lactobacillus-free mice colonized with enterococci than in samples from lactobacillus-free mice colonized with lactobacilli. Bile salt hydrolase activity in the duodena, jejuna, ilea, and ceca of reconstituted lactobacillus-free mice colonized by lactobacilli was similar to that in samples from the intestinal tracts of conventional mice. We conclude from these studies that lactobacilli are the main contributors to total bile salt hydrolase activity in the murine intestinal tract.     

Lactobacilli and bile salt hydrolase in the murine intestinal tract

Mice that have a complex intestinal microflora but that do not harbor lactobacilli were used to determine the contribution of lactobacilli to the total bile salt hydrolase activity in the murine intestinal tract. Bile salt hydrolase activity in the ileal contents of these mice was reduced 86% in the absence of lactobacilli and by greater than 98% in the absence of lactobacilli and enterococci compared with samples from conventional mice. Bile salt hydrolase activities were lower in ileal and cecal contents from lactobacillus-free mice colonized with enterococci than in samples from lactobacillus-free mice colonized with lactobacilli. Bile salt hydrolase activity in the duodena, jejuna, ilea, and ceca of reconstituted lactobacillus-free mice colonized by lactobacilli was similar to that in samples from the intestinal tracts of conventional mice. We conclude from these studies that lactobacilli are the main contributors to total bile salt hydrolase activity in the murine intestinal tract.     

Bacterial Responses to Different Dietary Cereal Types and Xylanase Supplementation in the Intestine of Broiler Chicken

Several studies were carried out to investigate the influence of dietary cereals differing in soluble non starch polysaccharides (NSP) content and a xylanase preparation on selected bacterial parameters in the small intestine of broiler chicken. Compared to a maize diet colony forming units (CFU) of mucosa associated bacteria were higher in a wheat/rye diet, most notably for enterobacteria and enterococci. Xylanase supplementation to the wheat/rye diet generally led to lower CFU, especially in the first week of life. However, xylanase supplementation also displayed higher in vitro growth potentials for enterobacteria and enterococci. Bacterial growth of luminal samples in minimal media supplemented with selected NSP showed that the wheat/rye diet enhanced bacterial capacities to utilize NSP only in ileal samples. The xylanase application generally shifted respective maximum growth to the proximal part of the small intestine. The presence of soluble NSP from wheat or rye in the diet per se did not enhance bacterial NSP hydrolyzing enzyme activities in the small intestine, but xylanase supplementation resulted in higher 1,3-1,4- g - glucanase activity. Compared to a maize diet the activity of bacterial bile salt hydrolases in samples of the small intestine was not increased due to inclusion of wheat/rye or triticale to the diet. However, xylanase supplementation led to a reduction with a corresponding increase of lipase activity. It was concluded that dietary cereals producing high intestinal viscosities lead to increased overall bacterial activity in the small intestine. The supplementation of a xylanase to cereal based diets producing high intestinal viscosity, changes composition and metabolic potential of bacterial populations and may specifically influence fat absorption in young animals.     

Gastric emptying time and absorption of simple and complex carbohydrates in Macaca mulatta

Recent studies of nonhuman primates have shown that the amount and type of dietary carbohydrate influences the development of atherosclerotic lesions. In an effort to examine the gastrointestinal handling of simple and complex carbohydrates, measurements of gastric emptying times and uptake of C¹⁴ labeled carbohydrates were made for six female Macaca mulatta. A significantly smaller volume of sucrose solutions remained in the stomach after 60 minutes than solutions containing an equal amount by weight of dextrin. Similarly, when solutions of equal osmotic pressures but unequal weights of sucrose and dextrin were used, a significantly smaller volume of the sucrose solution remained in the stomach after 60 minutes. These findings are consistent with the idea that breakdown of dextrin by pancreatic amylase results in an increase in osmotic pressure of the intestinal contents and a slowing of gastric emptying. No significant differences in the levels of glucose or total C¹⁴ labeled carbohydrates were found in the peripheral blood following ingestion of C¹⁴ labeled sucrose or dextrin. This finding suggests that although an initial difference in the rate of gastric emptying of these carbohydrates exists, other factors, such as the rate of production and activity of pancreatic amylase, activity of intestinal disaccharidases, and intestinal mucosal transport, tend to equalize the rate of absorption of these sugars.     

Substrate specificity of the ileal and the hepatic Na(+)/bile acid cotransporters of the rabbit. II. A reliable 3D QSAR pharmacophore model for the ileal Na(+)/bile acid cotransporter

To design a reliable 3D QSAR model of the intestinal Na(+)/bile acid cotransporter, we have used a training set of 17 inhibitors of the rabbit ileal Na(+)/bile acid cotransporter. The IC(50) values of the training set of compounds covered a range of four orders of magnitude for inhibition of [(3)H]cholyltaurine uptake by CHO cells expressing the rabbit ileal Na(+)/bile acid cotransporter allowing the generation of a pharmacophore using the CATALYST algorithm. After thorough conformational analysis of each molecule, CATALYST generated a pharmacophore model characterized by five chemical features: one hydrogen bond donor, one hydrogen bond acceptor, and three hydrophobic features. The 3D pharmacophore was enantiospecific and correctly estimated the activities of the members of the training set. The predicted interactions of natural bile acids with the pharmacophore model of the ileal Na(+)/bile acid cotransporter explain exactly the experimentally found structure;-activity relationships for the interaction of bile acids with the ileal Na(+)/bile acid cotransporter (Kramer et al. 1999. J. Lipid. Res. 40: 1604;-1617). The natural bile acid analogues cholyltaurine, chenodeoxycholyltaurine, or deoxycholyltaurine were able to map four of the five features of the pharmacophore model: a) the five-membered ring D and the methyl group at position 18 map one hydrophobic site and the 21-methyl group of the side chain maps a second hydrophobic site; b) one of the alpha-oriented hydroxyl groups at position 7 or 12 fits the hydrogen bond donor feature; c) the negatively charged side chain acts as hydrogen bond acceptor; and d) the hydroxy group at position 3 does not specifically map any of the five binding features of the pharmacophore model. The 3-hydroxy group of natural bile acids is not essential for interactions with ileal or hepatic Na(+)/bile acid cotransporters. A modification of the 3-position of a natural bile acid molecule is therefore the preferred position for drug targeting strategies using bile acid transport pathways.     

Hydrolysis of retinyl palmitate by enzymes of rat pancreas and liver. Differentiation of bile salt-dependent and bile salt-independent, neutral retinyl ester hydrolases in rat liver

Previous studies have demonstrated that homogenates of the livers of rats contain a neutral retinyl ester hydrolase activity that requires millimolar concentrations of bile salts for maximal in vitro activity. The enzymatic properties of this neutral, bile salt-dependent retinyl ester hydrolase activity in liver homogenates are nearly identical to those observed in the present report for the in vitro hydrolysis of retinyl palmitate by purified rat pancreatic cholesteryl ester hydrolase (EC 3.1.1.13). Moreover, anti-rat pancreatic cholesteryl ester hydrolase IgG completely inhibits the bile salt-dependent retinyl ester hydrolase activity of rat liver homogenates whereas normal rabbit IgG does not. We also show that liver homogenates contain a neutral, bile salt-independent retinyl ester hydrolase activity that differs from the bile salt-dependent activity in that 1) its absolute activity does not vary markedly among individual rats, 2) it is not inhibited by antibodies to pancreatic cholesteryl ester hydrolase, and 3) it is localized in the microsomal fraction of liver homogenates. Subfractionation of microsomes demonstrates that the neutral, bile salt-independent retinyl ester hydrolase activity is associated with liver cell plasma membranes and thus may play a role in the hydrolysis of retinyl esters delivered to the liver by chylomicron remnants.     

Large doses of zinc oxide increases the activity of hydrolases in rats

The effect of pharmacological doses of zinc oxide (1000; 2500; 5000 mg per kg diet) and two levels of dietary protein on pancreatic and intestinal hydrolase activity in rats were studied. It was hypothesized that ZnO would increase intestinal and pancreatic hydrolase enzyme activity. Male Wistar rats, averaging 64 g body weight, were randomly allocated to dietary treatments (chow diets- meeting all NRC requirements) containing 10% or 15% protein supplemented with additional ZnO (above 100 mg/kg ZnSO(4)) as follows: 0.0; 0.1; 0.25; 0.5% w/w. Water and food were provided ad libitum. Animals were fed the diets for 10 days and body weights were recorded; after decapitation blood and organ samples were collected. Amylase, lipase, trypsin, and total protease activity of pancreatic homogenates and small intestinal contents were determined. ZnO supplementation dose dependently increased the plasma Zn concentration and significantly increased amylase, lipase, trypsin and total protease activity in pancreatic homogenates and small intestinal contents. The statistical analysis showed significant protein and ZnO interaction on the activity of amylase in the pancreas, and amylase, trypsin and total-protease in the small intestinal content. Therefore ZnO at high dietary concentration may influence the digestion of nutrients via increased hydrolase activity.     

Comparison of Lactobacillus strains with respect to bile salt hydrolase activity, colonization of the gastrointestinal tract, and growth rate of the murine host.

The significance of bile salt hydrolase production by lactobacilli in the microecology of the murine intestinal tract has not been extensively studied previously. Assays of bile salt hydrolase (sodium taurocholate as substrate) associated with cell extracts of five Lactobacillus strains of murine origin gave a range of activities (from 915 nmol of cholate released per mg of protein per 30 min to none detected). All of the strains tested colonized the murine gastrointestinal tract equally well. The growth rates of mice were not affected by colonization of their intestinal tracts by lactobacilli whether or not the bacteria produced bile salt hydrolase.     

Diet-induced adaptation of intestinal fructose absorption in the rat.

The effect of dietary sucrose, fructose and glucose on the intestinal absorption of fructose and glucose was investigated in adult rats in vivo: Glucose absorption was not affected by the type of dietary carbohydrate, while the absorption of fructose was increased by the ingestion of the sucrose or fructose diet, as compared with the glucose diet. An almost maximal increase of fructose absorption was already observed when the quarter of the total dietary carbohydrates was replaced by fructose. Faecal fructose elimination declined during the feeding experiment. The enhanced intestinal absorption of the fructose load in rats fed the fructose diet was manifested by higher concentrations of fructose, but also of glucose and lactate in the hepatic portal blood.     

Identification of the bile acid-binding site of the ileal lipid-binding protein by photoaffinity labeling, matrix-assisted laser desorption ionization-mass spectrometry, and NMR structure

The ileal lipid-binding protein (ILBP) is the only physiologically relevant bile acid-binding protein in the cytosol of ileocytes. To identify the bile acid-binding site(s) of ILBP, recombinant rabbit ILBP photolabeled with 3-azi- and 7-azi-derivatives of cholyltaurine was analyzed by a combination of enzymatic fragmentation, gel electrophoresis, and matrix-assisted laser desorption ionization (MALDI)-mass spectrometry. The attachment site of the 3-position of cholyltaurine was localized to the amino acid triplet His(100)-Thr(101)-Ser(102) using the photoreactive 3,3-azo-derivative of cholyltaurine. With the corresponding 7,7-azo-derivative, the attachment point of the 7-position could be localized to the C-terminal part (position 112-128) as well as to the N-terminal part suggesting more than one binding site for bile acids. By chemical modification and NMR structure of ILBP, arginine residue 122 was identified as the probable contact point for the negatively charged side chain of cholyltaurine. Consequently, bile acids bind to ILBP with the steroid nucleus deep inside the protein cavity and the negatively charged side chain near the entry portal. The combination of photoaffinity labeling, enzymatic fragmentation, MALDI-mass spectrometry, and NMR structure was successfully used to determine the topology of bile acid binding to ILBP.     

The level of dietary protein and carbohydrate has a different effect on intestinal uptake of hexoses and lipids in rabbits with an ileal resection than in those with an intact intestinal tract

Either high protein, low carbohydrate (HP) or low protein, high carbohydrate (LP) diets were fed for 6 weeks to rabbits with or without resection of the distal half of the small intestine. Control and resected rabbits fed HP consumed more food and gained more weight than the animals fed LP. The level of dietary protein has a different effect on intestinal transport in animals with an ileal resection than in those with an intact small intestine. With 0.5 mM glucose, the in vitro uptake in control rabbits was greater for the LP than HP diet but was unchanged in resected rabbits; uptake of 0.5 mM galactose and 3-O-methyl glucose was unaffected by HP and LP, whereas in rabbit uptake was lower in LP than HP. The uptake of 40 mM glucose was greater with the LP than HP diet in control rabbits, but lower with LP than HP in resected rabbits. In control rabbits, the uptake of aluric acid was lower on the LP than HP diet; the uptake of octanoic acid, decanoic acid, and cholesterol was unchanged; and the uptake of each fatty acid and cholesterol was greater in resected rabbits fed LP than HP. Feeding a low protein diet to animals with an ileal resection is associated with lower jejunal uptake of high concentrations of glucose, but the higher uptake of galactose and enhanced permeability to fatty acids result in superior weight gain. Thus, recommendations for alterations in dietary protein and carbohydrate levels following ileal resection must be made with the knowledge that these changes may influence intestinal transport function as well as body weight gain.     

Effect of Carbohydrate Demand on the Remobilization of Starch in Stolons and Roots of White Clover (Trifolium repens L.) after Defoliation

White clover plants were grown from stolon tips in growth cabinetsand then defoliated. Thereafter, changes in the contents ofnon-structural carbohydrates such as starch, sucrose, glucose,fructose, maltose, and pinitol in stolons and roots were monitored.Initial contents of carbohydrate reserves, photosynthetic supplyof new carbohydrates and carbohydrate demand after defoliationwere varied by growing the plants at various CO₂ partial pressures,by varying the extent of defoliation and by removing eitherroots or stolon tips at the time of defoliation. Remobilization of carbohydrate reserves in stolons increasedproportionally to their initial contents and was greater whenplants had been severely defoliated, suggesting that carbohydrateswere remobilized according to availability and demand. Starchwas the predominant reserve carbohydrate. Starch degradationwas associated with decreased contents of sucrose, glucose andfructose in young stolon parts and roots but not in old stolonparts suggesting that starch degradation was not strictly controlledby the contents of these sugars. A decrease in the demand forcarbohydrates by removal of roots did not decrease starch degradationbut increased the contents of sucrose, glucose, and fructose.Removal of stolon tips decreased starch degradation and contentsof sucrose, glucose, and fructose. The results suggest thatstarch degradation was controlled by a factor other than sucrose,glucose, and fructose which was exported from stolon tips, e.g.gibberellin. Key words: White clover, storage carbohydrates, remobilization, regrowth     

Rye Affects Bacterial Translocation,Intestinal Viscosity,Microbiota Composition and Bone Mineralization in Turkey Poults

Previously, we have reported that rye significantly increased both viscosity and Clostridium perfringens proliferation when compared with corn in an in vitro digestive model. Two independent trials were conducted to evaluate the effect of rye as a source of energy on bacterial translocation, intestinal viscosity, gut microbiota composition, and bone mineralization, when compared with corn in turkey poults. In each experiment, day-of-hatch, turkey poults were randomly assigned to either a corn or a rye diet (n = 0 /group). At 10 d of age, in both experiments, 12 birds/group were given an oral gavage dose of fluorescein isothiocyanate dextran (FITC-d). After 2.5 h of oral gavage, blood and liver samples were collected to evaluate the passage of FITC-d and bacterial translocation (BT) respectively. Duodenum, ileum and cecum gut sections were collected to evaluate intestinal viscosity and to enumerate gut microbiota. Tibias were collected for observation of bone parameters. Broilers fed with a rye diet showed increased (p<0.05) intestinal viscosity, BT, and serum FITC-d. Bacterial enumeration revealed that turkey poults fed with rye had increased the number of total lactic acid bacteria (LAB) in all three sections of the gastrointestinal tract evaluated when compared to turkey poults fed with corn. Turkey poults fed with rye also had significantly higher coliforms in duodenum and ileum but not in the ceca, whereas the total number of anaerobes increased only in duodenum. A significant reduction in bone strength and bone mineralization was observed in turkey poults fed with rye when compared with corn fed turkey poults. In conclusion, rye evoked mucosal damage in turkey poults that increased intestinal viscosity, increased leakage through the intestinal tract, and altered the microbiota composition and bone mineralization. Studies to evaluate dietary inclusion of selected Direct-Fed Microbial (DFM) candidates that produce exogenous enzymes in rye fed turkey poults are currently being evaluated.     

Age-related topographical metabolic signatures for the rat gastrointestinal contents

Symbiotic gut microbiota is essential for mammalian physiology and analyzing the metabolite compositions of gastrointestinal contents is vital for understanding the microbiome-host interactions. To understand the developmental dependence of the topographical metabolic signatures for the rat gastrointestinal contents, we systematically characterized the metabolite compositional variations of the contents in rat jejunum, ileum, cecum, and colon for two age-groups using (1)H NMR spectroscopy and multivariate analysis. Significant topographical metabolic variations were present for the jejunal, ileal, cecal, colonic contents, and feces, reflecting the absorption functions for each intestinal region and the gut microbiota therein. The concentrations of amino acids, lactate, creatine, choline, bile acids, uracil and urocanate decreased drastically from jejunal to ileal contents followed with steady decreases from cecal content to feces. Short-chain fatty acids (SCFAs) and arabinoxylan-related carbohydrates had highest levels in cecal content and feces, respectively. Such topographical metabolic signatures for the intestinal contents varied with animal age highlighted by the level changes for lactate, choline, taurine, amino acids, carbohydrates, keto-acids, and SCFAs. These findings provided essential information for the topographical metabolic variations in the gastrointestinal tract and demonstrated metabolic profiling as a useful approach for understanding host-microbiome interactions and functional status of the gastrointestinal regions.     

Conjugated bile acid uptake by Xenopus laevis oocytes induced by microinjection with ileal Poly A+ mRNA.

Apical membranes of ileal enterocytes contain the major Na+/bile acid cotransporter activity in mammals. Microinjection of guinea pig ileal mucosal Poly A+ mRNA (25 ng) into Xenopus oocytes resulted in 22,23-3H-cholyltaurine uptake at day 3 after injection (453 fmol/oocyte-hr), while control viral mRNA (25 ng) gave an uptake rate of 133 fmol/oocyte-hr. The transport rate increased in direct relationship to the concentration of injected mRNA, cholyltaurine, or Na+ in the incubation media. Uptake of cholyltaurine using rabbit ileal mucosal Poly A+ mRNA was 3891 fmole/oocyte-hr compared to rabbit jejunal-mucosa Poly A+ mRNA (control) injections inducing 728 fmol/oocyte-hr. Such expression of the ileal Na+/bile acid cotransporter may facilitate cloning of this key mammalian gene.     

The role of hepatic, renal and intestinal gluconeogenic enzymes in glucose homeostasis of juvenile rainbow trout

Rainbow trout is unable to utilize high levels of dietary carbohydrates and experiences hyperglycemia after consumption of carbohydrate-rich meals. Carbohydrates stimulate hepatic glycolytic activity, but gene expression of the rate-limiting gluconeogenic enzymes glucose-6-phosphatase (G6Pase), fructose-1,6-bisphosphatase (FBPase) and phosphoenolpyruvate carboxykinase (PEPCK) remains high. Although there is significant mRNA expression and activity of gluconeogenic enzymes in trout intestine and kidney, the regulation of these enzymes by diet is not known. We tested the hypothesis that dietary carbohydrate modulates intestinal and renal G6Pase, FBPase and PEPCK. Fish were either fasted or fed isocaloric carbohydrate-free (CF) or high carbohydrate (HC) diets for 14 days. As expected, fish fed HC exhibited postprandial hyperglycemia and enhanced levels of hepatic glucokinase mRNA and activity. Dietary carbohydrates had no significant effect on the expression and activity of PEPCK, FBPase and G6Pase in all three organs. In contrast, fasting enhanced the activity, but not the mRNA expression of both hepatic and intestinal PEPCK, as well as intestinal FBPase. Therefore, the activity of rate-limiting gluconeogenic enzymes in trout can be modified by fasting, but not by the carbohydrate content of the diet, potentially causing hyperglycemia when fed high levels of dietary carbohydrates. In this species consuming low carbohydrate diets at infrequent intervals in the wild, fasting-induced increases in hepatic and intestinal gluconeogenic enzyme activities may be a key adaptation to prevent perturbations in blood glucose during food deprivation. Presented in part at Experimental Biology, April 2006, San Francisco, CA [Kirchner S., Panserat S., Kaushik S. and Ferraris R. FASEB-IUPS-2006 A667.6].     

Impact of dietary protein on microbiota composition and activity in the gastrointestinal tract of piglets in relation to gut health: a review

In pigs, the microbial ecosystem of the gastrointestinal tract (GIT) is influenced by various factors; however, variations in diet composition have been identified as one of the most important determinants. Marked changes in fermentation activities and microbial ecology may occur when altering the diet, for example, from milk to solid feed during weaning. In that way, access of pathogens to the disturbed ecosystem is alleviated, leading to infectious diseases and diarrhea. Thus, there is increasing interest in improving intestinal health by use of dietary ingredients suitable to beneficially affect the microbial composition and activity. For example, fermentable carbohydrates have been shown to promote growth of beneficial Lactobacillus species and bifidobacteria, thereby enhancing colonization resistance against potential pathogens or production of short-chain fatty acids, which can be used as energy source for epithelial cells. On the other hand, fermentation of protein results in the production of various potentially toxic products, such as amines and NH₃, and is often associated with growth of potential pathogens. In that way, excessive protein intake has been shown to stimulate the growth of potentially pathogenic species such as Clostridium perfringens, and to reduce fecal counts of beneficial bifidobacteria. Therefore, it seems to be a promising approach to support growth and metabolic activity of the beneficial microbiota by developing suitable feeding strategies. For example, a reduction of dietary CP content and, at the same time, dietary supplementation with fermentable carbohydrates have proven to successfully suppress protein fermentation. In addition, the intestinal microbiota seems to be sensible to variations in dietary protein source, such as the use of highly digestible protein sources may reduce growth of protein-fermenting and potentially pathogenic species. The objective of the present review is to assess the impact of dietary protein on microbiota composition and activity in the GIT of piglets. Attention will be given to studies designed to determine the effect of variations in total protein supply, protein source and supplementation of fermentable carbohydrates to the diet on composition and metabolic activity of the intestinal microbiota.