Biochemical intestinal parameters in pigs reared outdoors and indoors,and in germ-free pigs

Intestinal microbial functions reflect cross-talk between a host and its flora, and external factors may influence these functions. The aim of this investigation was to follow the development of six biochemical microbial-related functions of piglets, raised outdoors (OPs) or indoors (IPs), from birth to slaughter age. The following parameters (microflora-associated characteristic; MAC) were consecutively measured at five different ages: production of short-chain fatty acids (SCFA), conversion of cholesterol to coprostanol and of bilirubin to urobilinogens, inactivation of trypsin, degradation of beta-aspartylglycine and of mucin. Additionally, four parameters (production of SCFA. conversion of cholesterol to coprostanol, inactivation of trypsin, degradation of beta-aspartylglycine) were investigated in faecal samples from germ-free minipigs. The differences in MAC patterns between OPs and IPs were most pronounced at 20 days of age. Differences were found in the total amount of SCFAs, proportions of the acetic, propionic and butyric acids, conversion of bilirubin to urobilinogens, degradation of faecal tryptic activity and degradation of mucin. The values found in the minipigs were within the range of a germ-free animal characteristic (GAC) pattern. Our results show that environmental factors influence the development of some intestinal microbial functions in pigs.     

Influence of bacitracin on microbial functions in the gastrointestinal tract of horses

This study investigated the influence of zinc bacitracin on the intestinal flora of horses. The functionally active intestinal flora was examined in 6 horses during treatment with zinc bacitracin. Utilising gas chromatography, spectrophotometry, gel electrophoresis and paper chromatography, samples were analysed on biochemical markers reflecting the action of parts of the intestinal flora. The following 5 flora-related functions were studied in faecal samples and intestinal samples from different sections of the hindgut: conversion of cholesterol to coprostanol and of bilirubin to urobilinogens, degradation of mucin and of beta-aspartylglycine and inactivation of tryptic activity. Conversion to coprostanol, conversion to urobilinogens and degradation of mucin were affected by treatment of zinc bacitracin and conversion to coprostanol was most sensitive. All functions were normalised in a short time, in contrast to man and rats. Differences in environmental exposures are probably the reason for a more rapid normalisation of the intestinal flora functions in horses.     

Biochemical intestinal parameters in germ-free minipigs and rats and in ex-germ-free minipigs and rats monoassociated with Escherichia coli

Intestinal contents of newborn and young germ-free minipigs and germ-free rats were investigated for the following biochemical parameters - conversion of cholesterol to coprostanol, degradation of beta-aspartylglycine, level of tryptic activity, formation of urobilinogen and the profile of short-chain fatty acids. Additionally, germ-free minipigs and germ-free rats were monoassociated with non-pathogenic strains of Escherichia coli and were investigated for the same biochemical parameters. The conversion of cholesterol to coprostanol, degradation of beta-aspartylglycine, tryptic activity and the short-chain fatty acid profile were similar to those found in previous studies in germ-free animals. Slightly higher amounts of urobilinogen than in the other species investigated so far were found in samples from germ-free and monoassociated minipigs. Except for the total amount of short-chain fatty acids in rats, monoassociation with E. coli did not alter any of the parameters either in the minipigs or in the rats.     

Influence of Zinc Bacitracin and Bacillus licheniformis on Microbial Intestinal Functions in Weaned Piglets

The influence of zinc bacitracin (ZB) and of Bacillus licheniformis on host microbial-related functions in young piglets was investigated by applying the concept of microflora-associated characteristics. Six biochemical parameters were determined before and after weaning in faecal samples from piglets in four litters having access to a diet containing ZB, to a diet containing B. licheniformis, to a diet with both additives, or to a diet with no additives, from 3 weeks of age. Statistically significant differences were found in three of the intestinal functions investigated: formation of short-chain fatty acids (at 7 and 10 weeks of age), degradation of mucin (at 7 and 10 weeks of age) and conversion of bilirubin to urobilins (at 7 weeks of age). We also found age-dependent influences on the formation of short-chain fatty acids, on conversion of cholesterol to coprostanol and on conversion of bilirubin to urobilins. We conclude that a functional approach is appropriate for measuring exogenous influence(s) on the microbial intestinal metabolisms in weaned piglets.     

Effect of diet on markers of intestinal health in dogs

The effects of two popular commercial pet foods on faecal markers of microbial metabolism were investigated. Adult dogs were fed a dry, extruded diet and a moist, canned diet in a randomly assigned crossover design. Fresh faecal samples were collected for chemical and enzyme activity assays. Relative to the canned diet, the dry food resulted in decreased faecal pH and faecal indole, sulphide and ammonia concentrations and increased total short-chain fatty acid, acetic and propionic acid concentrations. Faecal beta-glucosidase, beta-glucuronidase, beta-galactosidase and nitroreductase activities were decreased in dogs fed the dry diet. These changes in microbial metabolic activity are consistent with beneficial effects of the dry diet on colonic health.     

Macronutrient digestibility, nitrogen balance, plasma indicators of protein metabolism and mineral absorption in horses fed a ration rich in sugar beet pulp

In a cross-over study with six mature horses, the effect of iso-energetic replacement of dietary glucose by beet pulp on macronutrient digestibility, nitrogen metabolism and mineral absorption was studied. The test ration contained 25% beet pulp in the total dietary dry matter. Beet pulp feeding significantly lowered crude fat and non-structural carbohydrate digestibility, but had no significant effect on digestibility of other macronutrients, faecal and urinary nitrogen excretion and the faecal to urinary nitrogen excretion quotient. However, on the beet pulp diet, plasma ammonia and creatinin concentrations were significantly lower than on the glucose diet. No diet effect on magnesium absorption was observed. It is suggested that dietary beet pulp stimulates the conversion of ammonia into urea.     

Effect of sodium phytate supplementation on fat digestion and cholesterol metabolism in female rats

The effects of sodium phytate supplementation on fat digestion and cholesterol metabolism were investigated in female rats. On the basis of an in vitro experiment showing that phytate raised the solubility of bile acids, it was predicted that phytate feeding would depress faecal bile acid excretion, raise apparent fat digestibility and elevate serum cholesterol concentrations. The experimental diets with or without sodium phytate were either cholesterol-free or cholesterol-rich and had a normal calcium concentration. Rats fed on the cholesterol-rich diet with sodium phytate showed enhanced faecal bile acid excretion, but there was no effect on fat digestibility. In rats fed the cholesterol-free diets, phytate did neither affect fat digestion nor bile acid excretion. Sodium phytate inclusion in the cholesterol-rich diet raised serum cholesterol concentrations, but reduced liver cholesterol concentration. Thus, the in vivo data do not agree with the in vitro observations. Both phytate and cholesterol feeding influenced mineral and trace element metabolism. Liver zinc concentrations were raised by phytate feeding. Cholesterol consumption reduced hepatic concentrations of copper, iron and zinc. Both phytate and cholesterol feeding reduced the apparent absorption of calcium, magnesium and phosphorus.     

Dietary carbohydrates affect caecal fermentation and modify nitrogen excretion patterns in rats. I. Studies with protein-free diets.

In a two-factorial experiment on 96 young male rats, the effects of substituting 10% raw potato starch (PS), pectins (PEC), or cellulose (CEL) for corn starch (CS) were studied using an unsupplemented protein-free (PF) diet or a PF diet supplemented either with DL-methionine or urea. The pH and the short chain fatty acids (SCFA) content in caecal digesta, as well as caecal digesta and tissue weights were determined and used as the criteria of caecal fermentation intensity. Blood urea level, amount of N excreted via faeces and urine, DAPA content, and amino acid composition of faecal protein were analyzed as indices of protein metabolism. A 10-day adaptation period to the carbohydrates fed with the casein diet preceded the experimental period of feeding the respective carbohydrates with protein-free diets. Dietary carbohydrates significantly influenced total and individual SCFA content in caecal digesta, as well as other parameters related to the intensity of fermentation. Potato starch and pectins were more intensively fermented than cellulose. Supplementation of the PF diet with methionine and urea affected only caecal isobutyric and valeric acid content in a way dependent on the carbohydrates present in the diet. Carbohydrates significantly altered the routes of N excretion. Faecal excretion was increased by all carbohydrates studied compared to corn starch, pectins had the most marked effect. Urinary excretion was significantly increased by cellulose (as compared with the PEC and PS groups) and decreased by pectins as compared with all other groups. There was an interaction between the effects of carbohydrates and type of protein-free diet on faecal and urinary excretion. The sum of amino acids in faecal protein was the lowest on the PEC diet, but the amino acid composition expressed as a per cent of total amino acid content was similar in all groups. It can be concluded that dietary carbohydrates alter the excretion patterns of endogenous nitrogen in rats in different ways and that this effect is related to the intensity of their fermentation in the hind gut.     

Inter-breed diversity and temporal dynamics of the faecal microbiota in healthy horses

Understanding gut microbiota similarities and differences across breeds in horses has the potential to advance approaches aimed at personalized microbial modifications, particularly those involved in improving sport athletic performance. Here, we explore whether faecal microbiota composition based on faecal 16S ribosomal RNA gene sequencing varies across six different sport breeds at two time points 8 months apart within a cohort of 189 healthy horses cared for under similar conditions. Lusitano horses presented the smallest and Hanoverians the greatest bacterial diversity. We found subtle but significant differences in β-diversity between Lusitano, Anglo Arabian and the central European breeds, and we reproduced these results across the two time points. Repeat sampling of subjects showed community to be temporally more stable in Lusitano and Anglo Arabian breeds. Additionally, we found that 27 genera significantly varied in abundance across breeds. Overall, 33% of these taxa overlapped with previously identified taxa that were associated with genetic variation in humans or other species. However, a non-significant correlation was observed between microbial composition and the host pedigree-based kinship. Despite a notable variation in the diversity and composition of the faecal microbiota, breed exerted limited effects on the equine faecal microbiota.     

Effect of intestinal microflora on digestible energy and fiber digestion in chickens fed a high-fiber diet.

The effect of intestinal microflora on digestible energy (DE) value and fiber digestion was studied in single-comb White Leghorn chickens fed a low-fiber diet (experiment 1) or a high-fiber diet with low or adequate metabolizable energy (ME) value (experiment 2). Fecal energy excretion was calculated from the difference between total energy excretion in urinary and fecal droppings and urinary energy excretion, which was estimated from the energy values for individual urinary nitrogenous compounds extracted with Li2CO3. When the birds were fed the low-fiber diet, no differences in growth, DE, or ME were observed between germ-free and conventional environments. Of birds fed the high-fiber diet, growth of those in the conventional environment was similar to that of the birds in the germ-free environment at the adequate ME value, whereas birds in the conventional environment grew faster than the birds in the germ-free environment at the low ME value. Changes in observed dietary ME values of the high-fiber diets, being higher in birds in the conventional environment than in birds in the germ-free environment (experiment 2), were almost entirely accounted for by those in dietary DE values, most of which was contributed by crude fiber digestion. It was concluded, therefore, that by means of fiber digestion, the intestinal microflora may benefit the host bird by supplying extra energy, which would result in growth promotion, particularly when the bird is deficient in energy.     

Physiological performance of quails that underwent dietary and pharmacological manipulation of cholesterol

The present work evaluated whether dietary and pharmacological interference on cholesterol synthesis were capable of inducing alterations in blood and yolk cholesterol levels and the secretion of corticosterone metabolites. Forty‐five 40‐day‐old quails were divided into three experimental groups: vegetal fat diet, 2% beef fat (tallow) diet and vegetal fat diet with simvastatin administration (3.13 mg/kg/day). During all experiments, the animal weights and food consumption were recorded and blood and faecal samples (days 0, 15, 30, 45 and 60), as well as eggs (days 30, 45 and 60), were collected. Analysis of serum and yolk cholesterol was performed and faecal corticosterone levels were measured. No differences were observed on blood cholesterol or faecal corticosterone between all treatments, despite a tendency of increased cholesterol in the group with the animal fat diet. However, quails submitted to an animal fat diet displayed an increase in yolk cholesterol at day 30 of the treatment and the egg yolks of quails treated with simvastatin exhibited a decrease in cholesterol content by the end of the treatment at 60 days. These results improved the knowledge regarding the physiology of quails and offered support to other studies concerning the consequences of the pharmacological treatment and the dietary manipulation of cholesterol levels.     

Copper and zinc balance in exercising horses fed 2 forms of mineral supplements

Studies comparing the absorption and retention of various forms of trace minerals in horses have yielded mixed results. The objective of this study was to compare Cu and Zn absorption and retention in exercising horses where the mineral was supplemented in the sulfate or organic chelate form. Nine mature horses were used in a modified switchback design experiment consisting of seven 28-d periods. Horses were fed a diet consisting of 50% concentrate and 50% hay that was balanced to meet the energy, protein, Ca, and P requirements for horses performing moderate-intensity exercise. Horses were subjected to a controlled mineral repletion-depletion diet sequence before feeding the experimental diet to standardize mineral status across horses. The experimental diet was designed to provide 90% of the 1989 NRC for Cu and Zn, with supplemental mineral provided in the inorganic sulfate form (CuSO(4) and ZnSO(4)) or the organic chelate form (Cu-Lys and Zn-Met). Feed, fecal, urine, and water samples collected during a total collection during the last 4 d of the experimental diet periods were analyzed to determine apparent absorption and retention of Cu and Zn from the 2 mineral forms. A formulation error caused horses receiving the organic chelate diet to consume about 3 times the amount of Cu and Zn compared with those fed the sulfate-supplemented diet. Copper and Zn intake and fecal excretion were greater (P < 0.05) for horses consuming the organic chelate-supplemented diet. Apparent absorption values for all horses were negative. Apparent Cu absorption and retention as a percentage of intake were greater for horses fed the organic chelate diet (P < 0.05). It is unknown why excretion of Cu and Zn by the horses during the total collection exceeded the mineral intake. Although Cu-Lys seemed to be better absorbed than CuSO(4) and absorption of Zn-Met and ZnSO(4) were not different, these results are tempered by the observation of abnormally high fecal and urinary excretion values for Cu and Zn in the present study.     

Dietary fish oil supplementation affects serum fatty acid concentrations in horses

Thirteen horses of Thoroughbred or Standardbred breeding were used to study the effect of dietary fish oil supplementation on blood lipid characteristics. Horses were assigned to either fish oil (n = 7) or corn oil (n = 6) treatment groups for 63 d. The fish oil contained 10.8% eicosapentaenoic acid (EPA) and 8% docosahexaenoic acid (DHA). Each horse received timothy hay and a mixed-grain concentrate at rates necessary to maintain BW. Oil (corn or fish) was top-dressed on the concentrate daily at a rate of 324 mg/ kg of BW. The n-6:n-3 ratio was approximately 3.6:1 for horses receiving the corn oil diet and 1.4:1 for horses receiving the fish oil diet. Horses were exercised 5 d/wk during the study. Before supplementation, there was no difference in the concentrations of any serum fatty acids between the 2 treatment groups. The mean basal concentrations of EPA and DHA on d 0 were 0.04 and 0.01 mg/mL, respectively. After 63 d, horses receiving the fish oil treatment, but not those receiving the corn oil treatment, had increased concentrations of EPA and DHA (P <0.05). Fish oil supplementation for 63 d also increased the concentrations of C22:0, C22:1, and C22:5 fatty acids (P <0.05). Overall, horses receiving fish oil had a decreased concentration of n-6 fatty acids (P <0.05) and a greater concentration of n-3 fatty acids (P <0.01), resulting in a lower n-6:n-3 fatty acid ratio after 63 d (P <0.05). Serum cholesterol concentrations increased (P <0.05) during the supplementation period in horses receiving the corn oil but not in horses receiving the fish oil. Compared with horses receiving corn oil, horses receiving fish oil had lower serum triglycerides at d 63 (P <0.05). These results demonstrate that 63 d of fish oil supplementation at 324 mg/kg of BW was sufficient to alter the fatty acid profile and blood lipid properties of horses receiving regular exercise.     

Effect of physical training on nutrient digestibility and faecal fermentative parameters in Standardbred horses

This study aimed at evaluating, in previously inactive Standardbreds horses, the effect of 5 weeks of an exercise training programme on nutrient digestibility and faecal fermentative parameters (FFPs). As an increase in digestibility had previously been reported in trained endurance horses, we hypothesized that similar results would be found in horses being trained for other types of exercise on a different type of diet. After 3 weeks of dietary adaptation, a digestibility trial (DT1) was undertaken, over 3 days, in eight untrained Standardbreds with a fresh faecal sample being collected on the second day for FFP determinations. Six of the eight horses undertook a training programme, and after 5 weeks of exercise, the DT and the FFP measurements were then repeated (DT2). DT2 began after 3 days of inactivity. The same natural meadow hay (H) and pelleted complementary feed (CF) were fed throughout. For 5 days before the DTs, horses were fed 2.1% BW on a dry matter basis (55:45 ratio H:CF). Body weight and body condition score remained constant. Apparent digestibility of dry matter, organic matter, neutral detergent fibre, hemicellulose, crude protein and gross energy, as well as faecal total volatile fatty acids (VFA), acetate and propionate concentrations were significantly (p < 0.05) higher at DT2 than at DT1. A 5‐week exercise programme had a positive impact on nutrient digestibility and FFP. Training may improve dietary energy supply, in particular via increased hindgut VFA production. The potential improvement of digestive efficiency with training should be taken into account when formulating nutritional recommendations for the exercising horse, particularly when performing light work, which is low‐intensity exercise for 1–3 h per week.     

Investigations of the maintenance system of the Konik Polski horse and its effects on fecal microbiota activity during the winter and summer seasons

Gastrointestinal microbiota play a key role in the nutrients digestion and hence maintaining animal health and welfare. The diet offered to the animals in captivity may differ considerably from that on natural pastures. In a stabled maintenance system, horses have a limited choice of habitat and feed. Time spend for feeding is relevant for equine welfare because the reduction of the time devoted for foraging may be responsible for inducing gastric inflammation and ulceration. Therefore, in the present study, it was hypothesized that fecal bacterial fermentative processes differ between free-roaming and stabled Konik Polski Horses (KPHs) with respect to microbial enzymatic activity, and thus the fecal concentration of short-chain fatty acids (SCFA), which can be further utilized for assessing the feeding behavior and welfare in free-roaming versus stabled horses. The SCFA concentration and profile, as well as the extracellular and intracellular activities of selected bacterial enzymes, were characterized in horse feces collected during the winter and summer feeding seasons. The results showed higher enzymatic activity and SCFA production in the feces excreted by free-roaming versus stabled horses, especially during summer. An increase in pasture plant diversity may be beneficial for the gastrointestinal microbiota and hence for maintaining health and welfare.     

Studies on digestive physiology and feed digestibilities in captive Indian rhinoceros (Rhinoceros unicornis)

We performed intake, digestibility and ingesta passage studies in 11 Indian rhinoceroses (Rhinoceros unicornis) from four zoological institutions, using total faecal collection for the quantification of faecal output. The regularly fed zoo ration of roughage and concentrates (ration RC) and a roughage-only ration (ration R) were used; the roughage source differed between the facilities and comprised grass hay, grass silage, straw and lucerne hay. Dry matter intake ranged between 0.8 and 1.3% of body weight on ration RC and 0.5-1.2% on ration R. Digestibility coefficients achieved were similar to those reported for horses on diets of comparable composition. Endogenous losses as determined by linear regression analysis were within the range reported for horses. Measurements of faecal volatile fatty acids, faecal lactate and faecal pH also showed similarity to similar measurements in horses. The mean retention times of fluids (Co-EDTA) and particles (Cr-mordanted fibre <2 mm) in the whole gastrointestinal tract averaged 42 and 61 h, respectively, and were the longest ever recorded in a monogastric ungulate with this marker system. The results suggest that the horse is a useful model animal for designing diets for Indian rhinoceroses. Why digestive parameters are similar between these species in spite of enormous differences in body weight and retention times remains to be answered.     

The concentration of plasma triacylglycerols in horses fed diets containing either medium chain triacylglycerols or an isoenergetic amount of starch or cellulose.

In a Latin square design, six horses were fed hay and concentrates with isoenergetic amounts of either starch, cellulose or medium chain triacylglycerols (MCT). The dietary variables provided on average 22% of total dietary net energy. Plasma triacylglycerols and other variables of lipid metabolism were determined. The experimental periods lasted 21 days. Blood samples were taken just before the morning meal and three and six hours later. The diet rich in MCT significantly raised the plasma level of triacylglycerols when compared to either the starch- or cellulose-rich diet. The plasma concentrations of 3-hydroxybutyrate, total cholesterol and phospholipids were significantly higher when the horses were fed the ration with MCT instead of either cellulose or starch. Postprandial insulin concentrations were lowest for the MCT diet, and concentrations of free fatty acids were highest. Lipoprotein lipase activity was not significantly different for the three diets. Our study does not support the idea that cellulose feeding generates sufficient acetic acid in the caecum and colon, so that it would enhance the provision of cytosolic acetyl-CoA which in turn would stimulate hepatic fatty acid synthesis and then raise plasma triacylglycerols.     

The effect of oligofructose on urea metabolism and faecal odour components in cats

The effect on urea metabolism by the supplementation of oligofructose to a reduced protein diet was evaluated in cats by the use of labelled urea. The effect on faecal odour was also evaluated. Four cats were tested in a crossover study with two treatments: control and fructo-oligosaccharide (FOS). The FOS was supplemented at 3.11% on dry matter (DM) basis to a reduced protein diet (28.9% DM). After an adaptation period of 3 weeks, faeces and urine were collected during a 5-day collection period. Fresh faecal samples were collected for determination of odour components. On the first day of the collection period, labelled urea was injected subcutaneously. Urine production was estimated by potassium excretion. The fresh faecal samples were incubated in an air-closed recipient with two solid-phase micro extraction (SPME) fibres to bind the produced sulphur (S)-containing components. The reduced protein diet decreased plasma urea concentration but FOS supplementation had no effect. The tendency for a higher faecal output by FOS supplementation was the consequence of both an increased moisture content and faecal DM production. Supplementation of FOS showed tendencies to increase total faecal nitrogen (N) excretion and faecal (15)N excretion and tended to decrease urinary (15)N excretion. Twenty-seven different odour components were detected but were not affected by FOS supplementation.     

Study of crib-biting and gastric inflammation and ulceration in young horses

Nineteen young horses that had recently started to perform the stereotypy of crib-biting were compared with 16 non-stereotypic horses for 14 weeks. After initial observations of their behaviour and an endoscopic examination of the condition of their stomachs, the horses were randomly allocated to a control or an antacid diet At the start of the trial, the stomachs of the crib-biting foals were significantly more ulcerated and inflamed than the stomachs of the normal foals. In addition, the faecal pH of the crib-biting foals (6.05) was significantly lower than that of the normal foals (6.58). The antacid diet resulted in a significant improvement in the condition of the horses' stomachs. The crib-biting behaviour declined in most of the foals, regardless of their diet, but tended to decline to a greater extent in the foals on the antacid diet.     

Effects of dietary cation-anion balance on blood PH, acid base parameters, serum and urine mineral levels, and parathyroidhormone (PTH) in weanling horses.

This study demonstrated that the feeding of treatment diets with calculated dietary cation-anion balances (DCAB) of +370.43 (H) and -25.69 (L) did not have significant effects on blood pH, pCO₂, and HCO₃-. Serum Ca²⁺, P, Na⁺, and Cl^- as well as plasma PTH did not differ (P > .05) between the two treatment groups. Serum K⁺ was higher (P< .05) in horses fed diet H rather than diet L. The DCAB of the diet significantly affected urinary Ca²⁺, P, Na⁺, K⁺, and Cl^- excretion in the young growing horse. Urine Ca²⁺ and Cl- levels were higher (P < .01) in horses fed diet H versus diet L. Furthermore, levels of P, Na⁺, and K⁺ in the urine were higher (P < .01) in horses on diet H as opposed to diet L. Results of this study indicate that horses were able to maintain acid-base status regardless of diet. However, these data imply that growing horses consuming diets low in DCAB may be predisposed to abnormal bone mineralization due to the increase in calcium excretion which could lead to a weakening of the skeletal system.