The impact of phosphorus on the immune system and the intestinal microbiota with special focus on the pig.

There is increasing interest in dietary ingredients that are appropriate to support digestive and immune functions, but also maintain a stable microbial ecosystem in the gastrointestinal tract (GIT), particularly in weaned pigs. P is an essential nutrient for both microbes and their host, as it is involved, for example, in bone formation, energy metabolism, cellular signalling and stabilisation of cell membranes. Non-ruminant animals have limited access to phytate, the main storage form of P in plant seeds. The release of P bound to phytate requires phytase activity of plant or microbial origin, resulting in the formation of variable phosphorylated inositol phosphates (InsPs). The present review focuses on interactions between variations in dietary P supply, the immune system of the host, and the intestinal microbial ecosystem. Although results on the interaction between P and the immune system are inconsistent, several studies in different species have shown a positive impact of dietary P and phytase addition on the adaptive immune response. Recent studies with pigs suggest that P supply may influence intestinal microbial composition and activity. Individual InsPs or phosphate may also affect properties of pathogenic micro-organisms, such as metabolism or virulence. In conclusion, P may be considered as part of an integrated approach to support immune functions and maintain a stable microbial ecosystem in the GIT, thereby providing a barrier against potential pathogens. Within this regard, differences in phytate-P content and intrinsic phytase activity of plant feedstuffs, as well as the formation of individual InsPs, have to be taken into account.

Effect of increasing dietary fermentable fiber on diet nutrient digestibility and estimation of endogenous phosphorus losses in growing pigs.

Fermentable fiber may increase endogenous losses of phosphorus (EPL) and amino acids (AA), thereby reducing apparent nutrient digestibility. Acacia gum with medium-to-high fermentability and low viscosity was increasingly included in diets to investigate its effect on apparent ileal digestibility (AID) and apparent total tract digestibility (ATTD) of nutrients, gross energy (GE), and standardized total tract digestibility (STTD) of P in growing pigs. A control diet (49% cornstarch; 18% bovine plasma protein) was formulated to measure basal EPL. Three additional diets were formulated to include 2.5%, 5.0%, or 7.5% acacia gum at the expense of cornstarch. Diets contained 16.1% to 17.4% CP and 0.31% to 0.33% total P (DM-basis). The four diets were fed to eight ileal-cannulated barrows (initial BW, 54.6 kg) for four 9 d periods in a double 4 × 4 Latin square. Apparent hindgut fermentation (AHF) was calculated as ATTD minus AID. Feeding increasing acacia gum quadratically affected (P < 0.05) AID of DM, GE, linearly decreased (P < 0.05) ATTD of DM, crude protein (CP), GE, digestible (DE) and predicted net energy (NE) value of diets, and linearly increased (P < 0.001) AHF of DM and GE. Increasing acacia gum did not affect AID and standardized ileal digestibility (SID) of CP and AA. Basal EPL was 377 mg/kg DM intake (DMI) and increasing acacia gum linearly increased (P < 0.05) total tract EPL. Increasing acacia gum linearly decreased (P < 0.05) diet ATTD of P, and STTD of P based on either the calculated EPL or NRC (2012) recommended value (190 mg P/kg DMI). Increasing acacia gum did not affect AID and ATTD of Ca of diets. In conclusion, feeding increasing dietary fermentable, low-viscous acacia gum decreased diet AID and ATTD of DM and GE, but did not affect AID or SID of CP and AA. Increasing acacia gum decreased ATTD of P, which might have been due to increased specific endogenous losses of P in the total tract of growing pigs.

Fermentable fiber may increase endogenous losses of phosphorus (EPL) and amino acids (AA), thereby reducing nutrient digestibility. This study assessed effects of feeding increasing acacia gum with medium-to-high fermentability and low viscosity. Eight barrows cannulated at the terminal ileum were fed a control diet to measure basal endogenous P losses and three diets including 2.5%, 5.0%, or 7.5% acacia gum. Increasing acacia gum quadratically decreased diet ileal digestibility of dry matter (DM), gross energy (GE), linearly decreased diet total tract digestibility of DM, crude protein (CP), GE, and linearly increased hindgut fermentation of DM and GE. Increasing acacia gum linearly increased total tract endogenous P losses. Increasing acacia gum linearly decreased diet total tract digestibility of P, and standardized total tract digestibility of P calculated based on the calculated endogenous P losses or table values (NRC, 2012; 190 mg P/kg DMI). In conclusion, increasing dietary fermentable, low-viscous acacia gum decreased diet ileal and total tract digestibility of DM and GE, but did not affect ileal digestibility of CP and most AA. Increasing acacia gum inclusion reduced total tract digestibility of P, which might have been due to increased specific endogenous losses of P in the total tract of growing pigs.

Effect of dietary calcium concentration and exogenous phytase on inositol phosphate degradation, mineral digestibility, and gut microbiota in growing pigs.

Variations in the dietary Ca concentration may affect inositol phosphate (InsP) degradation, and thereby, P digestibility in pigs. This study assessed the effects of dietary Ca concentration and exogenous phytase on InsP degradation, nutrient digestion and retention, blood metabolites, and microbiota composition in growing pigs with ileal cannulation. In a completely randomized row-column design with four periods, eight ileal-cannulated barrows (initial body weight 27 kg) were fed four corn-soybean- and rapeseed meal-based diets containing 5.5 or 8.5 g Ca/kg dry matter (DM), with or without 1,500 FTU of an exogenous hybrid-6-phytase/kg diet. No mineral P was added and the P concentration in the feed was 4.8 g P/kg DM. Prececal InsP6 disappearance in pigs fed diets containing exogenous phytase was lower (P = 0.022) with additional Ca than without. Concentrations of InsP2-4 isomers and myo-inositol in the distal ileal digesta and prececal P digestibility were greater (P < 0.001) with exogenous phytase than without exogenous phytase. In feces, InsP6 disappearance was lower (P < 0.002) and concentration of InsP5 and InsP4 isomers was higher (P ≤ 0.031) with additional Ca compared to without additional Ca. The prececal amino acid digestibility, energy digestibility, and hindgut disappearance of energy did not differ. The Shannon diversity index of the microbiota in the distal ileal digesta and feces was similar among the diets but was lower in the distal ileal digesta than in the feces (P < 0.001). Permutation analysis of variance revealed no dietary differences between the bacterial groups within the ileal digesta and fecal samples (P > 0.05). In conclusion, additional Ca reduced the effect of exogenous phytase on prececal InsP6 degradation. Endogenous InsP degradation was impaired by additional Ca only in the hindgut but the abundance of bacterial genera in feces was not affected.

The dietary calcium concentration can influence the release of phosphorus from phytate in growing pigs. This study assessed the effects of dietary calcium and exogenous phytase on inositol phosphate (InsP) degradation and nutrient digestibility in ileal-cannulated, growing pigs. The phosphorus, calcium, and myo-inositol concentrations in the blood, microbiota composition in the ileal digesta and feces, and volatile fatty acid concentrations in the feces were also evaluated. Additional dietary calcium decreased prececal inositol hexakisphosphate (InsP6) disappearance, but only with exogenous phytase. Concentrations of InsP2-4 isomers and myo-inositol in the ileal digesta and prececal phosphorus digestibility were greater with exogenous phytase, but not affected by dietary calcium concentration. In contrast, fecal InsP6 disappearance was lower and the concentration of InsP4-5 isomers in feces was greater with additional dietary calcium. Regarding microbiota, the Shannon diversity index was lower in ileal digesta than in feces but was unaffected by dietary calcium concentration or exogenous phytase. In conclusion, dietary calcium concentration is relevant for phytate disappearance in feces, but not in the ileal digesta. However, when exogenous phytase is used, the dietary calcium concentration is important because prececal phytate degradation is changed.

Cereal grain fiber composition modifies phosphorus digestibility in grower pigs.

Increased fermentable carbohydrates (e.g., ß-glucan, amylose) may increase endogenous losses including for P, and thereby reduce apparent total tract digestibility (ATTD) of P. The present study assessed effects of barley cultivars varying in fermentable starch and fiber on apparent ileal digestibility (AID) and ATTD of P, myo-inositol 1,2,3,4,5,6-hexakis (dihydrogen phosphate; InsP6) and Ca, and standardized total tract digestibility (STTD) of P and the presence of lower inositol phosphates (InsP) compared to wheat. In a 6 (period) × 5 (diet) Youden square, seven ileal-cannulated barrows (initial BW, 27.7 kg) were fed diets containing 80% of one of five cereal grains differing in amylose, ß-glucan, and fiber content: 1) high-fermentable, high-ß-glucan, hull-less barley (HFB); 2) high-fermentable, high-amylose, hull-less barley (HFA); 3) moderate-fermentable, hull-less barley (MFB); 4) low-fermentable, hulled barley (LFB); and 5) low-fermentable, Canadian Western Red Spring wheat (LFW). On dry matter (DM) basis, cereal grains contained between 0.32% to 0.53% total P and 0.24% to 0.50% InsP6-P. The InsP6-2-P was calculated as the sum of all detected InsP-P (InsP6-P to InsP2-P) in the sample. The P release of degraded InsP-P was calculated by using the following equation: sum InsP6-2-Pdiet (g/kg DM) × (AID or ATTD sum InsP6-2-P (%)/ 100). Data were analyzed using a mixed model with diet as fixed effect, and pig and period as random effects. On DM basis, diets contained 41.4% to 50.6% starch, 0.88% to 8.54% ß-glucan, 0.81% to 0.89% total P, and 0.19% to 0.35% InsP6-P. The MFB, LFB, and LFW had greater (P < 0.05) diet AID of P than HFB and HFA, and MFB had greater (P < 0.05) diet ATTD and STTD of P than HFB. The ATTD of InsP6-P was greater (P < 0.05) for HFB than LFB and the ATTD of the sum InsP6-2-P was greater (P < 0.05) for HFB and HFA than LFB. Total tract P release was greater (P < 0.001) for HFB, HFA, and LFW than MFB and LFB. The LFW had greater (P < 0.05) ATTD of Ca than LFB. Diet ß-glucan content was not correlated with STTD of P (R2 = 0.03) or ATTD of InsP6 (R2 = 0.05). In conclusion, cereal grains high in fermentable fiber, e.g., amylose and ß-glucans included in specific hull-less barley cultivars, had lower diet AID, ATTD, and STTD of P, but greater ATTD of InsP6-P and sum InsP6-2-P. Carbohydrate fermentation, thus, results in greater total tract P release from InsP-P hydrolysis.

Increased fermentable carbohydrates (e.g., ß-glucan, amylose) may increase intestinal endogenous phosphorus (P) losses and thereby reduce P digestibility. The study assessed effects of cereal grains varying in fermentable carbohydrates on non-phytate-P and phytate-P. Phytate is the major binding form of P in plant seed and is incompletely degraded. Seven barrows cannulated at the terminal ileum were fed diets containing 80% of one of five cereal grains: 1) high-fermentable, high-ß-glucan, hull-less barley; 2) high-fermentable, high-amylose, hull-less barley; 3) moderate-fermentable, hull-less barley; 4) low-fermentable, hulled barley; and 5) low-fermentable wheat. Diet ileal digestibility of P was greater for low- and moderate-fermentable grain than high-fermentable grain, and diet total tract digestibility of P was greater for moderate-fermentable barley than high ß-glucan barley. Total tract digestibility of phytate-P was greater for high ß-glucan than low-fermentable barley. Total tract P release was greater for high-fermentable barley, and wheat than moderate- and low-fermentable barley. In conclusion, cereal grains high in fermentable fiber had lower diet ileal and total tract digestibility of P resulting in greater excretion of P, but greater total tract digestibility of phytate-P. Carbohydrate fermentation, thus, increases total tract P release from phytate-P degradation.

Effect of Fiber Fermentation and Protein Digestion Kinetics on Mineral Digestion in Pigs.

Nutrient kinetic data and the timing of nutrient release along the gastrointestinal tract (GIT), are not yet widely used in current feed formulations for pigs and poultry. The present review focuses on interactions between fermentable substrates (e.g., starch, fiber, and protein) and selected minerals on nutrient digestion and absorption to determine nutritional solutions to maximize animal performance, principally in the grower-finisher phase, with the aim of minimizing environmental pollution. For phosphorus (P), myo-inositol 1,2,3,4,5,6-hexakis (dihydrogen phosphate) (InsP6), copper (Cu), and zinc (Zn), no standardized methodologies to assess in vitro mineral digestion exist. The stepwise degradation of InsP6 to lower inositol phosphate (InsP) forms in the GIT is rare, and inositol phosphate4 (InsP4) might be the limiting isomer of InsP degradation in diets with exogenous phytase. Furthermore, dietary coefficients of standardized total tract digestibility (CSTTD) of P might be underestimated in diets with fermentable ingredients because of increased diet-specific endogenous P losses (EPL), and further clarification is required to better calculate the coefficients of true total tract digestibility (CTTTD) of P. The quantification of fiber type, composition of fiber fractions, their influence on digestion kinetics, effects on digesta pH, and nutrient solubility related to fermentation should be considered for formulating diets. In conclusion, applications of nutrient kinetic data should be considered to help enhance nutrient digestion and absorption in the GIT, thereby reducing nutrient excretion.

Effect of feeding acidified or fermented barley using Limosilactobacillus reuteri with or without supplemental phytase on diet nutrient digestibility in growing pigs.

Fermentation of cereal grains may degrade myo-inositol 1,2,3,4,5,6-hexakis (dihydrogen phosphate) (InsP6) thereby increasing nutrient digestibility. Effects of chemical acidification or fermentation with Limosilactobacillus (L.) reuteri with or without phytase of high ß-glucan hull-less barley grain on apparent ileal digestibility (AID) and apparent total tract digestibility (ATTD) of nutrients and gross energy (GE), standardized ileal digestibility (SID) of crude protein (CP) and amino acids (AAs), and standardized total tract digestibility (STTD) of P were assessed in growing pigs. Pigs were fed four mash barley-based diets balanced for water content: 1) unfermented barley (Control); 2) chemically acidified barley (ACD) with lactic acid and acidic acid (0.019 L/kg barley grain at a ratio of 4:1 [vol/vol]); 3) barley fermented with L. reuteri TMW 1.656 (Fermented without phytase); and 4) barley fermented with L. reuteri TMW 1.656 and phytase (Fermented with phytase; 500 FYT/kg barley grain). The acidification and fermentation treatments occurred for 24 h at 37 °C in a water bath. The four diets were fed to eight ileal-cannulated barrows (initial body weight [BW], 17.4 kg) for four 11-d periods in a double 4 × 4 Latin square. Barley grain InsP6 content of Control, ACD, Fermented without phytase, or Fermented with phytase was 1.12%, 0.59%, 0.52% dry matter (DM), or not detectable, respectively. Diet ATTD of DM, CP, Ca, and GE, digestible energy (DE), predicted net energy (NE) value, and urinary excretion of P were greater (P < 0.05) for ACD than Control. Diet ATTD of DM, CP, Ca, GE, DE and predicted NE value, urinary excretion of P was greater (P < 0.05), and diet AID of Ca and ATTD and STTD of P tended to be greater (P < 0.10) for Fermented without phytase than Control. Diet ATTD of GE was lower (P < 0.05) and diet ATTD and STTD of P, AID and ATTD of Ca was greater (P < 0.05) for Fermented with phytase than Fermented without phytase. Acidification or fermentation with/without phytase did not affect diet SID of CP and AA. In conclusion, ACD or Fermented without phytase partially degraded InsP6 in barley grain and increased diet ATTD of DM, CP, and GE, but not SID of CP and most AA in growing pigs. Fermentation with phytase entirely degraded InsP6 in barley grain and maximized P and Ca digestibility, thereby reducing the need to provide inorganic dietary P to meet P requirements of growing pigs.

Nutrient digestibility of extruded canola meal in ileal-cannulated growing pigs and effects of its feeding on diet nutrient digestibility and growth performance in weaned pigs.

Canola meal (CM) contains less crude protein (CP) and more fiber and anti-nutritional factors such as glucosinolates than soybean meal (SBM) and consequently has a lower nutrient digestibility. Therefore, processing strategies that may increase the feeding value of CM warrant study. In two experiments, the effects of extrusion of Brassica napus CM on apparent (AID) and standardized ileal digestibility (SID) of amino acids (AA), apparent total tract digestibility (ATTD) of gross energy (GE) in growing pigs, and growth performance and diet digestibility in weaned pigs were assessed. Solvent-extracted CM was extruded using a single-screw extruder at three screw speeds: 250 (CM-250), 350 (CM-350), or 450 (CM-450) rpm. In exp. 1, in a double 4 × 4 Latin square, eight ileal-cannulated barrows (initial body weight [BW], 68.1 kg) were fed corn starch-based diets containing 50% CM or extruded CM. The CM sample contained 43.2% CP, 33.2% total dietary fiber (TDF), and 8.9 µmol of total glucosinolates/g on a dry matter (DM) basis. Extrusion increased (P < 0.05) the AID of CP, reduced (P < 0.05) apparent hindgut fermentation of CP, and decreased (P < 0.05) predicted net energy (NE) value of diets. Extrusion increased diet AID and CM SID of most indispensable AA by 3.1 to 5.3%-units. In exp. 2, 200 weaned pigs (initial BW, 8.3 kg) were fed diets containing 20% SBM, CM, or extruded CM starting 2 wk postweaning for 3 wk. The CM sample contained 42.7% CP, 28.3% TDF, and 5.3 µmol total glucosinolates/g DM. Wheat-based diets provided 2.3 Mcal NE/kg and 5.1 g SID Lys/Mcal NE. Dietary inclusion of extruded CM replacing SBM decreased (P < 0.05) diet ATTD of DM, GE and CP, and DE value. Average daily feed intake, average daily gain (ADG), and gain:feed (G:F) of pigs did not differ between extruded CM and SBM diets and were not affected by extrusion, but increasing extruder screw speed linearly increased (P < 0.05) ADG for day 1 to 7 and G:F for the entire trial. In conclusion, extrusion increased diet AID and CM SID of AA but not DE and predicted NE values of CM. However, increasing extruder speed did not further increase the SID of most of the AA of CM in growing pigs. Dietary inclusion of 20% CM or extruded CM did not affect the growth performance in weaned pigs.

Digestibility of branched and linear α-gluco-oligosaccharides in vitro and in ileal-cannulated pigs.

Isomalto-oligosaccharides (IMOs) may promote health by modulating intestinal microbiota. We hypothesized that the proportion of α-(1 → 6) linkages in IMOs determines their digestibility. Ileal-cannulated pigs were fed diets containing IMO, IMO-DP3 with a greater DP and more α-(1 → 4) linkages, and digestible or resistant maltodextrins. Oligosaccharides were analysed by high-performance anion-exchange chromatography. Compared to IMO, IMO-DP3 contained more panose (18.6 vs. 10.3%) but less isomaltose (7.5 vs. 22.3%) and isomaltotriose (6.1 vs. 12.6%). The apparent ileal digestibility of dry matter were 3% greater for IMO-DP3 and digestible maltodextrin than resistant maltodextrin; the digestibility of IMO was not different from other oligosaccharides. Ileal propionate, isovalerate, and total SCFA was greater for IMO-DP3 and digestible maltodextrin than IMO. In conclusion, IMO was less digestible than IMO-DP3. Structural properties of IMOs are important determinants of their functional properties within the porcine digestive tract.

Dietary changes in nutritional studies shape the structural and functional composition of the pigs' fecal microbiome-from days to weeks.

BACKGROUND: The possible impact of changes in diet composition on the intestinal microbiome is mostly studied after some days of adaptation to the diet of interest. The question arises if a few days are enough to reflect the microbial response to the diet by changing the community composition and function. The present study investigated the fecal microbiome of pigs during a time span of 4 weeks after a dietary change to obtain insights regarding the time required for adaptation. Four different diets were used differing in either protein source (field peas meal vs. soybean meal) or the concentration of calcium and phosphorus (CaP). RESULTS: Twelve pigs were sampled at seven time points within 4 weeks after the dietary change. Fecal samples were used to sequence the 16S rRNA gene amplicons to analyse microbial proteins via LC-MS/MS and to determine the SCFA production. The analysis of OTU abundances and quantification values of proteins showed a significant separation of three periods of time (p = 0.001). Samples from the first day are used to define the 'zero period'; samples of weeks 1 and 2 are combined as 'metabolic period' and an 'equilibrium period was defined based on samples from weeks 3 and 4. Only in this last period, a separation according to the supplementation of CaP was significantly detectable (p = 0.001). No changes were found based on the corn-soybean meal or corn-field peas administration. The analysis of possible factors causing this significant separation showed only an overall change of bacterial members and functional properties. The metaproteomic approach yielded a total of about 9700 proteins, which were used to deduce possible metabolic functions of the bacterial community. CONCLUSIONS: A gradual taxonomic and functional rearrangement of the bacterial community has been depicted after a change of diet composition. The adaptation lasts several weeks despite the usually assumed time span of several days. The obtained knowledge is of a great importance for the design of future nutritional studies. Moreover, considering the high similarities between the porcine and human gastrointestinal tract anatomy and physiology, the findings of the current study might imply in the design of human-related nutritional studies.

Impact of a High-Fat or High-Fiber Diet on Intestinal Microbiota and Metabolic Markers in a Pig Model.

To further elaborate interactions between nutrition, gut microbiota and host health, an animal model to simulate changes in microbial composition and activity due to dietary changes similar to those in humans is needed. Therefore, the impact of two different diets on cecal and colonic microbial gene copies and metabolic activity, organ development and biochemical parameters in blood serum was investigated using a pig model. Four pigs were either fed a low-fat/high-fiber (LF), or a high-fat/low-fiber (HF) diet for seven weeks, with both diets being isocaloric. A hypotrophic effect of the HF diet on digestive organs could be observed compared to the LF diet (p < 0.05). Higher gene copy numbers of Bacteroides (p < 0.05) and Enterobacteriaceae (p < 0.001) were present in intestinal contents of HF pigs, bifidobacteria were more abundant in LF pigs (p < 0.05). Concentrations of acetate and butyrate were higher in LF pigs (p < 0.05). Glucose was higher in HF pigs, while glutamic pyruvic transaminase (GPT) showed higher concentrations upon feeding the LF diet (p < 0.001). However, C-reactive protein (CRP) decreased with time in LF pigs (p < 0.05). In part, these findings correspond to those in humans, and are in support of the concept of using the pig as human model.