Plant-oriented microbiome inoculum modulates age-related maturation of gut-mucosal expression of innate immune and barrier function genes in suckling and weaned piglets.

In the immediate time after weaning, piglets often show symptoms of gut inflammation. The change to a plant-based diet, lack of sow milk, and the resulting novel gut microbiome and metabolite profile in digesta may be causative factors for the observed inflammation. We used the intestinal loop perfusion assay (ILPA) to investigate jejunal and colonic expression of genes for antimicrobial secretion, oxidative stress, barrier function, and inflammatory signaling in suckling and weaned piglets when exposed to "plant-oriented" microbiome (POM) representing postweaning digesta with gut-site specific microbial and metabolite composition. Two serial ILPA were performed in two replicate batches, with 16 piglets preweaning (days 24 to 27) and 16 piglets postweaning (days 38 to 41). Two jejunal and colonic loops were perfused with Krebs-Henseleit buffer (control) or with the respective POM for 2 h. Afterward, RNA was isolated from the loop tissue to determine the relative gene expression. Age-related effects in jejunum included higher expression of genes for antimicrobial secretions and barrier function as well as reduced expression of pattern-recognition receptors post- compared to preweaning (P < 0.05). Age-related effects in the colon comprised downregulation of the expression of pattern-recognition receptors post- compared to preweaning (P < 0.05). Likewise, age reduced the colonic expression of genes encoding for cytokines, antimicrobial secretions, antioxidant enzymes, and tight-junction proteins post- compared to preweaning. Effect of POM in the jejunum comprised an increased the expression of toll-like receptors compared to the control (P < 0.05), demonstrating a specific response to microbial antigens. Similarly, POM administration upregulated the jejunal expression of antioxidant enzymes (P < 0.05). The POM perfusion strongly upregulated the colonic expression of cytokines and altered the expression of barrier function genes, fatty acid receptors and transporters, and antimicrobial secretions (P < 0.05). In conclusion, results indicated that POM signaled via altering the expression of pattern-recognition receptors in the jejunum, which in turn activated the secretory defense and decreased mucosal permeability. In the colon, POM may have acted pro-inflammatory via upregulated cytokine expression. Results are valuable for the formulation of transition feeds for the immediate time after weaning to maintain mucosal immune tolerance towards the novel digesta composition.

After weaning, piglets often show symptoms of gut inflammation and reduced performance. The plant-based diet, lack of sow milk, and the resulting novel gut microbiome and metabolite composition in digesta may be causative. However, the acute response of the gut mucosa when exposed to the novel digesta composition has not been fully elucidated. Here, we used the intestinal loop perfusion assay to characterize the immediate effect of a plant-oriented microbiome inoculum (POM) representing postweaning digesta composition on gene expression related to innate immune pathways and barrier function at the jejunal and colonic mucosa in suckling and weaned piglets. Results showed that the recognition of microbial components and barrier function changed in the jejunal and colonic mucosa from pre- to postweaning, indicating age-related maturation and priming by digesta compounds prior to the intestinal loop perfusion assay. In the jejunum, exposure to POM increased expression of receptors recognizing microbial components. In the colon, POM exposure upregulated the expression of genes for pro-inflammatory cytokines and other components of the first line of defense. Results have implications for the formulation of transition feeds for the immediate time after weaning. Inclusion of bioactive porcine milk components may help maintain mucosal immune tolerance towards the novel digesta composition.

Exposure to plant-oriented microbiome altered jejunal and colonic innate immune response and barrier function more strongly in suckling than in weaned piglets.

Weaning often leaves the piglet vulnerable to gut dysfunction. Little is known about the acute response of a gut mucosa primed by a milk-oriented microbiome before weaning to a plant-oriented microbiome (POM) after weaning. We evaluated the epithelial structure, secretory response and permeability in the small and large intestines of piglets receiving a milk-based (i.e., preweaning) or plant-based diet (i.e., postweaning) to POM inocula using intestinal loop perfusion assays (ILPA). The POM were prepared from jejunal and colonic digesta of four 7 week-old weaned (day 28 of life) piglets, having gut-site specific microbial and metabolite composition. Two consecutive ILPA were performed in 16 piglets pre- (days 24 to 27) and 16 piglets postweaning (days 38 to 41) in two replicate batches. Two jejunal and colonic loops per piglet were perfused with Krebs-Henseleit buffer (control) or the respective POM. The outflow fluid was analyzed for antimicrobial secretions. Jejunal and colonic loop tissue were collected after each ILPA for histomorphology and electrophysiology using Ussing chambers. ANOVA was performed using the MIXED procedure in SAS. The POM stimulated the secretory response by increasing mucin in the jejunal and colonic outflow by 99.7% and 54.1%, respectively, and jejunal IgA by 19.2%, whereas colonic lysozyme decreased 25.6% compared to the control (P < 0.05). Fittingly, the POM raised the number of goblet cells by 96.7% in jejunal and 56.9% in colonic loops compared to control loops (P < 0.05). The POM further flattened jejunal villi by 18.3% and reduced crypt depth in jejunal and colonic loops by 53.8% and 9.0% compared to the control (P < 0.05); observations typically made postweaning and indicative for mucosal recognition of 'foreign' compounds. The POM altered the jejunal and colonic net ion flux as indicated by 22.7% and 59.2% greater short-circuit current compared to control loops, respectively; the effect being stronger postweaning (P < 0.05). Colonic barrier function improved with age (P < 0.05), whereas POM perfusion compromised the mucosal barrier as suggested by 17.7% and 54.1% greater GT and mucosal-to-serosal flux of fluorescein-isothiocyanate dextran, respectively, compared to the control (P < 0.05). In conclusion, results demonstrated that the preweaning gut epithelium acutely responds to novel compounds in postweaning digesta by upregulating the first line of defense (i.e., mucin and lysozyme secretion) and impairment of the structural integrity.

Creep feed is offered during the suckling period to prepare the piglet's gut for the dietary transition from a milk- to a plant-based diet at weaning. Nevertheless, the discontinuation of sow milk consumption after weaning can lead to disturbed interactions between the host mucosa and the gut microbiota. Little information is available on the immediate mucosal response towards the altered microbial and metabolite composition in digesta. Therefore, the main objective of this study was to evaluate the immediate effect of the exposure of the jejunal and colonic mucosa to a plant-oriented microbiome (POM), prepared from intestinal digesta of weaned pigs, on the mucosal structure, secretory response, and permeability in piglets before and after weaning using the intestinal loop perfusion assay. The perfusion with POM stimulated the host's secretory response, altered the gut structure and decreased the epithelial integrity before and after weaning. Effects were less strong postweaning, indicating that adaptation processes at the gut epithelium occurred from pre- to postweaning which increased the tolerance towards the POM inoculum.

Evaluation of serum parameters to predict the dietary intake of calcium and available phosphorus in growing pigs.

Adequate provision of calcium (Ca) and phosphorus (P) is essential for bone formation and high growth performance in pigs. Nevertheless, reliable serum biomarkers for pig's Ca and P intake are still missing. Here, we used phytase supplementation to alter the dietary available P (aP) level in order to investigate the effect of differences in dietary aP levels on serum parameters related to the Ca and P homeostasis in pigs. Moreover, we assessed whether serum parameters can be used to predict the Ca, total P (tP), and aP intake in barrows and gilts throughout the fattening period. In total, 216 pigs (115 gilts and 101 barrows) were randomly allotted to one of the two diets in three replicate batches, each lasting 56 d (n = 108/diet). Pigs had free access to the diets without (Con) or with phytase (Phy; 650 phytase units/kg) via a transponder-based feeding system. Blood samples were collected on days 2, 23, and 52, and serum parameters were correlated with the daily Ca, tP, and aP intake. The intake of tP, aP, and Ca was overall 14.2%, 13.8%, and 14.2% higher in barrows compared with gilts, respectively (P < 0.001). Concurrently, phytase decreased the intake of tP and Ca by 8.4% and 6.7%, respectively, whereas it raised the intake of aP by 16.3% compared with the Con diet (P < 0.001). Serum levels of fibroblast growth factor 23, alkaline phosphatase (ALP), vitamin D (VitD), and osteocalcin (OCN) decreased with age (P < 0.05). The higher aP intake of pigs fed the Phy diet increased serum P on days 2 and 23 but decreased it on day 52 compared with the Con diet (P = 0.004). Pigs fed the Phy diet had higher serum ALP compared with pigs fed the Con diet on days 23 and 52 (P < 0.05). Correlation analysis between serum parameters and Ca, tP, and aP intake showed age- and sex-related associations. With 12 wk of age, serum P in both sexes, serum VitD in barrows, and serum OCN and ALP in gilts correlated with aP intake (|r| > 0.38), whereas serum OCN correlated with Ca in both sexes' intake (r > 0.50). At 20 wk, serum Ca and ALP in gilts correlated with aP intake, whereas serum P, Ca, and VitD correlated with Ca intake in both sexes (|r| > 0.39). In conclusion, the present results showed that the daily Ca and aP intake could be most reliably estimated from serum parameters for an approximate age of 12 and 20 wk. Serum P and the Ca:P ratio at 12 wk of age and serum VitD at 20 wk of age may be used to predict pig's daily aP intake in both sexes.

Dietary Phytase- and Lactic Acid-Treated Cereals Caused Greater Taxonomic Adaptations than Functional Adaptations in the Cecal Metagenome of Growing Pigs.

Phosphorus (P) is an essential nutrient for the gut bacteria and the host. Nevertheless, little information exists that indicates to what extent an improved level of P availability in the small intestine leads to functional adaptations in bacterial metabolic pathways in the large intestine. Therefore, we investigated the changes in the taxonomic and functional bacterial metagenome in cecal digesta of growing pigs fed diets containing phytase and/or cereals treated with 2.5% lactic acid (LA) for 19 days (n = 8/diet) using shotgun metagenome sequencing. The phytase supplementation resulted in strikingly distinct bacterial communities, affecting almost all major bacterial families, whereas functional changes were less dramatic among the feeding groups. While phytase treatment decreased predominant Prevotellaceae levels, it seemed that Clostridiaceae, Ruminococcaceae, and Lachnospiraceae filled the opening metabolic niches (P < 0.05). The LA-treated cereals mediated reduced levels of Bacteroidaceae and increased levels of Veillonellaceae, but those results were mainly seen when the cereals were fed as a single treatment (P < 0.05). In association with the taxonomic alterations, phytase caused changes within the major functional pathways corresponding to amino acid metabolism; translation; membrane transport; folding, sorting, and degradation; and energy metabolism, whereas the LA treatment of cereals resulted in decreased enzymatic capacities within the carbohydrate metabolism and energy metabolism pathways (P < 0.05). Metabolic dependencies corresponding to the starch and sucrose metabolism, glycolysis/gluconeogenesis, and citrate cycle pathways were indicated by diet-associated changes in enzymatic capacities related to short-chain fatty acid, methane, vitamin, and bacterial antigen synthesis. Accordingly, the present results support the idea of the importance of the availability of intestinal P for bacterial metabolism. However, the functional profiles were less different than the taxonomic profiles among the dietary treatment results, indicating a certain degree of metabolic plasticity within the cecal metagenome.IMPORTANCE Dietary strategies (e.g., phytase supplementation and lactic acid [LA] treatment of cereals) used to improve the availability of phytate-phosphorus (P) from pig feed reduce the amount of P flowing into the large intestine, whereas LA treatment-induced changes in nutrient fractions alter the substrate being available to the microbiota. In ruminants, lower intestinal P availability compromises the fibrolytic activity of the microbiome. Here, we report that the functional capacities were less dramatically affected than the taxonomic composition by phytase-supplemented and LA-treated cereals. The bacterial community appeared to be partly capable of functionally compensating for the altered flow of P by replacing taxa with higher P needs by those with lower P needs. Therefore, by acting as mucosal immune stimulants, alterations in microbiota-associated molecular patterns (MAMPs) due to the taxonomic shifts may play a greater role for host physiology and health than functional differences caused by differing intestinal P availabilities, which merits further research.

Dietary Phytase and Lactic Acid-Treated CerealGrains Differently Affected Calcium and PhosphorusHomeostasis from Intestinal Uptake to SystemicMetabolism in a Pig Model.

High intestinal availability of dietary phosphorus (P) may impair calcium (Ca)homeostasis and bone integrity. In the present study, we investigated the effect of phytasesupplementation in comparison to the soaking of cereal grains in 2.5% lactic acid (LA) on intestinalCa and P absorption; intestinal, renal, and bone gene expression regarding Ca and P homeostasis;bone parameters; and serum levels of regulatory hormones in growing pigs. Thirty-two pigs wererandomly assigned to one of four diets in a 2 × 2 factorial design in four replicate batches for 19days. The diets comprised either untreated or LA-treated wheat and maize without and withphytase supplementation (500 phytase units/kg). Although both treatments improved the Pbalance, phytase and LA-treated cereals differently modulated gene expression related to intestinalabsorption, and renal and bone metabolism of Ca and P, thereby altering homeostatic regulatorymechanisms as indicated by serum Ca, P, vitamin D, and fibroblast growth factor 23 levels.Moreover, phytase increased the gene expression related to reabsorption of Ca in the kidney,whereas LA-treated cereals decreased the expression of genes for osteoclastogenesis in bones,indicating an unbalanced systemic availability of minerals. In conclusion, high intestinalavailability of dietary P may impair Ca homeostasis and bone integrity.

A multicomponent mycotoxin deactivator modifies the response of the jejunal mucosal and cecal bacterial community to deoxynivalenol contaminated feed and oral lipopolysaccharide challenge in chickens1.

Mycotoxin deactivators are a widely used strategy to abrogate negative effects of mycotoxin-contaminated feed. It has not been adequately evaluated whether these deactivators may detoxify bacterial toxins in the intestinal lumen and subsequently lower the inflammatory response in chickens. The present objective was to study the effect of a multicomponent mycotoxin deactivator (B), containing a bentonite and a bacterial strain capable to enzymatically biotransform trichothecenes especially deoxynivalenol (DON), when supplemented to a DON-contaminated feed in combination with an oral lipopolysaccharide challenge on visceral organ size, expression of innate immune genes and mucosal permeability in the small intestine as well as on the cecal bacterial composition and metabolites in broiler chickens. Eighty 1-d-old male chickens were randomly allotted to four treatment groups in two replicate batches (n = 10/treatment/replicate): 1) basal diet without DON (CON), 2) CON diet supplemented with B (2.5 mg B/kg feed) (CON-B), 3) CON diet contaminated with 10 mg DON/kg feed (DON), and 4) DON diet supplemented with 2.5 mg B/kg feed (DON-B). In half of the chickens per treatment, effects were assessed under nonchallenge conditions, whereas in the other half of birds, to increase their intestinal bacterial toxin load, effects were tested after an oral challenge with 1 mg LPS/kg BW from Escherichia coli O55:B5 on the day before sampling. DON reduced (P < 0.05) the weight of bursa fabricii and thymus. DON increased the expression level of intestinal alkaline phosphatase at the duodenal mucosa (P = 0.027) but did not modify jejunal gene expression and mucosal permeability. The LPS challenge decreased the jejunal MUC2 expression but increased ZO1 and IL6 expression compared to the unchallenged animals (P < 0.05). DON × B interactions indicated lower expression of IL10 in duodenum and NFKB in jejunum with the B diet but higher expression with the DON-B diet (P = 0.050). Furthermore, the B lowered jejunal expression of NFKB and IL6 but only in LPS-challenged chickens (P < 0.05). Alterations in the cecal microbiota composition and VFA profile were likely associated with alterations in host physiology in the small intestine caused by DON, B, and LPS. According to the present data, B appeared to have potential to detoxify antigens other than DON in the intestinal lumen of chickens, whereby the toxin load may limit the efficacy of B to modify the intestinal and systemic response as indicated by interactions of DON, B, and LPS.

Alterations of the Viable Ileal Microbiota of the Gut Mucosa-Lymph Node Axis in Pigs Fed Phytase and Lactic Acid-Treated Cereals.

The gut-lymph node axis is a critical player in the symbiotic relationship between gut microbiota and the host. However, little is known about the impact of diet-related bacterial shifts in the gut lumen on bacterial translocation into lymph nodes. Here, we (i) characterized changes in the viable microbiota composition along the ileal digesta-mucosa-lymph node axis and (ii) examined the effect of dietary phytase supplementation and lactic acid (LA) soaking of cereals on the bacterial taxonomy along this axis, together with their effect on the mucosal expression of innate immune and barrier function genes in pigs (n = 8/diet). After 18 days on diets, ileal digesta, mucosa, and ileocecal lymph nodes (ICLNs) were collected for RNA isolation and 16S rRNA-based high-resolution community profiling. Bacterial communities were dominated by Lactobacillaceae and Clostridiaceae, with clearly distinguishable profiles at the three sampling sites. Specific bacterial subsampling was indicated by enrichment of the ICLNs with Lactobacillaceae, Lachnospiraceae, Veillonellaceae, and Methanobacteriaceae and less Clostridiaceae, Pasteurellaceae, Helicobacteraceae, and Enterobacteriaceae compared to that of the mucosa. LA treatment of cereals reduced proteolytic taxa in the lumen, including pathobionts like Helicobacteraceae, Campylobacteraceae, and Fusobacteriaceae When combined, phytase- and LA-treated cereals largely increased species richness, while the single treatments reduced Actinobacteria and Bacteroidetes in ICLNs and increased mucosal MUC2 expression. In contrast, phytase reduced mucosal CDH1 expression, indicating altered barrier function with potential effects on bacterial translocation. Overall, both treatments, although often differently, changed the viable microbiome along the digesta-mucosa-lymph node axis in the ileum, probably due to altered substrate availability and microbial-host interactions.IMPORTANCE A host's diet largely determines the gut microbial composition and therefore may influence bacterial translocation into ICLNs. Due to its importance for cell metabolism, the intestinal phosphorus availability, which was modified here by phytase and LA treatment of cereals, affects the intestinal microbiota. Previous studies mainly focused on bacteria in the lumen. The novelty of this work resides mainly in that we report diet-microbe effects along the digesta-mucosa-ICLN axis and linked those effects to mucosal expression of barrier function genes as crucial components for host health. Lymph nodes can serve as reservoir of pathobionts; therefore, present diet-microbiome-host interactions have implications for food safety.

Lactic Acid Treatment of Cereals and Dietary Phytase Modified Fecal Microbiome Composition Without Affecting Expression of Virulence Factor Genes in Growing Pigs.

Besides the major nutrients, phosphorus (P) is an essential mineral for both the host animal and the porcine gut microbiota. Different strategies including phytase supplementation and more recently lactic acid (LA) are used to enhance the P availability from cereals in pig diets; however, their impact on the gut microbiota has been rarely related to fecal shedding of opportunistic pathogens. The present study investigated the effect of phytase supplementation and the treatment of dietary cereals with 2.5% LA on the fecal microbiome composition of metabolic active bacteria and expression of virulence factor genes of enterotoxigenic Escherichia coli and Clostridium perfringens in growing pigs. Phytase supplementation reduced the fecal abundance of the most abundant Lactobacillaceae family, whereas the LA-treatment of cereals had a stronger impact on the bacterial community, reducing amylolytic, pullulanolytic and hemicellulolytic Lactobacillaceae, Lachnospiraceae and Ruminococcaceae as well as the fecal bacterial species richness (Chao1) and diversity (Shannon index). Mainly the family Clostridiaceae benefited from the decline in the aforementioned families, being enriched by both dietary treatments. Multigroup data integration using sparse partial least squares-discriminant analysis showed that among the most discriminative operational taxonomic units (OTU) especially two unclassified Clostridiaceae-OTUs, one Prevotella copri-like OTU and one OTU within the vadinCA11 group were associated with calcium and P levels but were negatively linked with complex carbohydrates in feces. Heat-stable toxin A (Sta) of enterotoxigenic E. coli and Stx2e of Shiga-toxin producing E. coli were expressed in feces but were similar among feeding groups. Without modifying the total bacterial gene copies and virulence factor expression of E. coli, both dietary phytase supplementation and LA-treatment of cereals drastically altered the bacterial community composition in pig feces. Results thereby allowed for the characterization of bacterial nutrient dependencies, indicating a link between fecal P availability, complex carbohydrate composition and alterations in the predominant genera.

Improvement of Feed Efficiency in Pigs through Microbial Modulation via Fecal Microbiota Transplantation in Sows and Dietary Supplementation of Inulin in Offspring.

As previous studies have demonstrated a link between the porcine intestinal microbiome and feed efficiency (FE), microbiota manipulation may offer a means of improving FE in pigs. A fecal microbiota transplantation procedure (FMTp), using fecal extracts from highly feed-efficient pigs, was performed in pregnant sows (n = 11), with a control group (n = 11) receiving no FMTp. At weaning, offspring were allocated, within sow treatment, to (i) control (n = 67; no dietary supplement) or (ii) inulin (n = 65; 6-week dietary inulin supplementation) treatments. The sow FMTp, alone or in combination with inulin supplementation in offspring, reduced offspring body weight by 8.1 to 10.6 kg at ∼140 days of age, but there was no effect on feed intake. It resulted in better FE, greater bacterial diversity, and higher relative abundances of potentially beneficial bacterial taxa (Fibrobacter and Prevotella) in offspring. Due to the FMTp and/or inulin supplementation, relative abundances of potential pathogens (Chlamydia and Treponema) in the ileum and cecal concentrations of butyric acid were significantly lower. The maternal FMTp led to a greater number of jejunal goblet cells in offspring. Inulin supplementation alone did not affect growth or FE but upregulated duodenal genes linked to glucose and volatile fatty acid homeostasis and increased the mean platelet volume but reduced ileal propionic acid concentrations, granulocyte counts, and serum urea concentrations. Overall, the FMTp in pregnant sows, with or without dietary inulin supplementation in offspring, beneficially modulated offspring intestinal microbiota (albeit mostly low-relative-abundance taxa) and associated physiological parameters. Although FE was improved, the detrimental effect on growth limits the application of this FMTp-inulin strategy in commercial pig production.IMPORTANCE As previous research suggests a link between microbiota and FE, modulation of the intestinal microbiome may be effective in improving FE in pigs. The FMTp in gestating sows, alone or in combination with postweaning dietary inulin supplementation in offspring, achieved improvements in FE and resulted in a higher relative abundance of intestinal bacteria associated with fiber degradation and a lower relative abundance of potential pathogens. However, there was a detrimental effect on growth, although this may not be wholly attributable to microbiota transplantation, as antibiotic and other interventions were also part of the FMT regimen. Therefore, further work with additional control groups is needed to disentangle the effects of each component of the FMTp in order to develop a regimen with practical applications in pig production. Additional research based on findings from this study may also identify specific dietary supplements for the promotion/maintenance of the microbiota transferred via the maternal FMTp, thereby optimizing pig growth and FE.

Deoxynivalenol in the Diet Impairs Bone Mineralization in Broiler Chickens.

Deoxynivalenol (DON) is one of the most abundant and important trichothecene mycotoxins produced by Fusarium species. In chickens, DON intake causes feed refusal, impairs performance, gut barrier function, and immunity, and raises oxidative stress. To determine the effect of DON on bone mineralization and serum calcium and phosphorus, 80 newly-hatched chickens were fed 4 diets with 0, 2.5, 5, and 10 mg DON/kg feed in this pilot study. In week 5, chickens were euthanized, femur and tibiotarsus bones were separated from the meat, and after incineration ash composition, as well as serum calcium and phosphorus, were determined using clinical biochemistry. Dietary DON reduced chicken dry matter, calcium, and phosphorus intake, and subsequently body and leg weight. DON affected bone density and composition of the tibiotarsus more drastically than of the femur. However, lower mineral intake did not solely explain our observations of the quadratically lower tibiotarsus density and ash content, as well as linearly decreased Ca content in the femur and tibiotarsus with increasing DON levels. Linearly decreasing serum phosphorus concentrations with increasing DON levels further supported impaired mineral homeostasis due to DON. In conclusion, already low dietary DON contamination of 2.5 mg/kg feed can compromise bone mineralization in chickens.

Lactic acid treatment of by-products and phosphorus level in the diet modulate bacterial microbiome and the predicted metagenome functions using the rumen simulation technique.

This study used a rumen simulation technique to evaluate the effects of soaking of by-product-rich concentrate (BPC) in 5% lactic acid (LAC; vol/vol) on the rumen microbiota, predicted metagenome, fermentation characteristics, and nutrient degradation without or with supplemented P. The diet was supplemented with 1.6 g of P in the form of monocalcium phosphate per kilogram of dry matter in addition to 284 mg of inorganic P/d per fermentor via artificial saliva. Fermentor fluid was collected for analyses of short-chain fatty acids, fermentation gases, redox potential, and microbiota and feed residues for calculation of nutrient degradation. The microbiota composition was assessed using paired-end Illumina (Illumina Inc., San Diego, CA) MiSeq sequencing of the V3 to V5 region of the 16S rRNA gene. Soaking in LAC reduced the contents of crude protein, neutral and acid detergent fibers, and organic matter fractions as well as ash and P content of the BPC. Both the LAC treatment of BPC and the inorganic P modified the relative bacterial abundances mainly within the predominant orders Bacteroidales and Clostridiales. Supervised DIABLO N-integration networking supported that operational taxonomic units related to BS11, Ruminococcaceae, Christensenellaceae, Eubacterium, and Selenomonas were the most discriminant for the LAC-treated BPC, whereas other operational taxonomic units related to BS11, RFN20, Ruminococcus, and Succiniclasticum were best correlated with the inorganic P supplementation. Integration networking also showed that carbohydrate and pyruvate metabolism, biosynthesis of unsaturated fatty acids, and degradation of several xenobiotics were stimulated by the LAC treatment of BPC. Those data supported the enhanced fermentation activity as indicated by increased total short-chain fatty acid concentration, especially propionate and butyrate, and methane, but decreased ruminal crude protein degradation, with the LAC-treated compared with control-treated BPC. In contrast, despite an increased abundance of imputed functions, such as inositol phosphate metabolism, phosphatidylinositol signaling, and fructose and mannose metabolism, the reduced abundance of the imputed Kyoto Encyclopedia of Genes and Genomes pathway "transcription machinery" as well as the decrease in total short-chain fatty acids and nutrient degradation indicated reduced bacterial metabolic activity with the inorganic P supplementation. In conclusion, soaking of BPC in LAC may favor the proliferation of certain fibrolytic bacterial taxa and stimulate their metabolic activity, whereas the supplemented P to a diet already meeting ruminal P needs may impair ruminal nutrient utilization.

Graded replacement of maize grain with molassed sugar beet pulp modulated ruminal microbial community and fermentation profile in vitro.

BACKGROUND: Molassed sugar beet pulp (Bp) is a viable alternative to grains in cattle nutrition for reducing human edible energy input. Yet little is known about the effects of high inclusion rates of Bp on rumen microbiota. This study used an in vitro approach and the quantitative polymerase chain reaction technique to establish the effects of a graded replacement of maize grain (MG) by Bp on the ruminal microbial community, fermentation profile and nutrient degradation. RESULTS: Six different amounts of Bp (0-400 g kg-1 ), which replaced MG in the diet, were tested using the in vitro semi-continuous rumen simulation technique. The increased inclusion of Bp resulted in greater dietary content and degradation of neutral detergent fibre (P < 0.01). Further, Bp feeding enhanced (P < 0.01) the abundance of genus Prevotella and shifted (P < 0.01) the short-chain fatty acid patterns in favour of acetate and propionate and at the expense of butyrate. A total replacement of MG with Bp resulted in an increased daily methane production (P < 0.01). CONCLUSION: Results suggest positive effects of the replacement of MG by Bp especially in terms of stimulating ruminal acetate and propionate fermentation. However, high replacement rates of Bp resulted in lowered utilization of ammonia and higher ruminal methane production. © 2017 Society of Chemical Industry.

Changing dietary calcium-phosphorus level and cereal source selectively alters abundance of bacteria and metabolites in the upper gastrointestinal tracts of weaned pigs.

Several dietary ingredients may affect the bacterial community structure and metabolism in the porcine gut and may therefore influence animals' health and performance. This study investigated the effects of cereal source and calcium-phosphorus (CaP) level in the diet on bacterial microbiota and metabolites, nutrient intake, and gut environment in weaned pigs. Pigs (n=8/treatment) were fed wheat-barley- or corn-based diets with an adequate or high CaP level for 14 days. Effects on microbiota in the stomach, ileum, and midcolon were assessed using quantitative PCR. Data showed that Enterobacteriaceae, Campylobacter spp., and Helicobacter spp., which all contain highly immune reactive lipopolysaccharide (LPS), were abundant at all gut sites. Diet effects on bacteria and metabolites were moderate and occurred mainly in the upper gut, whereas no effects on bacteria, fermentation products, and LPS could be observed in the colon. Differences in carbohydrate intake with corn versus wheat-barley diets selectively stimulated Bifidobacterium in the stomach and ileum. There was a growth advantage for a few bacterial groups in the stomach and ileum of pigs fed the high versus adequate CaP level (i.e., gastric Enterobacteriaceae and ileal Enterococcus, Bacteroides-Prevotella-Porphyromonas, and Campylobacter). Interestingly, gastrointestinal pH was not affected by dietary CaP level. The present findings demonstrate the stability of the bacterial community and gut environment toward dietary changes even in young pigs. The results on stimulation of gastric and ileal Bifidobacterium by corn diets may be employed in nutritional strategies to support gut health after weaning.

Oat ß-glucan and dietary calcium and phosphorus differentially modify intestinal expression of proinflammatory cytokines and monocarboxylate transporter 1 and cecal morphology in weaned pigs.

Physiologic effects of dietary oat ß-glucan and low and high dietary calcium-phosphorus (CaP) on intestinal morphology and gene expression related to SCFA absorption, mucus production, inflammation, and peptide digestion have not been established in weaned mammals. We therefore randomized 32 weaned pigs into 4 equal groups that received a cornstarch-casein-based diet with low (65% of the Ca and P requirement) and high (125 and 115% of the Ca and P requirement, respectively) CaP levels and low- and high-CaP diets supplemented with 8.95% oat ß-glucan concentrate for 14 d. High-CaP diets downregulated duodenal expression of IL-1ß (P < 0.05) by 30% compared with low-CaP diets. Furthermore, high-CaP diets reduced (P < 0.05) cecal crypt depth by 14% compared with low-CaP diets. Dietary ß-glucan upregulated the expression of cecal MCT1 (P < 0.05) by 40% and that of colonic IL-6 (P < 0.05) by 142% compared with the control diet. Correlation analysis indicated that cecal MCT1 (r = 0.99, P < 0.001) and colonic IL-6 (r = 0.84, P < 0.05) expression was positively related to luminal butyrate and total SCFA, respectively, indicating that ß-glucan may partly modify gene expression via increased SCFA generation. In conclusion, ß-glucan and CaP levels modulated the expression of selected genes and morphology in the postweaning period, but effects were specific to intestinal segment. The present results further indicate that, in addition to being essential nutrients for bone accretion, dietary CaP level may modify the intestinal tissue response in young pigs.

Dietary calcium phosphate content and oat ß-glucan influence gastrointestinal microbiota, butyrate-producing bacteria and butyrate fermentation in weaned pigs.

This study aimed to evaluate the effects of oat ß-glucan in combination with low- and high-dietary calcium phosphate (CaP) content on gastrointestinal bacterial microbiota, prevalence of butyrate-production pathway genes and fermentation end-products in 32 weaned pigs allocated to four diets: a cornstarch-casein-based diet with low [65% of the calcium (Ca) and phosphorous (P) requirement] and high CaP content (125% and 115% of the Ca and P requirement, respectively); and low and high CaP diets supplemented with 8.95% of oat ß-glucan concentrate. Pigs were slaughtered after 14 days, and digesta were collected for quantitative PCR analysis, and quantification of short-chain fatty acids and lactate. The high CaP content reduced gastric lactate and streptococci and propionate in the large intestine. Oat ß-glucan distinctly raised gastric bacterial numbers, and colonic lactobacilli and bifidobacteria. Although not reflected by gene copies of butyrate-production pathway genes, oat ß-glucan also increased gastric, caecal and colonic butyrate concentrations, which may be favourable for intestinal development in weaned pigs. Thus, a high CaP content negatively affected the intestinal abundance of certain fermentation end-products, whereas oat ß-glucan generally enhanced bacterial numbers and activity. The results emphasize the importance of the stomach for bacterial metabolism of oat ß-glucan in weaned pigs.

Effects of betaine, organic acids and inulin as single feed additives or in combination on bacterial populations in the gastrointestinal tract of weaned pigs.

A study was carried out to investigate whether blends of betaine, organic acids and inulin may improve their efficacy to modulate intestinal bacterial populations in weaned pigs compared with the single application of these additives. Moreover, potential postprandial diurnal variations in ileal bacterial numbers were determined in piglets fed the control diet. Twenty-four piglets in two consecutive experiments received a wheat-barley-soybean meal control diet (Con) or the Con diet supplemented with betaine (BET; 0.2%), an organic acid blend (AC; 0.4%) or inulin (IN; 0.2%) as single additives or in combination. Ileal bacterial numbers of total bacteria, lactobacilli, bifidobacteria (p < 0.05) and enterobacteria (p < 0.10) showed a postprandial diurnal variation, thus spot sampling of ileal digesta for the determination of bacterial numbers may not be representative. There were only small effects of BET, AC and IN on ileal and faecal bacterial populations. BET + AC increased total ileal bacterial numbers compared to the Con and AC treatments. BET reduced lactobacilli numbers in faeces, whereas BET + IN increased ileal numbers of bifidobacteria compared to AC and IN. There is evidence that BET, AC, IN and their combinations may affect proliferation of beneficial bacterial populations, although this has to be confirmed in further studies.