Acinetobacter in fresh faecal samples is related to sperm motility: A case report in Piétrain boars housed on different deep-litter bedding materials.

The swine gastrointestinal tract contains a great variety of microbes, forming a complex and dynamic ecosystem. Various internal and external factors (e.g. age, breed and diet) may influence its composition. This study aimed to investigate the gut microbial diversity of German Piétrain boars housed on different deep-litter bedding materials (regional wood shavings, linen, hemp, spelt husks, and wood shavings) via 16S-rDNA sequencing. Additionally, short-chain fatty acids were analysed using gas chromatography. Fresh faecal samples (n = 80) from 40 Piétrain boars were collected twice during the trial. Although it can be assumed that boars ingest bedding orally, no differences in the microbiome composition could be found. The main phyla were Firmicutes and Bacteroides. Acinetobacter was identified as a biomarker for sperm quality differences (total sperm motility) in breeding boars.

Influence of high- and low-fermentable dietary fibres in sows' diet on the colostrum potential against Clostridioides difficile toxin-induced effects in IPEC-J2 cells.

Sow colostrum has been reported to protect the IPEC-J2 cells and piglet colon tissues from detrimental effect of Clostridioides difficile toxins. Since dietary fibre can influence the colostrum composition in sows, we hypothesised that it can also differentially affect the colostrum potential against C. difficile toxin-induced effects in IPEC-J2. IPEC-J2 were incubated with colostrum from sows fed either high-fermentable sugar beet pulp (SBP) or low-fermentable lignocellulose (LNC) fibres and in combination with the toxins and analysed by trans-epithelial electrical resistance (TEER) and cell viability using propidium iodide in flow cytometry. Toxins drastically decreased the integrity of IPEC-J2. Colostrum from the sows fed either SBP or LNC exerted protective effect against toxins on IPEC-J2 integrity and this effect was numerically superior in the SBP group. Differences in the percentages of TEER between different treatments were noted after 2 h (p = 0.043), 3 h (p = 0.017) and 4 h (p = 0.017) of incubation and a tendency for differences was noted after 5 h of incubation (p = 0.071). Colostrum from either SBP- or LNC-fed sows did not protect the IPEC-J2 from toxin-induced death. Colostrum of the sows fed either high-fermentable or low-fermentable fibres has a potential to protect IPEC-J2 from the loss of integrity, which may be important in protection from C. difficile-infection development in neonatal piglets.

Fiber Composition in Sows' Diets Modifies Clostridioides difficile Colonization in Their Offspring.

Dietary fiber has a potential to modulate the gut microbiota in sows. We hypothesized that a maternal diet rich in either high- or low-fermentable fiber during gestation and lactation influences Clostridioides difficile gut colonization in suckling piglets. Twenty sows were fed gestation and lactation diets enriched with either high-fermentable sugar beet pulp (SBP) or low-fermentable lignocellulose (LNC) fibers. C. difficile, toxin B (TcdB), fecal score, microbial abundance (16S-rDNA sequencing) and metabolites were measured in the feces from the sows and their piglets. C. difficile concentration was higher in piglets from the sows fed LNC than SBP along the study (P ≤ 0.05). Higher prevalence of C. difficile was noted in three-week-old piglets from sows fed LNC vs. SBP (45% vs. 0%, P = 0.001). TcdB prevalence was higher in six-day-old piglets from the sows fed LNC vs. SBP (60% vs. 17%, P = 0.009). In sows, fecal microbial metabolites were higher in SBP than LNC, while C. difficile concentration showed no difference. Higher microbial diversity Shannon index was noted in sows from SBP vs. LNC one week before parturition and at the parturition (P ≤ 0.05). Piglets from SBP vs. LNC tended to have higher microbial diversity Shannon index at two and three weeks of age. Diets enriched with high-fermentable fiber compared to low-fermentable fiber in sows reduced C. difficile colonization in their piglets. Susceptibility to colonization by C. difficile in neonatal piglets can be modulated by the sows' diet, supporting the hypothesis of the early microbial programming in the offspring and the importance of the sow-piglet couple.

A Preliminary Survey of the Distribution of Segmented Filamentous Bacteria in the Porcine Gastrointestinal Tract.

Segmented filamentous bacteria (SFB) are present in various animal species including pigs. The aim of this work was to analyze the occurrence of SFB in different parts of the gastrointestinal tract of piglets of different ages. A total of 377 DNA extracts from stomach, jejunum, ileum, cecum and colon digesta, and from feces collected on different time points, originating from 155 animals, were screened by qPCR method with primers specific for the SFB. SFB sequences were detected in 74 of 377 samples (19.6%) from 155 animals in total. SFB were most abundant in ileum (50.0%), cecum (45.0%), and colon (37.0%), followed by feces (14.6%). SFB prevalence in sows was 12.9% (13/101) and 75.9% (41/54) in individual piglets. Of the 41 SFB-positive piglets, only two samples were from pre-weaning animals, while the rest of samples originated from post-weaning piglets. SFB sequences are abundant in post-weaning piglets, but not in suckling or adult animals. They are most abundant in the ileum and cecum of pigs. Further studies are warranted to reveal the role of SFB in pigs.

Effects of dietary cellobiose on the intestinal microbiota and excretion of nitrogen metabolites in healthy adult dogs.

In order to evaluate the potential prebiotic effects of cellobiose, 10 healthy adult research beagle dogs received a complete diet containing 0, 0.5 and 1 g cellobiose/kg bodyweight (BW)/day. At the end of each feeding period, faeces, urine and blood of the dogs were collected. The results demonstrated a significant increase of faecal lactate concentrations, indicating a bacterial fermentation of cellobiose in the canine intestine. Along with this, a dose-dependent linear increase of the relative abundance of Lactobacillaceae in the faeces of the dogs was observed (p = 0.014). In addition, a dose-dependent increase (p < 0.05) of Alloprevotella, Bacteroides and Prevotella, and a linear decrease for unidentified Lachnospiraceae (p = 0.011) was observed when cellobiose was added to the diet, although the relative abundance of these genera was low (<1%) among all groups. The faecal pH was not affected by dietary cellobiose. Cellobiose seemed to modulate the excretion of nitrogen metabolites, as lower concentrations of phenol (p = 0.034) and 4-ethylphenol (p = 0.002) in the plasma of the dogs were measured during the supplementation periods. Urinary phenols and indoles, however, were not affected by the dietary supplementation of cellobiose. In conclusion, cellobiose seems to be fermented by the intestinal microbiota of dogs. Although no effect on the faecal pH was detected, the observed increase of microbial lactate production might lower the pH in the large intestine and consecutively modulate the intestinal absorption of nitrogen metabolites. Also, the observed changes of some bacterial genera might have been mediated by increased intestinal lactate concentrations or a higher relative abundance of lactobacilli. Whether these results could be considered as a prebiotic effect and used as a dietetic strategy in diseased animals to improve gut function or hepatic and renal nitrogen metabolism should be evaluated in future studies.

Impact of the dietary inclusion of dried food residues on the apparent nutrient digestibility and the intestinal microbiota of dogs.

The use of food residues for animal nutrition might imply ecological and economic advantages; however, their effects as a potential ingredient have not yet been evaluated in dogs. In the present study, four diets with 0, 5, 10 and 15% dried food residues (DFR), derived from hotel catering, were fed to 10 healthy adult dogs. At the end of each three-week feeding period, faeces and blood were collected. The apparent nutrient digestibility was calculated by the dietary inclusion of titanium dioxide as an inert marker. The results demonstrated that the apparent crude protein digestibility and ether extract digestibility decreased with increasing amounts of DFR in the diets (p < 0.05). In addition, an increase of the faecal concentrations of acetic acid, propionic acid, n-butyric acid and total short-chain fatty acids (SCFA) was observed (p < 0.05). Faecal ammonium and lactate concentrations, as well as plasma phenol and indole concentrations, were not linearly affected by the dietary inclusion of DFR. The relative abundance of Fusobacteria in the faeces of the dogs decreased, and the relative abundance of Actinobacteria and Bacteroidetes increased with increasing amounts of DFR in the diets (p < 0.05). In conclusion, the DFR seemed to be intensively fermented by the intestinal microbiota of the dogs, as indicated by the increased faecal SCFA concentrations and the shifts in the composition of the faecal microbiota. Dietary inclusion levels of up to 5% can be recommended based on our results, as the observed lower apparent crude protein and ether extract digestibility might limit the use of food residues for dogs at higher amounts.

In vitro conjugation kinetics of AmpC, broad spectrum and extended-spectrum beta-lactamase-producing Escherichia coli donors and various Enterobacteriaceae recipients.

BACKGROUND: Extended spectrum beta-lactamase (ESBL)-producing enterobacteria pose a major hazard to public health. Due to the possibility of genetic transfer, ESBL genes might spread to pathogenic enterobacterial strains. Thus, information on possible genetic transfer between enterobacteria is of high interest. It was therefore the aim of this in vitro study to screen the capacity of a wide range of Enterobacteriaceae for differences in conjugation at different time points with five ESBL-producing Escherichia coli strains. RESULTS: Conjugation frequencies for five potential E. coli donor strains producing the enzymes CTX-M-1, CTX-M-15, SHV-12, TEM-1, TEM-52 and CMY-2, and six potential recipient strains commonly detected in the gastrointestinal tract of poultry (E. coli, Serratia marcescens subsp. marcescens, Enterobacter cloacae, Salmonella (S.) enterica serovar Typhimurium and Proteus mirabilis) were obtained. Different combinations of donor and recipient strains were co-incubated for between 0 and 22 h and spread on selective agar. Conjugation frequencies were calculated as transconjugants per donor. Some donor and recipient strain combinations did not perform plasmid transfer within 22 h. Hence, the recipient Proteus mirabilis did not accept plasmids from any of the given donors and the E. coli ESBL10716 donor was unable to transfer its plasmid to any recipient. Enterobacter cloacae only accepted the plasmids from the donors E. coli ESBL10708 and E. coli ESBL10716 while E. coli ESBL10708 did not transfer its plasmid to Serratia marcescens subsp. marcescens. E. coli IMT11716 on the other hand did not perform conjugation with the donor E. coli ESBL10689. The remaining mating pairs differed in conjugation frequency, ranging from 10- 5 to 10- 9 transconjugants/donor. The earliest conjugation events were detected after 4 h. However, some mating pairs turned positive only after 22 h of coincubation. CONCLUSION: A suitable mating pair for future in vivo studies to combat transfer of antibiotic resistance to pathogenic bacteria in broiler chicken was determined. The results of this study also suggest that the kinetic of conjugation differs between mating pairs and is independent of species origin. This should be considered when performing conjugation experiments.

Distinct patterns of microbial metabolic fingerprints in sows and their offspring: a pilot study.

Microbial metabolism and growth in the intestinal tract depend on the composition of substrates present in the digesta and their ability to be metabolised by the microorganisms. The aim of this pilot study was to characterise potential hindgut microbial activity during perinatal period in sows and their offspring. Rectal samples from three sows (1-3 weeks before and after birth) and three of their piglets (1-5 weeks after birth), were subjected to assays using BIOLOG GEN III microplates to produce metabolic fingerprints for each animal. The number of metabolised substrates of the sow hindgut microbiota was stable during the pregnancy and lactation periods, as assessed by the richness index. In piglets, the richness was stable during the suckling period and at beginning of weaning, however, it decreased when the piglets were 5 weeks old (P ≤ 0.05). Analysis of associations between the sows and the piglets and the microbial metabolic potential showed that microbial metabolism was strongly associated with the catabolism of carbohydrates especially in sows. Only 5-week-old weaned piglets clustered together with the sows regarding the microbial catabolism of substrates, but not suckling piglets. The association analyses clustered all the piglets in two groups distinctive for litter. The analysis of metabolic fingerprints via microbial growth with different substrates can be useful to positively influence microbial community function such as selectively enhancing desirable active microbial populations to benefit health of the gut and the animal.

Lythrum salicaria Ellagitannins Stimulate IPEC-J2 Cells Monolayer Formation and Inhibit Enteropathogenic Escherichia coli Growth and Adhesion.

Lythrum salicaria herb (LSH) was applied in diarrhea therapy since ancient times. Despite empirically referenced therapeutic effects, the bioactivity mechanisms and chemical constituents responsible for pharmacological activity remain not fully resolved. Taking into consideration the historical use of LSH in treatment of diarrhea in humans and farm animals, the aim of the study was to examine in vitro the influence of LSH and its C-glycosylic ellagitannins on processes associated with maintaining intestinal epithelium integrity and enteropathogenic Escherichia coli (EPEC) growth and adhesion. LSH was not only inhibiting EPEC growth in a concentration dependent manner but also its adhesion to IPEC-J2 intestinal epithelial cell monolayers. Inhibitory activity toward EPEC growth was additionally confirmed ex vivo in distal colon samples of postweaning piglets. LSH and its dominating C-glycosylic ellagitannins, castalagin (1), vescalagin (2), and salicarinins A (3) and B (4) were stimulating IPEC-J2 monolayer formation by enhancing claudin 4 production. Parallelly tested gut microbiota metabolites of LSH ellagitannins, urolithin C (5), urolithin A (6), and its glucuronides (7) were inactive. The activities of LSH and the isolated ellagitannins support its purported antidiarrheal properties and indicate potential mechanisms responsible for its beneficial influence on the intestinal epithelium.

Optimization of Production Parameters for Probiotic Lactobacillus Strains as Feed Additive.

In animal nutrition, probiotics are considered as desirable alternatives to antibiotic growth promoters. The beneficial effects of probiotics primarily depend on their viability in feed, which demands technical optimization of biomass production, since processing and storage capacities are often strain-specific. In this study, we optimized the production parameters for two broiler-derived probiotic lactobacilli (L. salivarius and L. agilis). Carbohydrate utilization of both strains was determined and preferred substrates that boosted biomass production in lab-scale fermentations were selected. The strains showed good aerobic tolerance, which resulted in easier scale-up production. For the freeze-drying process, the response surface methodology was applied to optimize the composition of cryoprotective media. A quadratic polynomial model was built to study three protective factors (skim milk, sucrose, and trehalose) and to predict the optimal working conditions for maximum viability. The optimal combination of protectants was 0.14g/mL skim milk/ 0.08 g/mL sucrose/ 0.09 g/mL trehalose (L. salivarius) and 0.15g/mL skim milk/ 0.08 g/mL sucrose/ 0.07 g/mL (L. agilis), respectively. Furthermore, the in-feed stabilities of the probiotic strains were evaluated under different conditions. Our results indicate that the chosen protectants exerted an extensive protection on strains during the storage. Although only storage of the strains at 4 °C retained the maximum stability of both Lactobacillus strains, the employed protectant matrix showed promising results at room temperature.

In vitro evaluation of the effects of Yucca schidigera and inulin on the fermentation potential of the faecal microbiota of dogs fed diets with low or high protein concentrations.

This in vitro study aimed to evaluate the effects of Yucca schidigera powder (YSP) and inulin (IN) on protein fermentation metabolites (short-chain fatty acids [SCFA] and branched-chain fatty acids, phenolic and indolic compounds, biogenic amines, ammonia and pH) by using faecal inocula from dogs fed either a low (L) or a high (H) protein diet (crude protein 201 or 377 g/kg as fed). Four treatments for each diet were evaluated in an in vitro batch culture system over 24 h: (1) control with no addition of substrates; (2) 4 g YSP/l; (3) 5 g IN/l; (4) 4 g YSP/l in combination with 5 g IN/l of faecal culture of dogs fed Diet L or H. Several changes in fermentation metabolites were analysed. Samples incubated with the faecal inocula of dogs fed Diet L produced higher concentrations of total SCFA, propionate (p = 0.001), acetate (p ≤ 0.001), d-lactate (p = 0.041) and indole (p = 0.003), whereas pH (p = 0.004) was decreased. Supplementation of IN increased the content of putrescine, d- and l-lactate, total SCFA, acetate, propionate, n-butyrate (p ≤ 0.001) and n-valerate (p = 0.003), while i-valerate, indole and pH (p ≤ 0.001) were reduced. Ammonia was lower (p = 0.013) in samples with faecal inocula from dogs fed Diet H and further reduced by the addition of IN (p ≤ 0.001). Samples with faecal inocula from dogs fed Diet L had a fewer quotient of ammonia and total SCFA (p = 0.040). Supplementation of YSP (p = 0.021), IN (p ≤ 0.001) and YSP in combination with IN (p = 0.047) led to a higher reduction of the quotient of ammonia and total SCFA. In conclusion, dietary protein concentration and the supplementation of IN resulted in a stimulation of fermentation while YSP appeared to have only minor effects.

Formula Feeding Predisposes Neonatal Piglets to Clostridium difficile Gut Infection.

Spontaneous outbreaks of Clostridium difficile infection (CDI) occur in neonatal piglets, but the predisposing factors are largely not known. To study the conditions for C. difficile colonization and CDI development, 48 neonatal piglets were moved into isolators, fed bovine milk-based formula, and infected with C. difficile 078. Analyses included clinical scoring; measurement of the fecal C. difficile burden, toxin B level, and calprotectin level; and postmortem histopathological analysis of colon specimens. Controls were noninfected suckling piglets. Fecal specimens from suckling piglets, formula-fed piglets, and formula-fed, C. difficile-infected piglets were used for metagenomics analysis. High background levels of C. difficile and toxin were detected in formula-fed piglets prior to infection, while suckling piglets carried about 3-fold less C. difficile, and toxin was not detected. Toxin level in C. difficile-challenged animals correlated positively with C. difficile and calprotectin levels. Postmortem signs of CDI were absent in suckling piglets, whereas mesocolonic edema and gas-filled distal small intestines and ceca, cellular damage, and reduced expression of claudins were associated with animals from the challenge trials. Microbiota in formula-fed piglets was enriched with Escherichia, Shigella, Streptococcus, Enterococcus, and Ruminococcus species. Formula-fed piglets were predisposed to C. difficile colonization earlier as compared to suckling piglets. Infection with a hypervirulent C. difficile ribotype did not aggravate the symptoms of infection. Sow-offspring association and consumption of porcine milk during early life may be crucial for the control of C. difficile expansion in piglets.

The Spore Coat Protein CotE Facilitates Host Colonization by Clostridium difficile.

Clostridium difficile infection (CDI) is an important hospital-acquired infection resulting from the germination of spores in the intestine as a consequence of antibiotic-mediated dysbiosis of the gut microbiota. Key to this is CotE, a protein displayed on the spore surface and carrying 2 functional elements, an N-terminal peroxiredoxin and a C-terminal chitinase domain. Using isogenic mutants, we show in vitro and ex vivo that CotE enables binding of spores to mucus by direct interaction with mucin and contributes to its degradation. In animal models of CDI, we show that when CotE is absent, both colonization and virulence were markedly reduced. We demonstrate here that the attachment of spores to the intestine is essential in the development of CDI. Spores are usually regarded as biochemically dormant, but our findings demonstrate that rather than being simply agents of transmission and dissemination, spores directly contribute to the establishment and promotion of disease.

Physical Pre-Treatment Improves Efficient DNA Extraction and qPCR Sensitivity from Clostridium Difficile Spores in Faecal Swine Specimens.

A considerable fraction of the faecal microbiota is spore-forming. Molecular quantification of bacteria may be underestimated if preceded with nucleic acid extraction without special treatment to extract recalcitrant bacterial spores. The objective of this study was to improve the DNA extraction regarding the presence of Clostridium difficile spores in faecal swine specimens. Sow faeces were inoculated with spores of C. difficile (10(6) CFU), frozen at - 30 °C overnight and subjected to DNA extraction. As a preceding step to a standard DNA extraction method (QIAamp DNA stool Mini kit), different physical treatments such as microwave oven heating and repeated bead-beating techniques and a combination of both were applied and compared with each other by means of qPCR. Using a standard DNA extraction method only, C. difficile spores were quantified at 4.96 log copy number/200 mg of faeces. A repeated bead-beating at 6 m/s for 10 min followed by a standard DNA extraction resulted in 5.77 log copy number of spores in inoculated faeces. Heating in a microwave oven at 800 W for 1, 3, 5 and 10 min followed by a standard DNA extraction resulted in a gene quantification of up to 4.89 log copy number. A combination of both methods resulted in the bacterial gene quantity of 5.37 log copy number. Pre-treatment with repeated bead-beating led to the highest quantification of bacteria, and therefore it can be applied for more efficient DNA extraction from spores of C. difficile in faecal specimens.

Determination of the extent of Clostridium difficile colonisation and toxin accumulation in sows and neonatal piglets.

Clostridium difficile is an important spore-forming, opportunistic pathogen in animal husbandry and health care. In pig farming, only neonatal piglets are affected, and diarrhoea and necrotising lesions are common symptoms leading to dehydration and in some cases death. This study aimed at the assessment of the quantitative development of C. difficile colonisation in neonatal piglets by determining the shedding of spores and C. difficile toxins A (TcdA) and B (TcdB) concentrations in sow (n = 5-6) and piglet pen faeces (n = 5-6) at different time points. Spores were quantified on selective agar plates and toxins using ELISA method. C. difficile was not detected in the faeces of all but one sow during the perinatal period. Faeces of 2- and 4-day-old piglets contained 0.65 log cells/g and 5.88 log cells/g of C. difficile, respectively. Toxins were detected on day 4 at a concentration of 2.13 log ng/g (TcdA) and 2.06 log ng/g (TcdB). On day 6, concentration of C. difficile reached 6.14 log CFU/g and toxins 2.02 log ng/g (TcdA) and 2.20 log ng/g (TcdB). Two-week-old piglets showed 4.72 log CFU/g of C. difficile but toxins could not be detected. At 21 days of age, both C. difficile and toxins were undetectable. The concentration and the prevalence of C. difficile were positively associated with the prevalence of toxins in piglets. A very short time window for colonisation by C. difficile, including toxin-producing strains can be observed in neonatal piglets. The significance for animal health and the risk of a carrier status need to be addressed in future studies.

Down-regulation of monocarboxylate transporter 1 (MCT1) gene expression in the colon of piglets is linked to bacterial protein fermentation and pro-inflammatory cytokine-mediated signalling.

The present study investigated the influence of bacterial metabolites on monocarboxylate transporter 1 (MCT1) expression in pigs using in vivo, ex vivo and in vitro approaches. Piglets (n 24) were fed high-protein (26 %) or low-protein (18 %) diets with or without fermentable carbohydrates. Colonic digesta samples were analysed for a broad range of bacterial metabolites. The expression of MCT1, TNF-α, interferon γ (IFN-γ) and IL-8 was determined in colonic tissue. The expression of MCT1 was lower and of TNF-α and IL-8 was higher with high-protein diets (P< 0·05). MCT1 expression was positively correlated with l-lactate, whereas negatively correlated with NH3 and putrescine (P< 0·05). The expression of IL-8 and TNF-α was negatively correlated with l-lactate and positively correlated with NH3 and putrescine, whereas the expression of IFN-γ was positively correlated with histamine and 4-ethylphenol (P< 0·05). Subsequently, porcine colonic tissue and Caco-2 cells were incubated with Na-butyrate, NH4Cl or TNF-α as selected bacterial metabolites or mediators of inflammation. Colonic MCT1 expression was higher after incubation with Na-butyrate (P< 0·05) and lower after incubation with NH4Cl or TNF-α (P< 0·05). Incubation of Caco-2 cells with increasing concentrations of these metabolites confirmed the up-regulation of MCT1 expression by Na-butyrate (linear, P< 0·05) and down-regulation by TNF-α and NH4Cl (linear, P< 0·05). The high-protein diet decreased the expression of MCT1 in the colon of pigs, which appears to be linked to NH3- and TNF-α-mediated signalling.

Dietary inulin affects the intestinal microbiota in sows and their suckling piglets.

BACKGROUND: Several studies have focused on the effects of dietary inulin on the intestinal microbiota of weaned piglets. In the present study, inulin was added to a diet for gestating and lactating sows, expecting not only effects on the faecal microbiota of sows, but also on the bacterial cell numbers in the gastrointestinal tract of their piglets during the suckling period. Sows were fed a diet without (n = 11) or with (n = 10) 3% inulin, and selected bacterial groups were determined in their faeces ante and post partum. Suckling piglets, 8 per group, were euthanised on day 10 after birth to analyse digesta samples of the gastrointestinal tract. RESULTS: Dietary inulin increased the cell numbers of enterococci, both, in the faeces of the sows during gestation and lactation, and in the caecum of the piglets (P ≤ 0.05). Moreover, higher cell numbers of eubacteria (stomach) and C. leptum (caecum), but lower cell numbers of enterobacteria and L. amylovorus (stomach) were detected in the digesta of the piglets in the inulin group (P ≤ 0.05). CONCLUSIONS: In conclusion, inulin seems to have the potential to influence the gastrointestinal microbiota of suckling piglets through the diet of their mother, showing the importance of the mother-piglet couple for the microbial development. Early modulation of the intestinal microbiota could be especially interesting with regard to the critical weaning time.

A standardised challenge model with an enterotoxigenic F4+ Escherichia coli strain in piglets assessing clinical traits and faecal shedding of fae and est-II toxin genes.

This study evaluated the effect of five feed additives on post weaning diarrhoea (PWD) in piglets challenged 3 d after weaning with an enterotoxigenic Escherichia coli strain (ETEC). In three experimental runs, a total of 84 piglets was weaned at 21 days of age and randomly assigned to seven treatments. As dietary treatment, piglets were fed a basal diet or diets with addition of bovine colostrum (0.2%), pineapple stem extract containing bromelain (0.2%), an autolysed yeast preparation (Saccharomyces cerevisiae) (0.1%), a combination of organic acids (0.7%) and a phytogenic product with thyme essential oil (0.015%). A porcine ETEC, serotype O149:K91:K88ac was given twice via oral infection on day 3 after weaning at 10(10) colony forming units/animal. One group of piglets was fed the basal diet without ETEC challenge. Traits included clinical sores, body temperature, faecal scoring and determination of faecal dry matter and the shedding of fae and est-II ETEC toxin genes. After weaning, non-challenged control piglets did not show signs of diarrhoea or impaired health, while the majority of infected piglets had a drop in body temperature, signs of diarrhoea and impaired general health. Mortality, the decrease of faecal dry matter and shedding of the toxin genes fae and est-II were not affected by the different additives. In conclusion, the ETEC challenge model induced distinct clinical signs of PWD in piglets, but the tested feed additives had no preventive effect under these conditions.

Interaction between dietary protein content and the source of carbohydrates along the gastrointestinal tract of weaned piglets.

Although fermentable carbohydrates (CHO) can reduce metabolites derived from dietary protein fermentation in the intestine of pigs, the interaction between site of fermentation and substrate availability along the gut is still unclear. The current study aimed at determining the impact of two different sources of carbohydrates in diets with low or very high protein content on microbial metabolite profiles along the gastrointestinal tract of piglets. Thirty-six piglets (n = 6 per group) were fed diets high (26%, HP) or low (18%, LP) in dietary protein and with or without two different sources of carbohydrates (12% sugar beet pulp, SBP, or 8% lignocellulose, LNC) in a 2 × 3 factorial design. After 3 weeks, contents from stomach, jejunum, ileum, caecum, proximal and distal colon were taken and analysed for major bacterial metabolites (D-lactate, L-lactate, short chain fatty acids, ammonia, amines, phenols and indols). Results indicate considerable fermentation of CHO and protein already in the stomach. HP diets increased the formation of ammonia, amines, phenolic and indolic compounds throughout the different parts of the intestine with most pronounced effects in the distal colon. Dietary SBP inclusion in LP diets favoured the formation of cadaverine in the proximal parts of the intestine. SBP mainly increased CHO-derived metabolites such as SCFA and lactate and decreased protein-derived metabolites in the large intestine. Based on metabolite profiles, LNC was partly fermented in the distal large intestine and reduced mainly phenols, indols and cadaverine, but not ammonia. Multivariate analysis confirmed more diet-specific metabolite patterns in the stomach, whereas the CHO addition was the main determinant in the caecum and proximal colon. The protein level mainly influenced the metabolite patterns in the distal colon. The results confirm the importance of CHO source to influence the formation of metabolites derived from protein fermentation along the intestinal tract of the pig.

A high amount of dietary zinc changes the expression of zinc transporters and metallothionein in jejunal epithelial cells in vitro and in vivo but does not prevent zinc accumulation in jejunal tissue of piglets.

High dietary zinc concentrations are used to prevent or treat diarrhea in piglets and humans, but long-term adaptation to high zinc supply has yet not been assessed. Intestinal zinc uptake is facilitated through members of zinc transporter families SLC30 (ZnT) and SLC39 (ZIP). Whereas in rodents, regulation of zinc homeostasis at low or adequate zinc supply has been described, such mechanisms are unclear in piglets. A total of 54 piglets were fed diets containing 57 [low dietary zinc (LZn)], 164 [normal dietary zinc (NZn)], or 2425 [high dietary zinc (HZn)] mg/kg dry matter zinc. After 4 wk, 10 piglets/group were killed and jejunal tissues taken for analysis of zinc transporters SLC30A1 (ZnT1), SLC30A2 (ZnT2), SLC30A5 (ZnT5), SLC39A4 (ZIP4), divalent metal transporter 1 (DMT1), and metallothionein-1 (MT). Weight gain was higher (P < 0.05) in pigs fed HZn than in the LZn and NZn groups during the first 2 wk. Food intake did not differ between groups. The digesta and jejunal tissue zinc concentrations were higher (P < 0.05) in the HZn pigs than in NZn and LZn pigs. Expression of ZnT1 was higher (P < 0.05) and ZIP4 lower (P < 0.05) in HZn pigs than in the 2 other groups, whereas expression of ZnT5 and DMT1 did not differ between treatments. Expression of ZnT2 was lower (P < 0.05) in the LZn group than in the HZn and NZn groups. The mRNA expression and protein abundance of MT was higher (P < 0.05) in the HZn group than in the NZn and LZn groups. Studies with intestinal porcine cell line intestinal epithelial cell-J2 confirmed the dose-dependent downregulation of ZIP4 and upregulation of ZnT1 and MT (P < 0.05) with increasing zinc concentration within 24 h. In conclusion, high dietary zinc concentrations increase intracellular zinc, promote increased zinc export from intestinal tissues into extracellular compartments, and decrease zinc uptake from the gut lumen. The adaptive process appears to be established within 24 h; however, it does not prevent tissue zinc accumulation.