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Abstract In the last decades, gut microbiota and its role in mammal host development and health have been increasingly investigated. Metabolites produced by gut microbiota can affect intestinal homeostasis and immune system maturity and activation, and in turn, they can influence the health and growth performance of livestock. Therefore, a better understanding of the functional metabolic capability of the gut microbiota would be appreciated by the scientific community. In this study, the BiologTM Ecoplates technology was applied for studying the metabolic potential of the aerotolerant microbial community of pig fecal samples, evaluating the interference of different storage conditions and cell concentrations. The length of time for which a fecal sample maintained detectable and unchanged microbial metabolic activity was also investigated. Two assays aimed to evaluate differences in the metabolic activities between fresh and snap-frozen fecal samples at different dilutions and at different lengths of times of preservation at −80°C were carried out. The biodiversity and the predicted functionality of the entire bacterial community through a targeted metagenomic approach were also explored. The results highlighted that snap freezing of fecal samples preserved the metabolic activity of the microbial community when compared to fresh feces. Sample storage at −80 °C did not significantly affect the metabolic activity of the microbial community, which was stable for 150 days. Furthermore, the highest metabolic activity was detected with 1:2 to 1:5 dilutions of the stock suspension. BiologTM Ecoplates technology is a rapid and useful method to explore microbial communities’ metabolism in animal fecal samples contributing to investigate host animal physiology. Key points • Freezing of samples can preserve the functional activity of the aerotolerant microbial community for 150 days. • The concentration of microbial cells strongly influences metabolic activity detection. • Sequencing coupled with the BiologTMEcoplates could be a strategy to evaluate the metabolic potential of the microbiota of the fecal sample. Graphical abstract

A microbiome is defined as a complex collection of microorganisms and their genetic material. Studies regarding gut microbiomes of different animals have provided ecological and evolutionary information showing a strong link between health and disease. Very few studies have compared the gut microbiota of animals housed under controlled conditions and those in wild habitats. Little research has been performed on the reptile gut microbiota, and what studies do exist are mainly focused on carnivorous reptiles. The aim of this study was first to describe the overall microbiota structure of Aldabra giant tortoises (Aldabrachelys gigantea) and, second, to compare the microbiota of tortoises living under natural conditions and tortoises living in controlled environments, such as zoological and botanical parks, in Italy and in the Seychelles. Seventeen fecal samples were collected from giant tortoises located on Curieuse Island (CI, n = 8), at the Botanical Garden (BG, n = 3) in Mahé (Seychelles Islands) and at Parco Natura Viva–Garda Zoological Park (PNV, n = 6) in Verona (Italy). The V3-V4 region of the 16S rRNA gene was amplified in order to characterize the gut microbiota profile. Overall, the major phyla identified were Bacteroidetes 42%, Firmicutes 32%, and Spirochaetes 9%. A higher microbial diversity (alpha indices) was observed for the BG samples as compared to the PNV samples (Shannon: 5.39 vs. 4.43; InvSimpson: 80.7 vs. 25; Chao1: 584 vs. 377 p < 0.05). The results in the present study showed a significant difference in beta diversity between the samples from CI, BG, and PNV (p = 0.001), suggesting a different bacterial fecal profile of giant tortoises at the different habitats. This study provided novel insights into the effects of different environmental conditions on the gut microbial communities of giant tortoises. In particular, differences were reported regarding the bacterial gut community structure between tortoises in natural and in controlled environments. These results could help to improve the management of giant tortoises under human care, thus enhancing ex-situ conservation efforts far from the species geographic range.

Antibiotic resistance is a public health problem of growing concern. Animal manure application to soil is considered to be a main cause of the propagation and dissemination of antibiotic residues, antibiotic-resistant bacteria (ARB), and antibiotic resistance genes (ARGs) in the soil-water system. In recent decades, studies on the impact of antibiotic-contaminated manure on soil microbiomes have increased exponentially, in particular for taxonomical diversity and ARGs’ diffusion. Antibiotic resistance genes are often located on mobile genetic elements (MGEs). Horizontal transfer of MGEs toward a broad range of bacteria (pathogens and human commensals included) has been identified as the main cause for their persistence and dissemination. Chemical and bio-sanitizing treatments reduce the antibiotic load and ARB. Nevertheless, effects of these treatments on the persistence of resistance genes must be carefully considered. This review analyzed the most recent research on antibiotic and ARG environmental dissemination conveyed by livestock waste. Strategies to control ARG dissemination and antibiotic persistence were reviewed with the aim to identify methods for monitoring DNA transferability and environmental conditions promoting such diffusion.

The objective of the study was to test the associations between backfat fatty acid composition (FAC) in a sample of 536 Italian Large White pigs and single nucleotide polymorphisms (SNPs) located in candidate genes, using univariate and multivariate approaches. The strongest associations were identified for the SNP AY183428 c.265T>C in Fatty acid synthase (FASN) gene, with the T allele linked to lower backfat contents of stearic (P = 0.003) and arachidic (P

International audience; Enterotoxigenic Escherichia coli (ETEC) is the aetiological agent of postweaning diarrhoea (PWD) in piglets. The SNPs located on the Mucine 4 (MUC4) and Fucosyltransferase 1 (FUT1) genes have been associated with the susceptibility to ETEC F4 and ETEC F18, respectively. The interplay between the MUC4 and FUT1 genotypes to ETEC infection and the use of amoxicillin in modifying the intestinal microbiota during a natural infection by multiresistant ETEC strains have never been investigated. The aim of this study was to evaluate the effects of the MUC4 and FUT1 genotypes and the administration of amoxicillin through different routes on the presence of diarrhoea and the faecal microbiota composition in piglets naturally infected with ETEC. Seventy-one piglets were divided into three groups: two groups differing by amoxicillin administration routes-parenteral (P) or oral (O) and a control group without antibiotics (C). Faecal scores, body weight, presence of ETEC F4 and F18 were investigated 4 days after the arrival in the facility (T0), at the end of the amoxicillin administration (T1) and after the withdrawal period (T2). The faecal bacteria composition was assessed by sequencing the 16S rRNA gene. We described that MUC4 and FUT1 genotypes were associated with the presence of ETEC F4 and ETEC F18. The faecal microbiota was influenced by the MUC4 genotypes at T0. We found the oral administration to be associated with the presence of diarrhoea at T1 and T2. Furthermore, the exposure to amoxicillin resulted in significant alterations of the faecal microbiota. Overall, MUC4 and FUT1 were confirmed as genetic markers for the susceptibility to ETEC infections in pigs. Moreover, our data highlight that group amoxicillin treatment may produce adverse outcomes on pig health in course of multiresistant ETEC infection. Therefore, alternative control measures able to maintain a healthy faecal microbiota in weaners are recommended.

We evaluated the potential of multi-strain probiotic (Bifidobacterium longum subsp. infantis CECT 7210 and Lactobacillus rhamnosus HN001) with or without galacto-oligosaccharides against enterotoxigenic Escherichia coli (ETEC) F4 infection in post-weaning pigs. Ninety-six piglets were distributed into 32 pens assigned to five treatments: one non-challenged (CTR+) and four challenged: control diet (CTR−), with probiotics (>3 × 1010 CFU/kg body weight each, PRO), prebiotic (5%, PRE), or their combination (SYN). After 1 week, animals were orally inoculated with ETEC F4. Feed intake, weight, and clinical signs were recorded. On days 4 and 8 post-inoculation (PI), one animal per pen was euthanized and samples from blood, digesta, and tissues collected. Microbiological counts, ETEC F4 real-time PCR (qPCR) quantification, fermentation products, serum biomarkers, ileal histomorphometry, and genotype for mucin 4 (MUC4) polymorphism were determined. Animals in the PRO group had similar enterobacteria and coliform numbers to the CTR+ group, and the ETEC F4 prevalence, the number of mitotic cells at day 4 PI, and villus height at day 8 PI were between that observed in the CTR+ and CTR− groups. The PRO group exhibited reduced pig major acute-phase protein (Pig-MAP) levels on day 4 PI. The PRE diet group presented similar reductions in ETEC F4 and Pig-MAP, but there was no effect on microbial groups. The SYN group showed reduced fecal enterobacteria and coliform counts after the adaptation week but, after the inoculation, the SYN group showed lower performance and more animals with high ETEC F4 counts at day 8 PI. SYN treatment modified the colonic fermentation differently depending on the MUC4 polymorphism. These results confirm the potential of the probiotic strains and the prebiotic to fight ETEC F4, but do not show any synergy when administered together, at least in this animal model.

The establishment and maintenance of a balanced gut microbiota in early life play a pivotal role in pigs. This study aims to evaluate the effect of the administration of a single early-life probiotic on piglet faecal microbiota and growth performance until weaning. Forty-eight hours after birth (d0), 820 piglets were allocated into 4 groups (205 piglets/16 litters/group) and orally inoculated as follows: 1) Control (CO: 4 mL of pure water); 2) Saccharomyces (SA: 4 mL containing a total of 1 × 1010 CFU of Saccharomyces cerevisiae var. boulardii CNCM-1079; 3) Enterococcus (EF: 4 mL containing a total of 1 × 1010 CFU of Enterococcus faecium lactiferm WS200); 4) a mix of the two probiotics at the same doses (SAEF). At d7 and d18, the piglets were weighed, and faeces from the piglets (18 piglets/group from 6 sows/group) and their mothers were analysed for a microbial profile by sequencing the v3-v4 regions of the 16S rRNA gene. Data were arranged in a 2x2 factorial design. The probiotic supplement improved piglet ADG in the periods d7-d18 (p

Simple Summary Nucleotides represent a group of bioactive compounds essential for the development of the gastrointestinal tract and immune function. This study aimed to evaluate the short-term effect of oral administration of nucleotides before and after weaning on growth performance, health, development of the intestinal immunity and microbiome of piglet. A nucleotide-based product (NU) was orally given four times before weaning and once after to one group of piglets, while a second group was used as a control (CO). The NU pigs did not grow more than the control until 12 days post-weaning but had increased hemoglobin and hematocrit values. At weaning, feces of NU piglets had a microbial profile more typical of growing pigs, while those of CO were more representative of suckling pigs. The upregulation of genes in the blood of control pigs at weaning was indicative of more activation towards an inflammatory response, while genes of erythropoiesis were more active in NU pigs post-weaning. NU supplementation stimulated genes for proliferative activity in the intestinal immune system, a sign of possible anticipated maturation. NU supplementation did not influence the growth performance of piglets but may have expressed a positive effect on pig microbiota anticipating its maturation at weaning, with possible immunostimulant activity on the intestinal immune system. Abstract Nucleotides are essential for the development of the gastrointestinal tract and immune function, but their intake with milk by piglets could be insufficient. The effect of nucleotides on growth and health was tested on 98 piglets divided into two groups: NU, orally administrated with 4 mL of a nucleotide-based product (SwineMOD®) at 10, 15, 18, 21, 27 days, or not (CO). Blood and feces were sampled at weaning (26 d, T1), and at 38 d (T2). Per each group and time-point, eight piglets were slaughtered and jejunal Peyer’s patches (JPPs) were collected. NU increased hemoglobin content and hematocrit, but not growth. At weaning, the NU fecal microbiota was characterized by the abundance of Campylobacteraceae, more typical of the growing phase, compared to CO, with a greater abundance of Streptococcaceae. For the blood transcriptome, an initial greater inflammatory activation was seen in CO, while at T2, NU enriched gene sets related to erythropoiesis. The activation of gene groups ranging from epigenetic response to transcriptional regulation evidenced an intense proliferative activity in NU JPPs. NU supplementation did not influence the growth performance of piglets but could have expressed a positive effect on pig microbiota anticipating its maturation at weaning. This immunostimulant activity in the JPPs could moderate the inflammation in the immediate pre-weaning.

Background Scarce is knowledge on the process regulating the development of acid secretion, orexigenic signaling, and chemosensing in the stomach of young pigs. Changes of early microbial encounters by suckling pigs can interact with the gut maturation, by the induction of different molecular signaling. Our goal was to assess if the age of offspring and the maternal environment, influenced by sow antibiotic treatment peripartum, could affect gastric morphology and the expression of genes involved in the control of hydrochloric secretion, feed intake, taste, and inflammation in offspring stomach. Methods 84 pigs from sows fed a diet with amoxicillin (on –d10 to +d21 from farrowing, ANT) or without (CON) were sacrificed at d14, d21, d28 (weaning) or d42. Samples of oxyntic (OXY), pyloric (PY) and cardiac mucosae close to OXY were collected and parietal and enteroendocrine cells (EECs) were counted. Relative gene expression of a set of 11 key genes (ATP4A, SSTR2, GAST, GHRL, MBOAT4, PCSK1, GNAT1, TAS1R1, TAS1R3, IL8 and TNF) was assessed by qRT-PCR. In addition, 40 offspring obtained from the same ANT and CON sows were offered a normal or a fat-enriched diet for 4 weeks between 140 and 169 d of age, and then OXY and PY were sampled. Results The number of parietal and EECs increased with age (P ATP4A increased with age (within suckling, P = 0.043, post-weaning vs. suckling, P P GHRL increased during suckling (P = 0.012), and post-weaning as a trend (P = 0.088). MBOAT4 tended to increase during suckling (P = 0.062). TAS1R1 increased from suckling to post-weaning period (P =0.001) and was lower in ANT offspring (P = 0.013). GNAT1 in PY was higher in ANT offspring (P = 0.041). Antibiotic treatment of sows peripartum increased expression of GHRL and MBOAT4 in OXY of growing-finishing offspring aged 5 months. Conclusions Data show that sensing for umami taste and ghrelin regulation can be affected by maternal environment, but the development of acid secretion, orexigenic signaling and taste perception in the stomach are mostly developmentally controlled.

Background Oligosaccharides (OS) are indigestible carbohydrates naturally found in milk. The composition of porcine colostrum OS may influence the growth and the health of the neonate and consuming optimal concentrations of OS may reduce piglet susceptibility to illness. In this manner, targeted supplementation of animal feed with OS is being explored as a health management tool in the livestock industry. The variation in OS composition between different breeds of pig and its association with the litter performance is currently unknown. The aim of this study was to characterize the colostrum OS composition from sows of different breed and parity and correlate this data with sow maternal traits. Methods Eighty-three colostrum samples from parities 1 to 8 were gathered from 3 different breeds of sow: 44 Large White sows, 27 Landrace sows and 12 Duroc sows. Samples were taken between the birth of the first and the last piglet from sows that were not pharmacologically induced to farrow. OS were purified from the samples and analysed by MALDI-ToF mass spectrometry (21 OS compositions detected). The farrowing season and the maternal data were recorded for each sow, including the number of live piglets and the litter body weight at birth, at day (d) 3 and at weaning. Results Five OS compositions, including isomers of the bifidogenic Sialyllactose, Lacto-N-Tetraose and Lacto-N-Hexaose series, were detected in all the samples. Twelve other OS were identified in at least 50% of samples, and their abundances were affected by breed (P P P = 0.063) and at weaning (P Conclusions OS composition of colostrum may partially explain the variability in maternal performance within and between different breeds of sow. The obtained OS data can provide useful information for the development of novel prebiotic food supplements for suckling and weaning pigs.

Simple Summary Acidifiers, especially formic acid and its salts, have long been used for improving the gastrointestinal health and growth performance of pigs, especially piglets. Although, some biological mechanisms explaining their effects are already known, a lack of consensus is still reported on their effects on the gastrointestinal microbiome. The present review provides an overview and evaluation of the effects of formic acid and its salts on the gastrointestinal microbiota and on the growth performance of pigs. Abstract Out of the alternatives to antibiotics and zinc oxide, organic acids, or simply acidifiers, play significant roles, especially in ensuring gut health and the growth performance of pigs. Regarding acidifiers, formic acid and its salts have shown very promising results in weaning, growing and finishing pigs. Although it is known that the main mechanisms by which acidifiers can improve livestock performance and health are related to the regulation of gastrointestinal pH, an improvement in intestinal digestibility and mineral utilization, and their antimicrobial properties against specific pathogens has been observed, while poor consensus remains in relation to the effect of acidifers on bacteria and the complex microbiome. Therefore, the aim of the present review was to critically evaluate the effects of formic acid and its salts on the performance and the gastrointestinal microbiota balance of pigs.

Exogenous enzymes can favour the release of shorter polymers of the dietary fibre, favouring the development of a beneficial digestive microflora. The addition of bacterial xylanase to a weaner pig diet was tested for its impact on the intestinal microbiota and digestive homeostasis. Thirty-two pigs genetically susceptible to enterotoxigenic Escherichia coli (ETEC), equally divided into two experimental groups, were used to increase the risk of diarrhoea and test the response of xylanase under conditions representing those severe situations which are frequently present on farms. Pigs, weaned at 25 ± 1 days, were fed a corn/barley standard diet without (Group CO) or with (Group XY) 100 g/t xylanase from BELFEED NV, Belgium. Blood samples (for measuring the reactive oxygen metabolites) and faeces were taken 14 and 28 days from the beginning of the trial. On day 28, the pigs were euthanised and jejunal samples were collected.The faecal bacteria16S rRNA gene was sequenced using a MiSeq Reagent Kit V3-V4 on a MiSeq-Illumina platform. The pigs had diffuse diarrhoea starting from day 4. On the morning of day 8 and for the two following days, all the pigs were treated with Enrofloxacin intramuscularly. The efficacy of the Enrofloxacin was confirmed using the ETEC F18 growth inhibition test. Four animals in each treatment group died or were suppressed to reduce pain. The diet did not change growth, the faecal score or the reactive oxygen metabolites in the blood. The XY treatment trended to increase villus length in the jejunum (p = 0.066). The operational taxonomic unit (OTU) distribution was fairly homogeneous, the microbial diversity indices were not changed by the treatment, and the per phylum abundances were homogenous among the diets and were dominated by Bacteroidetes and Firmicutes. The beneficial xylose-fermenting Lactobacillus reuteri persisted after weaning in the XY treatment group (P < 0.05). The Beta Diversity was clusterised for the time of sampling (P = 0.003). The supplementation with xylanase did not improve growth or protection against ETEC, but the effect on some beneficial bacteria species is merits additional study.

F4- and F18-positive enterotoxigenic Escherichia coli (ETEC) are well-known pathogens able to cause a severe swine disease with a worldwide economically importance in pig production. Vaccination of piglets with an oral live bivalent F4/F18 Escherichia coli vaccine has shown some promise and efficacy results in preventing ETEC post weaning diarrhoea (PWD); however, currently, there is a lack of knowledge on its effect on the intestinal microbial community. Furthermore, is not known if the effect of vaccination on gut eubiosis can vary according to pig genetic susceptibility to ETEC F4 and F18. The aim of this study is to evaluate the effect of an E. coli bivalent vaccine and the potential influence of the host genetic susceptibility to ETEC growth performance and faecal microbial profile of healthy piglets around weaning. A total of 288 healthy piglets at 26 days of age (d0; weaning) were divided into two different groups balanced for body weight: i) vaccine (VAX: 6.33 kg) and control (CO: 6.45 kg). At d0 VAX group was orally inoculated with 2 mL of live non-pathogenic E. coli O8:K87 (F4ac-positive) and O141:K94 (F18ac positive) vaccine (Coliprotec F4/F18; Prevtec Microbia an Elanco Company), while CO was orally inoculated with 2 mL of sterile water. At d0 and 18 days later (d18), piglets were individually weighed, and a faecal sample was collected for microbial profile of V3-V4 regions of the 16S rRNA gene using MiSeq Illumina platform. Bristles were collected to determine the genetic ETEC susceptibility by genotyping analysis of MUC4 and FUT1 polymorphisms. No effect of vaccination and genetic susceptibility was observed on growth performance. The vaccination modulated the faecal microbial composition, reducing the alpha diversity indices (P

Arginine plays an important role during reproduction, however, the amount of supplementation in sow diet is still uncertain. The aim of the study was to verify the efficacy of a gestating diet enriched or not with a low dose of L-arginine (Arg) on sow productive performance in terms of numbers and weight of piglets at birth and at weaning, frequency of intrauterine growth retardation (IUGR) and piglets’ mortality, sow placenta weight and salivary humoral immunity and intestinal microbial balance of the sows. 205 sows (Landrace x Large White) were divided into two experimental groups: a control group (CON) (102 sows) and a group supplemented with 0.25% of Arg (ARG) for the whole pregnancy period. Saliva and faecal samples were collected two days before farrowing and used for immunoglobulins and microbial analysis, respectively. Arg improved the number of total born piglets (p = .043) and tended to improve the number of total born alive (p = .086) and to reduce IUGR % (p = .090) and dead piglets at d0–d3 (p = .088). The weight of placenta and humoral immunity were not influenced by Arg. Arg did not modify the faecal microbial structure (alpha and beta indices) but increased the relative abundance of Bacteroidaceae family and Bacteroides genera (p = .0001). The results support the knowledge that Arg plays a key role in nutrition and physiology of pregnant sows without compromising gut eubiosis.HighlightArg supplementation of sows’ gestation diet increased the number of total bornsArg supplementation of sows’ gestation diet did not negatively affect the sows’ gut eubiosisArg plays a significant role in the nutrition of pregnant sows. Arg supplementation of sows’ gestation diet increased the number of total borns Arg supplementation of sows’ gestation diet did not negatively affect the sows’ gut eubiosis Arg plays a significant role in the nutrition of pregnant sows.

Background Probiosis is considered a potential strategy to reduce antibiotics use and prevent post-weaning diarrhea (PWD). This study investigated the effect of Bacillus amyloliquefaciens DSM25840 or Bacillus subtilis DSM25841 supplementation on growth, health, immunity, intestinal functionality and microbial profile of post-weaning pigs after enterotoxigenic E. coli (ETEC) F4 challenge. Methods Sixty-four post-weaning piglets (7748 g ± 643 g) were randomly allocated to four groups: control basal diet (CO); CO + 1.28 × 106 CFU/g of B. amyloliquefaciens (BAA); CO + 1.28 × 106 CFU/g feed of B. subtilis (BAS); CO + 1 g colistin/kg of feed (AB). At day (d) 7, animals were challenged with 105 CFU/mL of ETEC F4ac O149 and then followed for fecal score and performance until d 21. Blood was collected at d 6, d 12 and d 21 for immunoglobulins, at d 8 for acute phase proteins, at d 8 and d 21 for metabolomics analysis. Jejunum was sampled for morphometry, quantification of apoptosis, cell proliferation, neutral and acid mucine and IgA secretory cells, and microarray analysis at d 21. Jejunum and cecum contents were collected for microbiota at d 21. Results AB and BAS reduced the fecal score impairment compared to CO (P Bacillus groups and CO. AB improved BW at d 7, d 14 and d 21, ADWG ADFI and G:F from d 0 to d 7 (P P P = 0.06). Conclusion Bacillus subtilis DSM25841 treatment may reduce ETEC F4ac infection in weaned piglets, decreasing diarrhea and influencing mucosal transcriptomic profile.

The study proposed an exploratory functional analysis on differential gene expression of the jejunum and of cecum in chickens. For this study, 150 Ross 308 male chickens were randomly allotted in six pens (25 birds/pen) and fed the same commercial diet. From 19 birds of 42 days of age, jejunum and cecum mucosae were collected for RNA extraction for transcriptome microarray analysis. Differentially expressed genes (DEGs) submitted to DAVID (Database for Annotation, Visualization, and Integrated Discovery) and Gene Set Enrichment Analysis (GSEA) software evidenced enriched gene clusters for biological functions differentiated in the tissues. DAVID analysis in the jejunum showed enriched annotations for cell membrane integral components, PPAR (peroxisome proliferator-activated receptor) signaling pathway, and peroxisome and lipid metabolism, and showed DEGs for gluconeogenesis, not previously reported in chicken jejunum. The cecum showed enriched annotations for disulfide bond category, cysteine and methionine metabolism, glycoprotein category, cell cycle, and extracellular matrix (ECM). GSEA analysis in the jejunum showed peroxisome and PPAR signaling pathway-related gene sets, as found with DAVID, and gene sets for immune regulation, tryptophan and histidine metabolism, and renin&ndash, angiotensin system, like in mammals. The cecum showed cell cycle and regulation processes, as well as ECM receptor interaction and focal adhesion-related gene sets. Typical intestinal functions specific for the gut site and interesting functional genes groups emerged, revealing tissue-related key aspects which future studies might take advantage of.

Organic acids (OA), especially benzoic acid, have long been used for their wide antimicrobial activity and have shown to promote intestinal health and increase average daily gain (ADG), feed intake (FI) and feed conversion ratio (FCR) of pigs. On the other side, the European Commission decided to ban the use of pharmacological levels of zinc oxide (ZnO) in diets for this animal species from 2022. The aim of this study was to investigate the effect of a blend of protected aromatic compounds including benzoic acid (PBA) as a substitute of supra-nutritional levels of ZnO in piglet diets on growth performance and on gut microbiota profile. The trial was conducted in 4 farms. For each farm, piglets were allocated to one of the two experimental groups: i) piglets receiving a diet with 2.5 g/kg of ZnO; ii) piglets receiving the same diet but with 2.5 g/kg of PBA. Piglets were blocked to each diet based on body weight and sex. 1720 piglets were allocated to the ZnO treatment and 1728 to the PBA. The animals received the treatment for a total of 6 weeks (4 weeks during suckling and 2 weeks post-weaning). Cumulative performance from weaning to 6 weeks post-weaning were calculated. Faecal pool samples were collected from 8 box replicates per group from each farm to characterize the microbial profile. Piglets receiving PBA showed higher ADG (P

Background Colostrum is the first secretion produced by mammary glands during the hours immediately preceding and succeeding parturition. This secretion differs from milk and represents an essential vehicle of passive immunity, prebiotic compounds and growth factors involved in intestinal development. Most of the literature concerning colostrum composition refers mainly to human and cow; and little is known about pig colostrum metabolome and how it varies between pig breeds and different farrowing parity. Thus, the aim of the present research is to provide new information about pig colostrum composition and the associations between metabolites, the sows’ breed and the survival and growth rates of their litters. Results Colostrum samples were gathered from 58 parturitions of sows belonging to three different breeds chosen for their importance in Italian heavy pig production: 31 Large White, 15 Landrace and 12 Duroc respectively. The defatted and ultrafiltered colostrum samples were analysed using 1H–NMR spectroscopy. Principal Components Analysis (PCA) was assessed on the obtained spectra. In addition, using a Stepwise Regression and a Linear Regression analyses the metabolites named after the signals assignment were tested for their associations with piglets’ performances. Twenty-five metabolites were identified, comprehending monosaccharides, disaccharides (such as lactose), organic acids (lactate, citrate, acetate and formate), nitrogenous organic acids (such as creatine) and other compounds, including nucleotides. PCA results evidence a clustering due to breed and season effects. Lactose was the main compound determining the assignment of the samples into different clusters according to the sow breed. Furthermore, some metabolites showed to be associated with piglets’ performance and survival traits: acetate and taurine were positively related to litter weight gain and piglets’ survival rate, respectively, while dimethylamine and cis-aconitate were linked to new-borns’ impaired ability to survive. Conclusions The results obtained suggest that colostrum composition is affected by breed, which, together with environmental conditions, may cause changes in colostrum metabolites content with possible consequences on piglets’ performances. Among the identified metabolites, acetate, taurine, dimethylamine and cis-aconitate showed consistent associations with piglets’ survival rate and litter weight gain, implying that these compounds may affect new-borns’ ability to survive. Electronic supplementary material The online version of this article (10.1186/s40104-018-0237-1) contains supplementary material, which is available to authorized users.

Studies regarding the transfer of mycotoxins from sow feed to colostrum are scarce. A sample of in-house produced lactation feed and one of colostrum were collected from two or three sows per farm (total 49) from 19 farms. The feed contents of aflatoxins (AFs), fumonisins (FUs), deoxynivalenol (DON) and zearalenone (ZEA) were assessed using ELISA and confirmed by liquid chromatography-mass spectrometry (LC-MS), The values were very low (10, 12, 17 and 2 positive samples for AFs, FUs, DON and ZEA, respectively), except for two samples (one AF, one DON). Based on feed values, colostrum samples from 13 farms were tested for at least one mycotoxin (Total 35). Aflatoxins were not found in any sample. A signal for FUs was observed in 5 of 11 colostra, despite low feed values, DON was frequently present in the colostrum (10/14). On the farm where the feed exceeded the DON suggested limits, a higher colostrum content was seen, 10.9 &micro, g/kg, approximately 1/69 of the value showing toxicity in young pigs. The absence of reference values for neonate pigs, and the risk of higher and longer ingestion of DON by sows suggested considering routine checks of sow feed, more research on DON transfer and toxicity in piglets is needed.

Putative genetic markers have been associated with ETEC F4 (Mucine 4 [MUC4]; MUC4 GG;CG as susceptible; MUC4 CC as resistant) and F18 (Fucosyltransferase 1 [FUT1]; FUT1 GG;AG as susceptible; FUT1 AA as resistant) resistances respectively. In this study, 71 post-weaning pigs were followed from d0 (35days old) to d42 (77days of age) to investigate the effect of MUC4 or FUT1 genotypes on the mid-jejunal microbiota composition, pigs expression of genes related to inflammation (IL8, GPX2, REG3G, TFF3, CCL20 and LBPI) and glycomic binding pattern profile (Ulex europaeus agglutinin I [UEA] fucose-binding lectin and peanut agglutinin [PNA] galactose-specific), and on blood plasma targeted metabolomics profile, faecal score and performance parameters of growing healthy pigs. The MUC4 and FUT1 resistant genotypes improved the pigs’ growth performance and had firmed faecal score susceptible genotypes in d0–d21 period. Pigs with MUC4 GG genotype had a higher jejunal expression of genes relate to immune function (CCL20 and REG3G) than MUC4 CG and MUC4 CC pigs (p&nbsp

The host-microbiota interplay is recognized as a key factor for the homeostatic maintenance in animals. In pigs, the weaning transition represents a drastic changes event leading to high risk of gut dysbiosis, which in most cases results in economic losses for swine industry. The blood type antigens expressed on mucosal surfaces can act as receptors for bacterial adhesion and the hypothesis of possible associations between blood groups and intestinal microbial profiles has been tested in human with contrasting results. Nevertheless, no studies testing the blood type as possible shaping factor for gut microbiota are available for pigs. The results of our previous study suggested the porcine AO blood types system as a possible factor influencing the microbiota composition. In the present study, the changes in fecal microbiota of 12 piglets were followed from 7 days after birth to 2 weeks post-weaning, testing the hypothesis that blood types may impact on its structure. No effects attributable to the difference in blood groups were detected, however, the sampling site (faeces) and the low statistical power might have masked the hypothesized impact. The data clearly showed the rearrangement of the bacterial ecosystem triggered by weaning transition; mainly consisting of a shift from a Bacteroidaceae-Enterobacteriaceae dominated community, to a Prevotellaceae-Ruminococcaceae dominated community. The functional analysis by metagenomic predictions suggested a role of the high levels of long-chain fatty acid in swine milk as energy source for Enterobacteriaceae (E. coli), in suckling piglets. This study provides a first insight for further investigations; indicating the need for larger sample size, preferably derived from intestinal mucosa, to test the potential effect of blood groups on gut microbiota profiles, and for analyses aimed at assessing the long-chain fatty acids degradation activity within the intestinal microbiota of suckling piglets, with particular attention to the role of E. coli.

One mechanism through which S. cerevisiae may improve the performance of pigs is by altering the composition of the gut microbiota, a response that may be enhanced by early postnatal supplementation of probiotics. To test this hypothesis, newborn piglets (16 piglets/ group) were treated with either S. cerevisiae yeast (5 x 109 cfu/pig: Low) or (2.5 x 1010 cfu/ piglet: High) or equivalent volume of sterile water (Control) by oral gavage every other day starting from day 1 of age until weaning (28±1 days of age). Piglet body weight was recorded on days 1, 3, 7, 10, 17, 24 and 28 and average daily gain (ADG) calculated for the total period. At weaning, piglets were euthanized to collect cecum content for microbial profiling by sequencing of the 16S rRNA gene. ADG was higher in both Low and High yeast groups than in Control group (P

This study describes the response of piglet microbiota to weaning in various environments, and investigates whether microbiota composition is associated with the robustness of piglets. Faecal samples and growth data were collected just before and 7 days after weaning in 288 piglets from 16 commercial farms characterised by their pathogenic status and antimicrobial use. The effect of weaning on the most abundant microbial families of faecal microbiota was roughly the same in all farms and in agreement with previous findings. Four enterotypes, ubiquitous in all farms, were identified, for which the most discriminating genera werePrevotella,Faecalibacterium,Roseburia, andLachnospira.They corresponded to a gradual maturational shift from pre to post-weaning microbiota. Besides antimicrobial use, the pathogenic status of the farm had a major influence on the post-weaning microbial evolution of apparently healthy piglets. Regarding individual characteristics, piglets whose growth was less perturbed by weaning had moreBacteroidetes(P < 0.01) and lessProteobacteria (P < 0.001).In response to weaning, they showed a greater increase inPrevotella(P < 0.01),Coprococcus(P< 0.01) andLachnospira(P < 0.05) than piglets that grew more slowly. Thus, the microbiota of robust piglets shares common characteristics regardless the living environment of animals.

Pig weaning cause transient stress and reduction of feed intake and intestinal villi. Providing feed with solid texture could stimulate explorative interest, relieving weaning drawbacks and improving welfare. Feed blocks (0.8 kg each) were produced and 8 formulas were preliminarily tested for consumption on a pig farm (ingredients: wheat by-products, dried milk whey, calcium carbonate, oil, molasses). Feed consumption and growth within 3 days after weaning were assessed on penned litters fed the normal feed ad libitum, and one of the 3 best block formulas or a control (wooden pieces). Block consumption was relevant, but growth and carcase quality on a sub-sample reared to commercial maturation were not changed. These formulations were also tested against control on 72 weaned pigs on which behaviour related to feeding and social activities was evaluated by means of surveys using cameras. After 4 days, pigs were slaughtered and the small intestines were sampled for mucosa morphology. In general, block consumption was additive with the consumption of normal feed. Growth was not affected. One formulation (major ingredients: wheat middlings, cane molasses, milk whey and coconut oil) increased the mucosal surface area of the intestinal villi by 7.9% (p p Ingredient composition may influence the disappearance of the feed block, as a variable combination of disaggregation and intake by the piglets.The formulation based on wheat middlings, cane molasses, milk whey and coconut oil may improve growth and favour better welfareBlock supplementation may increase the absorbent surface of the proximal jejunum. Ingredient composition may influence the disappearance of the feed block, as a variable combination of disaggregation and intake by the piglets. The formulation based on wheat middlings, cane molasses, milk whey and coconut oil may improve growth and favour better welfare Block supplementation may increase the absorbent surface of the proximal jejunum.

Porcine colostrum lipid classes and fatty acids (FA) were characterized in 6 pools (from 69 samples) from 3 sow breeds (Italian Large White, Italian Landrace, and Italian Duroc) and different parity orders (only Large White). Triacylglycerols (TAG; 94.44 expressed as g/100 g of fat) were the most abundant lipid class, followed by diacylglycerols (DAG; 3.36 g/100 g of fat), free fatty acids (FFA; 0.98 g/100 g of fat), and cholesterol (0.84 g/100 g of fat). The main FAs found in swine colostrum were palmitic (27.29%, expressed as g/100 g of total FA), oleic (28.81%), and linoleic (23.39%) acids. Both the breed of sow and parity order affected the FA and lipid composition. The results suggest that the FA composition of swine colostrum is similar to that of human colostrum and could represent a new source of nutrients for human infants, after further assessment of hygienic and quality aspects. The swine model could be an opportunity for a better understanding of colostrum effects on newborns.

Plant ingredients contain low Ca levels and 1/3 may be bound to P phytate. Therefore, Ca has to be supplemented with animal or mineral sources, such as limestone or calcium phosphates. However, growth performance and feed intake has been reported by other authors to decrease linearly with increasing dietary Ca. To better understand the possible mechanisms underlying detrimental effect of including limestone in weanling diets, two experimental trials were performed to evaluate its possible impact on growth performance, colonic microbiota and, jejunal gene expression. In Trial 1, a total of 240 piglets (26 ± 2 d old, average body weight (BW) = 7.7 ± 1.04 kg) were distributed into 3 dietary treatments: 0% limestone, 0.8% limestone, and 1.6% limestone to provide 0.6, 0.9 and 1.2% total Ca, respectively. Feed intake and individual BW were registered during the pre-starter phase (d 0–14 post-weaning). Pigs fed diet with 1.6% of limestone (1.2% of total Ca) had lower BW, average daily gain (ADG), and gain to feed ratio (G:F) compared with pigs fed diets with 0.8 and 0% limestone (0.9 and 0.6% total Ca, respectively; P < 0.05). In Trial 2, a total of 18 pigs (28 ± 0 d old, average BW = 7.2 ± 0.24 kg) were allocated individually in cages and assigned to 2 dietary treatments: 0% limestone and 1.4% limestone to provide 0.6 and 1.2% total Ca, respectively. Piglets were fed for 14 d and then euthanized to obtain jejunum tissue for gene expression and colon digesta for 16SRNA gene microbiota analyses. Pigs fed 1.4% limestone (1.2% of total Ca) showed a higher beta-diversity and an increase on the Bacteroides genera in colon (P < 0.05). These animals also showed an up regulation of gene sets related to the cell cycle regulation, DNA and RNA transcription and inflammatory response in the jejunum. In conclusion, high levels of limestone inclusion in weaning diets, decreases growth performance without affecting feed intake in weaned pigs. Also, addition of limestone to diets for 14 d after weaning can upregulate the expression of genes related to the inflammatory response, and enlarge colonic beta-diversity with an increased Bacteroides genera.

Background The stomach is an underestimated key interface between the ingesta and the digestive system, affecting the digestion and playing an important role in several endocrine functions. The quality of starter microbiota and the early life feeding of medium chain triglycerides may affect porcine gastric maturation. Two trials (T1, T2) were carried out on 12 and 24 cesarean-delivered piglets (birth, d0), divided over two microbiota treatments, but slaughtered and sampled at two or three weeks of age, respectively. All piglets were fed orally: sow serum (T1) or pasteurized sow colostrum (T2) on d0; simple starter microbiota (Lactobacillus amylovorus, Clostridium glycolicum and Parabacteroides spp.) (d1-d3); complex microbiota inoculum (sow diluted feces, CA) or a placebo (simple association, SA) (d3-d4) and milk replacer ad libitum (d0-d4). The The T1 piglets and half of the T2 piglets were then fed a moist diet (CTRL); the remaining half of the T2 piglets were fed the CTRL diet fortified with medium chain triglycerides and 7% coconut oil (MCT). Total mRNA from the oxyntic mucosa was analyzed using Affymetrix©Porcine Gene array strips. Exploratory functional analysis of the resulting values was carried out using Gene Set Enrichment Analysis. Results Complex microbiota upregulated 11 gene sets in piglets of each age group vs. SA. Of these sets, 6 were upregulated at both ages, including the set of gene markers of oxyntic mucosa. In comparison with the piglets receiving SA, the CA enriched the genes in the sets related to interferon response when the CTRL diet was given while the same sets were impoverished by CA with the MCT diet. Conclusions Early colonization with a complex starter microbiota promoted the functional maturation of the oxyntic mucosa in an age-dependent manner. The dietary fatty acid source may have affected the recruitment and the maturation of the immune cells, particularly when the piglets were early associated with a simplified starter microbiota.

The aim of the study was to characterize the soluble metabolomics profile of defatted colostrum of sows at different parity number (PA) and to correlate the metabolomics profile with the Brix percentage estimate of colostrum immunoglobulin G (IgG) and sow productive traits. A total of 96 Meidam (crossbreed Large White &times, Meishan) sows of PA from 1&ndash, 4 (PA1: 28, PA2:26, PA3:12, PA4:26) were included, and their productive traits were recorded at 10 days post-farrowing. Colostrum IgG was quantified using a Brix refractometer, and metabolomics profile was assessed using 1H-NMR spectroscopy. Sows&rsquo, PA slightly influenced the metabolomics profile of colostrum. lactose and glycine were higher in PA1 compared with PA4 (p 0.05) and N-acetylglucosamine (GlcNAc) tended to be higher in PA2 than PA3 and PA4 (p &lt, 0.10). The Brix percentage of IgG was negatively associated with lactose and positively with creatine, myo-inositol, and O-phosphocholine (p &lt, 0.05). Taurine was positively related to litter weight at birth. GlcNAc and myo-inositol were linked to piglet mortality at day 10 with a negative and positive trend, respectively. In conclusion, colostrum of gilts and multiparous sows had a similar metabolomics profile. Specific metabolites contributed to explanation of the variability in colostrum Brix percentage estimate of IgG concentration and the sows&rsquo, productive performance.

Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.anifeedsci.2017.06.015.; The use of organic acids in diets for livestock animals is interest due to their being potentially beneficial in maintaining gut homeostasis and controlling pathogenic bacterial growth. The objective of this study was to evaluate the influence of two doses of formic acid supplementation for six weeks to weaner pigs. The long-term effects of formic acid were tested for growth performance, microbiota composition in the jejunum using 163 rRNA, expression of marker genes of inflammation in the jejunum, and quantification of acid-secreting cells in the gastric oxyntic mucosa using immunohistochemistry. Three diets containing no (control; CO), 1.4 g/kg (low formic acid; LFA) or 6.4 g/kg (high formic acid; HFA) free formic acid were fed to 36 pigs for 42 d from 7 d post-weaning (7.1 +/- 1.0 kg live weight) until the end of the trial (27.7 +/- 5.5 kg live weight); they were housed in individual cages for six weeks. During the first three weeks, formic acid supplementation, at any of the dosage above, increased average daily gain (ADG) (P = 0.004), tended to increase average daily feed intake (ADFI) (P = 0.08) and tended to decrease g of feed to g of gain (F:G) (P = 0.08) while no effect was observed during the last three weeks. At the end of the trial, formic acid supplementation had only a minor effect on the microbiota composition in the jejunum. A higher microbiota diversity (Chaol index P < 0.05) was found in the HFA group than in the CO group. The Control group showed a higher abundance (P < 0.05) of Gemella, Lactobacillus and Parvimimoas than the HFA and LFA groups, higher levels of Acinetobacter, Fusobactrium, Leuconostoc respect to the HFA group and of Turcibacter as compared to the LFA group. The abundance of Streptococcus was lower in the CO group than in the HFA group and higher than in the LFA group. The jejunal expression of C-C Motif Chemokine Ligand 2 (CCL20) was higher in the HFA group respect to the LFA group. Formic acid intake did not affect the thickness of the gastric mucosa, the number of parietal cells and stomach weight. This study showed that the addition of formic acid to piglet diets improved growth performance during the first period after weaning and that the long-term supplementation of formic slightly affected the microbiota composition according to the dose.

The yeast populations of natural whey starter (NWS) samples from dairies of PDO Parmigiano Reggiano cheese production area were enumerated and characterised. Yeasts were detected with an average value higher than 3 log cfu mL −1 and in a case study we highlighted the correlation between the yeast growth and incorrect management of NWS fermentation. Isolated strains showed the typical Kluyveromyces marxianus PCR-RFLP pattern and D1/D2 sequences obtained displayed similarity values ranging from 99% to 100% for the same species. RAPD-PCR fingerprinting revealed 14 biotypes. The ability of several strains to grow at 52 °C demonstrates that the yeasts may be perfectly integrated into the ecosystem of NWS. This work confirms that yeasts are part of the NWS secondary microbiota, composed by wild strains with relevant biotechnological interest.

The ability of live yeasts to modulate pig intestinal cell signals in response to infection with Escherichia coli F4ac (ETEC) has not been studied in-depth. The aim of this trial was to evaluate the effect of Saccharomyces cerevisiae CNCM I-4407 (Sc), supplied at different times, on the transcriptome profile of the jejunal mucosa of pigs 24 h after infection with ETEC. In total, 20 piglets selected to be ETEC-susceptible were weaned at 24 days of age (day 0) and allotted by litter to one of following groups: control (CO), CO+ colistin (AB), CO +5 × 1010 colony-forming unit (CFU) Sc/kg feed, from day 0 (PR) and CO+ 5 × 1010 CFU Sc/kg feed from day 7 (CM). On day 7, the pigs were orally challenged with ETEC and were slaughtered 24 h later after blood sampling for haptoglobin (Hp) and C-reactive protein (CRP) determination. The jejunal mucosa was sampled (1) for morphometry; (2) for quantification of proliferation, apoptosis and zonula occludens (ZO-1); (3) to carry out the microarray analysis. A functional analysis was carried out using Gene Set Enrichment Analysis. The normalized enrichment score (NES) was calculated for each gene set, and statistical significance was defined when the False Discovery Rate % was

The development of an early complex gut microbiota may play an important role in the protection against intestinal dysbiosis later in life. The significance of the developed microbiota for gut barrier functionality upon interaction with pathogenic or beneficial bacteria is largely unknown. The transcriptome of differently perfused jejunal loops of 12 caesarian-derived pigs, neonatally associated with microbiota of different complexity, was studied. Piglets received pasteurized sow colostrum at birth (d0), a starter microbiota (Lactobacillus amylovorus (LAM), Clostridium glycolicum, and Parabacteroides) on d1-d3, and a placebo inoculant (simple association, SA) or an inoculant consisting of sow's diluted feces (complex association, CA) on d3-d4. On d 26-37, jejunal loops were perfused for 8 h with either enterotoxigenic Escherichia coli F4 (ETEC), purified F4 fimbriae, LAM or saline control (CTRL). Gene expression of each intestinal loop was analyzed by Affymetrix Porcine Gene 1.1_ST array strips. Gene Set Enrichment Analysis was performed on expression values. Compared to CTRL, 184 and 74; 2 and 139; 2 and 48 gene sets, were up- and down-regulated by ETEC, F4 and LAM, respectively. ETEC up-regulated networks related to inflammatory and immune responses, RNA processing, and mitosis. There was a limited overlap in up-regulated gene sets between ETEC and F4 fimbriae. LAM down-regulated genes related to inflammatory and immune responses, as well as to cellular compound metabolism. In CA pigs, 57 gene sets were up-regulated by CA, while 73 were down-regulated compared to SA. CA up-regulated gene sets related to lymphocyte modulation and to cellular defense in all loop perfusions. In CA pigs, compared to SA pigs, genes for chemokine and cytokine activity and for response to external stimuli were down-regulated in ETEC-perfused loops and up-regulated in CTRL. The results highlight the importance of the nature of neonatal microbial colonization in the response to microbial stimuli later in life.

Genome-wide association study results are presented for intramuscular fat in Italian Large White pig breed. A total of 886 individuals were genotyped with PorcineSNP60 BeadChip. After quality control performed with plink software and in R environment, 49208 markers remained for the association analysis. The genome-wide association studies was conducted using linear mixed model implemented in GenABEL. We detected seven new SNPs of genes till now not found associated to intramuscular fat (IMF). Three markers map in a wide intergenic region rich of QTL linked to fat traits, one map 388kb upstream the gene SDK1, one map inside PPP3CA gene, one inside SCPEP1 gene and the last is not mapped in the porcine genome yet. Associations here presented indicate a moderate effect of these genes on IMF. In particular, PPP3CA, that is involved in the oxidative metabolism of skeletal muscle, could be considerated as an interesting candidate gene for IMF content in pigs. However, further studies are needed to clarify the role of these genes on the physiological processes involved in IMF regulation. These results may be useful to control this trait that is important in terms of nutritional, technological and organoleptic characteristics of fresh meat and processed products.

Here, we investigated the possible linkages among geophagy, soil characteristics, and gut mycobiome of indri (Indri indri), an endangered lemur species able to survive only in wild conditions. The soil eaten by indri resulted in enriched secondary oxide-hydroxides and clays, together with a high concentration of specific essential micronutrients. This could partially explain the role of the soil in detoxification and as a nutrient supply. Besides, we found that soil subject to geophagy and indris’ faeces shared about 8.9% of the fungal OTUs. Also, several genera (e.g. Fusarium, Aspergillus and Penicillium) commonly associated with soil and plant material were found in both geophagic soil and indri samples. On the contrary, some taxa with pathogenic potentials, such as Cryptococcus, were only found in indri samples. Further, many saprotrophs and plant-associated fungal taxa were detected in the indri faeces. These fungal species may be involved in the digestion processes of leaves and could have a beneficial role in their health. In conclusion, we found an intimate connection between gut mycobiome and soil, highlighting, once again, the potential consequent impacts on the wider habitat.