Post-weaning Diarrhoea In Piglets Research Articles

Preparation of Bispecific IgY-scFvs Inhibition Adherences of Enterotoxigenic Escherichia coli (K88 and F18) to Porcine IPEC-J2 Cell.

Enterotoxigenic Escherichia coli (ETEC) strains are significant contributors to postweaning diarrhea in piglets. Of the ETEC causing diarrhea, K88 and F18 accounted for 92.7%. Despite the prevalence of ETEC K88 and F18, there is currently no effective vaccine available due to the diversity of these strains. This study presents an innovative approach by isolating chicken-derived single-chain variable fragment antibodies (scFvs) specific to K88 and F18 fimbrial antigens from chickens immunized against these ETEC virulence factors. These scFvs effectively inhibited adhesion of K88 and F18 to porcine intestinal epithelial cells (IPEC-J2), with the inhibitory effect demonstrating a dose-dependent increase. Furthermore, a bispecific scFv was designed and expressed in Pichia pastoris. This engineered construct displayed remarkable potency; at a concentration of 25.08 μg, it significantly reduced the adhesion rate of ETEC strains to IPEC-J2 cells by 72.10% and 69.11% when challenged with either K88 or F18 alone. Even in the presence of both antigens, the adhesion rate was notably decreased by 57.92%. By targeting and impeding the initial adhesion step of ETEC pathogenesis, this antibody-based intervention holds promise as a potential alternative to antibiotics, thereby mitigating the risks associated with antibiotic resistance and residual drug contamination in livestock production. Overall, this study lays the groundwork for the development of innovative treatments against ETEC infections in piglets.

Towards identification of new genetic determinants for post-weaning diarrhea in piglets.

Post-weaning diarrhea in pigs is a considerable challenge in the pig farming industry due to its effect on animal welfare and production costs, as well as the large volume of antibiotics, which are used to treat diarrhea in pigs after weaning. Previous studies have revealed loci on SSC6 and SSC13 associated with susceptibility to specific diarrhea causing pathogens. This study aimed to identify new genetic loci for resistance to diarrhea based on phenotypic data. In depth clinical characterization of diarrhea was performed in 257 pigs belonging to two herds during the first 14 days post weaning. The daily diarrhea assessments were used for the classification of pigs into case and control groups. Pigs were assigned to case and control groups based only on the incidence of diarrhea in the second week of the study in order to differentiate between differences in etiology. Genome-wide association studies and metabolomics association analysis were performed in order to identify new biological determinants for diarrhea susceptibility. With the present work, we revealed a new locus for diarrhea resistance on SSC16. Furthermore, studies of metabolomics in the same pigs revealed one metabolite associated with diarrhea.

Lactobacillus johnsonii Improves Intestinal Barrier Function and Reduces Post-Weaning Diarrhea in Piglets: Involvement of the Endocannabinoid System

Probiotic intervention is a well-established approach for replacing antibiotics in the management of weaning piglet diarrhea, which involves a large number of complex systems interacting with the gut microbiota, including the endocannabinoid system; nevertheless, the specific role of the endocannabinoid system mediated by probiotics in the piglet intestine has rarely been studied. In this study, we used antibiotics (ampicillin) to perturb the intestinal microbiota of piglets. This resulted in that the gene expression of the intestinal endocannabinoid system was reprogrammed and the abundance of probiotic Lactobacillus johnsonii in the colon was lowered. Moreover, the abundance of Lactobacillus johnsonii was positively correlated with colonic endocannabinoid system components (chiefly diacylglycerol lipase beta) via correlation analysis. Subsequently, we administered another batch of piglets with Lactobacillus johnsonii. Interestingly, dietary Lactobacillus johnsonii effectively alleviated the diarrhea ratio in weaning piglets, accompanied by improvements in intestinal development and motility. Notably, Lactobacillus johnsonii administration enhanced the intestinal barrier function of piglets as evidenced by a higher expression of tight junction protein ZO-1, which might be associated with the increased level in colonic diacylglycerol lipase beta. Taken together, the dietary Lactobacillus johnsonii-mediated reprogramming of the endocannabinoid system might function as a promising target for improving the intestinal health of piglets.

Feed supplementation with Ligilactobacillus salivarius PS21603 optimises intestinal morphology and gut microbiota composition in weaned piglets.

Post-weaning diarrhoea in piglets remains an important cause of economic losses for swine producers. Feed supplementation with probiotics is one of the alternatives to antibiotics used to reduce the impact of such gastrointestinal disease. The aim of the present study was to evaluate the effect of Ligilactobacillus salivarius PS21603 supplementation on the intestinal structure and the gut microbiota composition of weaned piglets. Safety and tolerance of L. salivarius PS21603 were previously evaluated in a 28-days study using 384 weaned piglets (28± 2 days old and 7.5± 1.5kg) divided in three treatment groups: T1: Basal diet+ L. salivarius PS21603 109 cfu/day, T2: Basal diet+ L. salivarius PS21603 107 cfu/day, and T3: Basal diet (control group). For the present study, 16 piglets per treatment group were randomly selected and faecal samples were collected on day 0 (weaning) and 28 of study. At the end of study, three males and three females per treatment were euthanised. Intestinal morphometric values were measured after necropsy. Faecal counts of Escherichia coli were evaluated by culture techniques, and faecal microbiota composition was assessed by high-throughput sequencing. All data were analysed and compared between treatment groups. Supplementation with L. salivarius PS21603 caused an increase in the intestine length of piglets from T1 and in the villous height:crypt ratio of piglets from T2 (P< 0.05) compared to T3 on day 28. According to the Shannon Diversity Index, microbiota diversity increased on day 28 compared to day 0, with no significant differences observed between treatments. The main changes in the relative abundance of bacteria at the phylum, family, and genus levels were observed between different sampling time points. However, piglets from T1 and T2 had lower faecal E. coli counts than T3 on day 28 (P< 0.05). Moreover, supplementation with L. salivarius PS21603 modulated gut microbiota through a more optimal composition, reducing Escherichia and increasing Bifidobacterium relative abundance in piglets from T1 (P< 0.05) from the beginning to the end of the study. Therefore, the strain L. salivarius PS21603 has shown probiotic properties to be used as feed additive in the pig industry, along with good hygiene and farm management practices, for the prevention and/or treatment of post-weaning diarrhoea in piglets.

Developing and Evaluating a Weaned Piglet Model of Enterotoxigenic Escherichia Coli Infection

Enterotoxigenic Escherichia Coli (ETEC) remains the main pathogen associated with post-weaning diarrhea in piglets. Here, we introduce a reliable method to develop an ETEC-infected weaned piglet model. For the first 24 h post-birth, neonatal piglets were separated from dams so that they did not receive colostrum. Afterwards, piglets were returned to the care of their mothers, until weaning (21-day-old). On days 25, 26 and 27 post-birth, piglets were orally challenged with 1011 Colony-Forming Units (CFU) of virulent ETEC strain incubated in colony-forming antigen medium and delivered in chitosan-coated capsules. In all infected piglets, severe diarrhea and typical body weight loss were observed post-infection. To simplify the work, for the newly developed method, the number of challenging ETEC cells was lowered to 1010 CFU/head and the incubation medium was changed to tryptic soy. These modifications influenced neither the diarrheal defecation ratio nor the body weight loss in piglets. To induce experimentally ETEC infection in future work, we recommend that 1010 CFU/head of ETEC cell grown in tryptic soy broth be delivered in chitosan-coated capsules to colostrum restricted piglets, because this methodology stably caused 100% diarrheal defecation and growth reduction.

Effects of a Natural Polyphenolic Product from Olive Mill Wastewater on Oxidative Stress and Post-Weaning Diarrhea in Piglets

The present study aimed to investigate the effects of a commercial phytogenic feed additive (PFA) on the prevention of post-weaning diarrhea and oxidative stress in piglets. The concentrations of thiobarbituric acid reactive substances (TBARS) and protein carbonyls (CARBS) were investigated as biomarkers for oxidative damage, as were the health and performance parameters of weaned piglets. In total, 100 weaned piglets were divided into two groups: a control group (T1), which was fed regular weaning feed; an experimental group (T2), which was fed regular weaning feed supplemented with a phenolic feed additive (PFA) for 3 weeks. The TBARS and CARBS concentrations in plasma samples from 20 piglets per group were measured at 45 and 65 days of age. Fecal samples were collected from 24 weaned piglets per group using FTA ELUTE cards. Diarrhea score, body weight (BW) at weaning, and average daily weight gain (ADWG) were recorded. The TBARS (p &lt; 0.001) and CARBS (p = 0.001) concentrations were significantly higher in the T1 group compared to those in the T2 group. The lowest diarrhea score was noted in the T2 group for the age groups of 45 (p &lt; 0.001) and 65 days (p = 0.008). In conclusion, the use of a phenolic PFA in the current study had beneficial antioxidative and antimicrobial effects on weaned piglets, which improved their health and growth performance.

The Early Life Microbiota Is Not a Major Factor Underlying the Susceptibility to Postweaning Diarrhea in Piglets.

Postweaning diarrhea (PWD) in piglets impair welfare, induce economic losses and lead to overuse of antibiotics. The early life gut microbiota was proposed to contribute to the susceptibility to PWD. The objective of our study was to evaluate in a large cohort of 116 piglets raised in 2 separate farms whether the gut microbiota composition and functions during the suckling period were associated with the later development of PWD. The fecal microbiota and metabolome were analyzed by 16S rRNA gene amplicon sequencing and nuclear magnetic based resonance at postnatal day 13 in male and female piglets. The later development of PWD was recorded for the same animals from weaning (day 21) to day 54. The gut microbiota structure and α-diversity during the suckling period were not associated with the later development of PWD. There was no significant difference in the relative abundances of bacterial taxa in suckling piglets that later developed PWD. The predicted functionality of the gut microbiota and the fecal metabolome signature during the suckling period were not linked to the later development of PWD. Trimethylamine was the bacterial metabolite which fecal concentration during the suckling period was the most strongly associated with the later development of PWD. However, experiments in piglet colon organoids showed that trimethylamine did not disrupt epithelial homeostasis and is thus not likely to predispose to PWD through this mechanism. In conclusion, our data suggest that the early life microbiota is not a major factor underlying the susceptibility to PWD in piglets. IMPORTANCE This study shows that the fecal microbiota composition and metabolic activity are similar in suckling piglets (13 days after birth) that either later develop post-weaning diarrhea (PWD) or not, which is a major threat for animal welfare that also causes important economic losses and antibiotic treatments in pig production. The aim of this work was to study a large cohort of piglets raised in separates environments, which is a major factor influencing the early life microbiota. One of the main findings is that, although the fecal concentration of trimethylamine in suckling piglets was associated with the later development of PWD, this gut microbiota-derived metabolite did not disrupt the epithelial homeostasis in organoids derived from the pig colon. Overall, this study suggests that the gut microbiota during the suckling period is not a major factor underlying the susceptibility of piglets to PWD.

Selected Nutrition and Management Strategies in Suckling Pigs to Improve Post-Weaning Outcomes.

Weaning is a critical period in a pig's life. Piglets are confronted with abrupt changes to their physical and social environment, as well as management and nutritional changes. Weaning has always been associated with a growth check and is frequently accompanied by post-weaning diarrhoea in piglets. However, rapid increases in litter size in the last decade have increased within-litter piglet weight variation, with piglets now generally lighter at weaning, making the challenges associated with weaning even greater. Many interventions can be employed during the suckling period to ease the weaning transition for piglets. Pre-weaning strategies such as supervised farrowing (assistance with suckling and oxytocin provision), the provision of pain relief to sows around farrowing, split-suckling, early oral supplementation with glucose, bovine colostrum, faecal microbiota transplantation, feed additives and solid and liquid creep feeding (milk and liquid feed) have all been investigated. The objective of these strategies is to stimulate earlier maturation of the digestive tract, improve immunity, reduce latency to the first feed post-weaning and increase early post-weaning feed intake and growth. This review focuses in particular on: (1) pain relief provision to sows around farrowing, (2)split-suckling of piglets, (3) pre-weaning provision of supplementary milk and/or liquid feed, (4) other strategies to stimulate earlier enzyme production (e.g., enzyme supplementation), (5) other nutritional strategies to promote improved gut structure and function (e.g., L-glutamine supplementation), and (6) other strategies to modulate gut microbiota (e.g., probiotics and prebiotics). Correctly implementing these strategies can, not only increase post-weaning growth and reduce mortality, but also maximise lifetime growth in pigs.

Araceae root and citrus fibers tend to decrease Escherichia coli adhesion and myeloperoxidase levels in weaned piglets.

Weaning is a stressful experience in the piglet's life, and it often coincides with impaired gut health. Post-weaning diarrhea in piglets is frequently caused by enterotoxigenic Escherichia coli (E. coli). The first step of an E. coli infection is the adhesion to host-specific receptors present on enterocytes, leading to pro-inflammatory immune responses. The aim of this study was to examine if specific fiber fractions in the piglet diet can prevent E. coli adhesion and subsequent immune responses. The trial included 200 piglets (Danbred × Piétrain): 10 piglets/pen × 10 pens/dietary treatment × 2 dietary treatments. From weaning until 14 days (d14) post-weaning, piglets were fed a control diet or test diet with 2 kg/ton of a mixture of specific fiber fractions derived from Araceae root and citrus. Afterwards, 1 piglet per pen was euthanized, a section was taken at 75% of small intestinal length and E. coli colonization on the mucosal epithelium was quantified by scraping and conventional plating. From the same small intestinal section, histo-morphological indices were assessed, and mucosal scrapings were analyzed for gene expression of pro- and anti-inflammatory cytokines, and NF-kB. Analyses of specific intestinal bacteria and SCFA were performed on samples of intestinal content (small intestine, caecum, colon). Fecal samples were taken to measure myeloperoxidase (MPO), calprotectin and PAP/RAG3A as biomarkers for intestinal inflammation. Piglets fed the fiber mixture tended to have decreased E. coli colonization to the mucosal epithelium (5.65 vs. 4.84 log10 CFU/g; P = 0.07), less E. coli in the caecum (8.91 vs. 7.72 log10 CFU/g; P = 0.03) and more Lachnospiraceae in the colon (11.3 vs. 11.6 log10 CFU/g; P = 0.03). Additionally, the fiber mixture tended to increase cecal butyric acid (10.4 vs. 19.1 mmol/kg; P = 0.07). No significant effect on histo-morphological indices and on gene expression of pro- and anti-inflammatory cytokines and NF-kB was observed. The fecal MPO concentration tended to decrease (20.2 vs. 10.4 ng/g; P = 0.07), indicating less intestinal inflammation. In conclusion, this study showed that specific fiber fractions from Araceae root and citrus in piglet weaner diets may decrease the risk of pathogen overgrowth by reducing E. coli adhesion and intestinal inflammation.

Piperine as potential therapy of post-weaning porcine diarrheas: an in vitro study using a porcine duodenal enteroid model

Post-weaning diarrhea in piglets is a major problem, resulting in a significant loss in pig production. This study aimed to investigate the effects of piperine, an alkaloid abundantly found in black peppers, on biological activities related to the pathogenesis of post-weaning diarrhea using a porcine duodenal enteroid model, a newly established intestinal stem cell-derived in vitro model recapitulating physiology of porcine small intestinal epithelia. Porcine duodenal enteroid models were treated with disease-relevant pathological inducers with or without piperine (8 μg/mL and/or 20 μg/mL) before measurements of oxidative stress, mRNA, and protein expression of proinflammatory cytokines, nuclear factor-kappa B (NF-κB) nuclear translocation, barrier leakage, and fluid secretion. We found that piperine (20 μg/mL) inhibited H2O2-induced oxidative stress, TNF-α-induced mRNA, and protein expression of proinflammatory cytokines without affecting NF-κB nuclear translocation, and prevented TNF-α-induced barrier leakage in porcine duodenal enteroid monolayers. Importantly, piperine inhibited fluid secretion induced by both forskolin and heat-stable toxins (STa) in a three-dimensional model of porcine duodenal enteroids. Collectively, piperine possesses both anti-inflammatory and anti-secretory effects in porcine enteroid models. Further research and development of piperine may provide novel interventions for the treatment of post-weaning porcine diarrhea.

Short Communication: Dietary butyrate and valerate glycerides impact diarrhea severity and immune response of weaned piglets under ETEC F4-ETEC F18 coinfection conditions.

Enterotoxigenic Escherichia coli (ETEC) causes post-weaning diarrhea in piglets, significantly impacting animal welfare and production efficiency. The two primary ETEC pathotypes associated with post-weaning diarrhea are ETEC F4 and ETEC F18. During the post-weaning period, piglets may be exposed to both ETEC F4 and ETEC F18. However, the effects of coinfection by both strains have not been studied. Short chain fatty acid (SCFA) feed additives, such as butyrate and valerate, are being investigated for their potential to improve animal performance and disease resistance. Therefore, this pilot experiment aimed to test the effects of butyrate glycerides or valerate glycerides on growth performance, diarrhea incidence, and immune responses of piglets under ETEC F4-ETEC F18 coinfection conditions. Twenty piglets were individually housed and assigned to one of three dietary treatments immediately at weaning (21-24 d of age). The dietary treatments included control (basal diet formulation), control supplemented with 0.1% butyrate glycerides or 0.1% valerate glycerides. After a seven-day adaptation, all pigs were inoculated with ETEC F4 and ETEC F18 (0.5 × 109 CFU/1.5mL dose for each strain) on three consecutive days. Pigs and feeders were weighed throughout the trial to measure growth performance. Fecal cultures were monitored for hemolytic coliforms, and blood samples were collected for whole blood and serum analysis. Pigs fed valerate glycerides tended (P = 0.095) to have higher final body weight compared with control. The overall severity of diarrhea was significantly (P < 0.05) lower in both treatment groups than control. Pigs fed valerate glycerides tended (P = 0.061) to have lower neutrophils and had significantly (P < 0.05) lower serum TNF-α on d 4 post-inoculation. This pilot experiment established an appropriate experimental dose for an ETEC F4-ETEC F18 coinfection disease model in weaned piglets. Results also suggest that butyrate glycerides and valerate glycerides alleviated diarrhea and regulated immune responses in piglets coinfected with ETEC F4 and ETEC F18.

Effects of maternal supplementation of fish oil during late gestation and lactation on growth performance, fecal microbiota structure and post-weaning diarrhoea of offspring piglets.

Homeostasis of gut microbiota is a critical contributor to growth and health in weaned piglets. Fish oil is widely reported to benefit health of mammals including preventing intestinal dysfunction, yet its protective effect during suckling-to-weaning transition in piglets remains undetermined. Low (30 g/d) and high (60 g/d) doses of n-3-rich fish oil were supplemented in sows from late gestation to lactation. Serum indicators and gut microbiota were determined to evaluate the effects of maternal fish oil on growth performance, immunity and diarrhea of piglets. DHA and EPA in the colostrum as well as serum of suckling and 1-week post-wean piglets were significantly and linearly increased by maternal supplementation of fish oil (P < 0.05). IGF1 and T3 in nursing and weaned piglets were significantly elevated by maternal fish oil (P < 0.05), and the increase of IGF1 was concerning the dosage of fish oil. Colostrum IgG, plasma IgG, IgM in suckling piglets, IgG, IgM and IgA in weaned piglets were significantly increase as maternal replenishment of fish oil increased (P < 0.05). Additionally, cortisol was significantly reduced in weaned pigs (P < 0.05), regardless of dosage. 16S rRNA sequencing revealed that α-diversity of fecal microbiota in nursery piglets, and fecal Lactobacillus genus, positively correlated with post-weaning IgA, was significantly increased by high dosage. Collectively, maternal fish oil during late pregnancy and lactation significantly promoted growth, enhanced immunity, and reduced post-weaning diarrhea in piglets, therefore facilitated suckling-to-weaning transition in piglets, which may be partially due to the altered gut microbial community.

Identification of a 5-Methylcytosine Site (mC-7) That May Inhibit CXCL11 Expression and Regulate E. coli F18 Susceptibility in IPEC-J2 Cells.

The primary pathogen causing post-weaning diarrhea in piglets is Escherichia coli F18 (E. coli F18), hence it is essential to investigate the mechanism governing E. coli F18 resistance in native pig breeds. Based on the previous RNA-seq results of the duodenum from E. coli F18-resistant and -susceptible Meishan piglets, CXCL11, an important functional gene, was preliminarily screened. In this investigation, in order to further examine the expression regulation mechanism of E. coli F18 in intestinal porcine epithelial cells (IPEC-J2) against E. coli F18 infection, CXCL11 gene expression on IPEC-J2 cells infected by E. coli F18 was detected, which was significantly downregulated (p &lt; 0.01). Secondly, the overexpression on the IPEC-J2 cell line was successfully structured, and a relative quantification method of the PILIN, bacteria enumeration, and immunofluorescence assay indicated that the CXCL11 overexpression significantly reduced the ability of E. coli F18 to interact with IPEC-J2 in vitro. The promoter region of the CXCL11 gene was predicted to contain a CpG island (-619 ~ -380 bp) of which 13 CpG sites in the sequencing region were methylated to varying degrees, and the methylation level of one CPG site (mC-7) positively linked negatively with the expression of the CXCL11 gene (p &lt; 0.05). Meanwhile, a dual luciferase assay detected the mutation of the mC-7 site that significantly inhibited the luciferase activity of the CXCL11 gene promoter (p &lt; 0.01). Transcription factor prediction and expression verification indicated that mC-7 is located in the OSR1-binding domain, and that its expression level is related to E. coli F18 susceptibility. We speculated that methylation modification of the mC-7 site of the CpG island in the promoter region of the CXCL11 gene might inhibit the binding of transcription factor OSR1 with the mC-7 site, and then affect its expression level to regulate the susceptibility to E. coli F18.

Review on Preventive Measures to Reduce Post-Weaning Diarrhoea in Piglets.

Simple SummaryPost-weaning diarrhoea (PWD) is a major challenge in pig production, which results in the high use of ZnO for its prevention and the utilisation of antibiotics for its treatment. There is a global agenda to decrease the use of antibiotics in order to reduce the risk of antimicrobial resistance (AMR); moreover, from June 2022, administering medical levels of ZnO has been banned in many countries due to its negative impact on the environment and AMR. Many feeding strategies and feed additives have been found to be tools to reduce PWD risk, which affect the gut microbiota and the host through different modes of action. Many feeding strategies and additives show positive but variable effects on PWD. A combination of such interventions tailored according to the specific conditions is most probably the best strategy.In many countries, medical levels of zinc (typically as zinc oxide) are added to piglet diets in the first two weeks post-weaning to prevent the development of post-weaning diarrhoea (PWD). However, high levels of zinc constitute an environmental polluting agent, and may contribute to the development and/or maintenance of antimicrobial resistance (AMR) among bacteria. Consequently, the EU banned administering medical levels of zinc in pig diets as of June 2022. However, this may result in an increased use of antibiotic therapeutics to combat PWD and thereby an increased risk of further AMR development. The search for alternative measures against PWD with a minimum use of antibiotics and in the absence of medical levels of zinc has therefore been intensified over recent years, and feed-related measures, including feed ingredients, feed additives, and feeding strategies, are being intensively investigated. Furthermore, management strategies have been developed and are undoubtedly relevant; however, these will not be addressed in this review. Here, feed measures (and vaccines) are addressed, these being probiotics, prebiotics, synbiotics, postbiotics, proteobiotics, plants and plant extracts (in particular essential oils and tannins), macroalgae (particularly macroalgae-derived polysaccharides), dietary fibre, antimicrobial peptides, specific amino acids, dietary fatty acids, milk replacers, milk components, creep feed, vaccines, bacteriophages, and single-domain antibodies (nanobodies). The list covers measures with a rather long history and others that require significant development before their eventual use can be extended. To assess the potential of feed-related measures in combating PWD, the literature reviewed here has focused on studies reporting parameters of PWD (i.e., faeces score and/or faeces dry matter content during the first two weeks post-weaning). Although the impact on PWD (or related parameters) of the investigated measures may often be inconsistent, many studies do report positive effects. However, several studies have shown that control pigs do not suffer from diarrhoea, making it difficult to evaluate the biological and practical relevance of these improvements. From the reviewed literature, it is not possible to rank the efficacy of the various measures, and the efficacy most probably depends on a range of factors related to animal genetics and health status, additive doses used, composition of the feed, etc. We conclude that a combination of various measures is probably most recommendable in most situations. However, in this respect, it should be considered that combining strategies may lead to additive (e.g., synbiotics), synergistic (e.g., plant materials), or antagonistic (e.g., algae compounds) effects, requiring detailed knowledge on the modes of action in order to design effective strategies.

Lactoferrin Decreases Enterotoxigenic Escherichia coli-Induced Fluid Secretion and Bacterial Adhesion in the Porcine Small Intestine.

Enterotoxigenic Escherichia coli (ETEC) infections are one of the most prevalent causes of post-weaning diarrhea in piglets, resulting in morbidity, mortality and elevated use of antibiotics. The emergence and further spread of antimicrobial resistance together with the growing demand for high quality animal protein requires the identification of novel alternatives for antimicrobials. A promising alternative is lactoferrin, as we previously showed that it can both inhibit the growth and degrade bacterial virulence factors of porcine ETEC strains in vitro. Aiming to confirm these findings in vivo, we performed a small intestinal segment perfusion experiment in piglets. Here, we showed that lactoferrin could not only decrease ETEC-induced fluid secretion, but also their ability to colonize the small intestinal epithelium. Furthermore, while ETEC infection induced pro-inflammatory cytokine mRNA expression in this experiment, lactoferrin was not able to counteract these responses. In addition, a bacterial motility assay showed that lactoferrin can reduce the motility of ETEC. Our findings further support the use of lactoferrin as an alternative for antimicrobials and also show its potential for the prevention of ETEC infections in pigs.

The Role of Probiotics in Alleviating Postweaning Diarrhea in Piglets From the Perspective of Intestinal Barriers.

Early weaning of piglets is an important strategy for improving the production efficiency of sows in modern intensive farming systems. However, due to multiple stressors such as physiological, environmental and social challenges, postweaning syndrome in piglets often occurs during early weaning period, and postweaning diarrhea (PWD) is a serious threat to piglet health, resulting in high mortality. Early weaning disrupts the intestinal barrier function of piglets, disturbs the homeostasis of gut microbiota, and destroys the intestinal chemical, mechanical and immunological barriers, which is one of the main causes of PWD in piglets. The traditional method of preventing PWD is to supplement piglet diet with antibiotics. However, the long-term overuse of antibiotics led to bacterial resistance, and antibiotics residues in animal products, threatening human health while causing dysbiosis of gut microbiota and superinfection of piglets. Antibiotic supplementation in livestock diets is prohibited in many countries and regions. Regarding this context, finding antibiotic alternatives to maintain piglet health at the critical weaning period becomes a real emergency. More and more studies showed that probiotics can prevent and treat PWD by regulating the intestinal barriers in recent years. Here, we review the research status of PWD-preventing and treating probiotics and discuss its potential mechanisms from the perspective of intestinal barriers (the intestinal microbial barrier, the intestinal chemical barrier, the intestinal mechanical barrier and the intestinal immunological barrier) in piglets.

P194. Dietary indigestible protein increases post-weaning diarrhoea in piglets irrespective of sanitary housing conditions

P194. Dietary indigestible protein increases post-weaning diarrhoea in piglets irrespective of sanitary housing conditions

Lower abundance of Bacteroides and metabolic dysfunction are highly associated with the post-weaning diarrhea in piglets.

Growing evidences show a direct link between diarrhea and disorders of gut microbiota in pigs. However, whether there are microbial markers associated with post-weaning diarrhea remains unknown. In the current study, we compared the microbial community, functions and metabolites between healthy weaned piglets (group H, n=7) and piglets with post-weaning diarrhea (group D, n=7), in order to find out diarrhea associated microbial markers. Each of 7 fecal samples was collected from H and D piglets (weaned at 21 d and sampled at 26 d). The metagenomic and untargeted metabolomic analysis revealed that the microbial composition, function and metabolic profile in D pigs was considerably reshaped, including the reduced abundance and number of Bacteroides, which significantly correlated with the diarrhea status of host. The carbohydrate metabolism, biosynthesis and metabolism, lipid metabolism, amino acid metabolism, and the activity of glycan and carbohydrates digestion related enzymes showed extensively down-regulated in D pigs compared with H pigs. Diarrhea significantly changed the metabolic profiles of fecal microbiota, and most of the altered metabolites were negatively or positively correlated with the change in the abundance of Bacteroides. In conclusion, the lower abundance of Bacteroides and its associated metabolic dysfunction may be regarded as microbial markers of physiological post-weaning diarrhea in piglets.

Longitudinal study of the mcr-1 gene prevalence in Spanish food-producing pigs from 1998 to 2021 and its relationship with the use of polymyxins

BackgroundResistance to colistin was an uncommon phenomenon traditionally linked to chromosome point mutations, but since the first description of a plasmid-mediated colistin-resistance in late 2015, transmissible resistance to colistin has become a Public Health concern. Despite colistin is considered as a human last resort antibiotic, it has been commonly used in swine industry to treat post-weaning diarrhoea in piglets. However, the progressively increase of colistin resistance during the last decade led to the Spanish Medicines and Healthcare Products Agency (AEMPS) to launch a strategic and voluntary plan aimed to reduce colistin consumption in pig production. Our longitudinal study (1998–2021) aimed to evaluate the trend of colistin resistance mediated through the mcr-1 mobile gene in Spanish food-producing pig population and compare it with published polymyxin sales data in veterinary medicine to assess their possible relationships.ResultsThe first mcr-1 positive sample was observed in 2004, as all samples from 1998 and 2002 were mcr-1 PCR-negative. We observed a progressive increase of positive samples from 2004 to 2015, when mcr-1 detection reached its maximum peak (33/50; 66%). From 2017 (27/50; 54%) to 2021 (14/81; 17%) the trend became downward, reaching percentages significantly lower than the 2015 peak (p < 0.001). The abundance of mcr-1 gene in PCR-positive samples showed a similar trend reaching the highest levels in 2015 (median: 6.6 × 104mcr-1 copies/mg of faeces), but decreased significantly from 2017 to 2019 (median 2.7 × 104, 1.2 × 103, 4.6 × 102mcr-1 copies/mg of faeces for 2017, 2018 and 2019, respectively), and stabilizing in 2021 (1.6 × 102mcr-1 copies/mg of faeces) with similar values than 2019.ConclusionsOur study showed the decreasing trend of colistin resistance associated to mcr-1 gene, after a previous increase from among 2004–2015, since the European Medicines Agency and AEMPS strategies were applied in 2016 to reduce colistin use in animals, suggesting a connection between polymyxin use and colistin resistance. Thus, these plans could have been effective in mcr-1 reduction, reaching lower levels than those detected in samples collected 17 years ago, when resistance to colistin was not yet a major concern.

Gut microbiome modifications over time when removing in-feed antibiotics from the prophylaxis of post-weaning diarrhea in piglets.

Post-weaning diarrhea (PWD) in pigs has mainly an infectious basis and control strategies are centred on antibiotics added to the diet. Given concerns on the spread of multi-resistant bacteria, it is necessary to develop alternative prophylactic approaches to control PWD in piglets. The most promising alternative strategies are based on substances that act indirectly on the bacteria by stimulating the immune system or by improving gut health. The aim of this study was to evaluate the effect on the gut microbiota of feed supplemented with a mixture of essential oils (garlic and oregano) in weaning piglets, compared to traditional PWD management (in-feed antibiotics) and to a control group without any diet supplementation. The study involved 197 piglets from 18 litters in a single farm. The piglets were followed from birth to day 58 of age and were weaned at day 26. During the experimental period, the animals were monitored for weight and growth, average daily gain, morbidity and mortality. For the metataxonomics analysis, rectal samples were collected from 17 piglets from the three experimental groups at 4 different time-points (days 1, 12, 26 and 58). Results revealed that the gut microbiota in pre- and post-weaning piglets was dominated by the phyla Firmicutes (51%), Bacteroidetes (25%) and Proteobacteria (16%), which together make up for over 90% of the entire piglet core gut microbiota. The core microbiota comprised 10 taxa before weaning and 43 taxa after weaning, with 7 taxa overlapping between timepoints: two of them (Prevotella 9, p-value = 0.00095; Solobacterium p-value = 0.00821) were significantly more abundant after weaning. All alpha diversity indexes were significantly different between pre- and post-weaning, while only Shannon and Simpson diversity and equitability were significantly different between treatments. Based on the matrix of Bray-Curtis dissimilarities, samples showed clear clustering per timepoint (before and after weaning, p-value < 0.001) and between treatments by timepoint (p-value = 0.0086). The oil-diet group showed a consistently higher F:B ratio at all timepoints. These results show that the pig gut microbiota changes significantly with weaning, and suggest that the use of essential oils as feed supplementation to control PWD does not seem to alter sgnificantly the microbiota nor the growth parameters of piglets, however modifications of specific taxa may occur.