Diarrhea In Pigs Research Articles

Rapamycin-induced autophagy restricts porcine epidemic diarrhea virus infectivity in porcine intestinal epithelial cells

Porcine epidemic diarrhea virus (PEDV) invades porcine intestinal epithelial cells (IECs) and causes diarrhea and dehydration in pigs. In the present study, we showed a suppression of PEDV infection in porcine jejunum intestinal epithelial cells (IPEC-J2) by an increase in autophagy. Autophagy was activated by rapamycin at a dose that does not affect cell viability and tight junction permeability. The induction of autophagy was examined by LC3I/LC3II conversion. To confirm the autophagic-flux (entire autophagy pathway), autophagolysosomes were examined by an immunofluorescence assay. Pre-treatment with rapamycin significantly restricted not only a 1 h infection but also a longer infection (24 h) with PEDV, while this effect disappeared when autophagy was blocked. Co-localization of PEDV and autophagosomes suggests that PEDV could be a target of autophagy. Moreover, alleviation of PEDV-induced cell death in IPEC-J2 cells pretreated with rapamycin demonstrates a protective effect of rapamycin against PEDV-induced epithelial cell death. Collectively, the present study suggests an early prevention against PEDV infection in IPEC-J2 cells via autophagy that might be an effective strategy for the restriction of PEDV, and opens up the possibility of the use of rapamycin in vivo as an effective prophylactic and prevention treatment.

Endemic course of epidemic diarrhea of pigs in the stabilized focus of infection

Porcine epidemic diarrhea virus (PEDV) has been circulating in Ukraine since 2014 and induces an especially dangerous viral infection with a lethal diarrheal syndrome in newborn piglets, with the initial appearance at the focus of infection. The number of infected cases and lethality among diseased piglets of 1–5 days of age can reach 100%, which together with the forced anti-epizootic measures brings significant economic losses. PED can spread to all pigs, but the emergent quality of infectious pathology appears in newborn piglets. No effective and biologically safe means of specific antiviral prophylaxis, which substantially halts the epizootic process is registered, and etiopathogenetic therapy is not developed, therefore PED is an emergent infection which is difficult tocontrol. Over time there appear stationary foci of infection, where evolutionary changes in relationships in the host-parasite system take place fairly rapidly, since pigs are prolific and fast maturing animals able to replace each generation up to three times each year. This leads to a significant variability in interpopulation relationships and the induction of biodiversity in the molecular mechanisms of adaptation and processing of the viral genome. Clinically, genetic modifications of local variants of PEDV – populations are manifested in the form of changes in epizootic peculiarities in the course of infectious pathology in different age groups of animals. Modifications of PEDV may be accompanied by a slight weakening of the intensity of the infectious process, a decrease in mortality and a decrease in the severity of the pathogenesis of diarrheal syndrome. At the same time, the age range of severe abdominal lesions expands from newborn piglets to fattening animals of older age groups of 28, 32, 70 days. Using a set of measures to combat the PED, including “reverse feeding” recycled infected biomaterial from convalescent pigs, eradication of the pathogen from the environment of the host macroorganisms through a total disinfection regime and strict compliance with veterinary and sanitary rules of animal husbandry provide temporary positive results, but in theory this approach is incorrect, since contamination of animals leads to the dispersal of the virus and the formation of endemic foci of infection. The persistence of the virus in convalescent organisms is not fixed, the external inanimate environment can only be a mechanical factor in transmission of the pathogen preserving the viability of PEDV over time. Stabilization of the epizootic foci of infection is possible due to three factors: a) dissemination of the virus in “reverse feeding”; b) preservation of the virus in the external environment as a result of poor-quality disinfection; c) occurrence of a non-immune element among the convalescent young gilts, who as a result of juvenile insufficiency of the immune system have a low titer accumulation of colostral antibodies to the virus received in the biomaterial through reverse feeding. Due to the lack of “lactogenic immunity”, neonatal pigs as biological indicators for the presence of PEDV in the environment begin reproducing the virus in the enterocytes and develop a typical diarrheal syndrome PED.

Novel plasmid-mediated colistin resistance mcr-4 gene in Salmonella and Escherichia coli, Italy 2013, Spain and Belgium, 2015 to 2016.

A novel mcr colistin resistance gene was identified in a strain of Salmonella enterica, monophasic variant of serovar Typhimurium (4,5,12:i:- ), isolated from a pig at slaughter in Italy in 2013, and in Escherichia coli strains collected during routine diagnostic of post-weaning diarrhoea in pigs from Spain and Belgium in 2015 and 2016. Immediate implementation of mcr-screening including this novel gene variant is required for Salmonella and E. coli from humans and food-producing animals in Europe.

Efficacy of a single oral dose of a live bivalent E. coli vaccine against post-weaning diarrhea due to F4 and F18-positive enterotoxigenic E. coli

F4- and F18-positive enterotoxigenic E. coli strains (F4-ETEC and F18-ETEC) are important causes of post-weaning diarrhea (PWD) in pigs. F4 (antigenic variant ac) and F18 (ab and ac) fimbriae are major antigens that play an important role in the early stages of infection. Herein, the efficacy of a live oral vaccine consisting of two non-pathogenic E. coli strains, one F4ac- and one F18ac-positive, was evaluated using F4ac-ETEC and F18ab-ETEC challenge models. A randomized, masked, placebo-controlled, block design, parallel-group confirmatory study with two different vaccination-challenge intervals (7 and 21 days) was conducted for each challenge model. The vaccine was administered in one dose, to ≥18-day-old piglets via drinking water. Efficacy was assessed by evaluating diarrhea, clinical observations, weight gain and fecal shedding of F4-ETEC or F18-ETEC. Anti-F4 and anti-F18 immunoglobulins in blood were measured.The vaccination resulted in significant reductions in clinical PWD and fecal shedding of F4-ETEC and F18-ETEC after the 7- and 21-day-post-vaccination heterologous challenges, except for after the 21-day-post-vaccination F4-ETEC challenge, when the clinical PWD was too mild to demonstrate efficacy. A significant reduction of mortality and weight loss by vaccination were observed following the F18-ETEC challenge. The 7-day protection was associated with induction of anti-F4 and anti-F18 IgM, whereas the 21-day protection was mainly associated with anti-F4 and anti-F18 IgA. The 7-day onset and 21-day duration of protection induced by this vaccine administered once in drinking water to pigs of at least 18days of age were confirmed by protection against F4-ETEC and F18-ETEC, and induction of F4 and F18-specific immunity. Cross protection of the vaccine against F18ab-E. coli was demonstrated for both the 7- and 21-day F18-ETEC challenges.

Supplementation with Lactobacillus paracasei or Pediococcus pentosaceus does not prevent diarrhoea in neonatal pigs infected with Escherichia coli F18.

Infectious diarrhoea is a worldwide problem in newborns. Optimal bacterial colonisation may enhance gut maturation and protect against pathogenic bacteria after birth. We hypothesised that lactic acid bacteria (LAB) administration prevents pathogen-induced diarrhoea in formula-fed newborns. Newborn caesarean-delivered, colostrum-deprived term piglets on parenteral nutrition for the first 15 h, were used as models for sensitive newborn infants. A commercially available probiotic strain, Lactobacillus paracasei F19 (LAP, 2·6×108 colony-forming units (CFU)/kg per d) and a novel LAB isolate, Pediococcus pentosaceus (PEP, 1·3×1010 CFU/kg per d), were administered for 5 d with or without inoculation of the porcine pathogen, Escherichia coli F18 (F18, 1010 CFU/d). This resulted in six treatment groups: Controls (n 9), LAP (n 10), PEP (n 10), F18 (n 10), F18-LAP (n 10) and F18-PEP (n 10). The pathogen challenge increased diarrhoea and density of F18 in the intestinal mucosa (P<0·05). LAB supplementation further increased the diarrhoea score, relative to F18 alone (P<0·01). Intestinal structure and permeability were similar among groups, whereas brush border enzymes were affected in variable intestinal regions with decreased activities in most cases after F18 and LAB inoculation. Bacterial density in colon mucosa increased after F18 inoculation (P<0·05) but was unaffected by LAB supplementation. In colon contents, acetic and butyric acids were increased by PEP (P<0·05). The LAB used in this study failed to reduce E. coli-induced diarrhoea in sensitive newborn pigs. In vulnerable newborns there may be a delicate balance among bacterial composition and load, diet and the host. Caution may be required when administering LAB to compromised newborns suffering from enteric infections.

Zinc oxide enriched peat influence Escherichia coli infection related diarrhea, growth rates, serum and tissue zinc levels in Norwegian piglets around weaning: five case herd trials

BackgroundZinc oxide (ZnO), commonly used to control post-weaning diarrhea in piglets, has been highlighted as of potential concern from an environmental perspective. The aim of this field trial was to examine effects of different sources and levels of ZnO added to peat on average daily weight gain (ADG), fecal score in pens and serum and tissue zinc (Zn) levels around time of weaning in order to reduce the environmental impact without loss of the beneficial effect of ZnO on intestinal health and growth. Five case herds with enterotoxic colibacillosis challenges were included. The piglets entered the study aged three or five weeks. All piglets received a commercial diet containing <150 mg Zn/ per kg of complete feed. Four treatment groups received commercial peat added A: uncoated ZnO, B: lipid microencapsulated ZnO, C: solely commercial peat or D: no peat (Farms 2 and 3).ResultsAt Farms 1, 2 and 3, a significant effect of treatment was identified for fecal score (P < 0.05). Treatment A led to lower fecal scores compared to treatments C (P < 0.05) and D (P < 0.01). At Farms 2 and 3, there was a significant difference in individual average daily weight gain (iADG) between treatment A and D (P < 0.05). The iADG of piglets receiving treatment B did not differ significantly from treatment A.ConclusionsIn 2016, The European Medicines Agency’s Committee on Veterinary Medicinal Products concluded that the benefits of ZnO for the prevention of diarrhea in pigs do not outweigh the risks to the environment. Effective alternative measures to reduce the accumulation of Zn in the environment have not been identified. Our results imply that peat added low concentration of both coated and uncoated ZnO influences the gut health of weaned piglets reflected by enhanced weight gain and reduced occurrence of diarrhea. This preventive approach certainly represents a favourable alternative in the “One Health” perspective. It will also contribute to reduced antibiotic use in pig farming while diminishing the environmental consequences caused by ZnO.

A Naturally Occurring Recombinant Enterovirus Expresses a Torovirus Deubiquitinase.

Enteroviruses (EVs) are implicated in a wide range of diseases in humans and animals. In this study, a novel enterovirus (enterovirus species G [EVG]) (EVG 08/NC_USA/2015) was isolated from a diagnostic sample from a neonatal pig diarrhea case and identified by using metagenomics and complete genome sequencing. The viral genome shares 75.4% nucleotide identity with a prototypic EVG strain (PEV9 UKG/410/73). Remarkably, a 582-nucleotide insertion, flanked by 3Cpro cleavage sites at the 5' and 3' ends, was found in the 2C/3A junction region of the viral genome. This insertion encodes a predicted protease with 54 to 68% amino acid identity to torovirus (ToV) papain-like protease (PLP) (ToV-PLP). Structural homology modeling predicts that this protease adopts a fold and a catalytic site characteristic of minimal PLP catalytic domains. This structure is similar to those of core catalytic domains of the foot-and-mouth disease virus leader protease and coronavirus PLPs, which act as deubiquitinating and deISGylating (interferon [IFN]-stimulated gene 15 [ISG15]-removing) enzymes on host cell substrates. Importantly, the recombinant ToV-PLP protein derived from this novel enterovirus also showed strong deubiquitination and deISGylation activities and demonstrated the ability to suppress IFN-β expression. Using reverse genetics, we generated a ToV-PLP knockout recombinant virus. Compared to the wild-type virus, the ToV-PLP knockout mutant virus showed impaired growth and induced higher expression levels of innate immune genes in infected cells. These results suggest that ToV-PLP functions as an innate immune antagonist; enterovirus G may therefore gain fitness through the acquisition of ToV-PLP from a recombination event.IMPORTANCE Enteroviruses comprise a highly diversified group of viruses. Genetic recombination has been considered a driving force for viral evolution; however, recombination between viruses from two different orders is a rare event. In this study, we identified a special case of cross-order recombination between enterovirus G (order Picornavirales) and torovirus (order Nidovirales). This naturally occurring recombination event may have broad implications for other picornaviral and/or nidoviral species. Importantly, we demonstrated that the exogenous ToV-PLP gene that was inserted into the EVG genome encodes a deubiquitinase/deISGylase and potentially suppresses host cellular innate immune responses. Our results provide insights into how a gain of function through genetic recombination, in particular cross-order recombination, may improve the ability of a virus to evade host immunity.

Effect of Tetracycline Dose and Treatment Mode on Selection of Resistant Coliform Bacteria in Nursery Pigs.

This study describes the results of a randomized clinical trial investigating the effect of oxytetracycline treatment dose and mode of administration on the selection of antibiotic-resistant coliform bacteria in fecal samples from nursery pigs. Nursery pigs (pigs of 4 to 7 weeks of age) in five pig herds were treated with oxytetracycline for Lawsonia intracellularis-induced diarrhea. Each group was randomly allocated to one of five treatment groups: oral flock treatment with a (i) high (20 mg/kg of body weight), (ii) medium (10 mg/kg), or (iii) low (5 mg/kg) dose, (iv) oral pen-wise (small-group) treatment (10 mg/kg), and (v) individual intramuscular injection treatment (10 mg/kg). All groups were treated once a day for 5 days. In all groups, treatment caused a rise in the numbers and proportions of tetracycline-resistant coliform bacteria right after treatment, followed by a significant drop by the time that the pigs left the nursery unit. The counts and proportions of tetracycline-resistant coliforms did not vary significantly between treatment groups, except immediately after treatment, when the highest treatment dose resulted in the highest number of resistant coliforms. A control group treated with tiamulin did not show significant changes in the numbers or proportions of tetracycline-resistant coliforms. Selection for tetracycline-resistant coliforms was significantly correlated to selection for ampicillin- and sulfonamide-resistant strains but not to selection for cefotaxime-resistant strains. In conclusion, the difference in the dose of oxytetracycline and the way in which the drug was applied did not cause significantly different levels of selection of tetracycline-resistant coliform bacteria under the conditions tested.IMPORTANCE Antimicrobial resistance is a global threat to human health. Treatment of livestock with antimicrobials has a direct impact on this problem, and there is a need to improve the ways that we use antimicrobials in livestock production. We hypothesized that antibiotic resistance development following treatment of diarrhea in nursery pigs could be reduced either by lowering the dose of oxytetracycline or by replacing the commonly used practice of flock treatment with individual or small-group treatments, since this would reduce the number of pigs treated. However, the study showed no significant difference between treatment groups with respect to the number or proportion of tetracycline-resistant coliforms selected. The most important conclusion is that under practical field conditions, there will be no added value, in terms of lowering resistance development, by exchanging flock treatment for individual or small-group treatment of nursery pigs. The reason for the lack of an effect of single-animal treatment is probably that such animals share the environment with treated animals and take up resistant bacteria from the environment.

Infection of porcine colon explants with "Brachyspira hampsonii" leads to increased epithelial necrosis and catarrhal exudate.

Mucohemorrhagic diarrhea in pigs caused by Brachyspira spp. has a global distribution, and an economic impact on affected farms due to poor performance of animals. Demonstrations that "Brachyspira hampsonii" is pathogenic have been achieved using in vivo animal models, but a critical knowledge gap exists regarding the pathogenic mechanisms employed by Brachyspira. Here, we used in vitro organ culture of porcine colon to investigate interactions between "B. hampsonii" and explants during the first 12 h of contact. Explants were either inoculated with "B. hampsonii" or sterile culture broth. Responses to infection were evaluated by optical microscopy and quantitative PCR. Significantly greater numbers of necrotic crypt cells and thicker catarrhal exudate were observed on infected explants compared to controls. Spirochaetes were observed in the mucus layer, in contact with necrotic exfoliated cells, in crypts and the lamina propria. Statistical differences were observed in mRNA levels between inoculated and control explants for IL-1α, TNF-α and ZO-1 using a Bayesian analysis, but not observed using the ΔΔCq method. These results provide a demonstration of a porcine colon explant model for investigating interactions of Brachyspira with its host and show that initial effects on the host are observed within the first 12 h of contact.

A multivariate dynamic linear model for early warnings of diarrhea and pen fouling in slaughter pigs

We present a method for providing early, but indiscriminant, predictions of diarrhea and pen fouling in grower/finisher pigs. We collected data on dispensed feed amount, water flow, drinking bouts frequency, temperature at two positions per pen, and section level humidity from 12 pens (6 double pens) over three full growth periods. The separate data series were co-modeled at pen level with time steps of one hour, using a multivariate dynamic linear model. The step-wise forecast errors of the model were unified using Cholesky decomposition. An alarm was raised if the unified error exceeded a set threshold a sufficient number of times, consecutively. Using this method with a 7day prediction window, we achieved an area under the receiver operating characteristics curve of 0.84. Shorter prediction windows yielded lower performances, but longer prediction windows did not affect the performance.

α1,2-Fucosyllactose Does Not Improve Intestinal Function or Prevent Escherichia coli F18 Diarrhea in Newborn Pigs.

Infectious diarrhea, a leading cause of morbidity and deaths, is less prevalent in breastfed infants compared with infants fed infant formula. The dominant human milk oligosaccharide (HMO), α-1,2-fucosyllactose (2'-FL), has structural homology to bacterial adhesion sites in the intestine and may in part explain the protective effects of human milk. We hypothesized that 2'-FL prevents diarrhea via competitive inhibition of pathogen adhesion in a pig model for sensitive newborn infants. Intestinal cell studies were coupled with studies on cesarean-delivered newborn pigs (n = 24) without (control) or with inoculation of enterotoxigenic Escherichia coli F18 (7.5 × 10/day for 8 days) fed either no (F18) or 10 g/L 2'-FL (2FL-F18). In vitro studies revealed decreased pathogen adhesion to intestinal epithelial cells with 2'-FL (5 g/L; P < 0.001). F18 pigs showed more diarrhea than control pigs (P < 0.01). Administration of 2'-FL to F18 pigs failed to prevent diarrhea, although the relative weight loss tended to be reduced (-19 vs -124 g/kg, P = 0.12), higher villi were observed in the distal small intestine (P < 0.05), and a trend toward increased proportion of mucosa and activities of some brush border enzymes in the proximal small intestine. In situ abundance of α-1,2-fucose and E coli was similar between groups, whereas sequencing showed higher abundance of Enterobacteriaceae in F18, Enterococcus in control and Lachnospiraceae in 2FL-F18 pigs. 2'-FL inhibited in vitro adhesion of E coli F18 to epithelial cells, but had limited effects on diarrhea and mucosal health in newborn pigs challenged with E coli F18.

Porcine kobuvirus 1 in healthy and diarrheic pigs: Genetic detection and characterization of virus and co-infection with rotavirus A

The porcine kobuvirus 1 (PKV-1) is believed to be an enteric virus. To investigate the prevalence of PKV-1 in pigs, virus was detected by RT-PCR in rectal swabs originating from 414 healthy and diarrheic pigs of different age categories on farms in Slovakia. Among all ages of animals, PKV-1 was detected equally in diarrheic (63.8%) and clinically healthy (62.9%) pigs. PKV-1 was more often detected in diarrheic (74.6%) than in healthy (64.4%) suckling piglets (<28days) but data was not statistically significant. Results in weaned (28–70days) and fattening (>70days) of both healthy and diarrheic pigs were inconsistent ranging in interval 56.2% to 67.9%. This study did not confirm a clear relationship of PKV-1 infection with diarrhea in pigs. Rotavirus A infection was detected among the same animals in 39% diarrheic and 9.2% healthy suckling piglets (p<0.001) confirming rotavirus as a causative agent of diarrhea in this age group. The difference was not significant in older pigs with both diarrheic and healthy pigs being infected within a range of 0% to 12.2%. Co-infection with PKV-1 and rotavirus A was detected overall in 5.6% of healthy and in 13.5% of diarrheic pigs and was highest in suckling piglets (33.9%). The PKV-1sequences from pigs in Slovakia were analyzed at the genetic level in the partial 3D gene region for the first time. The viral sequences were grouped in phylogenetic clusters according to their farm of origin. When compared with 157 nucleotide sequences originating from pig samples of different countries around the world Slovakian PKV-1 sequences were clustered in the phylogenetic tree with Asian sequences but not with nucleotide sequences from the neighbouring countries of Czech Republic or Hungary.

The fecal presence of enterotoxin and F4 genes as an indicator of efficacy of treatment with colistin sulfate in pigs

BackgroundEnterotoxigenic Escherichia coli (ETEC) strains producing multiple enterotoxins are important causes of post-weaning diarrhea (PWD) in pigs. The aim of the present study was to investigate the fecal presence of ETEC enterotoxin as well as F4 and F18 genes as an indicator of colistin sulfate (CS) efficacy for treatment of PWD in pigs. Forty-eight piglets were weaned at the age of 21 days, and were divided into four groups: challenged treated, challenged untreated, unchallenged treated, and unchallenged untreated. Challenge was performed using 109 CFU of an ETEC: F4 strain, and treatment was conducted using oral CS at the dose of 50,000 IU/kg. The fecal presence of genes encoding for STa, STb, LT, F4 and F18 was detected using PCR.ResultsThe PCR amplification of ETEC virulence genes showed that nearly 100% of pigs excreted genes encoding for STa and STb toxins in the feces before the challenge. These genes, in the absence of the gene encoding F4, were considered as a marker for F4-negative ETEC. One day after ETEC: F4 oral challenge pigs in the two challenged groups excreted the genes encoding LT and F4 in the feces. These genes were considered as a marker for F4-positive ETEC. However, the gene encoding F18 was not detected in any fecal samples of the 4 groups throughout the experiment. After only 3 days of successive oral treatment with CS, a significant reduction in both the F4-positive and negative ETEC populations was observed in the challenged treated group compared to the challenged untreated group (p < 0.0001).ConclusionsOur study is among the first to report that under controlled farming conditions, oral CS treatment had a significant effect on both fecal F4-positive and F4-negative ETEC in pigs. However, CS clinical efficiency was correlated with non-detection of F4-positive ETEC in the feces. Furthermore the fecal presence of F4-negative ETEC was not associated with clinical symptoms of post-weaning diarrhea in pigs.

A comparative study of the efficacy of Detach® versus zinc oxide to control post-weaning diarrhoea in pigs

Animal Production Science (continuing Australian Journal of Experimental Agriculture) publishes original research into applied agriculture including animal production, animal-plant interactions, pasture and fodder crops, field crops, extension and horticulture

Antimicrobial Activity, Cytotoxicity and Phytochemical Analysis of Sanguinaria canadensis Native to South Dakota

A methanolic extract of Sanguinaria canadensis rhizome contains a mixture of benzophenanthridine alkaloids with the major component sanguinarine. American Indians used this rhizome in the treatment of rheumatism, asthma, bronchitis, lung ailments, fever, and warts. Lethal diarrhea in neonatal and post-weaned pigs due to enterotoxigenic Escherchia coli (ETEC) is a major problem among the swine producers of South Dakota region. The increasing resistance of ETEC towards synthetic antibiotics is a matter of concern. Our aims in this study were I) to determine the minimum inhibitory concentration (MIC) of methanol extract of S. canadensis against ETEC II) to evaluate the cytotoxicity activity against porcine intestinal epithelial cell lines and III) to quantify the amount of sanguinarine in the rhizome of S. canadensis native to South Dakota. The MIC value was found to be 6.25 mg/mL against ETEC whereas the 100% inhibition of IPEC-J2 cells was found to be at 0.312 mg/mL. The total amount of alkaloids in this plant was found to be 1247 mg in 100 gm of fresh plant material whereas the amount of sangunarine was 559.64 mg in 100 gm of fresh plant material. The percentage of sangunarine in fresh plant material was 0.55% and in total alkaloid is 44.8%. Our results showed that S. canadensis has antibacterial effect against E. coli but it is toxic to IPECJ2 cells in vitro. It is important to carry out an in vivo animal study to confirm its further use as feed supplements.

Genomic investigation of piglet resilience following porcine epidemic diarrhea outbreaks

SummaryPorcine epidemic diarrhea virus (PEDV) belongs to the Coronaviridae family and causes malabsorptive watery diarrhea, vomiting, dehydration and imbalanced blood electrolytes in pigs. Since the 1970s, PED outbreaks have become a source of problems in pig producing countries all over the world, causing large economic losses for pig producers. Although the infection in adults is not fatal, in naïve suckling piglets mortality is close to 100%. In this study, we investigated genome‐wide differences between dead and recovered suckling piglets from commercial farms after PED outbreaks. Samples from 262 animals (156 dead and 106 recovered) belonging to several commercial lines were collected from five different farms in three different countries (USA, Canada and Germany) and genotyped with the porcine 80K SNP chip. Mean F st value was calculated in 1‐Mb non‐overlapping windows between dead and recovered individuals, and the results were normalized to find differences within the comparison. Seven windows with high divergence between dead and recovered were detected—five on chromosome 2, one on chromosome 4 and one on chromosome 15—in total encompassing 152 genes. Several of these genes are either under‐ or overexpressed in many virus infections, including Coronaviridae (such as SARS‐CoV). A total of 32 genes are included in one or more Gene Ontology terms that can be related to PED development, such as Golgi apparatus, as well as mechanisms generally linked to resilience or diarrhea development (cell proliferation, ion transport, ATPase activity). Taken together this information provides a first genomic picture of PEDV resilience in suckling piglets.

Genetic diversity of coronaviruses in bats in Lao PDR and Cambodia

South-East Asia is a hot spot for emerging zoonotic diseases, and bats have been recognized as hosts for a large number of zoonotic viruses such as Severe Acute Respiratory Syndrome (SARS), responsible for acute respiratory syndrome outbreaks. Thus, it is important to expand our knowledge of the presence of viruses in bats which could represent a risk to humans. Coronaviruses (CoVs) have been reported in bat species from Thailand, China, Indonesia, Taiwan and the Philippines. However no such work was conducted in Cambodia or Lao PDR. Between 2010 and 2013, 1965 bats were therefore sampled at interfaces with human populations in these two countries. They were tested for the presence of coronavirus by consensus reverse transcription-PCR assay. A total of 93 samples (4.7%) from 17 genera of bats tested positive. Sequence analysis revealed the presence of potentially 37 and 56 coronavirus belonging to alpha-coronavirus (αCoV) and beta-CoV (βCoV), respectively. The βCoVs group is known to include some coronaviruses highly pathogenic to human, such as SARS-CoV and MERS-CoV. All coronavirus sequences generated from frugivorous bats (family Pteropodidae) (n=55) clustered with other bat βCoVs of lineage D, whereas one coronavirus from Pipistrellus coromandra fell in the lineage C of βCoVs which also includes the MERS-CoV. αCoVs were all detected in various genera of insectivorous bats and clustered with diverse bat αCoV sequences previously published. A closely related strain of PEDV, responsible for severe diarrhea in pigs (PEDV-CoV), was detected in 2 Myotis bats. We highlighted the presence and the high diversity of coronaviruses circulating in bats from Cambodia and Lao PDR. Three new bat genera and species were newly identified as host of coronaviruses, namely Macroglossus sp., Megaerops niphanae and Myotis horsfieldii

Immunogenicity and protective efficacy of a single-dose live non-pathogenic Escherichia coli oral vaccine against F4-positive enterotoxigenic Escherichia coli challenge in pigs

Enterotoxigenic Escherichia coli strains expressing F4 (K88) fimbriae (F4-ETEC) are one of the most important causes of post-weaning diarrhea (PWD) in pigs. F4, a major antigen, plays an important role in the early steps of the infection. Herein, the efficacy of a live oral vaccine consisting of a non-pathogenic E. coli strain expressing F4 for protection of pigs against PWD was evaluated. Three blinded, placebo-controlled, block design, parallel-group confirmatory experiments were conducted, using an F4-ETEC PWD challenge model, each with a different vaccination-challenge interval (3, 7, and 21days). The pigs were vaccinated via the drinking water with a single dose of the Coliprotec® F4 vaccine one day post-weaning. Efficacy was assessed by evaluating diarrhea, clinical observations, intestinal fluid accumulation, weight gain, intestinal colonization and fecal shedding of F4-ETEC. The immune response was evaluated by measuring serum and intestinal F4-specific antibodies. The administration of the vaccine resulted in a significant reduction of the incidence of moderate to severe diarrhea, ileal colonization by F4-ETEC, and fecal shedding of F4-ETEC after the heterologous challenge at 7 and 21days post-vaccination. The 7-day onset of protection was associated with an increase of serum anti-F4 IgM whereas the 21-day duration of protection was associated with an increase of both serum anti-F4 IgM and IgA. Significant correlations between levels of serum and intestinal secretory anti-F4 antibodies were detected. Maternally derived F4-specific serum antibodies did not interfere with the vaccine efficacy. The evaluation of protection following a challenge three days after vaccination showed a reduction of the severity and the duration of diarrhea and of fecal shedding of F4-ETEC. The 7-day onset and the 21-day duration of protection induced by Coliprotec® F4 vaccine administered once in drinking water to pigs of at least 18days of age were confirmed by protection against F4-ETEC and induction of F4-specific protective immunity.

High Protective Efficacy of Probiotics and Rice Bran against Human Norovirus Infection and Diarrhea in Gnotobiotic Pigs.

Probiotics have been recognized as vaccine adjuvants and therapeutic agents to treat acute gastroenteritis in children. We previously showed that rice bran (RB) reduced human rotavirus diarrhea in gnotobiotic pigs. Human noroviruses (HuNoVs) are the major pathogens causing non-bacterial acute gastroenteritis worldwide. In this study, Lactobacillus rhamnosus GG (LGG) and Escherichia coli Nissle 1917 (EcN) were first screened for their ability to bind HuNoV P particles and virions derived from clinical samples containing HuNoV genotype GII.3 and GII.4, then the effects of LGG+EcN and RB on HuNoV infection and diarrhea were investigated using the gnotobiotic pig model. While LGG+EcN colonization inhibited HuNoV shedding, probiotic cocktail regimens in which RB feeding started 7 days prior to or 1 day after viral inoculation in the LGG+EcN colonized gnotobiotic pigs exhibited high protection against HuNoV diarrhea and shedding, characterized by significantly reduced incidence (89 versus 20%) and shorter mean duration of diarrhea (2.2 versus 0.2 days), as well as shorter mean duration of virus shedding (3.2 versus 1.0 days). In both probiotic cocktail groups, the diarrhea reduction rates were 78% compared with the control group, and diarrhea severity was reduced as demonstrated by the significantly lower cumulative fecal scores. The high protective efficacy of the probiotic cocktail regimens was attributed to stimulation of IFN-γ+ T cell responses, increased production of intestinal IgA and IgG, and maintenance of healthy intestinal morphology (manifested as longer villi compared with the control group). Therefore, probiotic cocktail regimens containing LGG+EcN and RB may represent highly efficacious strategies to prevent and treat HuNoV gastroenteritis, and potentially other human enteric pathogens.

0173 Comparison of strategies for combining dynamic linear models with artificial neural networks for detecting diarrhea in slaughter pigs

0173 Comparison of strategies for combining dynamic linear models with artificial neural networks for detecting diarrhea in slaughter pigs