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Differential expression of intestinal ion transporters and water channel aquaporins in young piglets challenged with enterotoxigenic Escherichia coli K88.

ABSTRACT: The study was to determine whether the expression of genes involved in intestinal water and ion transport would be affected by enterotoxigenic (ETEC) K88 both in vitro and in vivo. First, 36 male piglets (4 d old) were randomly allotted to either the control or the ETEC K88 group. Each group had 6 replicates with 3 piglets per replicate. All piglets were fed with the same diets for 17 d. On d 15, piglets in the ETEC K88 group were challenged with ETEC K88 (serotype O149:K91:K88ac) at 1 × 10 cfu per pig, whereas those in the control group received the same volume of sterile PBS. After being challenged with ETEC K88 for 72 h (d 18), 1 piglet from each replicate was selected for slaughter to collect samples from the jejunum, ileum, and colon. The mRNA expression and protein abundance of cystic fibrosis transmembrane conductance regulator (CFTR) in the ileum and colon were increased compared with that in the control group ( < 0.05). Furthermore, the mRNA expression of () in the ileum and colon was increased by ETEC K88 challenge ( < 0.05), whereas in the jejunum, both its mRNA and protein expression were increased by ETEC K88 treatment ( < 0.05). Additionally, an established porcine intestinal epithelial cell line (IPEC-J2) was used to investigate the effect and possible mechanism of ETEC K88 on expression of water channel aquaporins (AQP) and ion transporters. Cells (1.17 × 10 per well) were grown in 6-well plates and treated with ETEC K88 at a multiplicity of infection of 50:1 for 3 h. The mRNA expression of , , and () in IPEC-J2 cells was reduced after ETEC K88 treatment ( < 0.05). Further analyses using western blotting also demonstrated that ETEC K88 decreased the protein expression of AQP3, AQP9, and AQP11 in IPEC-J2 cells ( < 0.05). Moreover, the phosphorylation levels of protein kinase A (PKA) and cyclic adenosine monophosphate (cAMP)-response element binding protein (CREB) were decreased by ETEC K88 challenge ( < 0.05). The results indicate that ETEC K88 challenge induced differential expression of intestinal ion transporters and AQP in young piglets, probably by regulation of the cAMP-PKA signaling pathway. This study might provide new insights about the importance of fluid homeostasis in control of ETEC-induced diarrhea in young piglets.

SUBMITTER: Zhu C

PROVIDER: S-EPMC6292308 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Differential expression of intestinal ion transporters and water channel aquaporins in young piglets challenged with enterotoxigenic <i>Escherichia coli</i> K88.

Zhu C C Ye J L JL Yang J J Yang K M KM Chen Z Z Liang R R Wu X J XJ Wang L L Jiang Z Y ZY

Journal of animal science 20171201 12

The study was to determine whether the expression of genes involved in intestinal water and ion transport would be affected by enterotoxigenic (ETEC) K88 both in vitro and in vivo. First, 36 male piglets (4 d old) were randomly allotted to either the control or the ETEC K88 group. Each group had 6 replicates with 3 piglets per replicate. All piglets were fed with the same diets for 17 d. On d 15, piglets in the ETEC K88 group were challenged with ETEC K88 (serotype O149:K91:K88ac) at 1 × 10 cfu ...[more]

PMID: 29293799

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Project description:A high-quality protein substitute, Hermetia illucens (black soldier fly) larvae powder, is rich in protein and often used in animal feed. This study aimed to investigate the feasibility and optimal ratio of replacing fish meal with H. illucens larvae in weaned piglets and to demonstrate the effects on piglets' growth performance, intestinal microflora and immune performance. Forty-eight female weaned piglets were randomly classified into three groups. Each group consisted of eight pens (replicates), with two piglets per pen. Three groups containing different proportions of H. illucens larvae (0, 4, and 8%) were referred to as C, HI4, and HI8. We first designed a 28-day feeding experiment to detect growth performance; after that, the piglets were induced with oral gavage of enterotoxigenic Escherichia coli K88 (ETEC K88) and recording diarrhea on day 29 of the experiment. Samples were taken on the 32nd day to detect the effect of H. illucens larvae on the immune performance of the weaned piglets. H. illucens larvae replacement did not cause any obvious change in the growth performance nether in HI4 nor in HI8 of weaned piglets with 28 d feeding stage. H. illucens larvae could improve the intestinal health of weaned piglets by increasing the content of Lactobacillus and reducing the content of Streptococcus. Compared with C+K88 group, the diarrhea rate was attenuated for the H. illucens supplemented group. The integrity of ileum villi in HI4+K88 and HI8+K88 groups was better than that in C+K88 group, and the villi in C+K88 group were severely damaged. The expression of IL-10, Occludin and Claudin-3 in the intestinal mucosa of the HI4+K88 group and HI8+K88 group were significantly increased (P < 0.05), and the expression of TNF-α was significantly decreased (P < 0.05) compared with the C+K88 group. The results of immunoblotting also validated that the same ETEC K88 treatment of weaned piglets enhanced the expression of tight junction protein in the intestinal mucosa of the H. illucens addition group. ETEC-induced diarrhea will be reduced by the diet of weaned piglets containing H. illucens larvae, ameliorating the immune performance of piglets. Our results indicates that the optimal dosage of H. illucens replacement in weaned piglets is 4%.

Project description:Antibiotics are very effective for treating diarrhea in weaned pigs, but the global prohibition of antibiotics makes it urgent to find an alternative to antibiotics. An experiment was conducted to determine the antimicrobial activity of a linear trpzip-like β-hairpin antimicrobial peptide WK3 in vivo and to assess its effects on growth performance and intestinal health. Thirty-two piglets were weaned at 21 days and housed in individual metabolic cages, which were randomly divided into four groups and were maintained on a corn-soybean meal-based basal diet. Group 1 included a blank group. Groups 2, 3, and 4 were orally infected by feeding with Enterotoxigenic Escherichia coli (ETEC) K88, which was followed by saline treatment (group 2), enrofloxacin injection at a dose of 2.5 mg/kg (group 3), and WK3 injection at a dose of 2 mg/kg (group 4). The experiment lasted for 6 days, and feed and water were provided ad libitum. Both WK3 and enrofloxacin effectively attenuated diarrhea and improved growth performance of piglets. Compared with the control group, WK3 significantly improved the villus height in the ileum (P < 0.05) but did not affect the villus height in the duodenum or jejunum. Additionally, we did not observe any obvious difference in crypt depth or villus height/crypt depth among the duodenum, jejunum and ileum (P > 0.05). WK3 also reduced the numbers of Enterococcus spp (P < 0.01) in the cecal contents, and the number of Enterobacterium spp tended to decrease (0.05 < P < 0.1). Moreover, the jejunal mucosa of the WK3 group exhibited lower interleukin-1α (IL-1a; P < 0.01), toll-like receptors-4 (TLR-4; P < 0.05), and myeloid differentiation primary response 88 (MyD88; P < 0.01) messenger ribonucleic acid (mRNA) expression levels. The jejunum of the WK3 group also exhibited an increased antioxidant capacity, reduced concentration of malondialdehyde (MDA; P < 0.05), and enhanced superoxide dismutase (SOD) activity (P < 0.05). WK3 has the potential to replace antibiotics as a new generation feed additive.

Project description:BackgroundThis study aimed to investigate the effects of oral administration of Enterococcus faecium NCIMB 10415 (E. faecium) on intestinal development, immunological parameters and gut microbiota of neonatal piglets challenged with enterotoxigenic Escherichia coli K88 (ETEC). A total of 96 1-day-old sow-reared piglets were randomly assigned to 2 groups, with 48 piglets in each group. The piglets were from 16 litters (6 piglets each litter), and 3 piglets each litter were allocated to the E. faecium-supplemented (PRO) group, while the other 3 piglets were allocated to the control (CON) group. After colostrum intake, piglets in the PRO group were orally administrated with 3 × 109 CFU E. faecium per day for a period of one week. On day 8, one piglet per litter from each group was challenged (CON+ETEC, PRO+ETEC) or not (CON-ETEC, PRO-ETEC) with ETEC in a 2 × 2 factorial arrangement of treatments. On day 10 (2 days after challenge), blood and tissue samples were obtained from piglets.ResultsBefore ETEC challenge, there were no significant differences for the average daily gain (ADG) and fecal score between the two groups of piglets. After ETEC challenge, the challenged piglets had greater fecal score compared to the non-challenged piglets, whereas E. faecium administration was able to decrease the fecal score. Piglets challenged with ETEC had shorter villous height, deeper crypt depth, and reduced number of goblet cells in the jejunum and decreased mRNA abundance of claudin-1 in the ileum, whereas increased the percentage of lymphocytes, concentrations of IL-1β in the plasma and TNF-α in the ileal mucosa, as well as increased the mRNA abundances of innate immunity-related genes in the ileum tissue. These deleterious effects caused by ETEC were partly alleviated by feeding E. faecium. In addition, piglets in PRO-ETEC group had decreased the percentage of CD8+ T cells of the peripheral blood when compared to those in CON-ETEC group. Moreover, E. faecium administration increased Verrucomicrobia at phylum level and decreased Bilophila at genus level.ConclusionsThese results suggest that oral administration of E. faecium alleviated the intestinal injury and diarrhea severity of neonatal piglets challenged by ETEC, partly through improving the intestinal microbiota and immune response. This offers a potential strategy of dietary intervention against intestinal impairment by ETEC in neonatal piglets.

Project description:This study was conducted to investigate the effects of early supplementation during 4 to 18 d of age with Lactobacillus plantarum (LP) in liquid diets on intestinal innate immune response in young piglets infected with enterotoxigenic Escherichia coli (ETEC) K88. Seventy-two barrow piglets at 4 d old were assigned to basal or LP-supplemented liquid diet (5 × 1010 CFU·kg-1). On day 15, piglets from each group were orally challenged with either ETEC K88 (1 × 108 CFU·kg-1) or the same amount of phosphate-buffered saline. The intestinal mucosa, mesenteric lymph node (MLN), and spleen samples were collected on day 18. Here, we found that LP pretreatment significantly decreased the mRNA relative expression of inflammatory cytokines (interleukin [IL]-1β, IL-8, and tumor necrosis factor-α), porcine β-defensin 2 (pBD-2), and mucins (MUC1 and MUC4) in the jejunal mucosa in piglets challenged with ETEC K88 (P < 0.05). Moreover, LP significantly decreased the ileal mucosa mRNA relative expression of IL-8 and MUC4 in young piglets challenged with ETEC K88 (P < 0.05). Furthermore, the piglets of the LP + ETEC K88 group had lower protein levels of IL-8, secretory immunoglobulin A, pBD-2, and MUC4 in the jejunal mucosa than those challenged with ETEC K88 (P < 0.05). Besides, LP supplementation reduced the percentage of gamma/delta T cells receptor (γδTCR) and CD172a+ (SWC3+) cells in MLN and the percentage of γδTCR cells in the spleen of young piglets after the ETEC K88 challenge. Supplementation with LP in liquid diets prevented the upregulated protein abundance of toll-like receptor (TLR) 4, phosphorylation-p38, and phosphorylation-extracellular signal-regulated protein kinases in the jejunal mucosa induced by ETEC K88 (P < 0.05). In conclusion, LP supplementation in liquid diet possesses anti-inflammatory activity and modulates the intestinal innate immunity during the early life of young piglets challenged with ETEC K88, which might be attributed to the suppression of TLR4-mediated mitogen-activated protein kinase signaling pathways. Early supplementation with LP in liquid diets regulates the innate immune response, representing a promising immunoregulation strategy for maintaining intestinal health in weaned piglets.

Project description:BackgroundEnterotoxigenic Escherichia coli K88 (E. coli K88) are considered as a major cause of diarrhea and death in newly weaned piglets. Oral passive immunization with chicken egg yolk immunoglobulins (IgY) have attracted considerable attention for treatment of gastrointestinal infection due to its high specificity. In this study it was estimated the protective effect of anti-K88 fimbriae IgY against E. coli K88 adhesion to piglet intestinal mucus in vitro and to investigate the potential use of IgY for controlling E. coli-induced diarrhea in weaned piglets in vivo.ResultsE. coli K88 was incubated with IgY for 24 h, and the bacterial growth profiles showed that specific IgY with a concentration higher than 5 mg/mL was observed to significantly inhibit the growth of E. coli K88 compared to nonspecific yolk powder in a liquid medium. Moreover, pretreatment with 50 mg/mL of IgY was found to significantly decrease the adhesion ability of E. coli K88 to porcine jejunal and ileal mucus, further supported by the observations from our immunofluorescence microscopic analysis. In vivo, administration of IgY successfully protected piglets from diarrhea caused by E. coli K88 challenge. Additionally, IgY treatment efficiently alleviated E. coli-induced intestinal inflammation in piglets as the gene expression levels of inflammatory cytokines TNF-α, IL-22, IL-6 and IL-1β in IgY-treated piglets remained unchanged after E. coli K88 infection. Furthermore, IgY significantly prevented E. coli K88 adhering to the jejunal and ileal mucosa of piglets with E. coli infection and significantly decreased E. coli and enterotoxin expression in colonic contents.ConclusionOutcome of the study demonstrated that IgY against the fimbrial antigen K88 was able to significantly inhibit the growth of E. coli K88, block the binding of E. coli to small intestinal mucus, and protect piglets from E. coli-induced diarrhea. These results indicate that passive immunization with IgY may be useful to prevent bacterial colonization and to control enteric diseases due to E. coli infection. The study has great clinical implication to provide alternative therapy to antibiotics in E coli induced diarrhea.

Project description:Glucose oxidase (GOD) could benefit intestinal health and growth performance in animals. However, it is unknown whether GOD can protect piglets against bacterial challenge. This study aimed to evaluate the protective effects of GOD on growth performance, clinical symptoms, serum parameters, and intestinal health in piglets challenged by enterotoxigenic Escherichia coli (ETEC). A total of 44 male weaned piglets around 38 days old were divided into four groups (11 replicates/group): negative control (NC), positive control (PC), CS group (PC piglets +40 g/t colistin sulfate), and GOD group (PC piglets +200 g/t GOD). All piglets except those in NC were challenged with ETEC (E. coli K88) on the 11th day of the experiment. Parameter analysis was performed on the 21st day of the experiment. The results showed that the ETEC challenge elevated (p < 0.05) the rectal temperature and fecal score of piglets at certain time-points post-challenge, reduced (p < 0.05) serum glucose and IgG levels but increased (p < 0.05) serum alanine aminotransferase activity, as well as caused (p < 0.05) intestinal morphology impairment and inflammation. Supplemental GOD could replace CS to reverse (p < 0.05) the above changes and tended to increase (p = 0.099) average daily gain during the ETEC challenge. Besides, GOD addition reversed ETEC-induced losses (p < 0.05) in several beneficial bacteria (e.g., Lactobacillus salivarius) along with increases (p < 0.05) in certain harmful bacteria (e.g., Enterobacteriaceae and Escherichia/Shigella). Functional prediction of gut microbiota revealed that ETEC-induced upregulations (p < 0.05) of certain pathogenicity-related pathways (e.g., bacterial invasion of epithelial cells and shigellosis) were blocked by GOD addition, which also normalized the observed downregulations (p < 0.05) of bacterial pathways related to the metabolism of sugars, functional amino acids, nucleobases, and bile acids in challenged piglets. Collectively, GOD could be used as a potential antibiotic alternative to improve growth and serum parameters, as well as attenuate clinical symptoms and intestinal disruption in ETEC-challenged piglets, which could be associated with its ability to mitigate gut microbiota dysbiosis. Our findings provided evidence for the usage of GOD as an approach to restrict ETEC infection in pigs.

Project description:?-Conglycinin (?-CG), an anti-nutritional factor, is a major allergen in soybeans to induce intestinal dysfunction and diarrhea in neonatal animals, including piglets and human infants. This study with a piglet model determined the effects of N-acetylcysteine (NAC) on intestinal function and autophagy in response to ?-CG challenge. Twenty-four 12-day-old piglets (3.44?±?0.28 kg), which had been weaned at 7 days of age and adapted for 5 days after weaning, were randomly allocated to the control, ?-CG, and ?-CG?+?NAC groups. Piglets in the control group were fed a liquid diet containing 10% casein, whereas those in the ?-CG and ?-CG?+?NAC groups were fed the basal liquid diets containing 9.5% casein and 0.5% ?-CG for 2 days. Thereafter, pigs in the ?-CG?+?NAC group were orally administrated with 50 mg (kg BW)-1 NAC for 3 days, while pigs in the other two groups were orally administrated with the same volume of sterile saline. NAC numerically reduced diarrhea incidence (-?46.2%) and the concentrations of hydrogen peroxide and malondialdehyde, but increased claudin-1 and intestinal fatty-acid binding protein (iFABP) protein abundances and activities of catalase and glutathione peroxidase in the jejunum of ?-CG-challenged piglets. Although ?-CG challenge decreased the villus height, villus height/crypt depth ratio, and mRNA levels of claudin-1 and occludin, no significant differences were observed in these indices between the control and ?-CG?+?NAC groups, suggesting the positive effects of NAC supplementation on intestinal mucosal barrier function. Moreover, NAC increased the concentrations of citrulline and D-xylose in the plasma, as well as the expression of genes for aquaporin (AQP) 3, AQP4, peptide transporter 1 (PepT1), sodium/glucose co-transporter-1 (SGLT-1), potassium inwardly-rectifying channel, subfamily J, member 13 (KCNJ13), and solute carrier family 1 member 1 (SLC1A1) in the jejunum, demonstrating that NAC augmented intestinal metabolic activity and absorptive function. Remarkably, NAC decreased Atg5 protein abundance and the LC3II/LC3I ratio (an indicator of autophagy) in the jejunum of ?-CG-challenged piglets. Taken together, NAC supplementation improved intestinal function and attenuated intestinal autophagy in ?-CG-challenged piglets.

Dataset Information

Differential expression of intestinal ion transporters and water channel aquaporins in young piglets challenged with enterotoxigenic Escherichia coli K88.

Publications

Differential expression of intestinal ion transporters and water channel aquaporins in young piglets challenged with enterotoxigenic <i>Escherichia coli</i> K88.

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