Project description:To better understand the molecular effects of Enterotoxigenic Escherichia coli (ETEC) F4ab/ac infection, we performed a genome-wide comparison of the changes in DNA methylation in ETEC F4ab/ac infected porcine intestinal epithelial cells. Our data provides further insight into the epigenetic alterations of ETEC F4ab/ac infected porcine intestinal epithelial cells and may advance the identification of biomarkers and drug targets for predicting susceptibility to and controlling ETEC F4ab/ac induced diarrhea.
Project description:A GWAS study was then performed in 52 non-adhesive and 68 strong adhesive pigs for F4ab/ac ETEC originating from 5 Belgian farms. A new refined candidate region (chr13: 144,810,100-144,993,222) for F4ac ETEC susceptibility was identified with MUC13 adjacent to the distal part of the region. All pigs were phenotyped for the presence of the F4ab/ac receptor (F4ab/acR) using the in vitro villous adhesion assay with 4×108 F4ac E. coli (strain GIS26, serotype O149:K91, F4ac+) or F4ab E. coli (strain G7, serotype O8:K87, F4ab+) . A total of 120 F4ab/acR phenotyped pigs were genotyped using the Porcine SNP60 BeadChip (Illumina) containing 62,163 SNPs, according to the manufacturer’s protocol. The position of the SNPs was based on the current pig genome assembly (Sscrofa10.2).
Project description:A GWAS study was then performed in 52 non-adhesive and 68 strong adhesive pigs for F4ab/ac ETEC originating from 5 Belgian farms. A new refined candidate region (chr13: 144,810,100-144,993,222) for F4ac ETEC susceptibility was identified with MUC13 adjacent to the distal part of the region.
Project description:Enterotoxigenic Escherichia coli (ETEC) strains that produce both heat-stable (ST) and heat-labile (LT) enterotoxins cause severe post-weaning diarrhea in piglets. However, the relative importance of the individual enterotoxins to the pathogenesis of ETEC infection is poorly understood. In this study, we investigated the effect on virulence of an F4+ ETEC strain when removing some or all of its enterotoxins. Several isogenic mutant strains were constructed that lack the expression of LT in combination with one or both types of ST enterotoxins (STa and/or STb). Host early immune responses induced by these mutant strains 4h after infection were compared to the wild type strain GIS26 (O149:F4ac+, LT+ STa+ STb+). At the same time, the immune response of this wild type ETEC strain was compared to the mock-infected control, demonstrating the expression of porcine inflammatory response genes. For these purposes, the small intestinal segment perfusion (SISP) technique and microarray analysis were used and results were validated by qRT-PCR. We also measured net fluid absorption of pig small intestinal mucosa 4h after infection with wild type ETEC, the mutant strains and PBS (mock-infected). These data indicate an important role for STb in inducing small intestinal secretion early after infection. The microarray analysis of the different mutant strains also revealed an important role for STb in ETEC-induced immune response by the significant differential regulation of immune mediators like matrix metalloproteinase 3, interleukin 1 and interleukin 17. We conclude that STb can play a prominent role in ETEC-induced secretion and early immune response.
Project description:Enterotoxigenic Escherichia coli (ETEC) strains that produce both heat-stable (ST) and heat-labile (LT) enterotoxins cause severe post-weaning diarrhea in piglets. However, the relative importance of the individual enterotoxins to the pathogenesis of ETEC infection is poorly understood. In this study, we investigated the effect on virulence of an F4+ ETEC strain when removing some or all of its enterotoxins. Several isogenic mutant strains were constructed that lack the expression of LT in combination with one or both types of ST enterotoxins (STa and/or STb). Host early immune responses induced by these mutant strains 4h after infection were compared to the wild type strain GIS26 Â (O149:F4ac+, Â LT+ Â STa+ Â STb+). At the same time, the immune response of this wild type ETEC strain was compared to the mock-infected control, demonstrating the expression of porcine inflammatory response genes. For these purposes, the small intestinal segment perfusion (SISP) technique and microarray analysis were used and results were validated by qRT-PCR. We also measured net fluid absorption of pig small intestinal mucosa 4h after infection with wild type ETEC, the mutant strains and PBS (mock-infected). These data indicate an important role for STb in inducing small intestinal secretion early after infection. The microarray analysis of the different mutant strains also revealed an important role for STb in ETEC-induced immune response by the significant differential regulation of immune mediators like matrix metalloproteinase 3, interleukin 1 and interleukin 17. We conclude that STb can play a prominent role in ETEC-induced secretion and early immune response. In three pigs, 6 different treatments were performed. These treatments consisted of 4 mutant enterotoxigenic Escherichia coli GIS26 strains, GIS26 wild type strain, or PBS control. Per pig, the small intestine was divided into 6 loops with an interloop in between to avoid cross-contamination. In conclusion, every pig received each of the 6 treatments ad random.
Project description:We investigate the transcriptome response of porcine intestinal epitheliocyte cell line (PIE cells) to the challenge with heat-stable Enterotoxigenic Escherichia coli (ETEC) pathogen-associated molecular patterns (PAMPs) and, the changes induced by Lactobacillus jensenii TL2937 in that response. The transcriptome approach allowed us to obtain a global overview of the immune and immune related genes involved in response of PIE cells to heat-stable ETEC PAMPs. The most remarkable changes in PIE cells after heat-stable ETEC PAMPs challenge were observed in chemokines expressions, followed by cell adhesion molecules and, complement and coagulation cascades factors. We also confirmed that L. jensenii TL2937 differently modulates gene expression in ETEC PAMPs-challenged PIE cells. The microarray gene expression profiles clearly demonstrated that an anti-inflammatory effect was triggered by the immunobiotic strain in PIE cells. The main outcome from the study was the differential regulation of chemokines (CCL8, CXCL5, CXCL9, CXCL10 and CXCL11), complement factors (C1R, C1S, C3 and CFB) and, coagulation system proteins (Tissue factor) expression by L. jensenii TL2937. PIE cells treated with the negative control Lactobacillus plantarum TL2766 showed a transcriptomic response similar to ETEC PAMPs-challenged PIE cells.
Project description:Porcine deltacoronavirus (PDCoV) is an emerging pathogen of swine belonging to family Coronaviridae, genus Deltacoronavirus. PDCoV predominantly infects the porcine intestinal epithelial cells (IPECs) causing severe diarrhea and/or vomiting, dehydration, and death in piglets. However, there are no researches for clarifying the changes of proteins expression levels in the IPECs infected with PDCoV. To better understand the host response to PDCoV infection, in this study, an isobaric tags for relative and absolute quantification (iTRAQ) labeling combined with liquid chromatography-tandem mass spectrometry (LC-MS)-based quantitative proteomic analysis of PDCoV-infected IPEC-J2 cells were performed to investigate the differentially expressed cellular proteins in the PDCoV-infected IPEC-J2 cells. As a result, a total of 5,502 host proteins were quantified at 24 hours post-infection (hpi) in mock and infected cells, among which 78 cellular proteins were differentially expressed with 23 up-regulated proteins and 55 down-regulated proteins. Bioinformatics analysis demonstrated that most of these regulated proteins participated in immune system process and structural molecule activity. Further, expression levels of two representative proteins, ANAPC7 and IFIT1, were confirmed by relative real-time RT-PCR and western blot analysis. The data presented here will provide an overview of host cell response to PDCoV, which could benefit the development of potential antiviral research.
Project description:ETEC is an important human pathogen. Although the mechanism of diarrhea is known in ETEC, the regulatory networks are less understood. This study was conducted to understand the global expression of ETEC isolate E24377A under different growth and environemental conditions. ETEC isolate E24377A was grown in the presence of several chemical signals, including bile salts, glucose, and pre-conditioned media (PCM) from other enteric pathogens. E24377A was also grown to different densities, to see if a quorum sensing mechanism was in place
Project description:ETEC is an important human pathogen. Although the mechanism of diarrhea is known in ETEC, the regulatory networks are less understood. This study was conducted to understand the global expression of ETEC isolate E24377A under different growth and environemental conditions. ETEC isolate E24377A was grown in the presence of several chemical signals, including bile salts, glucose, and pre-conditioned media (PCM) from other enteric pathogens. E24377A was also grown to different densities, to see if a quorum sensing mechanism was in place The isolate was grown in different types of media, with different ammendments, and at different growth densities. The overall goal was to determine how expression gene expression changes in the presence of chemical signals; a special emphasis was placed on the expression of known and suspected virulence and colonization factors