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. 2014 May;63(5):761-70.
doi: 10.1136/gutjnl-2013-304739. Epub 2013 Jul 11.

Colonic mucosa-associated diffusely adherent afaC+ Escherichia coli expressing lpfA and pks are increased in inflammatory bowel disease and colon cancer (VSports注册入口)

Maelle Prorok-Hamon  1 , Melissa K Friswell, Abdullah Alswied, Carol L Roberts, Fei Song, Paul K Flanagan, Paul Knight, Caroline Codling, Julian R Marchesi, Craig Winstanley, Neil Hall, Jonathan M Rhodes, Barry J Campbell
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Colonic mucosa-associated diffusely adherent afaC+ Escherichia coli expressing lpfA and pks are increased in inflammatory bowel disease and colon cancer

Maelle Prorok-Hamon et al. Gut. 2014 May.
Free PMC article

"V体育2025版" Abstract

Objective: Colonic mucosa-associated Escherichia coli are increased in Crohn's disease (CD) and colorectal cancer (CRC). They variously haemagglutinate, invade epithelial cell lines, replicate within macrophages, translocate across M (microfold) cells and damage DNA. We investigated genes responsible for these effects and their co-association in colonic mucosal isolates. VSports手机版.

Design: A fosmid library yielding 968 clones was prepared in E coli EPI300-T1 using DNA from a haemagglutinating CRC isolate, and resulting haemagglutinating clones were 454-pyrosequenced. PCR screening was performed on 281 colonic E coli isolates from inflammatory bowel disease (IBD) (35 patients), CRC (21) and controls (24; sporadic polyps or irritable bowel syndrome). V体育安卓版.

Results: 454-Pyrosequencing of fosmids from the haemagglutinating clones (n=8) identified the afimbrial adhesin afa-1 operon. Transfection of afa-1 into E coli K-12 predictably conferred diffuse adherence plus invasion of HEp-2 and I-407 epithelial cells, and upregulation of vascular endothelial growth factor. E coli expressing afaC were common in CRC (14/21, p=0. 0009) and CD (9/14, p=0. 005) but not ulcerative colitis (UC; 8/21) compared with controls (4/24). E coli expressing both afaC and lpfA (relevant to M-cell translocation) were common in CD (8/14, p=0. 0019) and CRC (14/21, p=0. 0001), but not UC (6/21) compared with controls (2/24). E coli expressing both afaC and pks (genotoxic) were common in CRC (11/21, p=0. 0015) and UC (8/21, p=0 V体育ios版. 022), but not CD (4/14) compared with controls (2/24). All isolates expressed dsbA and htrA relevant to intra-macrophage replication, and 242/281 expressed fimH encoding type-1 fimbrial adhesin. .

Conclusions: IBD and CRC commonly have colonic mucosal E coli that express genes that confer properties relevant to pathogenesis including M-cell translocation, angiogenesis and genotoxicity. VSports最新版本.

Keywords: BACTERIAL ADHERENCE; BACTERIAL INTERACTIONS; BACTERIAL PATHOGENESIS; E. COLI; GUT INFLAMMATION. V体育平台登录.

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Figures (VSports最新版本)

Figure 1
Figure 1
Haemagglutinin-positive E coli in the fosmid clone library derived from E coli HM358 share a common region containing the afimbrial adhesin afa-1 operon. (A) Genomic organisation of the common region shared by haemagglutination (HA)-positive clones includes afaA (encoding a transciptional regulator); afaB (chaperone); afaC (usher); afaD (invasin); draP (linker element) and afaE-1 (an adhesin). (B) PCR for afaC. A 672 bp fragment is detected in all eight haemagglutination-positive clones and E coli HM358. E coli EPI300-T1/pCC1FOS and eight haemagglutination-negative fosmids lacked afaC.
Figure 2
Figure 2
The presence of the afa-1 operon in haemagglutinating E coli correlates with diffuse adherence and invasion to HEp-2 epithelial cells. Giemsa stain of HEp-2 cells infected with E coli strains. (A) All eight haemagglutinin-positive library clones showed diffuse adherence to cell cultures. (B) Eight haemagglutination (HA)-negative fosmid clones chosen at random from the library were non-adherent. (C) Colonic mucosally associated E coli HM358 exhibited a diffusely adherent pattern as per diffusely adherent E coli C1845. The E coli K12 plating strain EPI300T1 containing pCC1Fos was non-adherent. (D) The eight haemagglutination-positive fosmid library clones possessing the afa-1 gene cluster, exhibiting diffuse adherence, showed increased ability to invade Hep-2 cells compared to haemagglutination-negative clones. Invasion calculated as percentage of the original inoculums (multiplicity of infection 10) and expressed relative to E coli LF82 previously shown to be invasive in this cell line. * p<0.05 and *** p<0.001 when compared to the non-invasive plating strain EPI300-T1 containing pCC1Fos alone (mean±SEM; N=3 experiments, each performed with n=3 replicates; Kruskal–Wallis).
Figure 3
Figure 3
The presence of afa-1 in haemagglutinating E coli correlates with ability to adhere to and invade intestinal epithelial cells. Relative ability of eight haemagglutination-positive library clones possessing afa-1 (afaC+ as determined by PCR) and haemagglutination-negative clones (n=8) to (A) adhere to and (B) invade I-407 cells. Data mean (±SEM) relative to that observed by E coli HM358; n=4. Increased (C) adherence to, and (D) invasion of differentiated Caco2 cells by afa-1-possessing clones was observed. (E and F) E coli EPI300-T1/pCC1FOS transformed with afa-1 (pUCAfa) adheres to and invades I-407 cells. Data expressed relative to plating strain; N=3, each n=3–5 replicates). p Values determined by Kruskal–Wallis. Afa, afimbrial adhesin; afa-1, afimbrial adhesin operon 1; afaC, gene encoding afimbrial adhesin outer membrane usher protein.
Figure 4
Figure 4
The presence of afa in E coli upregulates vascular endothelial growth factor (VEGF) expression in cultured intestinal epithelial cells. Confluent serum-starved I-407 cells (8×105 cells/well) were infected for 4 h with either wild-type colorectal cancer adherent, invasive E coli HM358, Afa/Dr diffusely adhering E coli C1845 or E coli EPI300-T1/pCC1FOS transformed with afa-1 operon (pUCAfa) or vector alone (pUC18) and total RNA extracted. VEGF mRNA measured by quantitative PCR relative to β actin. Data mean (±SEM) relative to non-infected cells (set at 100%); N=2–4; each n=3 replicates). *p<0.05, *** p<0.001 determined by Kruskal–Wallis. AIEC, adherent, invasive E coli; DAEC, diffusely adherent E coli.
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References

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