Colonic mucosa-associated diffusely adherentafaC+ Escherichia coliexpressinglpfAandpksare increased in inflammatory bowel disease and colon cancer

Abstract

ObjectiveColonic 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.DesignA 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).Results454-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=0.022), 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.ConclusionsIBD and CRC commonly have colonic mucosal E coli that express genes that confer properties relevant to pathogenesis including M-cell translocation, angiogenesis and genotoxicity.

Pathogenesis of Crohn’s disease: Bug or no bug

“…As Friswell et al [44] highlight in their review of the role of bacteria in the pathogenesis of IBD, blockade of bacterial entry via M cells represents an important target for therapies. In a very recent paper [45] , certain isolates of E. coli have been shown to be related both with colorectal cancer and IBD. The authors suggest that interventions that either reduce colonization by diffusely adherent E. coli or block their interaction with the mucosa may have preventive or therapeutic effects in colon cancer and CD.…”

Selection and validation of reference house­keeping genes in the J774A1 macrophage cell line for quantitative real-time PCR

“…The murine macrophage lineage J774A1 cell line is a great in vitro model because of its remarkable plasticity (Franz et al, 2011;Hosokawa et al, 2011). Several recent genomic studies have used the J774A1 cell line to evaluate cytokine/chemokine activity (El Aamri et al, 2015), integrin modulation (Ferraz et al, 2016), inflammatory activity (Bohr et al, 2013), and cancer markers (Prorok-Hamon et al, 2014). These studies mainly utilized quantitative polymerase chain reaction (qPCR), a reliable and versatile technique, to evaluate gene expression under various biological conditions.…”

Commensal bacterial internalization by epithelial cells: An alternative portal for gut leakiness

“…64 Other studies have documented that mucosa-associated E.coli expressing pathogenicity islands of afimbrial adhesin (afa), long polar fimbriae (lpf) and polyketide synthase (pks) are frequently seen in IBD and colon cancer patients. 5,65 The afa and lpf operons are known to confer adhesive characteristics on E.coli for epithelial cells. 5,65 Recent studies in mouse models of colitis-associated colorectal cancer have demonstrated that pksencoding colibactin produced by mucosaassociated, adherent E.coli causes DNA damage and cell senescence, and drives tumor progression.…”

“…5,65 The afa and lpf operons are known to confer adhesive characteristics on E.coli for epithelial cells. 5,65 Recent studies in mouse models of colitis-associated colorectal cancer have demonstrated that pksencoding colibactin produced by mucosaassociated, adherent E.coli causes DNA damage and cell senescence, and drives tumor progression. 66,67 The newly acquired adherent and cytotoxic characteristics of enteric bacteria are likely to stimulate host immune response through pathogen-associated or damage-associated molecular patterns, triggering the onset of proinflammatory signaling.…”

“…Disruption of the gut barrier, followed by penetration of microbial products into the gut mucosa and circulatory bloodstream, predisposes the host to enterocolitis, systemic inflammation, or septic complications. 2,3 Abnormal bacterial adherence and internalization by epithelial cells has been reported in patients and experimental models with colorectal cancers, [4][5][6] inflammatory bowel disease (IBD), 7,8 celiac disease, 9,10 chronic psychological stress, 11,12 surgical manipulation, 13 intestinal obstruction, 14,15 and antibiotic-resistant bacterial infection. 16 Recent evidence suggests that mucosa-associated bacteria may contribute to pathological phenomena since many exhibit adherent and invasive characteristics, and are therefore termed 'pathobionts' (a commensal with the potential to cause disease; see review papers elsewhere).…”