Invasive ability of an Escherichia coli strain isolated from the ileal mucosa of a patient with Crohn's disease

Crohn's disease (CD) is an inflammatory bowel disease in which Escherichia coli strains have been suspected of being involved. We demonstrated previou sly that ileal lesions of CD are colonized by E. coli strains able to adher e to intestinal Caco-2 cells but devoid of the virulence genes so far descr ibed in the pathogenic E. coli strains involved in gastrointestinal infecti ons. In the present study we compared the invasive ability of one of these strains isolated from an ileal biopsy of a patient with CD, strain LF82, wi th that of reference enteroinvasive (EIEC), enteropathogenic (EPEC), entero toxigenic (ETEC), enteraggregative (EAggEC), enterohemorrhagic (EHEC), and diffusely adhering (DAEC) E. coli strains. Gentamicin protection assays sho wed that E. coli LF82 was able to efficiently invade HEp-2 cells. Its invas ive level was not significantly different from that of EIEC and EPEC strain s (P > 0.5) but significantly higher than that of ETEC (P < 0.03), EHEC (P < 0.005), EAggEC (P < 0.004) and DAEC (P < 0.02) strains. Strain LF82 also demonstrated efficient ability to invade intestinal epithelial cultured Cac o-2, Intestine-407, and HCT-8 cells, Electron microscopy examination of inf ected HEp-2 cells revealed the presence of numerous intracellular bacteria located in vacuoles or free in the host cell cytoplasm, In addition, the in teraction of strain LF82 with epithelial cells was associated with the elon gation of microvillar extensions that extruded from the host cell membranes and engulfed the bacteria. This internalization mechanism strongly resembl es Salmonella- or Shigella-induced macropinocytosis. The use of cytochalasi n D and colchicine showed that the uptake of strain LF82 by HEp-2 cells was mediated by both an actin microfilament-dependent mechanism and microtubul e involvement. In addition, strain LF82 survived for at least 24 h in HEp-2 and Intestine-407 cells and efficiently replicated intracellularly in HEp- 2 cells, PCR and hybridization experiments did not reveal the presence of a ny of the genetic determinants encoding EIEC, EPEC, or ETEC proteins involv ed in bacterial invasion, Thus, these findings show that LF82, which coloni zed the ileal mucosa of a patient with CD, is a true invasive E. coli strai n and suggest the existence of a new potentially pathogenic group of E. col i, which we propose be designated adherent-invasive E. coli.