ROLE OF INTIMIN AND BUNDLE-FORMING PILI IN ENTEROPATHOGENIC ESCHERICHIA-COLI ADHESION TO PEDIATRIC INTESTINAL TISSUE IN-VITRO

Attaching and effacing (A/E) lesion formation is central to enteropath ogenic Escherichia coli (EPEC) pathogenesis, In vitro experiments with human epithelial cell lines have implicated virulence plasmid-encoded bundle-forming pill (BFP) in initial binding and intimin in intimate attachment and A/E lesion formation. This study investigated the role of BFP and intimin in EPEC interactions with pediatric small intestina l biopsy tissue in in vitro organ culture. Organ culture infections (2 to 8 h) were performed with E2348/69 (a wild-type EPEC O127:H6 clinic al isolate) and E2348/69 derivatives including CVD206 (eae deficient), CVD206(pCVD438) (eae-complemented CVD206), CVD206(pCVD438/01) (expres sing intimin, which is nonfunctional due to a single amino acid substi tution), JPN15 (spontaneous EPEC adherence factor virulence plasmid-cu red E2348/69), and 31-6-1(1) (E2348/69 with a TnphoA insertion inactiv ation mutation in the virulence plasmid-encoded bfpA gene). Scanning a nd transmission electron microscopy revealed that after 8 h E2348/69 a nd CVD206 (pCVD438) (both Int(+) BFP+) adhered to all specimens, causi ng A/E lesions with surrounding microvillous elongation. JPN15 and 31- 6-1(1) (both Int(+) BFP-) adhered and caused A/E lesions although bact eria adhered in ''flat,'' two-dimensional groups, CVD206 and CVD206(pC VD438/01) (both Int(-) BFP+) did not adhere to any sample, and no path ological tissue changes were seen, Thus, in human intestinal organ cul ture, BFP do not appear to be involved in the initial stages of EPEC n onintimate adhesion but are implicated in the formation of complex, th ree-dimensional colonies via bacterium-bacterium interactions. Intimin appears to play an essential role in establishing colonization of EPE C on pediatric small intestinal tissue.