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Infections with verotoxin-producing Escherichia coli (VTEC) has resulted in increasing numbers of human illnesses annually. These illnesses usually result from the ability of VTEC to cause the attaching and effacing lesions (AE lesion). The AE phenotype is encoded by the locus of enterocyte effacement (LEE) pathogenicity island. A key adhesion factor involved is the outer membrane protein intimin, encoded by the eae gene within the LEE. Intimin types alpha, beta, gamma, delta, and epsilon have been described previously. Each intimin represents distinct phylogenetic lineages of LEE-positive strains. A new intimin type zeta was identified in a VTEC strain of the serotype O84:NM (nonmotile) that was isolated from a calf with diarrhea. zeta intimin showed the highest similarity (88%) of its amino acid sequence to the alpha intimin. For diagnostic purposes, we established a polymerase chain reaction (PCR) method for diagnosis of the key virulence traits of VTEC (i.e., verotoxins and intimins). This method also distinguishes between the toxins (VT1 and VT2) and the six intimin types. By applying the PCR method, intimin zeta in strains of other VTEC serotypes O84:H2, O92:NM, O119:H25, and O150:NM was identified. Because the intimin types represent distinctive phylogenetic E. coli lineages, application of the intimin subtyping PCR offers significant benefits. These include improving diagnosis of VTEC infection and increasing the understanding of evolution of attaching and effacing VTEC and other LEE-positive bacteria.

We describe a locus of enterocyte effacement (LEE) which is part of a new pathogenicity island (PAI) detected in the bovine Shiga toxin-producing Escherichia coli strain RW1374 (O103:H2). This PAI is at least 80 kb in size and inserted in the vicinity of the pheV tRNA gene at 67 min of the E. coli chromosome. Furthermore, the PAI differs from the previously described LEEs by unique flanking regions at both sides, which harbor one copy each of an insertion element in an inverted orientation that is 96% identical to insertion site (IS)629. In addition, a 5-kb PAI-specific sequence downstream of the LEE core region and adjacent to the E. coli K12 region is duplicated upstream of the LEE core region as well. The duplicated sequences are more than 80% identical to each other and consist partially of prophage sequences.