The virulence plasmid of Escherichia coli O157 strain EDL933 carries a 10-kb putative virulence gene designated toxB. Little is known about the distribution of this gene among E. coli O157 strains or its presence in other enterohemorrhagic E. coli (EHEC) and enteropathogenic E. coli (EPEC) strains. We developed PCR and hybridization tools for the detection of the entire toxB sequence and investigated its presence in a collection of EHEC O157 strains and other EHEC and EPEC strains belonging to different serogroups and isolated from different sources. The EHEC O157 strains reacted with all of the PCR primers and probes used, thus indicating the presence of a complete toxB gene regardless of the human or bovine origin of the isolates. Similar positive reactions were observed for about 50% of the EHEC O26 strains tested and a few other EHEC and EPEC strains. However, the size of the DNA fragments hybridizing with the toxB probes differed from that of the positive fragments from EHEC O157, suggesting a polymorphism in the toxB genes present in the different E. coli serogroups. Moreover, several EHEC and EPEC strains belonging to different serogroups reacted with only some of the genetic tools used, suggesting either the existence of major variants of toxB or the presence of fragments of the gene. Southern blotting analysis showed that toxB sequences were located on large plasmids in EHEC and EPEC O26 as well.