Escherichia coli O157 isolates obtained from 17 Danish cattle herds and from a national surveillance programme of cattle at slaughter were genotyped by pulsed-field gel electrophoresis (PFGE) and characterised with respect to presence of and variation in virulence factors. The characteristics of the cattle strains were compared to human clinical isolates from the same time period. All verocytotoxin (VT)-producing E. coli O157 (VTEC O157) from cattle possessed all typical VTEC O157:H7 virulence factors and had either the VT2c-variant alone or together with VT1. Among human isolates the dominant toxin profile was VT2 + VT2c. Only one PFGE group was represented on each farm, indicating that introduction and establishment of new E. coli O157 strains to these cattle farms is probably not common. Among E. coli O157 isolates from cattle, 22.8% were not VT-producing. The majority of these possessed the eae gene and all other genotypic and phenotypic traits typical for E. coli O157:H7. On the basis of the virulence characteristics, it is concluded that the VTEC O157 strains isolated from Danish cattle are potential human pathogens. However, the observed differences between cattle and human isolates with regard to VT-profile, genotype and antimicrobial resistance could be important, i.e. either Danish cattle might not be the most important reservoir for human infections or Danish cattle mostly harbour VTEC O157 isolates that are less likely to cause human disease.