OBJECTIVE: To evaluate the usefulness of different phenotypic and genotypic markers for epidemiological typing of enteroinvasive Escherichia coli 0124 (EIEC 0124). METHODS: Seven sporadic EIEC 0124 isolates and 22 isolates from two different outbreaks were characterized. Chromosomal deoxyribonucleic acid (DNA) macrorestriction analysis with XbaI resolved by pulsed-field gel electrophoresis (PFGE) and ribotyping with each of the three restriction endonucleases BglII EcoRI, and ClaI were compared with biotyping, antimicrobial susceptibility patterns and plasmid profiles. RESULTS: Biotypes and antimicrobial resistance profiles of the outbreak-associated strains showed considerable variation, thereby limiting the usefulness of such phenotypic markers. Only 57% of the sporadic isolates harbored plasmids. Three different ribotypes based on 5 to 7 bands were recorded among sporadic isolates whereas all outbreak-associated strains showed the same ribotype. BglII appeared to give the best discrimination whereas EcoRI and ClaI provided no additional information. Sporadic EIEC 0124 isolates showed a marked diversity of macrorestriction patterns (similarity coefficient 58 to 93%) and five different patterns were detected. In contrast, the outbreak isolates were closely related (similarity coefficient 90 to 100%). Genomic DNA macrorestriction analysis correlated well with ribotyping, but PFGE was more discriminating. CONCLUSIONS: PFGE is a useful method for epidemiological comparison and differentiation of EIEC 0124 isolates.