Pathogenicity O island 122 (OI-122) associated with the severity of Enteropathogenic Escherichia coli (EPEC) disease. The aim of this study was the characterization of OI-122 in Iranian EPEC isolates. The distribution patterns of OI-122 were investigated in 42 EPEC strains by detection of OI-122 genes and serogrouping, multi locus VNTR (MLVA) typing, and phylogenetic grouping. The complete OI-122 was identified in 18.75% of tEPEC and in 3.8% of aEPEC strains, an incomplete OI-122 with various combinations of genes was found in 75% of tEPEC and 30.76% of strains aEPEC, and OI-122 genes were absent in 6.25% of tEPEC and 65.3% of aEPEC strains. nleB (52.4%) was the most frequently detected, and the prevalence of pagC, efa1/lifA, and sen genes were 19%, 16.7%, and 14.3%, respectively. The most common phylogenetic group among tEPEC and aEPEC strains were B1 (68.75% and 80.76%, respectively). The common serogroups among EPEC isolates were O128, O111, O55, O127, and O44, and 45.2% of the isolates were untypeable. Forty-one MLVA types among 42 EPEC strains were classified into 7 clonal complexes. The diverse distribution of OI-122 genes among MLVA clonal groups provides evidence for dynamic evolution regarding the OI-122 pathogenicity island in tEPEC and aEPEC strains. The results also indicate that the acquisition of OI-122 gene contents occurs in a modular manner.