Shiga toxin (Stx) can be classified into two types, Stx1 and Stx2, and different subtypes. Stx2e is a subtype commonly causing porcine edema disease and rarely reported in humans. The purpose of this study was to analyze the prevalence and genetic characteristics of Stx2e-producing Escherichia coli (Stx2e-STEC) strains from humans compared to strains from animals and meats in China. Stx2e-STEC strains were screened from our STEC collection, and whole-genome sequencing was performed to characterize their genetic features. Our study showed a wide distribution of Stx2e-STEC among diverse hosts and a higher proportion of Stx2e-STEC among human STEC strains in China. Three human Stx2e-STEC isolates belonged to O100:H30, Onovel26:H30, and O8:H9 serotypes and varied in genetic features. Human Stx2e-STECs phylogenetically clustered with animal- and food-derived strains. Stx2e-STEC strains from animals and meat showed multidrug resistance, while human strains were only resistant to azithromycin and tetracycline. Of note, a high proportion (55.9%) of Stx2e-STEC strains, including one human strain, carried the heat-stable and heat-labile enterotoxin-encoding genes st and lt, exhibiting a STEC/enterotoxigenic E. coli (ETEC) hybrid pathotype. Given that no distinct genetic feature was found in Stx2e-STEC strains from different sources, animal- and food-derived strains may pose the risk of causing human disease.