Shigatoxigenic Escherichia coli (STEC) cause serious foodborne infections that lead to diarrheal disease and sequelae worldwide. In Burkina Faso, West Africa, STEC strains from environmental and human sources have not been isolated and characterized before. In this study, 21 STEC strains were isolated from food samples of animal origin and human feces using colony hybridization of the Shiga toxin gene stx. The STEC strains belonged to 15 different serotypes, including O43:H2, O8:H(-), and O2:H2. All strains were positive for stx(1) and 10 also for stx(2). The most common stx(1) subtype was stx(1a), and the most common stx(2) subtype was stx(2b). In five strains, stx(2) subtypes stx(2a) and/or stx(2c), which were previously associated with hemolytic uremic syndrome, were present. Some of the strains possessed the gene saa, encoding autoagglutinating adhesin. None of the strains possessed the gene eae, encoding intimin. Two STEC strains carried also an enterotoxigenic E. coli-associated gene estIa, encoding heat-stable enterotoxin. The STEC isolated from food in Burkina Faso are potentially pathogenic for humans based on the virulence gene combinations that they possess and phenotypes that they express.