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Purpose: To characterize the prevalence of hemolytic Shiga toxin-producing Escherichia coli (STEC) with a multidrug-resistant pattern in different age groups in Abeokuta, Nigeria. Methods: Nonrepetitive E. coli isolates were collected from 202 subjects with or without evidence of diarrhea. Each isolate was biochemically identified and antimicrobial susceptibility testing was performed using the disk diffusion method. A sorbitol fermentation test of all the E. coli isolates was done and the minimum inhibitory concentration of suspected STEC was measured by the standard broth microdilution method to determine antibiotic resistance. The genotypes of stx1, stx2, and hlyA were determined by polymerase chain reaction assay. Results: The majority of subjects were aged $40 years (41.6%) and were female (61.9%). Of the 202 subjects, 86.1% had STEC isolates (P,0.05). A high rate of STEC isolates resistant to amoxicillin (90.6%), cefotaxime (77.7%), and cefuroxime (75.7%) was observed. Resistance to amoxicillin, gentamicin, and cefotaxime was demonstrated with a minimum inhibitory concentration .16 μg/mL in 13.9%, 11.4%, and 10.4% of the isolates, respectively. The prevalence of stx1, stx2, and hlyA was 13.9%, 6.9%, and 2.0%, respectively; 5.5% of stx1 were in the 0–10-year-old age group, 3.5% of stx2 were aged $40 and above, and 1.0% of the hlyA isolates were in the 0–10-year-old age group. Conclusion: The prevalence of virulent STEC is a public health concern. The use of polymerase chain reaction assay should aid quick detection of this virulent serotype and help curb the severe epidemic of human diseases associated with STEC infections. [ABSTRACT FROM AUTHOR]