Escherichia coli (E. coli) O157:H7 is considered a significant food-borne microorganism that causes food poisoning infections in humans every year. E. coli O157:H7 has various virulence factors such as Shiga-toxin encoding (Stx1 and Stx2). Meat and its products are considered the best meals that consumers eat every day worldwide, but meat and its products are exposed to contamination through unhygienic processing, handling, and storage. The aim of the study was the isolation of E. coli O157:H7 and detection of the uidA, Stx1, and Stx2 genes. 504 samples of meat and butchers' shops were gathered from diverse areas in Mosul city. Classical and molecular biology techniques were used to isolate and identify E. coli O157:H7. The results appeared to indicate the total number of E. coli isolates in this study was 138 and the spread rate of E. coli O157:H7 isolated was 9.4% (13/138). The spread rate of E. coli O157:H7 was high in workers hands 4 (20%), while we did not detect E. coli O157:H7 in Machines. Additionally, all E. coli O157:H7 have the uidA, and Stx2 genes at 100%, while 92.3% of E. coli O157:H7 possess the Stx1 gene. The study concludes E. coli O157:H7 occurrences in meats and butchers' shops and that all equipment and tools used were capable of transmitting E. coli O157:H7 to meats. Meats and butchers' shops are a risk to humans who consume the uncooked meats.