Background: The increased rate of resistance among Gram-negative bacteria to 𝛽-lactam, quinolone, or aminoglycoside antibiotics has resulted in increased use of colistin antibiotic with the unavoidable risk of emerging resistance and spreading of colistin resistant strains. Objectives: Our study aimed to investigate urinary tract infections for detection of colistin resistance among E. coli strains causing urinary tract infections and isolated from two University hospitals in Upper Egypt; Assiut and Minia University hospitals. Methodology: One hundred isolates of E. coli strains were collected from urine specimens at both Assiut and Minia University Hospitals. Antibiotic sensitivity test was carried out using disc diffusion method. For analysis of colistin resistance, the broth microdilution technique was employed. In addition, the mcr-1 gene encoding an enzyme phosphoethanolamine transferase which has been identified recognized as a source of acquired resistance of colistin was detected by PCR assay. Results: The high resistance pattern among E. coli strains, as most strains were resistant to 13 antibiotics. mcr-1 gene was detected in (20.8%) and (23.1%) of Assiut and Minia University Hospital respectively E. coli isolates. In addition, in this report, we describe the detection of mcr-1 in E. coli isolates causing urinary tract disorders infections in Upper Egypt. Conclusion: Our study revealed a high rate of mcr-1 gene encoding a phosphoethanolamine transferase in E. coli isolates as a source of acquired resistance to colistin. The Emergence of mcr-1 in our isolates of E. coli is alarming. Therefore, broader surveillance of this resistance determinant would be recommended even among other members of Gram-negative bacilli.