The antibiotic resistance profiles of Escherichia coli isolated from three different sampling sites of the Gomti River at Lucknow city was evaluated. Water samples were collected and then analyzed for the presence of E. coli, using standard methods. Antibiotic susceptibility testing was performed by the disc diffusion method. Of the 77 E. coli isolates tested, marked antibiotic resistances (over 70%) were observed for amoxicillin, nitrofurozone, chloramphenicol, polymixin B, methicilin, ampicillin, nalidixic acid, cefpodoxime, erythromycin, penicillin, rifampicin and ofloxacin depending upon the sampling sites. All E. coli isolates also showed multiple resistance patterns in different combination of antibiotics. The MAR index ranges were found very high indicating the high risk of environmental contamination. The findings indicated that pollution of aquatic environments from different sources of the city may have a potential impact on the dissemination and survival of E. coli, as well as other pathogenic bacteria in the Gomti River water for public and animal health. This may result to a negative effect on antibiotic therapy for infectious diseases.