Backgrounds and Objectives: Total and Fecal coliforms (TC and FC), heterotrophic plate count (HPC), were counted by microbiological method and E.coli O157: H7 were detected through immunological and Real time PCR methods in water intake and all of units of Isfahan water treatment plant (IWTP).Materials and Methods: The microbial profile including TC, FC, and HPC, were monitored and turbidity and total organic carbon were analyzed in 8 locations of water intake, and unit operation and processes of IWTP, including, inlet, sedimentation, ozonation, and filtration and finished water. Immunological method through anti-serum kits and molecular method of RT-PCR were used to detect E.coli O157:H7 in the 8 locations and also the sludge of the sedimentation basin and filters backwash water of IWTP.Results: Survival of E.coli O157: H7 in sludge sample of sedimentation basin was indicated by formation of agglutination particles in immunological method and through indicator probes in the RT-PCR method. However, E.coli O157:H7 was not detected in water samples of other units of IWTP. The removal percent of TC, FC, and HPC were: 59.5, 49, and 54.8 % in sedimentation basin; 66, 45.8, and 57 % in ozonation; : 98.8, 98, and 78.8 in the filtration; and 96, 100, 91% in disinfection, respectively.Conclusion: This study approved the existence of the pathogenic coliform, E.coli O157:H7 in the sludge of sedimentation basin. Absent of E.coli O157:H7 in the finished water indicates that the existing units of IWTP could eliminate these pathogenic bacteria, before reaching the final units of the plant, including the filters and disinfection.