Abstract: Objective To explore the drug susceptibility and molecular epidemiological characteristics of clinical isolates of carbapenem-resistant Escherichia coli. Methods A total of 21 carbapenem-resistant E. coli strains were isolated from a hospital in 2015. The minimum inhibitory concentration (MIC) of these strains were detected by agar dilution method. Carba NP test was used to detect carbapenemases. Carbapenemase genes including blaNDM, blaKPC, blaIMP, blaVIM and blaOXA were detected by PCR followed by sequence alignment; Pulsed-field gel electrophoresis (PFGE) and multi locus sequence typing (MLST) were performed for the molecular subtyping. Results All the 21 carbapenem-resistant E. coli strains were resistant to imipenem, meropenem, piperacillin, cefepime, ceftriaxone, ceftazidime and levofloxacin. The resistant rate to trimethoprim-sulfamethoxazole was as high as 76.19%. The resistant rates to minocycline, amikacin and fosfomycin were 23.81%, 23.81% and 19.05%, respectively. Seventeen strains were positive in Carba NP test. PCR and sequences alignment confirmed that the 17 strains carried blaNDM-1 genes. The other four carbapenemase genes were not detected. MLST analysis showed that 21 strains could be divided into nine ST types, ST167 and ST540 were detected in the strains carrying blaNDM. Three strains from a urology ward had identical PFGE patterns and another two strains from different wards also had identical patterns, suggesting the spread of an E. coli clone in the ward. Conclusion blaNDM-1 gene was carried by E. coli strains with different ST types in the hospital. E. coli strains carrying the blaNDM-1 were highly resistant to common antimicrobial drugs. It is important to strengthen the detection and surveillance of transmission of E. coli strains carrying blaNDM-1 gene in hospitals.