Objective To investigate antimicrobial resistance and resistance genes of Escherichia coli from pet animals in Guangzhou.Method From July 2016 to July 2017, 319 samples were collected from cats and dogs at four animal hospitals in Guangzhou, including 127 samples from healthy animals and 192 samples from diseased animals. E. coli strains were isolated using the selective media, and were identified by MALDI-TOF mass spectrometry. The susceptibilities of E. coli isolates to 11 antimicrobial agents were determined by the agar dilution method. The presence of resistance genes was determined by PCR and sequencing.Result A total of 203 E. coli strains were isolated from 319 samples, including 109 E. coli isolates from diseased animals and 94 E. coli isolates from healthy animals. Among the 203 strains, 179 strains were resistant to at least one antimicrobial agent. The isolates showed the highest resistance rate against ampicillin (76.85%). The resistance rates against cefotaxime, tetracycline, doxycycline and sulfamethoxazole-trimethoprim were above 50%. The isolates were the most susceptible to amikacin with the resistance rate of only 10.84%. The isolates from diseased animals showed higher resistance rates against all antimicrobial agents compared with those from healthy animals. The resistance rates against all antimicrobial agents except amikacin, florfenicol and fosfomycin were significantly higher in the isolates from diseased animals compared with healthy animals (P < 0.01). The detection results of resistance genes showed that floR had the highest detection rate (34.97%), the detection rates of bla_CTX-M-9G, bla_CTX-M-1G, fosA3, rmtB and bla_CMY-2 genes were 22.66%, 20.19%, 17.73%,10.34% and 1.48% respectively. The genes bla_CTX-M-2G and bla_CTX-M-25G were not detected.Conclusion The antimicrobial resistance of E. coli isolates from pet animals in Guangzhou has become a serious problem, and some isolates carry several important resistance genes. Antimicrobial resistance in pet animals requires strict monitoring.