ObjectiveTo investigate antimicrobial resistance of Escherichia coli (E. coli) and Klebsiella pneumoniae(K. pneumoniae), antimicrobial use density(AUD), as well as relation between antimicrobial resistance and AUD in a tertiary firstclass hospital. MethodsAntimicrobial resistance rates of clinicallyisolated E. coli and K. pneumoniae, AUD of carbapenems and quinolones, as well as relation between resistance and AUD in 20132015 were statistically analyzed.ResultsCorrelation analysis of antimicrobial resistance of bacteria and AUD showed that the decrease in resistance rate of E. coli to levofloxacin was related to the decrease in the use density of quinolones（r=0.61，P=0.03）；increase in resistance rate of K. pneumoniae to imipenem was related to the increase in the use density of carbapenems（r=0.78，P＜0.01）. ConclusionAntimicrobial use is one of the causes of bacterial resistance, management on antimicrobial use needs to be strengthened to reduce the threat of bacterial resistance to human health.