Objective To analyze the distribution characteristics and antimicrobial resistance of pathogenic bacteria isolated from clinical urine specimens from hospitals in Gansu Province in 2021. Methods Pathogenic bacteria antimicrobial resistance monitoring data of clinical urine specimens from member units of the Gansu Province Antimicrobial Resistance Surveillance Network in 2021 were collected. Antimicrobial susceptibility was determined according to the Clinical and Laboratory Standards Institute (2021 edition). Data were analyzed using WHONET 5.6 software. Results In 2021, a total of 13 980 bacterial strains from urine specimens were included in the analysis, with Gram-negative and Gram-positive bacteria accounting for 76.5% (10 692 strains) and 23.5% (3 288 strains), respectively. 40.5% (5 656 strains) and 59.5% (8 324 strains) bacterial strains were isolated from male and female patients, respectively. Isolation rate of carbapenem-resistant Acinetobacter baumannii was relatively high, with a resistance rate of > 30% in male patients. Resistance rates of Escherichia coli and Proteus mirabilis to ampicillin were >75%. Resistance rate of Pseudomonas aeruginosa to carbapenems was < 10%. Resistance rate of Enterococcus to vancomycin, linezolid and teicoplanin was < 2%. Resistance rates of Enterococcus faecalis and Enterococcus faecium to ampicillin were < 20% and >85%, respectively. Conclusion The distribution and antimicrobial resistance rate of pathogens in urine specimens from different genders are varied. Clinical antimicrobial use can be stan-dardized and rationalized according to the local antimicrobial resistance monitoring data.