Objective To evaluate the improvement efficacy of pathogen detection management strategies before antimicrobial treatment, and provide evidence-based basis for standardizing the clinical application of antimicrobial agents. Methods Data reported from 568 sentinel hospitals nationwide from the first quarter of 2022 to the third quarter of 2023 were selected. The baseline survey stage was the first quarter of 2022, improvement strategies began to be implemented from the second quarter of 2022. Change in pathogen detection rate before antimicrobial treatment, pathogen detection rate related to healthcare-associated infection (HAI) diagnosis, pathogen detection rate before the combination use of key antimicrobial agents, percentage of blood culture, as well as length of hospital stay before and after improvement of strategies were compared. Results After implementing the improvement strategy (the third quarter of 2023), pathogen detection rate before antimicrobial treatment was 57.55%, which was higher than 39.08% before improvement (the first quarter of 2022) (P<0.001). Pathogen detection rate rela-ted to HAI diagnosis was 90.85%, which was higher than 80.33% before improvement (P<0.001). Pathogen detection rate before the combination use of key antimicrobial agents was 92.05%, which was higher than 87.75% before improvement (P<0.001). Percentage of blood culture was 12.35%, which was higher than 7.54% before improvement (P<0.001). In addition, the average length of hospital stay for patients shortened from 9.49 days to 8.24 days. Conclusion Comprehensive management strategies can effectively improve the pathogen detection rate and detection quality before antimicrobial treatment, shorten the length of hospital stay of patients, and provide practical basis for the rational use of antimicrobial agents as well as the prevention and control of antimicrobial resistance.