Background: Healthcare-associated INFECTIONS (HAIs) lead to serious problems such as antibiotic resistance and a high mortality rate. This study assessed the incidence of NOSOCOMIAL INFECTIONS (NIs), causative agents and their antimicrobial resistance patterns, and mortality rate in Isfahan province, Iran, by the national surveillance system. Methods: In a retrospective study, during 2017–2021, Iranian NOSOCOMIAL Infection Surveillance (INIS) software was used to extract data for inpatients with NIs in Isfahan province, Iran. The demographic and microbiologic findings were collected and analyzed. Results: From 2, 660, 073 hospitalized patients in 50 hospitals, the prevalence of NIs was 1. 46%. About 54. 5% of patients were male, and more than 62% were in the age group over 45 years. The total incidence was 24. 06%, 22. 22%, 17. 11%, 13. 25%, and 11. 32% for urinary tract infection (UTI), ventilator-associated events (VAE), skin and soft tissue infection (SSI), pneumonia (PNEU), and blood stream infection (BSI), respectively. During the study period, 15. 07% of deaths were recorded by NIs. The most common isolated bacteria from NIs were Acinetobacter spp. (12. 2%). Also, Acinetobacter spp. was the dominant isolated bacteria in VAE (17. 8%) and PNEU (4. 1%). The common isolated bacteria in UTI, BSI, and SSI were E. coli (27. 2%), S. epidermidis (22. 1%), and Klebsiella spp (5. 8%), respectively. Acinetobacter spp was the most resistant bacteria isolated. Conclusion: Following the low incidence rate of NIs in Iran compared to other parts of the world, there is a need for more accurate diagnosis techniques and better reporting methods. The available data from INIS need to be used correctly to make smarter decisions, more effective interventions, and appropriate antibiotic stewardship programs for antimicrobial resistance control.