Acinetobacter baumannii is a major causative agent of serious nosocomial infections. This study was carried out to investigate the molecular characterization of colistin resistant isolates of A. baumannii from hospitalized patients, based on multilocus sequence typing (MLST). A cross-sectional study was conducted to collect A. baumannii from clinical samples in Isfahan from 2021 to 2022. Isolates were identified as A. baumannii using biochemical tests and PCR of blaOXA-51. Antibiotic susceptibility testing was carried out using the Kirby-Bauer method and minimum inhibitory concentration (MIC) values were determined for colistin. Additionally, MLST was performed according to the Pasteur scheme to assess the relationship between colistin resistant A. baumannii. A total of 70 non-repetitive A. baumannii isolates were obtained from different clinical samples. MIC results showed that seven A. baumannii isolates were resistant to colistin. The antibiotic susceptibility pattern revealed that all seven colistin resistant strains were resistant to all tested antibiotics. Based on MLST analysis, the colistin resistant isolates were assigned to five unique STs namely, ST2 (3; 42.9%) followed by ST78 (1; 14.3%), ST1077 (1; 14.3%), ST415 (1; 14.3%) and ST391 (1; 14.3%). Among them ST2, ST391 and ST415 belong to clonal complex 2. Colistin resistant A. baumannii ST2 is the main circulating clone in clinical settings in Iran, but additionally ST415, ST391, and ST1077 are found for the first time in our country. Intensive control procedures and strict adherence to surveillance programs are recommended to decrease the spread of carbapenem and colistin resistant A. baumannii strain.  Kulcsszavak: Acinetobacter baumannii, colistin-resistant, multi-drug resistance, MLST