BACKGROUND AND ABJECTIVE Urinary tract infection (UTI) is one of the most common infections in humans which probably affects one-half of all people during their lifetimes. Uropathogenic Escherichia coli (UPEC), one of the members of the extra-intestinal pathogenic E. coli (ExPEC), is the main etiologic agent of UTI. The excessive and inappropriate use of antimicrobial agents specially β-lactam antibiotics for treatment of UTI infections has recently led to the emergence of multiple drug resistance (MDR) and increasing in Extended Spectrum β-lactamases (ESBL) producing E. coli, worldwide. The present study was conducted to investigate the presence of ESBLs among the UPESs isolated from patients with UTI referred to hospitals in Hamedan city using phenotypic and molecular methods. MATERIALS AND METHODS A total of 100 UPEC isolates, were used in this study. These strains were isolated from the patients with UTI referred to hospitals in Hamedan city, Hamedan, Iran. Bacterial isolation and identification was done using conventional microbiology methods and standard biochemical tests, additional identification was made using molecular methods. The UPEC isolates were screened for ESBLs production by the double-disk synergy test using Ceftazidime (30 μg) and Cefotaxime (30 μg) disks and confirmed by combined disk diffusion test using Clavulanic acid according to the guidelines recommended by the Clinical and Laboratory Standards Institute (CLSI, 2021). The presence of four β-lactamases gene including blaTEM, blaSHV, blaOXA, and blaCTX-M among ESBLs producer UPEC isolates (n= 40) was investigated using PCR RESULTS AND DISCUSSION From a total of 100 UPEC isolates, 40 (40%) isolates were phenotypically identified as ESBLs producers. Among 40 ESBLs producers UPEC isolate the most prevalent β-lactamases gene was blaCTX-M 19/40 (47.5%) followed by blaTEM 22.5% (9/40) and blaOXA 20 % (8/40). The blaSHV gene was not found in any tested isolates. CONCLUSION Due to relatively high pr
