Background and Aim: Selection and dissemination of plasmid-encoded extended-spectrum β-lactamase (ESBL) among Enterobacteriaceae confers resistance to beta-lactam antibiotics. The purpose of this study was to determine the prevalence and molecular characteristics of ESBL-producing organisms isolated from dairy cattle with a uterine infection. Materials and Methods: Bacterial isolates (n=62) were characterized by biochemical test for genus and species determination. Antimicrobial susceptibility tests were performed by Kirby–Bauer disk diffusion method using panel of antibiotics for initial screening of ESBL organism. Phenotypic confirmation of ESBL-suspected strains was done by combination disk method and double-disk method. Multiplex polymerase chain reaction (PCR) was carried out for phylogrouping of Escherichia coli isolates as well as for genotyping ESBL genes. Enterobacterial repetitive intergenic consensus-PCR method was used for genotypic characterization of isolates. Results: Antibiotic susceptibility profile of E. coli (n=40) isolates showed high rates of resistance for ampicillin (95.0%), cefpodoxime (97.5%), cefotaxime (87.5%), and ceftriaxone (70%). However, low rates of resistance were observed for cefoxitin (25%), amoxicillin/clavulanic acid (20%), ceftazidime (17.5%), gentamicin (10%), and ertapenem (7.5%). A total of 39/40 E. coli isolates were confirmed as ESBL with Epsilometer test as well as the genotypic method and 28 (70%) of them were multidrug-resistant. Genotype blaCTX-M was observed as a predominant beta-lactamase type with the preponderance of CTX-M Group 1. The following combinations were observed: blaTEM + blaCTX-M in 15 (36.2%) isolates, blaTEM / blaSHV in 8 (5.2%) isolates, and blaCTX-M / blaSHV in 6 (5.2%) isolates. The phylogenetic grouping of E. coli strains revealed the highest prevalence for B1 (22.0%) followed by A (20%). Conclusion: This report shows a high frequency of ESBL E. coli from cattle with postpartum uterine infections. These isolates showed reduced susceptibility to common antibiotics used for the treatment of uterine infections greater affecting the therapeutic outcome.