The objective of this study was to investigate the presence of multidrug-resistant (MDR) Escherichia coli strains isolated from clinical mastitis cases of cows in 42 different dairy farms located in the Bordj Bou Arreridj region of Algeria. Milk samples were cultured on Columbia blood agar and isolates were then identified by MALDI-TOF MS. A total of 200 samples were screened and 52 confirmed E. coli strains were obtained. The Antimicrobial susceptibility testing was performed by disc diffusion method. Antibiotic resistance genes, including extended-spectrum β-lactamases (ESBL) (i.e. bla TEM , bla SHV and bla CTX-M ), tetracyclines ( tetA , tetB , tetC and tetJ ), aminoglycosides ( aph(3’) , aac(3’) , aac(6’) , ant , aad and armA ) and quinolone ( qnrA and qnrB ) were amplified by polymerase chain reaction (PCR). DNA sequencing were then used to characterize the genotype. Genotyping analysis was performed using multilocus sequence typing (MLST). The most frequently observed resistance was amoxicillin (86.5%), followed by tetracycline (75%), amoxicillin/clavulanic Ac. (59.6%), trimethoprim/sulfamethoxazole (36.5%), doxycycline (13.5%) and ciprofloxacin (13.5%). Multidrug resistance was observed in 38.4% of isolates. Genotypic characterisation showed that tetA gene (44.2%) and   bla TEM-1 (30.7%) were the most prevalent. Screening for plasmid-mediated quinolone resistance (PMQR) genes demonstrated that seven isolates (13.5%) expressed the qnrB gene and one isolate (1.9%) harbored the qnrA gene. In addition, aminoglycosides resistance determinants including aadA 1 and aac (3)- Id were detected in 7 and 2 isolates respectively. The MLST revealed the presence of 3 different sequence types (ST162, ST371 and ST 949).