This study aimed at isolating uropathogenic Escherichia coli from urinary tract infections (UTIs) of human and cattle to examine the molecular diversity and phylogenetic relationship of the isolates. A total of 100 urine samples were collected from UTIs of human and cattle. The isolates identification was done using routine diagnostic methods and confirmed by Vitek2. Antimicrobial susceptibility was tested against 10 antimicrobials. Random amplified polymorphic DNA (RAPD)-polymerase chain reaction (PCR) was applied to identify the genetic diversity among E. coli isolates from human and animal origin by using five different octamer primers. The gelJ software for the phylogenetic analysis created Dendrograms. Out of 50 human urine samples, E. coli was isolated from 12 (24%) samples, and was positive in 5 out of 50 (10%) of cattle urine samples. Concerning the antimicrobial susceptibility test, both human and animal isolates revealed rather approximate results when tested mainly against imipenem, cefotaxime, and ciprofloxacin. These antimicrobial data might indicate presence of a degree of similarity between the human and animal isolates. Using RAPD-PCR, three of the primers produced polymorphic bands; therefore, they were used for further analysis of the results. Either of P1, P3 or P4 primers showed presence of similarity between human and cow isolates. To conclude, RAPD-PCR and gelJ software might be of attractive use to identify and analyze the occurrence of genetic relationships, as this could assist in controlling the routes and sources of infection transmission between human and animal in order to prevent zoonotic infections.