Background: Globally, urinary tract infections (UTIs) constitute the most prevalent health issue. UTIs are challenging to cure due to the emergence of antibiotics resistant bacterial strains. This research aimed to determine the incidence of UTIs, identify the causative bacteria, and evaluate antibiotic resistance profile and genes in Enterococcus faecalis. Materials and methods: In this study, twenty E. faecalis isolates from patients with urinary tract infection were identified; phenotypic and genotypic characteristics were investigated in Al-Najaf City, Iraq. Results: The data revealed different rates of resistance starting with high rate (65%) of isolates which were resistant to Nalidixic acid and (60%) of isolates resistant to Ciprofloxacin and Tetracycline. then ends with lower resistance which were (20%) of isolates have resistance to Imipenem and Fosfomycine. At the same respect, all isolates of E. faecalis displayed multi drug resistance (MDR). Antibiotics resistance genes bla-tem, emeA and gyrA were found in 100%, 95% and 55% of isolates, respectively. However, this research explained that none antibiotic was entirely working against all isolates. Conclusion: All E. faecalis isolates exhibited multi drug resistance (MDR), and the most potent antibiotics were imipenem and fosfomycin.