Background: Enterococcus faecalis is one of the human and animal gastrointestinal flora. They are very robust so that they can live in humid, salty, or acidic environments. Understanding the link between enteric and extra-enteric E. faecalis strains is interesting as it is widely distributed throughout a variety of different habitats. Objective: This research was carried out to investigate the percentage presence aggregation substance, cytolysin and hyaluronidase genes (asa-1, cyl A, hyl) in the E. faecalis isolates from UTI samples via the method of Polymerase Chain Reaction (PCR). Materials and Methods: In the current study, collected 64 Samples during the period from November 2021 to January 2022 at Medical City Hospital in Baghdad city, included 18 men and 46 women. The patients ages were ranged from (17-54) years. Urine samples cultured on Pfizer specific Enterococcus media, a selective medium for the isolation and growth of Enterococcus spp, and then molecular detection was done to detect the E. faecalis gene and all three-virulence gene. Results: Results demonstrated that the proportion of isolation for E. faecalis was 48%, while the percentage of cyl and asa-1 genes was 100% and 97% respectively, although hyl gene was not detected in any isolates of E. faecalis isolated from urine samples. Conclusion: Virulence genes may be prevalent is some isolates and absent from other depending on many factors such as the source of isolation.