Vancomycin resistant enterococci are challenging bacteria as they are difficult to be eradicated. Toxin-antitoxin (TA) systems are genetic elements located in most prokaryotic genomes. The mazEF TA system is harbored by a plasmid among Enterococcus faecalis (E. faecalis). To explore the relation between the existence of mazEF TA system and vancomycin resistance among clinical isolates of E. faecalis. Samples were collected from patients showing clinical picture of infection. Isolates of E. faecalis were identified by standard microbiological methods and their antimicrobial susceptibility patterns were detected by disk diffusion method. In addition, the E-test was used to confirm vancomycin resistant isolates. All the E. faecalis isolates were screened for the mazEF TA system by PCR. A total of 180 E. faecalis strains were identified with a vancomycin resistance rate of 30.6%. Vancomycin resistance was significantly associated with prolonged hospital stay (P= 0.04) and ICU setting (P= 0.001). The mazEF TA system was detected among 100% of vancomycin resistant isolates, while only 33.6% of the vancomycin sensitive isolates carried the system with a significant difference (P= 0.002). In addition, there was a significant association between the mazEF TA system-positive strains and the ICU setting (P= 0.02). A significant association was found between vancomycin resistance and the presence of the mazEF TA system among E. faecalis isolates. This association supports the current efforts to utilize the mazEF TA system as a possible target for novel antibacterial agents; however, further studies on a wider scale are necessary.