Background: Legionella pneumophila as a ubiquitous bacterium is inherently resistant to chlorine in tap water. It can easily enter water piping systems and get transmitted to immunocompromised populations and cause severe pneumonia. Owing to the fact that its presence in water sources doesn’t necessarily lead to onset of disease; therefore, several factors such as inhaled bacteria dose, virulence factors and diversity of serogroups can be considered as contributing factors. The main aim of current project was to investigate the contamination rate of hospital water systems with Legionella by culture and evaluate presence of major virulence factor genes as well as the ability to form biofilms among the Legionella isolates. Results: Twelve (12%) of the 100 water samples produced positive results in culture method. Additional confirmation was performed by PCR method with specific primers for Legionella genus (16SrRNA) and pneumophila species (mip). Fifty (5%) samples of 12 with positive culture have a colony forming unit higher than 1000cfu/100 ml. Legionella were isolated with a rate of 8%, 3% and 1% from shower heads, oxygen humidifier bottle and water bath, respectively. PCR assay for the virulence genes showed that all 12 (100%) isolates were positive for mip genes, 9 (75%) were positive for dot gene, 8 (66.66%) were positive for hsp, 6(50%) were positive for lvh and 4(33.33%) for rtx. Two of the isolates displayed higher ability to form biofilm in reference to the standard strain.Conclusion: Although the presence of Legionella pneumophila in hospital environment does not necessarily confer a threat to public health; continuous monitoring of water sources should be conducted in order to avoid elevated concentration of this bacterium and visible biofilm formation.