Introduction: Pseudomonas aeruginosa is an aerobic, motile, non-spore forming, straight or slightly curved, gram-negative bacilli and it can grow at 42° C. The incidence of P. aeruginosa has been increased in recent years and its virulence factors are various. The aim of this study was to compare the differences of the virulence factors in pigmented and non-pigmented P. aeruginosa isolates. Materials and methods: Isolates of patients that were identified as P. aeruginosa were included into the study. Strains of patients (n: 100) were divided into two groups according to pigment production. Antibiotic susceptibility tests of all strains were performed by disk diffusion method. DNase, protease, elastase, hemolysis and motility tests were performed as phenotypic tests. The presence of several virulence-associated genes encoding exotoxins T (exoT), S (exoS), U (exoU) and Y (exoY) and quorum sensing mediators (rhlA and rhlB) were assessed by Real-time PCR method for genotypic identification. The relationship between pigment production, antibiotic resistance and virulence factors were examined. Results: There was no significant difference between pigmented and non-pigmented isolates when elastolytic activity and mucus production were compared. Pigmented isolates produced significantly more (p<0.05) protease and hemolysis activity. Motility was present in pigmented isolates more frequently than in non-pigmented isolates. DNase activity was significant in pigmented isolates then non-pigmented isolates (p<0.05). Antibiotic resistance was present more frequently in non-pigmented isolates than pigmented isolates. Pigmented isolates had more frequently and significant more (p<0.05) virulence-associated genes rhlB, exoS, exoY (p<0.05). Conclusion: The results of this study suggest that both phenotypic and genotypic virulence factors may be associated with the pigment production in P. aeruginosa. Pigment production is easy to determine, which might be a good starting point to identify the virulence status of an isolate.