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Ureaplasma urealyticum is a causative agent of non-gonococcal urethritis and is implicated in the pathogenesis of prostatitis, epididymitis and infertility. The organism is more common in partners of infertile than fertile marriages. U. urealyticum infections not only jeopardize fertility but also pose a risk for infertility treatment and resulting pregnancies. The aim of this study was to determine the prevalence of U. urealyticum and Ureaplasma parvum in semen of infertile and healthy men by polymerase chain reaction (PCR). Semen samples were obtained from infertile patients and healthy controls and were subjected to the routine andrological analysis and PCR. DNA was extracted by the Cadieux method, and analysed by PCR protocol with specific primers for urease and multiple-banded antigen genes. Ureaplasmas were detected significantly by PCR in 12 of 100 (12%) semen specimens from infertile patients and in three of 100 (3%) healthy men. The volume of semen fluid, concentration of sperm cells, and sperm cell with normal morphology were significantly decreased in infertile men. In the group of infertile patients with PCR positive for Ureaplasmas, the volume, count and morphology of semen samples were lower than in the infertile patients with PCR-negative results. U. urealyticum species in semen of infertile men was found to be high (9%) than in healthy controls (1%). Detection rate for U. parvum was 3% in the infertile group and 2% in healthy men. The results indicate that U. urealyticum species is more common in specimens of infertile men. The percentage of normal sperm cells, the volume of semen and the percentage of sperm cells with motility in the PCR positive for U. urealyticum species group were lower than in the PCR positive for U. parvum group.