We have shown that patients with prostato-vesiculo-epididymitis (PVE) have the worst sperm output compared to patients with prostato-vesiculitis or prostatitis alone. The present study was undertaken to closely examine whether unilateral or bilateral PVE had a different impact on sperm parameters, seminal fructose levels and reactive oxygen species (ROS) overproduction. To accomplish this, 78 patients with persistent post-infectious inflammatory PVE, clearly identified by scrotal and transrectal ultrasonography, and 30 patients with asymptomatic post-infectious inflammatory prostatitis (control group) underwent semen analysis (including seminal leukocyte concentration and number of spermiophagies), seminal fructose measurement and sperm ROS production from 45 and 90% Percoll fractions. Fifty patients turned out to have PVE bilaterally, whereas the remaining 28 had unilateral PVE. Patients with bilateral PVE had sperm concentration and total sperm number significantly lower than those found in patients with unilateral PVE. The other sperm parameters, the physicochemical properties (hyperviscosity, the presence of nonspecific agglutination, delayed liquefaction), seminal fructose levels and ROS production in both 45 and 90% Percoll fractions turned out similar between the two groups. Patients with bilateral or unilateral PVE had sperm parameters, seminal fructose levels and ROS production significantly worst than those found in patients with prostatitis alone. In conclusion, although patients with bilateral PVE had a decreased number of spermatozoa, the other sperm parameters and seminal fructose levels did not reflect the extension of PVE. Therefore, the diagnosis of unilateral or bilateral involvement of this complicated form of male accessory gland infection relies on scrotal and transrectal ultrasonography.