Background: Trichomoniasis is a worldwide sexually transmitted disease (STD), and is associated with important public health problems, including enhancement of HIV transmission. The prevalence of the parasite, Trichomonas vaginalis, depends on host factors such as age, sexual activity, number of sexual partners and sexual behavior. The aim of the study was to evaluate the prevalence of trichomoniasis in high-risk behavior women, such as drug addicts and those who had multiple sexual partners, attending Gynecology Clinics in penitentiaries of Tehran province to help gynecologists with the diagnosis and treatment of the disease. Methods: Samples of posterior vaginal fornix discharges and urines of 450 women attending Gynecology Clinics of three prisons in Tehran province were collected. All samples were examined by direct smear and cultured in TYI-S-33 culture media. Results: 10.2% of subjects were positive for trichomoniasis. 82.7% of infected patients were symptomatic individuals who complained of vaginal discharge and itching and/or burning sensation. Most of the infected women in the high risk behavior group were drug users (54.3%). Physical examination showed that 50% of T. vaginalis positive subjects had a normal appearance of vagina and cervix. The peak prevalence (32.8%) of the disease occurred in the age range of 31-40 years. Conclusion: T. vaginalis infection is commonly associated with other STDs, and is a marker of high-risk sexual behavior. Due to the side effects of the drugs used to treat the infection, it is suggested that the treatment be performed after definite diagnosis using a diagnostic method with a higher sensitivity.