Trichomonas vaginalis (T. vaginalis) is a common pathogen with worldwide distribution. It is estimated that worldwide 180 million people are infected annually. Trichomoniasis is associated with vaginitis, cervicitis, low birth weight, and preterm delivery. PCR has the advantage of high sensitivity, shorter time for diagnosis and the ability to detect nonviable or defective organism. In this study we used these three methods for evaluation of PCR in comparison with conventional methods like wet mount and culture in the detection of T. vaginalis in vaginal discharge. Three vaginal swab specimens were obtained from each of 200 cases, of the age group 18-40years, both symptomatic and asymptomatic females attending Gynaecology OPD(50) and Family planning OPD(50) at Gandhi hospital, Secunderabadand two FSW(Female sex workers) clinics (100) in highly concentrated areas of them in Hyderabad, for validation of various forms of Trichomonas vaginalis diagnostic procedures. One swab was immediately examined by wetmount microscopy, a second swab was placed in Wittington’s medium for cultivation, and other swab is placed in 2SP transport medium for PCR for T.vaginalis. A total of 58 samples positive in one or more tests were identified: 11 (5.5%) infections were detected by wet mount microscopy, and 30 (15%) positives in culture respectively. PCR was positive in 50 (25%) samples. PCR appears to be the most sensitive method with high detection rate and method of choice for detection of genital infections with T. vaginalis.