Trichomonas vaginalis is a protozoan parasite that infects the human urogenital tract. This study determined the prevalence and risk factors of T. vaginalis infection amongst human immunodeficiency virus (HIV) infected patients in Limpopo Province using urine-based real time polymerase chain reaction (RT-PCR). Urine samples were collected from 155 patients attending three public hospitals and one private clinic in the Vhembe region. Demographic data, clinical and socioeconomic status were collected from the patients using a structured questionnaire. Total genomic DNA was isolated from urine samples using the Qiagen Blood Mini Kit and RT-PCR protocol was used for the detection of T. vaginalis. The overall prevalence of T. vaginalis in our study population was 21%. The prevalence was higher among patients less than 25 years compared to older patients. Patients who were taking antibiotics at the time of sample collection had less infection compared to patients who were not on antibiotics (24% vs. 7%; p=0.042). Low CD4 counts and early age of sexual debut appeared to be important risk factors. The high recovery rate obtained in this study demonstrates the importance of employing real-time PCR techniques in the diagnosis of the trichomoniasis in this population.