Evaluation of multiplex real-time PCR for detection of Haemophilus ducreyi, Treponema pallidum, herpes simplex virus type 1 and 2 in the diagnosis of genital ulcer disease in the Rakai District, Uganda.

OBJECTIVE: To develop a real-time PCR assay that reliably and accurately detects the predominant sexually transmitted aetiological agents of genital ulcer disease (GUD) (Haemophilus ducreyi, Treponema pallidum and herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2)) and to assess the use of real-time PCR diagnostic testing in a rural African field site. METHODS: Two multiplex real-time PCR reactions were used to detect H ducreyi/and HSV-1/HSV-2 in ulcer swabs from 100 people with symptomatic genital ulcers in rural Rakai, Uganda. Results were compared with syphilis, HSV-1 and HSV-2 serology. RESULTS: Of 100 GUD samples analysed from 43 HIV positive and 57 HIV negative individuals, 71% were positive for one or more sexually transmitted infection (STI) pathogens by real-time PCR (61% for HSV-2, 5% for T pallidum, 3% for HSV-1, 1% for H ducreyi and 1% for dual H ducreyi/HSV-2). The frequency of HSV in genital ulcers was 56% (32/57) in HIV negative individuals and 77% (33/43) in HIV positive individuals (p = 0.037). Assay reproducibility was evaluated by repeat PCR testing in the USA with 96% agreement (kappa = 0.85). CONCLUSIONS: STI pathogens were detected in the majority of GUD swab samples from symptomatic patients in Rakai, Uganda, by real-time PCR. HSV-2 was the predominant cause of genital ulcers. Real-time PCR technology can provide sensitive, rapid and reproducible evaluation of GUD aetiology in a resource-limited setting.

Aetiology of genital ulcer disease in female partners of male participants in a circumcision trial in Uganda.

HIV acquisition is associated with herpes simplex virus type 2 (HSV-2) infection and genital ulcer disease (GUD). Three randomized control trials demonstrated that male circumcision significantly decreases HIV, HSV-2, human papillomavirus and self-reported GUD among men. GUD is also decreased among female partners of circumcised men, but it is unknown whether male circumcision status affects GUD pathogens in female partners. For the evaluation of GUD aetiology, two separate multiplex assays were performed to detect Haemophilus ducreyi, Treponema pallidum, HSV-1 and HSV-2. Of all the female GUD swabs evaluated, 67.5% had an aetiology identified, and HSV-2 was the primary pathogen detected (96.3%). However, there was no difference in the proportion of ulcers due to HSV-2 or other pathogens between female partners of circumcised men (11/15, 73.3%) compared with uncircumcised men (15/25, 60.0%, P = 0.39). The seroprevalence of HSV-2 is high in this population and therefore most of the detected HSV-2 infections represent reactivation. Since GUD is associated with HIV acquisition and one-third of GUD in this study did not have an aetiological agent identified, further research is needed to better understand the aetiology of GUD in Africa, and its relationship to circumcision and HIV infection.