Clinical Application of Multiplex PCR Assay for the Diagnosis of the Etiology of Genital Ulcer Disease Among Patients Attending STD Clinics in Guangzhou, China

Objectives: To develop a method of simultaneous PCR detection of Haemophilus ducreyi, Treponema pallidum, andHerpes Simplex Virus Types 1 and 2 from genital ulcersamong patients attending STD clinics in Guangzhou, China;and evaluate the clinical application of multiplex PCR (M-PCR) assay for diagnosing the etiology of genital ulcerdiseases (GUD). Methods: 244 patients with a genital ulcer were evaluated.Clinical etiology of GUD was based on physical appearanceand microbiologic evaluations that included darkfieldmicroscopy examination (D-F) and serology test for syphilis(STS). Swabs of each genital ulcer were tested for HSVantigen by enzyme immunoassay (EIA) and processed in anM-PCR assay for simultaneous detection of T pallidum, HSVand H ducreyi. Results: The standard strains of T pallidum, HSV and Hducreyi were amplified by M-PCR, producing amplifiedproducts of 260bp,432bp,170bp, respectively. The sensitivityof M-PCR is 10~2 pg DNA. M-PCR assay for T.pallidum, HSVand H ducreyi showed good agreement en compared with D-F detection for T pallidum, STS, H ducreyi culture and EIAfor HSV antigen (Kappa scores are 0.774,0.704,0.793,0.756,respectively). Conclusions: The M-PCR is a convenient, accurate andreliable assay for the detection of T pallidum, HSV and Hducreyi from genital ulcers, and can be used as a method ofdiagnosing the etiology of GUD.

DETECTION OF PATHOGENS CAUSING GENITAL ULCER DISEASE BY MULTIPLEX POLYMERASE CHAIN REACTION

Objective To establish a multiplex polymerase chain reaction （M-PCR） assay for simultaneous detection of pathogens causing genital ulcer disease （GUD）. Mothods Based on the gene-specific region of the following pathogens： Chlamydia trachomatis omp l/ompb, herpes simplex virus （HSV） DNA polymerase, Treponema pollidum tpp47, Haemophilus ducreyi 16s rRNA, four sets of primers were designed and an M-PCR assay was developed to detect four pathogens in one test. The assay was evaluated with diagnostic result of golden standard for each pathogen.Results Of the 51 clinical samples, M-PCR showed slightly higher positive rate （47.1%） of HSV than cell culture （23.6%）. Meanwhile, the positive rate of T. pallidum detected by M-PCR and dark-field microscopy was 19.6% （10/51） and 15.7% （8/51）, respectively. Only one sample was positive for H. ducreyi and no sample was positive for C. trachomatis detected by both M-PCR assay and culture. Conclusion This primary study indicated that M-PCR assay can simultaneously and rapidly detect the four etiologic pathogens causing GUD.