Background Molecular assays based on PCR have become an important tool for the detection of herpes simplex virus-1&2 DNA in clinical specimens. Detection and typing of HSV can also be done by a monoclonal antibody based DFA. The present study was undertaken to standardise an in-house PCR for detection and typing of Herpes Simplex Virus (HSV) and compare it with Direct Fluorescent Antibody (DFA) test.

Methods 44 patients with genital herpes attending the STD clinic were studied. Specimens collected from genital lesions were placed in Viral transport medium (VTM) and stored at –70⁰C till tested. DNA extraction was done using QiaAmp DNA mini kit (Qiagen, USA), PCR was carried out in GeneAmp PCR system 9700 (Applied BioSystems). Post PCR analysis of PCR product was done by electrophoresis using 2% agarose gel. DFA (BioRad) was also done for identification and typing of HSV-1& 2.

Results By DFA, 4 specimens were positive for HSV 1, 19 were positive for HSV 2 while 7 were positive for both. By PCR, 5 were HSV-1 positive, 18 were HSV-2 positive while 6 were positive for both HSV-1 & 2. (kappa for HSV-1 = 0.879, HSV-2 = 0.63,) One HSV- 1 and 3 HSV-2 cases was positive by PCR but not by DFA. Four specimens that were positive by DFA but negative by PCR were retested in duplicate to rule out the presence of PCR inhibitors. Three of them were then found to have no inhibitors, while one had presence of PCR inhibitors.

Conclusions Our results show that although majority of cases studied were due to HSV-2, HSV-1 either alone or as a mixed infection with HSV-2 is not uncommon. PCR was found to be as sensitive as DFA for confirming the syndromic diagnosis, but some false negatives may occur due to presence of PCR infibitors.