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Open Access Highly Accessed Methodology article

Detection and subtyping of Herpes simplex virus in clinical samples by LightCycler PCR, enzyme immunoassay and cell culture

Julie Burrows3, Andreas Nitsche1, Belinda Bayly3, Elise Walker2, Geoff Higgins3 and Tuckweng Kok3*

Author Affiliations

1 TIB-MOLBIOL, Tempelhofer Weg 11-12, D-10829, Berlin, Germany

2 Roche Diagnostics Australia, Nunawading, Victoria 3131, Australia

3 Institute of Medical & Veterinary Science, Adelaide, South Australia

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BMC Microbiology 2002, 2:12  doi:10.1186/1471-2180-2-12

Published: 9 June 2002

Abstract

Background

Prompt laboratory diagnosis of Herpes simplex virus (HSV) infection facilitates patient management and possible initiation of antiviral therapy. In our laboratory, which receives various specimen types for detection of HSV, we use enzyme immunoassay (EIA) for rapid detection and culture of this virus. The culture of HSV has traditionally been accepted as the diagnostic 'gold standard'. In this study, we compared the use of real time PCR (LightCycler) for amplification, detection and subtyping of specific DNA with our in-house developed rapid and culture tests for HSV.

Results

The LightCycler PCR (LC-PCR) detected and subtyped HSV in 99% (66/67) of HSV positive specimens, compared to 81% (54/67) by rapid antigen EIA or 57% (36/63) by culture. A specimen was considered positive when two or more tests yielded HSV identifications or was culture positive. Discordant results were confirmed with an in-house developed PCR-ELISA or DNA sequence analysis. The typing results obtained with the LC-PCR and by culture amplified test were completely concordant.

Conclusions

This study showed that the LC-PCR provided a highly sensitive test for simultaneous detection and subtyping of HSV in a single reaction tube. In addition to increased sensitivity, the LightCycler PCR provided reduced turn-around-times (2 hours) when compared to enzyme immunoassay (4 hours) or culture (4 days).