Detection of herpes simplex virus DNA by real-time PCR

Molecular detection of herpes simplex virus (HSV) DNA is recognized as the reference standard assay method for the sensitive and specific diagnosis of central nervous system infections caused by HSV. In this study, a molecula r assay based on real time PCR on the LightCycler (LC) instrument was evalu ated and compared with a home-brew molecular assay. The detection limit of the LC assay was determined with 10-fold dilutions of plasmid pS4 with the San restriction fragment of the DNA polymerase gene and with the First Euro pean Union Concerted Action HSV Proficiency Panel. A total of 59 cerebrospi nal fluid (CSF) specimens were investigated for the comparative study. With plasmid pS4, the detection limit of the LC assay was found to be 10(4) cop ies per ml, i.e., 12.5 copies per run. When samples of the First European U nion Concerted Action HSV Proficiency Panel were tested, 2 x 10(3) to 5 x 1 0(3) HSV type 1 genome equivalents (GE) per mi, i.e., 2.5 ta 6.3 GE per run , could consistently be detected. There was a correlation between the LC as say and the home-brew assay in 55 of 59 specimens. In conclusion, the LC as say allows very rapid detection of HSV DNA in CSF. It was found to be labor saving and showed sufficient sensitivity.