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A real-time PCR assay
is a sensitive procedure for the rapid diagnosis of HSV in genital
lesions.
Rapid detection of
herpes simplex virus DNA in genital ulcers by real-time PCR using SYBR
Green I dye as the detection signal.
Aldea C, Alvarez CP, Folgueira L,
Delgado R, Otero JR.
Journal of Clinical Microbiology
2002;40:1060-1062.
Summary:
Question
How does a real-time PCR assay compare to conventional cell culture for
the detection of HSV in genital lesions?
Design
Direct comparison of a real-time PCR assay using SYBR Green I dye for
amplicon detection to conventional cell culture for the detection of HSV-1
and HSV-2 in genital swab samples from patients with genital lesions.
Participants
118 patients attending a sexually transmitted disease clinic in Madrid,
Spain who presented with genital lesions.
Description of Tests and Diagnostic
Standard
A swab was taken from the surface of the genital lesion and placed in
viral transport medium. Standard viral cultures were performed on MRC-5
and A-549 cells for up to 7 days. In samples considered positive for HSV,
cells were passaged to shell vials and stained with specific fluorescent
reagents to HSV-1 and HSV-2 (MicroTrak HSV 1-2 culture
identification/typing test, Dade-Behring, Marburg, Germany). PCR was
carried out on DNA extracted from the cell pellet of the transport medium
sample using a LightCycler and 2 sets of primers amplifying regions from
the thymidine kinase and DNA polymerase genes of HSV-1 and HSV-2. The HSV
PCR amplicons were detected by melting point analysis. Samples were
considered positive if they showed a specific signal for both sets of
primers and a quantification higher than 10 DNA copies.
Main Outcome Measures
The detection of HSV 1 and HSV 2 DNA by real-time PCR compared to
detection of HSV by conventional cell culture.
Main Results
HSV was isolated in 28 of 118 samples (24%) by cell culture, 10 samples
were positive for HSV-1, and 18 were positive for HSV-2. All cell culture
positive samples were positive by real-time PCR. Of 90 samples negative by
culture, 84 were negative and 6 were positive by PCR. HSV was detected
with both sets of primers for all PCR positive samples.
Authors' Conclusions
Real-time PCR using SYBR Green I as
detection signal is a sensitive procedure for the rapid diagnosis of HSV
in genital lesions.
Source of funding: In part, Fondo de
Investigacion Sanitaria.
For correspondence: Joaquin Otero,
Unidad de Virologia, Servicia de Microbiologia, Hospital 12 de Octubre,
Madrid 28041, Spain. E-mail address: jrotero@hdoc.insalud.es.
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