Real-time PCR for quantification of human herpesvirus 6 DNA from lymph nodes and saliva.
A real-time quantitative PCR assay has been developed to measure human herpesvirus 6 (HHV-6) DNA in biological specimens. The assay sensitivity was 10 copies of DNA per well, with a linear dynamic range of 10 to 10(7) copies of HHV-6 DNA. Intra- and interassay variations were, respectively, 0.88 and 0.8% for samples containing 10(2) DNA copies, 0.99 and 0.96% for samples containing 10(4) copies, and 0.76 and 0.9% for samples containing 10(6) copies. Among 34 saliva samples from healthy subjects, 26 were found to contain HHV-6 DNA (76.5%; median, 23,870 copies/ml), and following a single freeze-thaw cycle, 25 of the same samples were found to be positive for HHV-6 DNA, although at a statistically significantly lower concentration (median, 3,497 copies/ml). The assay enabled detection of HHV-6 DNA in lymph node biopsies from patients with Hodgkin's disease (HD) (13 of 37 patients [35.1%]), B-cell neoplasms (8 of 36 patients [22.2%]), and T- or NK-cell neoplasms (3 of 13 patients [23.1%]), with concentrations ranging from 100 to 864,640 HHV-6 copies per microg of DNA (HHV-6B being found in every case except two). All HD patients infected with HHV-6 presented clinically with the nodular sclerosis subtype of HD. The real-time quantitative PCR assay developed here was simple to perform and was sensitive over a wide range of HHV-6 concentrations. It therefore appears to be of potential value in clinical investigation or diagnosis of HHV-6 infection.
Collot
S
S
Department of Virology, Limoges University Teaching Hospital, Limoges, France.
Petit
B
B
Bordessoule
D
D
Alain
S
S
Touati
M
M
Denis
F
F
Ranger-Rogez
S
S
D000328:Adult; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000032:analysis* / Q000235:genetics*; D016679:Genome, Viral; D015654:Herpesvirus 6, Human / Q000235:genetics* / Q000302:isolation & purification*; D006689:Hodgkin Disease / Q000821:virology; D006801:Humans; D015448:Leukemia, B-Cell / Q000821:virology; D008198:Lymph Nodes / Q000821:virology*; D008223:Lymphoma / Q000821:virology; D016393:Lymphoma, B-Cell / Q000821:virology; D016399:Lymphoma, T-Cell / Q000821:virology; D008232:Lymphoproliferative Disorders / Q000821:virology; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards / Q000706:statistics & numerical data; D012015:Reference Standards; D019349:Roseolovirus Infections / Q000175:diagnosis / Q000821:virology; D012463:Saliva / Q000821:virology*; D012680:Sensitivity and Specificity
D017931:DNA Primers; D004279:DNA, Viral