Routine use of real-time quantitative PCR for laboratory diagnosis of Epstein-Barr virus infections.

Abstract

Real-time quantitative polymerase chain reaction (PCR) on the LightCycler instrument (LC-PCR) was developed to measure the Epstein-Barr virus (EBV) load in clinical samples. LC-PCR detected two copies of the EBV genome per 500 ng of DNA. Its specificity was confirmed by assays in EBV-negative cell lines, other human herpesviruses and EBV-seronegative individuals. Excellent inter-assay reproducibility of LC-PCR was obtained in 43 samples (r = 0.983). LC-PCR results were compared with a routinely used ELISA-PCR of 150 samples and a good correlation was found (r = 0.956). A total of 88 individuals were studied, including healthy EBV-seropositive adults (n = 32), patients with EBV-associated disease (n = 34), and HIV-infected patients (n = 22); 37.5% of PBMC samples from healthy individuals contained EBV DNA, while no serum sample was positive. The viral load was significantly higher in PBMCS and saliva specimens in patients recently infected with HIV (19 and 39,400 copies/microg DNA, respectively), as well as in AIDS patients (122 and 331,130 copies/microg DNA) than in the control population (0 and 35 copies/microg DNA). This study confirmed that EBV load measurement with LC-PCR is helpful in the management of EBV-related post-transplantation lymphoproliferative disorders and probably of EBV-associated primary central nervous system B-cell lymphoma.