Quantification of Epstein‐Barr virus load in peripheral blood of human immunodeficiency virus‐infected patients using real‐time PCR

Abstract

Epstein-Barr virus (EBV) reactivation is more likely to occur in immunocompromised patients with subsequent higher susceptibility to EBV-associated lymphoproliferations. In contrast to transplant recipients, limited data are available concerning the EBV load in HIV-infected patients, with or without AIDS-related non-Hodgkin's lymphomas. We developed a TaqMan real-time PCR assay, allowing both the EBV genome and a cellular gene to be quantified in order to obtain a reliable normalized measurement of the EBV load in peripheral blood mononuclear cells (PBMCs). With a wide 6-log10 quantification range and inter-assay variations of less than 24%, this quantitative PCR was sufficiently accurate and reproducible for routine follow-up. The EBV load was determined in PBMCs from 113 HIV-infected patients, 11 patients with primary HIV infection and 24 HIV-seronegative healthy controls. The rates of EBV detection were similar in the three groups. However, EBV loads were higher in the HIV-infected group (P < 0.00001) except for the patients with primary HIV infection. Unexpectedly, EBV loads were not correlated with the clinical stages of HIV infection or HIV replication, and did not depend on the degree of immunodepression, as judged by CD4+ counts. This study contributes towards the definition of the baseline EBV load during HIV infection and stresses the broad inter-individual variability of the EBV load in HIV-infected patients. Real-time PCR provides a useful tool that can be used in further longitudinal studies to assess the relevance of the EBV load to identify HIV-infected patients with a high risk of EBV-associated lymphoproliferations. J. Med. Virol. 65:543–552, 2001. © 2001 Wiley-Liss, Inc.