BackgroundReactivation of Epstein-Barr virus (EBV) infection may cause serious, life-threatening complications in immunocompromised individuals. EBV DNA is often detected in EBV-associated disease states, with viral load believed to be a reflection of virus activity. Two separate real-time quantitative polymerase chain reaction (QPCR) assays using SYBR Green I dye and a single quantification standard containing two EBV genes, Epstein-Barr nuclear antigen-1 (EBNA-1) and BamHI fragment H rightward open reading frame-1 (BHRF-1), were developed to detect and measure absolute EBV DNA load in patients with various EBV-associated diseases. EBV DNA loads and viral capsid antigen (VCA) IgG antibody titres were also quantified on a population sample.ResultsEBV DNA was measurable in ethylenediaminetetraacetic acid (EDTA) whole blood, peripheral blood mononuclear cells (PBMCs), plasma and cerebrospinal fluid (CSF) samples. EBV DNA loads were detectable from 8.0 × 102 to 1.3 × 108 copies/ml in post-transplant lymphoproliferative disease (n = 5), 1.5 × 103 to 2.0 × 105 copies/ml in infectious mononucleosis (n = 7), 7.5 × 104 to 1.1 × 105 copies/ml in EBV-associated haemophagocytic syndrome (n = 1), 2.0 × 102 to 5.6 × 103 copies/ml in HIV-infected patients (n = 12), and 2.0 × 102 to 9.1 × 104 copies/ml in the population sample (n = 218). EBNA-1 and BHRF-1 DNA were detected in 11.0% and 21.6% of the population sample respectively. There was a modest correlation between VCA IgG antibody titre and BHRF-1 DNA load (rho = 0.13, p = 0.05) but not EBNA-1 DNA load (rho = 0.11, p = 0.11).ConclusionTwo sensitive and specific real-time PCR assays using SYBR Green I dye and a single quantification standard containing two EBV DNA targets, were developed for the detection and measurement of EBV DNA load in a variety of clinical samples. These assays have application in the investigation of EBV-related illnesses in immunocompromised individuals.

Objectives: To identify Epstein-Barr virus (EBV) genotypes and strains in samples from individuals with and without a first diagnosis of central nervous system (CNS) demyelinating disease (a possible precursor to multiple sclerosis) and patients with EBV-associated diseases in Australia. Methods: Samples from 55 EBV DNA and serology positive subjects including individuals with (n = 17) and without (n = 21) a first clinical diagnosis of CNS demyelination and patients with EBV-related diseases (n = 17) were examined. EBV genotype and strain were identified by sequence mutations within the Epstein-Barr nuclear antigen-2 region (EBNA-2) using DNA sequence analysis. Results: Both EBV genotypes, A and B, were detected (genotype A, 54/55, 98.2%; genotype B, 1/55, 1.8%). Within genotype A, GD1 was the most commonly detected strain (42/54, 77.7%), followed by B95-8 (9/54, 16.7%) and M-ABA (3/54, 5.6%). Genotype B, strain AG876, was found in one individual with CNS demyelinating disease. Conclusions: EBV genotype A and the GD1 strain were the common EBV genotypes isolated from individuals with and without CNS demyelinating disease, and in subjects with various EBV-related diseases. Although disease-specific genotypes or strains were not identified, this study provides useful insights into the molecular epidemiology of EBV infection in Australia.

Primary infection with varicella zoster virus (VZV) occurs in immunocompromised and immunocompetent individuals. Clinical and asymptomatic reactivation with shedding of infectious virus and viremia may occur. The prevalence of VZV viremia is unknown. The aim of this study was to detect VZV viremia and quantify VZV DNA using quantitative polymerase chain reaction (qPCR) in blood from different populations. A qPCR-based method using EvaGreen® was used to quantify VZV DNA in 491 samples, including whole blood, plasma and buffy-coat, from patients hospitalized with varicella-associated disease (Group 1, n=10) and three groups with no VZV disease: individuals with a first clinical diagnosis of central nervous system demyelination (Group 2, n=213) with their age and sex-matched controls (Group 3, n=218); and HIV-infected individuals (Group 4, n=50). VZV-specific IgG antibody titres were measured in Group 3. The proportion positive for viremia and mean detectable VZV DNA load (copies/ml) were: Group 1: 100% (10/10) and 4.6 × 10(6) ± 1.4 × 10(7) ; Group 2: 4% (9/213) and 1.5 × 10(3) ± 1.8 × 10(4) ; Group 3: 8% (17/218) and 1.1 × 10(3) ± 7.8 × 10(3) ; Group 4: 12% (6/50) and 7.7 × 10(1) ± 2.8 × 10(2) . VZV DNA load and IgG titres were not significantly correlated (Group 3 only). VZV load in Group 1 was significantly elevated compared to Groups 2-4 (P<0.001); the latter were not significantly different from each other (P=0.05). VZV genotypes from clades 1-5 were identified in Group 1. VZV DNA was detected but at low frequency and viral load in both immunocompetent and immunocompromised individuals asymptomatic for VZV infection, compared to individuals with active VZV infection.