Toward Standardization of Epstein-Barr Virus DNA Load Monitoring: Unfractionated Whole Blood As Preferred Clinical Specimen

Overview

Journal J Clin Microbiol

Specialty Microbiology

Date 2001 Apr 3

PMID 11283029

Citations 42

Authors

S J Stevens

I Pronk

J M Middeldorp

Affiliations

Soon will be listed here.

Abstract

Epstein-Barr virus (EBV) DNA load monitoring in peripheral blood has been shown to be a useful tool for the diagnosis of aberrant EBV infections. In the present study we compared the relative diagnostic values of EBV DNA load monitoring in unfractionated whole blood and simultaneously obtained serum or plasma samples from Burkitt's lymphoma (BL) patients, transplant recipients, human immunodeficiency virus (HIV)-infected individuals, and infectious mononucleosis (IM) patients by a quantitative competitive PCR (Q-PCR). The EBV DNA load in BL patients was mainly situated in the cellular blood compartment (up to 4.5 x 10(6) copies/ml). EBV DNA loads in unfractionated whole blood and parallel serum samples showed no correlation. In transplant recipients, IM patients, and HIV-infected patients, the EBV burden in the circulation was almost exclusively restricted to the cellular blood compartment, because serum or plasma samples from these patients yielded negative results by Q-PCR, despite high viral loads in corresponding whole-blood samples. A 10-fold more sensitive but qualitative BamHI-W-repeat PCR occasionally revealed the presence of EBV at <2,000 copies of EBV DNA per ml of serum. Spiking of 100 copies of EBV DNA in samples with negative Q-PCR results excluded the presence of inhibitory factors in serum or plasma that influenced the Q-PCR result. Serum samples from all populations were often positive for beta-globin DNA, indicating cell damage in vivo or during serum preparation. We conclude that serum is an undesirable clinical specimen for EBV DNA load monitoring because it omits the presence of cell-associated virus and uncontrolled cell lysis may give irreproducible results or overestimation of the DNA load. Unfractionated whole blood is strongly preferred since it combines all blood compartments that may harbor EBV and it best reflects the absolute viral burden in the patient's circulation.

Citing Articles

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Worldwide Prevalence of Epstein-Barr Virus in Patients with Burkitt Lymphoma: A Systematic Review and Meta-Analysis.

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References

Lo Y, Chan L, Chan A, Leung S, Lo K, Zhang J . Quantitative and temporal correlation between circulating cell-free Epstein-Barr virus DNA and tumor recurrence in nasopharyngeal carcinoma. Cancer Res. 1999; 59(21):5452-5. View

Babcock G, Decker L, Freeman R, Thorley-Lawson D . Epstein-barr virus-infected resting memory B cells, not proliferating lymphoblasts, accumulate in the peripheral blood of immunosuppressed patients. J Exp Med. 1999; 190(4):567-76. PMC: 2195601. DOI: 10.1084/jem.190.4.567. View

Gustafsson A, Levitsky V, Zou J, Frisan T, Dalianis T, Ljungman P . Epstein-Barr virus (EBV) load in bone marrow transplant recipients at risk to develop posttransplant lymphoproliferative disease: prophylactic infusion of EBV-specific cytotoxic T cells. Blood. 2000; 95(3):807-14. View

Baldanti F, Grossi P, Furione M, Simoncini L, Sarasini A, Comoli P . High levels of Epstein-Barr virus DNA in blood of solid-organ transplant recipients and their value in predicting posttransplant lymphoproliferative disorders. J Clin Microbiol. 2000; 38(2):613-9. PMC: 86158. DOI: 10.1128/JCM.38.2.613-619.2000. View

Niesters H, van ESSER J, Fries E, Wolthers K, Cornelissen J, Osterhaus A . Development of a real-time quantitative assay for detection of Epstein-Barr virus. J Clin Microbiol. 2000; 38(2):712-5. PMC: 86184. DOI: 10.1128/JCM.38.2.712-715.2000. View

Rickinson A, Lane P . Epstein-Barr virus: Co-opting B-cell memory and migration. Curr Biol. 2000; 10(3):R120-3. DOI: 10.1016/s0960-9822(00)00308-0. View

Shotelersuk K, Khorprasert C, Sakdikul S, Pornthanakasem W, Voravud N, Mutirangura A . Epstein-Barr virus DNA in serum/plasma as a tumor marker for nasopharyngeal cancer. Clin Cancer Res. 2000; 6(3):1046-51. View

Gerna G, Baldanti F, Lilleri D, Parea M, Alessandrino E, Pagani A . Human cytomegalovirus immediate-early mRNA detection by nucleic acid sequence-based amplification as a new parameter for preemptive therapy in bone marrow transplant recipients. J Clin Microbiol. 2000; 38(5):1845-53. PMC: 86605. DOI: 10.1128/JCM.38.5.1845-1853.2000. View

Li J, Huang D, Sun B, Zhang X, Middeldorp J, Klamut H . Efficacy of ionizing radiation combined with adenoviral p53 therapy in EBV-positive nasopharyngeal carcinoma. Int J Cancer. 2000; 87(4):606-10. View

10.

Stevens S, Verschuuren E, Pronk I, van der Bij W, Harmsen M, The T . Frequent monitoring of Epstein-Barr virus DNA load in unfractionated whole blood is essential for early detection of posttransplant lymphoproliferative disease in high-risk patients. Blood. 2001; 97(5):1165-71. DOI: 10.1182/blood.v97.5.1165. View

11.

Yao Q, Ogan P, Rowe M, Wood M, Rickinson A . Epstein-Barr virus-infected B cells persist in the circulation of acyclovir-treated virus carriers. Int J Cancer. 1989; 43(1):67-71. DOI: 10.1002/ijc.2910430115. View

12.

Kwok S, Higuchi R . Avoiding false positives with PCR. Nature. 1989; 339(6221):237-8. DOI: 10.1038/339237a0. View

13.

BOOM R, Sol C, Salimans M, Jansen C, van der Noordaa J . Rapid and simple method for purification of nucleic acids. J Clin Microbiol. 1990; 28(3):495-503. PMC: 269651. DOI: 10.1128/jcm.28.3.495-503.1990. View

14.

Jiwa N, Kanavaros P, van der Valk P, Walboomers J, Horstman A, Vos W . Expression of c-myc and bcl-2 oncogene products in Reed-Sternberg cells independent of presence of Epstein-Barr virus. J Clin Pathol. 1993; 46(3):211-7. PMC: 501172. DOI: 10.1136/jcp.46.3.211. View

15.

van Grunsven W, Nabbe A, Middeldorp J . Identification and molecular characterization of two diagnostically relevant marker proteins of the Epstein-Barr virus capsid antigen complex. J Med Virol. 1993; 40(2):161-9. DOI: 10.1002/jmv.1890400215. View

16.

Fournie G, Martres F, Pourrat J, Alary C, Rumeau M . Plasma DNA as cell death marker in elderly patients. Gerontology. 1993; 39(4):215-21. DOI: 10.1159/000213536. View

17.

Savoie A, Perpete C, Carpentier L, Joncas J, Alfieri C . Direct correlation between the load of Epstein-Barr virus-infected lymphocytes in the peripheral blood of pediatric transplant patients and risk of lymphoproliferative disease. Blood. 1994; 83(9):2715-22. View

18.

Gan Y, Sullivan J, Sixbey J . Detection of cell-free Epstein-Barr virus DNA in serum during acute infectious mononucleosis. J Infect Dis. 1994; 170(2):436-9. DOI: 10.1093/infdis/170.2.436. View

19.

Riddler S, Breinig M, Mcknight J . Increased levels of circulating Epstein-Barr virus (EBV)-infected lymphocytes and decreased EBV nuclear antigen antibody responses are associated with the development of posttransplant lymphoproliferative disease in solid-organ transplant recipients. Blood. 1994; 84(3):972-84. View

20.

Yamamoto M, Kimura H, Hironaka T, Hirai K, Hasegawa S, Kuzushima K . Detection and quantification of virus DNA in plasma of patients with Epstein-Barr virus-associated diseases. J Clin Microbiol. 1995; 33(7):1765-8. PMC: 228265. DOI: 10.1128/jcm.33.7.1765-1768.1995. View