Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2002 Oct;76(19):9645-56.
doi: 10.1128/jvi.76.19.9645-9656.2002.

Identification of Epstein-Barr virus strain variants in hairy leukoplakia and peripheral blood by use of a heteroduplex tracking assay

Diane Sitki-Green  1 , Rachel H Edwards, Jennifer Webster-Cyriaque, Nancy Raab-Traub
Affiliations

Identification of Epstein-Barr virus strain variants in hairy leukoplakia and peripheral blood by use of a heteroduplex tracking assay

Diane Sitki-Green et al. J Virol. 2002 Oct.

Abstract

Epstein-Barr virus (EBV) strains can be distinguished by specific sequence variations in the LMP1 gene. In this study, a heteroduplex tracking assay (HTA) specific for LMP1 was developed to precisely identify the prototypic undeleted strain B958, other undeleted strains (Ch2, AL, NC, and Med-), and strains with the 30-bp deletion (Med+ and Ch1). This technique also provides an estimate of the relative abundance of strains in patient samples. In this study, EBV strains were identified in 25 hairy leukoplakia (HLP) biopsies and six matched peripheral blood samples and throat washes with the LMP1-HTA. To investigate the relationship of the virus found in the peripheral blood to that in the HLP lesion, the strain variants in the peripheral blood B lymphocytes and those present within the epithelial cells in the HLP lesion and in throat washes were identified. In many of the subjects, compartmental differences in the EBV strain profiles in the oral cavity and peripheral blood were readily apparent. The throat wash specimens usually had a strain profile similar to that within the corresponding HLP sample, which was distinct from the strain profile detected in the peripheral blood. These analyses reveal that the nature of EBV infection can be very dynamic, with changes in relative strain abundance over time as well as the appearance of new strains. The patterns of abundance in the blood and oral cavity provide evidence for compartmentalization and for the transmission of strains between the blood and oropharynx.

PubMed Disclaimer

Figures

FIG. 1.
FIG. 1.
EBV gene coordinates 168163 to 168427 used in this study to identify LMP1 strain variants. The locations of the primers that were used for PCR amplification are indicated. Sequence changes relative to the prototype B958 sequence are labeled for each strain. Signature changes for each strain are in bold. The slash marks at positions 265 to 294 denote the 30-bp deletion.
FIG. 2.
FIG. 2.
Schematic of the LMP1-HTA. The first step included mixing of the PCR-amplified LMP1 and the radiolabeled LMP1 probe of the same region. The mixture was denatured and allowed to reanneal. The resulting product contained heteroduplexes and homoduplexes. The asterisk denotes the radiolabeled strand of DNA.
FIG. 3.
FIG. 3.
Comparison of migration of heteroduplexes with undeleted and deleted probes. PCR-amplified control strains B958, Ch2, AL, NC, Med−, and Med+ were hybridized to (A) the undeleted Ch2 probe and (B) the deleted Med+ probe. Each lane contained the linearized vector, excess single-stranded (ss) probe, and probe homoduplex. The lanes labeled probe contained only probe in the hybridization reaction.
FIG. 4.
FIG. 4.
Migration patterns of undeleted control strain heteroduplexes bound to each strain probe. (A) B958 probe, (B) Med− probe, (C) NC probe, (D) AL probe, (E) Ch2 probe, and (F) Med+ probe. The Ch1 probe yielded a pattern identical to that with the Med+ probe.
FIG. 5.
FIG. 5.
Migration patterns of deleted strain heteroduplexes. The strains used as the probe for each panel were (A) B958, (B) Med+, and (C) Ch1. The Ch2, NC, and Med− probes produced a heteroduplex pattern for the deleted strains similar to that represented in panel A. The AL probe produced a heteroduplex pattern for the deleted strains similar to that represented in panel B.
FIG. 6.
FIG. 6.
Detection limitations for the Med+ and Ch2 probes were determined by mixing various strain DNAs at 1, 10, 50, 90, and 99% of the total template, which were then subjected to PCR and HTA. (A) The Ch2 probe hybridized to decreasing amounts of Ch2 and increasing amounts of Med+. (B) The Med+ probe hybridized to decreasing amounts of B958 and increasing amounts of Med+.
FIG. 7.
FIG. 7.
Examples of LMP1-HTA patterns for HLP biopsies from five subjects (12 to 16) analyzed with the Med+ probe. Only the regions of the gel that contained heteroduplexes are shown.
FIG. 8.
FIG. 8.
(A and B) LMP1-HTA (Med+ probe) of EBV strain profiles from the HLP lesion (H), the peripheral blood (P), and throat wash (T) samples of subjects 4 (A) and 6 (B). (C and D) EBV strain profiles from the HLP lesion (H) and the peripheral blood (P) samples of subjects 5 (C) and 3 (D).
FIG. 9.
FIG. 9.
LMP1-HTA (Med+ probe) of EBV strain profiles from two time points for the same subjects. (A) Subject 1. HLP (H), peripheral blood (P1), and throat wash (T1) samples from the first collection, and normal tongue scraping (TS2), peripheral blood (P2), and throat wash (T2) samples from the second collection 27 months later. (B) Subject 2. HLP (H), peripheral blood (P1), and oral lymphoma (L) samples from the first collection and peripheral blood (P2) sample 2 months later.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Abdel-Hamid, M., J. J. Chen, N. Constantine, M. Massoud, and N. Raab-Traub. 1992. EBV strain variation: geographical distribution and relation to disease state. Virology 190:168-175. - PubMed
    1. Anagnostopoulos, I., M. Hummel, C. Kreschel, and H. Stein. 1995. Morphology, immunophenotype, and distribution of latently and/or productively Epstein-Barr virus-infected cells in acute infectious mononucleosis: implications for the interindividual infection route of Epstein-Barr virus. Blood 85:744-750. - PubMed
    1. Babcock, G. J., L. L. Decker, M. Volk, and D. A. Thorley-Lawson. 1998. EBV persistence in memory B cells in vivo. Immunity 9:395-404. - PubMed
    1. Berger, C., D. van Baarle, M. J. Kersten, M. R. Klein, A. S. Al-Homsi, B. Dunn, C. McQuain, R. van Oers, and H. Knecht. 1999. Carboxy terminal variants of Epstein-Barr virus-encoded latent membrane protein 1 during long-term human immunodeficiency virus infection: reliable markers for individual strain identification. J. Infect. Dis. 179:240-244. - PubMed
    1. Bhatia, K., A. Raj, M. I. Guitierrez, J. G. Judde, G. Spangler, H. Venkatesh, and I. T. Magrath. 1996. Variation in the sequence of Epstein Barr virus nuclear antigen 1 in normal peripheral blood lymphocytes and in Burkitt's lymphomas. Oncogene 13:177-181. - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources