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  • Sequence variation of the Epstein-Barr virus nuclear antigen 1 (EBNA1) gene in chronic lymphocytic leukemia and healthy volunteer subjects

  • Zahra Vafapour,1,* Fatemeh Hosseini Tabatabaie,2 Seyed Younes Hosseini,3 Shirin Haghighat,4 Seyed Mohammad Ali Hashemi,5 Jamal Sarvari,6
    1. Department of Bacteriology and Virology, School of Medicine, Shiraz University of Medical Sciences
    2. Department of Bacteriology and Virology, School of Medicine, Shiraz University of Medical Sciences
    3. Department of Bacteriology and Virology, School of Medicine, Shiraz University of Medical Sciences
    4. Department of Hematology and Medical Oncology, Shiraz University of Medical Sciences, Shiraz, Iran
    5. Department of Bacteriology and Virology, School of Medicine, Shiraz University of Medical Sciences
    6. Department of Bacteriology and Virology, School of Medicine, Shiraz University of Medical Sciences


  • Introduction: Epstein-Barr virus (EBV) establishes persistent infection in B cells, and its nuclear antigen 1 (EBNA1) supports viral genome maintenance and gene regulation. Although EBNA1 sequence diversity has been investigated in several malignancies, its relevance to chronic lymphocytic leukemia (CLL) remains unclear. This study characterized variation in the C-terminal DNA-binding/dimerization and USP7-binding regions of EBNA1 in patients with CLL and healthy volunteers.
  • Methods: The study included 40 patients with CLL and 21 age- and sex-matched healthy volunteers recruited during 2019–2020. DNA extracted from buffy-coat samples was screened for EBV by PCR targeting BHRF1. EBV-positive samples underwent nested PCR amplification of the EBNA1 C-terminal region, followed by Sanger sequencing. Sequences were aligned and analyzed using MEGA11, with B95.8 as the reference strain. EBNA1 subtypes were classified according to amino acid residue 487. Group comparisons used Fisher’s exact test in SPSS version 26.
  • Results: EBV DNA was detected in 21/40 CLL samples and 14/21 control samples, without a significant difference between groups (P > 0.05). EBNA1 sequences were obtained from 13 patients and 12 controls. C-terminal sequence variants were found in 5/13 CLL samples (38.5%) and 2/12 controls (16.7%; P = 0.378). The synonymous G97320A substitution, corresponding to residue 553, occurred in five CLL samples and no controls (reported P = 0.039). Among CLL sequences, P-ala, P-thr, and V-val subtypes accounted for 10/13 (76.9%), 2/13 (15.4%), and 1/13 (7.7%), respectively. All 12 control sequences belonged to P-ala. The overall subtype distribution did not differ significantly between groups (P = 0.207).
  • Conclusion: EBNA1 sequence diversity was observed in both groups, with G97320A and the P-thr and V-val subtypes detected only in CLL samples in this cohort. These exploratory findings do not establish a causal role in CLL. Larger studies and functional analyses are needed to assess their reproducibility and biological significance.
  • Keywords: Epstein-Barr virus; CLL; Sequence variation; EBV-associated cancer

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