Integrated EBV and host DNA methylation profiling identifies distinct candidate EBV–host junction patterns in chronic active EBV infection

Chronic active Epstein–Barr virus (EBV) infection (CAEBV) is a potentially life-threatening lymphoproliferative disease with limited effective therapies. Host–virus interactions, including epigenetic regulation, may contribute to CAEBV progression. The methylation landscape of EBV and the host genome, and the genomic features of EBV–host junctions in CAEBV remain incompletely characterized. We profiled EBV DNA methylation in peripheral blood mononuclear cells (PBMCs) from patients with CAEBV and infectious mononucleosis (IM) using targeted bisulfite sequencing. Host DNA methylation patterns were assessed by reduced representation bisulfite sequencing in CAEBV, IM, healthy EBV carriers, and EBV-uninfected controls. In parallel, candidate EBV–host junctions were identified by liquid-phase hybridization capture sequencing. Because same-sample junction-specific PCR and Sanger sequencing were not available, the capture-derived junctions were regarded as candidate events. The EBV genome showed global hypermethylation in CAEBV compared with IM, together with distinct latency-associated promoter methylation patterns. CAEBV also exhibited a distinct host methylation signature relative to control groups, with differentially methylated genes enriched in immune-related processes and cancer-associated pathways. Candidate EBV–host junctions were detected more frequently in CAEBV than in IM, with viral-side regions showing recurrent candidate junction detection, including EBNA-LP , EBNA1 , and BZLF1 . In CAEBV, candidate junctions were enriched in intronic and promoter regions, common fragile sites, and repetitive elements. At the gene level, recurrently targeted host loci included genes implicated in genomic instability and cancer-related pathways. In exploratory longitudinal analyses, allogeneic hematopoietic stem cell transplantation was associated with lower EBV methylation levels and fewer detectable candidate EBV–host junctions, whereas chemotherapy alone was not associated with comparable changes. These data characterize EBV and host methylation landscapes together with candidate EBV–host junction patterns in CAEBV. The findings provide epigenetic and genomic clues to CAEBV pathogenesis and generate hypotheses regarding potential biomarkers of treatment response, but require validation in larger, independent cohorts.

IMPORTANCE

Chronic active Epstein–Barr virus (CAEBV) infection is a rare but severe disease that can progress to life-threatening inflammation or lymphoma, yet its molecular basis remains poorly understood. We found that both the EBV genome and the host genome exhibit distinct DNA methylation patterns in children with CAEBV compared with infectious mononucleosis. We also identified more frequent candidate EBV–host junction signals in CAEBV and observed that these signals became less detectable after successful hematopoietic stem cell transplantation. These findings improve our understanding of host–virus interactions in CAEBV and provide candidate molecular features that may be useful for future studies of disease mechanisms and treatment monitoring.