Abstract
Epstein-Barr virus (EBV) is strongly implicated as an essential environmental trigger of multiple sclerosis (MS), yet the host genetic mechanisms governing EBV activity and how infection triggers the disease are not known. We developed a pipeline to quantify EBV DNA from whole-genome sequencing data and applied it to population-scale cohorts. Using this pipeline, we performed a cross-ancestry genome-wide association study (GWAS) of EBV DNA positivity in 617,186 individuals and identified 39 independent susceptibility risk loci, with the strongest associations in the HLA region. We validated this finding in our independent cohort (N=94) and found that quantitative PCR (qPCR)-confirmed EBV DNA positive individuals were enriched in the top decile of EBV polygenic risk scores (PRS) containing newly discovered loci. A significant overlap with genetic variants associated with MS risk was observed. PRS and Mendelian randomization analyses further supported a causal role of EBV activity on MS risk, which was also seen in other autoimmune diseases. A meta-analysis of qPCR based case-control studies showed elevated EBV DNA positivity in MS. By establishing a single-cell RNA-seq method optimized for EBV detection, we identified EBV-infected B cells, primarily in memory B cells, atypical B cells and antibody-secreting cells from MS and healthy individuals. Notably, EBV-infected memory B cells and atypical B cells showed strong upregulation of cytokines and costimulatory signals that influence T cell activation, IFNg secreting Tregs, and regulators of B cell differentiation and survival. EBV-infected memory B cells also upregulated risk genes from both the EBV and MS GWAS, suggesting that EBV-infected B cells constitute a critical hub that modulates T cell responses while simultaneously activating MS susceptibility pathways within the B cell compartment. Together, these findings define a genetic and cellular framework linking EBV infection to the initiation of MS.