Epstein–Barr Virus in Multiple Sclerosis: Theory and Emerging Immunotherapies
Authors: Amit Bar-Or, Michael P. Pender, Rajiv Khanna, Lawrence Steinman, Hans-Peter Hartung, Tap Maniar, Ed Croze, Blake T. Aftab, Gavin Giovannoni, Manher A. Joshi
Journal: Trends in Molecular Medicine (2019)

Clinical studies show that depletion of B cells reduces disease burden in both relapsing-remitting and progressive multiple sclerosis (MS) patients.B cell-tropic viruses may trigger aberrant immune responses in MS in genetically susceptible individuals owing, in part, to a failure in viral surveillance and clearance.The most compelling data supporting an etiologic role for viral involvement in MS have emerged for Epstein–Barr virus (EBV).Targeting mechanisms by which EBV is thought to participate in MS pathogenesis provides an opportunity for new drug development in MS. New treatments for multiple sclerosis (MS) focused on B cells have created an atmosphere of excitement in the MS community. B cells are now known to play a major role in disease, demonstrated by the highly impactful effect of a B cell-depleting antibody on controlling MS. The idea that a virus may play a role in the development of MS has a long history and is supported mostly by studies demonstrating a link between B cell-tropic Epstein–Barr virus (EBV) and disease onset. Efforts to develop antiviral strategies for treating MS are underway. Although gaps remain in our understanding of the etiology of MS, the role, if any, of viruses in propagating pathogenic immune responses deserves attention. New treatments for multiple sclerosis (MS) focused on B cells have created an atmosphere of excitement in the MS community. B cells are now known to play a major role in disease, demonstrated by the highly impactful effect of a B cell-depleting antibody on controlling MS. The idea that a virus may play a role in the development of MS has a long history and is supported mostly by studies demonstrating a link between B cell-tropic Epstein–Barr virus (EBV) and disease onset. Efforts to develop antiviral strategies for treating MS are underway. Although gaps remain in our understanding of the etiology of MS, the role, if any, of viruses in propagating pathogenic immune responses deserves attention. MS is a chronic immune-mediated disease with a complex etiology involving a dysregulated immune system with bouts of peripherally mediated inflammation, as well as ongoing central nervous system (CNS)-compartmentalized inflammation leading to loss of neural tissue and worsening disability [1Hauser S.L. et al.Multiple sclerosis: prospects and promise.Ann. Neurol. 2013; 74: 317-327Crossref PubMed Scopus (120) Google Scholar,2Weiner H.L. Multiple sclerosis is an inflammatory T-cell-mediated autoimmune disease.Arch. Neurol. 2004; 61: 1613-1635Crossref PubMed Scopus (164) Google Scholar]. Intermittent waves of aberrant regulation and/or activation of immune cell subsets result in their trafficking and perivascular infiltration across the blood–brain barrier (BBB) into the CNS where immune cells become reactivated and impact on the underlying tissue resulting in disease relapses. The biological underpinnings of nonrelapsing progressive MS are not well understood but are thought, at least in part, to be driven by CNS-compartmentalized inflammation involving persistence of immune cells and their activation both around perivascular lesions and in the meninges. With respect to the etiology of MS, both genetic susceptibility and environmental exposures are thought to be involved [3Baranzini S.E. Oksenberg J.R. The genetics of multiple sclerosis: from 0 to 200 in 50 years.Trends Genet. 2017; 33: 960-970Abstract Full Text Full Text PDF PubMed Scopus (43) Google Scholar, 4Ikram M.A. et al.Genetic susceptibility to multiple sclerosis: brain structure and cognitive function in the general population.Mult. Scler. 2017; 23: 1697-1706Crossref PubMed Scopus (0) Google Scholar, 5Jokubaitis V.G. Butzkueven H. A genetic basis for multiple sclerosis severity: red herring or real?.Mol. Cell. Probes. 2016; 30: 357-365Crossref PubMed Scopus (10) Google Scholar]. The lifetime incidence of MS in the general population ranges from 2.0 to 9.6 per 100 000 patient-years. The concordance rate of MS in genetically identical twins, that ranges from ∼30% in northern populations to 15% or even 6% in countries such as Italy and France, highlights not only the contribution of genetic risk but also the importance of the environment acting on a genetically predisposed host. T cell and, more recently, B cell interactions have been shown to play a crucial role in driving new relapses [6Mulero P. et al.Ocrelizumab: a new milestone in multiple sclerosis therapy.Ther. Adv. Neurol. Disord. 2018; 11 (1756286418773025)Crossref PubMed Scopus (21) Google Scholar, 7Guan Y. et al.The role of Epstein–Barr virus in multiple sclerosis: from molecular pathophysiology to in vivo imaging.Neural Regen. Res. 2019; 14: 373-386Crossref PubMed Scopus (0) Google Scholar, 8Hauser S.L. et al.Ocrelizumab versus interferon beta-1a in relapsing multiple sclerosis.N. Engl. J. Med. 2017; 376: 221-234Crossref PubMed Scopus (387) Google Scholar, 9Bittner S. et al.Targeting B cells in relapsing-remitting multiple sclerosis: from pathophysiology to optimal clinical management.Ther. Adv. Neurol. Disord. 2017; 10: 51-66Crossref PubMed Scopus (19) Google Scholar, 10Bose T. Role of immunological memory cells as a therapeutic target in multiple sclerosis.Brain Sci. 2017; 7: E148Crossref PubMed Scopus (0) Google Scholar]. Highly effective disease control observed using the B cell-depleting antibody ocrelizumab reinforces interest in developing additional B cell-mediated treatments. Building on the success of these studies could include the design and execution of an approach targeting the implicit role of a B cell-tropic virus in MS. Specific environmental exposures are relevant to both triggering MS and modulating disease course. Virus infection is one crucial environmental factor. Of all viruses considered in MS pathogenesis, EBV, a highly B cell-tropic virus, is the best-studied (Table 1). Defective control of EBV is associated with infectious mononucleosis (IM) in addition to Hodgkin's lymphoma, Burkitt's lymphoma, gastric cancer, nasopharyngeal carcinoma, and conditions associated with HIV, such as hairy cell leukemia and other lymphomas [11Sheik-Ali S. Infectious mononucleosis and multiple sclerosis – updated review on associated risk.Mult. Scler. Relat. Disord. 2017; 14: 56-59Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar,12Endriz J. et al.Time correlation between mononucleosis and initial symptoms of MS.Neurol. Neuroimmunol. Neuroinflamm. 2017; 4: e308Crossref PubMed Scopus (6) Google Scholar]. There is also evidence that infection with EBV and associations with transcription factors, implicating EBV nuclear antigen 2 (EBNA2), operating across different disease loci are linked to a higher risk of autoimmune diseases other than MS, including systemic lupus erythematosus, rheumatoid arthritis, and Sjögren's syndrome [13Pender M.P. 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Genet. 2018; 50: 699-707Crossref PubMed Scopus (45) Google Scholar].Table 1Viruses Implicated in MS: Virus, Disease Involvement, and Association with MSVirusDisease involvementAssociation with MSEBV, HHV-4, lymphocryptovirusDouble-stranded (ds) DNA virus, neurotropicTropism: B cells, epithelial cellsCell latency: memory B cellsInfectious mononucleosis, Hodgkin and non-Hodgkin's lymphoma, Burkett's lymphoma, gastric and nasopharyngeal cancer, hairy cell leukemia, MSMononucleosis predisposes to MS, reduced activity of EBV-specific cytotoxic T cells (e.g., exhausted) in MS patients, EBV seropositive-epidemiological studies, virus present in MS brain, EBV prolongs the lifespan of B cellsHHV-6, roseolovirusdsDNA, neurotropicTropism: broad; hematopoietic and epithelial cellsCell latency: lymphocytes and monocytesExanthema subitum (roseola infantum) and pneumonitis, MSPresent in MS plaques, reactivation during relapses, high levels found in oligodendrocytes and areas of demyelination, elevated levels are found early in MS and during relapses/exacerbations, anti-HPV IgG and IgM titers are reported to predict relapsesCMV, betaherpesvirinaedsDNA, neurotropicTropism: broad; hematopoietic cells, smooth muscle, monocytes, epithelial and endothelial cells, fibroblasts, connective tissueCell latency: cells of the myeloid lineageRetinitis, hepatitis, colitis, pneumonia, encephalitis, MSBoth detrimental and beneficial properties reported, large meta-analysis MS versus controls did not yield a conclusive link between CMV and MSVaricella zoster virus (VZV), HHV-3dsDNA, neurotropicTropism: mononuclear cellsCell latency: sensory gangliaChickenpox, shingles, MSVirus is present during relapses, recent studies failed to show an increased risk of MS associated with varicella or zoster infectionsHERV-WTropism: cells of the nervous system, syncytiotrophoblast layer of the placentaCell latency: multipleMS, diabetes, autoimmune arthritis, and schizophrenia. In most cases the observed expression profiles of specific HERV-W sequences have not led to a definitive association with human disease pathologyPresent in infiltrating macrophages and activated MS lesions, MSRV Env protein is detected in blood of active MS patients, drives the expression of proinflammatory cytokines, reduces myelin protein, expression and kills oligodendrocyte precursors Open table in a new tab EBV appears to be involved across the clinical spectrum of MS, including early pediatric-onset MS, established relapsing-remitting (RRMS), and progressive forms (PMS), as well as in patients with both mild and severe disease course (Box 1). Viral-induced animal models of neuroinflammation, demyelination, and neurodegeneration provide additional proof of principle that viruses play a role in autoimmune disease (Box 2). Over the past 20 years, convergent studies from multiple investigators point to a link between EBV infection and the development (i.e., etiology) of MS [17Fernández-Menéndez S. et al.Epstein–Barr virus and multiple sclerosis. From evidence to therapeutic strategies.J. Neurol. Sci. 2016; 361: 213-219Abstract Full Text Full Text PDF PubMed Google Scholar, 18Pender M.P. Burrows S.R. Epstein–Barr virus and multiple sclerosis: potential opportunities for immunotherapy.Clin. Transl. Immunol. 2014; 3: e27Crossref PubMed Google Scholar, 19Pender M.P. The essential role of Epstein–Barr virus in the pathogenesis of multiple sclerosis.Neuroscientist. 2011; 17: 351-367Crossref PubMed Scopus (0) Google Scholar, 20Ascherio A. Munger K.L. Epstein–Barr virus infection and multiple sclerosis: a review.J. Neuroimmune Pharmacol. 2010; 5: 271-277Crossref PubMed Scopus (172) Google Scholar, 21Ascherio A. Munger K.L. Environmental risk factors for multiple sclerosis. Part I. The role of infection.Ann. Neurol. 2007; 61: 288-299Crossref PubMed Scopus (0) Google Scholar]. MS does not develop in the absence of exposure to EBV, and EBV is a 'required', but insufficient on its own, contributor in early disease pathophysiology. Independent support comes from an observed higher rate of EBV activation in MS [22Angelini D.F. et al.Increased CD8+ T cell response to Epstein–Barr virus lytic antigens in the active phase of multiple sclerosis.PLoS Pathog. 2013; 9: e1003220Crossref PubMed Scopus (0) Google Scholar,23Mechelli R. et al.Epstein–Barr virus nuclear antigen-1 B-cell epitopes in multiple sclerosis twins.Mult. Scler. 2011; 17: 1290-1294Crossref PubMed Scopus (0) Google Scholar], a role for EBV in triggering new relapses consistent with a reduced ability of EBV-specific CD8+ T cells (e.g., T cell exhaustion, see Glossary) to limit EBV reactivation in MS patients [24Cencioni M.T. et al.Programmed death 1 is highly expressed on CD8+ CD57+ T cells in patients with stable multiple sclerosis and inhibits their cytotoxic response to Epstein–Barr virus.Immunology. 2017; 152: 660-676Crossref PubMed Scopus (9) Google Scholar,25Pender M.P. et al.Defective T-cell control of Epstein–Barr virus infection in multiple sclerosis.Clin. Transl. Immunol. 2017; 6: e126Crossref PubMed Scopus (15) Google Scholar], and a strong correlation between the presence of anti-EBV antibodies in blood and disease onset [26Deeba E. et al.Evaluation of Epstein–Barr virus-specific antibodies in Cypriot multiple sclerosis patients.Mol. Immunol. 2019; 105: 270-275Crossref PubMed Scopus (2) Google Scholar, 27Mameli G. et al.Expression and activation by Epstein–Barr virus of human endogenous retroviruses-W in blood cells and astrocytes: inference for multiple sclerosis.PLoS One. 2012; 7: e44991Crossref PubMed Scopus (72) Google Scholar, 28Lünemann J.D. et al.Elevated Epstein–Barr virus-encoded nuclear antigen-1 immune responses predict conversion to multiple sclerosis.Ann. Neurol. 2010; 67: 159-169Crossref PubMed Scopus (0) Google Scholar, 29Ascherio A. et al.Epstein–Barr virus antibodies and risk of multiple sclerosis: a prospective study.JAMA. 2001; 286: 3083-3088Crossref PubMed Google Scholar]. Evidence of CNS involvement comes from reports of elevated anti-EBNA1 IgG serology associated with the appearance of new gadolinium (Gd)-enhancing brain lesions [30Tzartos J.S. et al.Association of innate immune activation with latent Epstein–Barr virus in active MS lesions.Neurology. 2011; 78: 15-23Crossref PubMed Scopus (75) Google Scholar] and the presence, in patients, of EBNA1-specific T cells recognizing myelin [31Lünemann J.D. et al.EBNA1-specific T cells from patients with multiple sclerosis cross react with myelin antigens and co-produce IFN-gamma and IL-2.J. Exp. Med. 2008; 205: 1763-1773Crossref PubMed Scopus (0) Google Scholar]. In addition to EBV-infected B cells in the periphery contributing to relapsing disease, EBV may play a crucial role in propagating CNS-compartmentalized inflammation and injury, potentially contributing to progressive (nonrelapsing) aspects of MS. B cells may traffic not only into the CNS but also out of the CNS into deep cervical lymph nodes [32Stern J.N. et al.B cells populating the multiple sclerosis brain mature in the draining cervical lymph nodes.Sci. Transl. Med. 2014; 6: 248ra107Crossref PubMed Scopus (180) Google Scholar,33von Büdingen H.C. et al.B cell exchange across the blood–brain barrier in multiple sclerosis.J. Clin. Invest. 2012; 122: 4533-4543Crossref PubMed Scopus (0) Google Scholar]. These B cells include those infected with EBV, which could then activate aberrant T cell responses in the periphery. One can speculate that EBV from CNS pools may contribute not only to CNS-compartmentalized inflammation but also to further disease relapses. In support of this contention is evidence of antibodies in cerebrospinal fluid (CSF) of MS patients that recognize EBV antigens [34Nociti V. et al.Epstein–Barr virus antibodies in serum and cerebrospinal fluid from multiple sclerosis, chronic inflammatory demyelinating polyradiculoneuropathy and amyotrophic lateral sclerosis.J. Neuroimmunol. 2010; 225: 149-152Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar], and evidence that EBV-infected B cells and plasma cells accumulate in MS brain in meningeal immune-cell collections (Box 3). Although the presence of such EBV-infected B cells in MS brain has not been observed in all studies [35Lassmann H. et al.Epstein–Barr virus in the multiple sclerosis brain: a controversial issue report on a focused workshop held in the Centre for Brain Research of the Medical University of Vienna, Austria.Brain. 2011; 134: 2772-2786Crossref PubMed Scopus (0) Google Scholar], the potential for such cells to participate in germinal center (GC)-like reactions in the meninges could support ongoing immune cell activation and the maintenance of a pool of pathogenic EBV-infected cells in the CNS.Box 1EBV Involvement across the MS SpectrumEvidence supporting an essential role for EBV in MS is derived from studies including pediatric and adult-onset patients with RRMS, patients with SPMS and PMS, as well as aggressive forms of MS. It is therefore worth reviewing the association of EBV across the broad spectrum of MS. In a cohort of 1047 clinically isolated syndrome (CIS) cases only one was seronegative for EBNA-1 [89Dobson R. et al.Epstein–Barr-negative MS: a true phenomenon?.Neurol. Neuroimmunol. Neuroinflamm. 2017; 4: e318Crossref PubMed Scopus (4) Google Scholar]. This study represents the largest EBV-CIS population evaluated to date. In a different study, including CIS, RRMS, and PMS, all had evidence of EBV serology and confirmed diagnosis of MS. Further support comes from a recent study by Gieß et al. [90Gieß R.M. et al.Epstein–Barr virus antibodies in serum and DNA load in saliva are not associated with radiological or clinical disease activity in patients with early multiple sclerosis.PLoS One. 2017; 12: e0175279Crossref PubMed Scopus (4) Google Scholar] reporting association of EBV infection with early-onset MS. EBV serology correlated with early diagnostic conversion from CIS to MS. However, neither EBNA-1 nor viral capsid antigen (VCA) IgG antibodies in serum, nor EBV DNA load in saliva, were associated with radiological or clinical disease activity. EBV infection is strongly associated with pediatric MS [91Nourbakhsh B. et al.Heterogeneity in association of remote herpesvirus infections and pediatric MS.Ann. Clin. Transl. Neurol. 2018; 5: 1222-1228Crossref PubMed Scopus (0) Google Scholar, 92Waubant E. et al.Common viruses associated with lower pediatric multiple sclerosis risk.Neurology. 2011; 76: 1989-1995Crossref PubMed Scopus (84) Google Scholar, 93Banwell B. et al.Clinical features and viral serologies in children with multiple sclerosis: a multinational observational study.Lancet Neurol. 2007; 6: 773-781Abstract Full Text Full Text PDF PubMed Scopus (212) Google Scholar, 94Pohl D. et al.High seroprevalence of Epstein–Barr virus in children with multiple sclerosis.Neurology. 2006; 67: 2063-2065Crossref PubMed Scopus (0) Google Scholar]. Herpes simples virus (HSV)-1 seropositivity was associated with pediatric MS cases negative for HLA-DRB1*15:01, highlighting the complex nature of viral exposure and genetic factors. Multivariate analysis in the same study revealed a reduction in the risk of developing MS associated with CMV infection and no influence on MS status associated with HSV-1 infection [91Nourbakhsh B. et al.Heterogeneity in association of remote herpesvirus infections and pediatric MS.Ann. Clin. Transl. Neurol. 2018; 5: 1222-1228Crossref PubMed Scopus (0) Google Scholar]. Taken together, a role for EBV in early MS is supported by convergent pediatric MS studies. As in adult MS, these studies are consistent with a role for EBV as 'required but insufficient', likely playing one or more key contributing roles across the MS spectrum, intersecting with genetic susceptibility and additional environmental factors.Box 2Virus-Induced Animal Models of Inflammation, Demyelination, and DegenerationAnimal models can be used to explore virus-specific mechanisms contributing to autoimmune and demyelinating diseases including MS [95Lassmann H. Bradl M. Multiple sclerosis: experimental models and reality.Acta Neuropathol. 2017; 133: 223-244Crossref PubMed Scopus (108) Google Scholar, 96Palumbo S. Pellegrini S. Experimental in vivo models of multiple sclerosis: state of the art.in: Zagon I.S. Mclaughlin P.J. Multiple Sclerosis: Perspectives in Treatment and Pathogenesis. Codon Publications, 2017Crossref Google Scholar, 97'tHart B.A. et al.EBV Infection and multiple sclerosis: lessons from a marmoset model.Trends Mol. Med. 2016; 22: 1012-1024Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar]. EBV itself does not infect mice, which has contributed to the challenge of studying the role of EBV in models of CNS inflammation including experimental autoimmune encephalomyelitis (EAE). Nevertheless, the EBV-like virus, murine gammaherpesvirus-68 (gHV-68), exacerbates EAE [98Márquez A.C. Horwitz M.S. The role of latently infected B cells in CNS autoimmunity.Front. Immunol. 2015; 6: 544-562Crossref PubMed Scopus (14) Google Scholar, 99Casiraghi C. et al.Gammaherpesvirus latency accentuates EAE pathogenesis: relevance to Epstein–Barr virus and multiple sclerosis.PLoS Pathog. 2012; 8: e1002715Crossref PubMed Scopus (0) Google Scholar, 100Peacock J.W. et al.Exacerbation of experimental autoimmune encephalomyelitis in rodents infected with murine gammaherpesvirus-68.Eur. J. Immunol. 2003; 33: 1849-1858Crossref PubMed Scopus (43) Google Scholar] and leads to a type I IFN-dependent increase in heparan sulfate and responsiveness to proliferation-inducing ligands, and inhibition of viral reactivation [101Jarousse N. et al.Virally-induced upregulation of heparan sulfate on B cells via the action of type I IFN.J. Immunol. 2011; 187: 5540-5547Crossref PubMed Scopus (6) Google Scholar]. The Theiler's murine encephalomyelitis virus (TMEV) model [95Lassmann H. Bradl M. Multiple sclerosis: experimental models and reality.Acta Neuropathol. 2017; 133: 223-244Crossref PubMed Scopus (108) Google Scholar] correlates infection with late-stage demyelination and entry of TMEV into the CNS [102Gilli F. et al.The effect of B-cell depletion in the Theiler's model of multiple sclerosis.J. Neurol. Sci. 2015; 359: 40-47Abstract Full Text Full Text PDF PubMed Google Scholar,103Martinez N.E. et al.Protective and detrimental roles for regulatory T cells in a viral model for multiple sclerosis.Brain Pathol. 2014; 24: 436-451Crossref PubMed Scopus (0) Google Scholar]. In contrast to MS, B cell depletion in the TMEV model caused worsening of disease, hinting that prolonged B cell depletion might worsen viral infection and progression of disability [102Gilli F. et al.The effect of B-cell depletion in the Theiler's model of multiple sclerosis.J. Neurol. Sci. 2015; 359: 40-47Abstract Full Text Full Text PDF PubMed Google Scholar]. The mouse hepatitis (corona) virus (MHV) model causes a chronic inflammatory demyelinating disease resembling MS [104Bender S.J. Weiss S.R. Pathogenesis of murine coronavirus in the central nervous system.J. Neuroimmune Pharmacol. 2010; 5: 336-354Crossref PubMed Scopus (38) Google Scholar]. In marmoset EAE, infection with endogenous viruses such as EBV or CMV alters immune responses and recruits intensely pathogenic T cells from the anti-effector memory cell population [97'tHart B.A. et al.EBV Infection and multiple sclerosis: lessons from a marmoset model.Trends Mol. Med. 2016; 22: 1012-1024Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar]. EBV-infected B cells mediate disease progression through MHC class Ib (Caja-E)-restricted cytotoxic T cells activated by gammaherpesvirus, causing demyelination of cortical grey matter [105'tHart B.A. et al.The primate autoimmune encephalomyelitis model; a bridge between mouse and man.Ann. Clin. Transl. Neurol. 2015; 2: 581-593Crossref PubMed Google Scholar]. Anti-CD20 antibody causes depletion of EBV-like CalHV3 from lymphoid organs, supporting a key role for CD20+ B cells in MS. The marmoset EAE model of MS suggests that EBV infection leads to increased citrullination of peptides in conjunction with autophagy during antigen presentation, allowing B cells to cross-present autoantigens to CD8+CD56+ T cells and leading to disease progression [97'tHart B.A. et al.EBV Infection and multiple sclerosis: lessons from a marmoset model.Trends Mol. Med. 2016; 22: 1012-1024Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar,106Morandi E. et al.EBV infection empowers human B Cells for autoimmunity: role of autophagy and relevance to multiple sclerosis.J. Immunol. 2017; 199: 435-448Crossref PubMed Scopus (18) Google Scholar]. EBV also upregulated the antigen-presenting machinery of infected B cells and facilitated cross-presentation of immunogenic MOG peptides to CD8+ T cells [107Dunham J. et al.Analysis of the crosstalk of Epstein–Barr virus-infected B cells with T cells in the marmoset.Clin. Transl. Immunology. 2017; 6: e127Crossref PubMed Google Scholar]. In a variety of animal models, EBV-like viruses and EBV itself lead to the development of autoimmune, neurodegenerative, and MS-like disease pathologies.Box 3EBV in MS BrainSeveral studies report detection of EBV-infected B cells and plasma cells in the brain of MS patients [30Tzartos J.S. et al.Association of innate immune activation with latent Epstein–Barr virus in active MS lesions.Neurology. 2011; 78: 15-23Crossref PubMed Scopus (75) Google Scholar,35Lassmann H. et al.Epstein–Barr virus in the multiple sclerosis brain: a controversial issue report on a focused workshop held in the Centre for Brain Research of the Medical University of Vienna, Austria.Brain. 2011; 134: 2772-2786Crossref PubMed Scopus (0) Google Scholar,46Magliozzi R. et al.B-cell enrichment and Epstein–Barr virus infection in inflammatory cortical lesions in secondary progressive multiple sclerosis.J. Neuropathol. Exp. Neurol. 2013; 72: 29-41Crossref PubMed Scopus (0) Google Scholar, 47Serafini B. et al.Dysregulated Epstein–Barr virus infection in the multiple sclerosis brain.J. Exp. Med. 2007; 204: 2899-2912Crossref PubMed Scopus (469) Google Scholar, 48Serafini B. et al.Detection of ectopic B-cell follicles with germinal centers in the meninges of patients with secondary progressive multiple sclerosis.Brain Pathol. 2004; 14: 164-174Crossref PubMed Google Scholar,108Hassani A. et al.Epstein–Barr virus is present in the brain of most cases of multiple sclerosis and may engage more than just B cells.PLoS ONE. 2018; 13: e0192109Crossref PubMed Scopus (0) Google Scholar, 109Veroni C. et al.Transcriptional profile and Epstein–Barr virus infection status of laser-cut immune infiltrates from the brain of patients with progressive multiple sclerosis.J. Neuroinflammation. 2018; 15: 18Crossref PubMed Scopus (9) Google Scholar, 110Moreno M.A. et al.Signature of Epstein–Barr virus infection in multiple sclerosis brain lesions.Neurol. Neuroimmunol. Neuroinflamm. 2018; 5: e466Crossref PubMed Scopus (13) Google Scholar, 111Angelini D.F. et al.Increased CD8+ T cell response to Epstein–Barr virus lytic antigens in the active phase of multiple sclerosis.PLoS Pathog. 2013; 9: e1003220Crossref PubMed Scopus (0) Google Scholar]. In earlier studies, meningeal B cells within specific structures, referred to as tertiary lymphoid follicles with a GC-like architecture, were described as major sites of EBV persistence in MS brain [46Magliozzi R. et al.B-cell enrichment and Epstein–Barr virus infection in inflammatory cortical lesions in secondary progressive multiple sclerosis.J. Neuropathol. Exp. Neurol. 2013; 72: 29-41Crossref PubMed Scopus (0) Google Scholar,47Serafini B. et al.Dysregulated Epstein–Barr virus infection in the multiple sclerosis brain.J. Exp. Med. 2007; 204: 2899-2912Crossref PubMed Scopus (469) Google Scholar]. More recently, the presence of EBV in both MS and healthy brains has been reported [108Hassani A. et al.Epstein–Barr virus is present in the brain of most cases of multiple sclerosis and may engage more than just B cells.PLoS ONE. 2018; 13: e0192109Crossref PubMed Scopus (0) Google Scholar, 109Veroni C. et al.Transcriptional profile and Epstein–Barr virus infection status of laser-cut immune infiltrates from the brain of patients with progressive multiple sclerosis.J. Neuroinflammation. 2018; 15: 18Crossref PubMed Scopus (9) Google Scholar, 110Moreno M.A. et al.Signature of Epstein–Barr virus infection in multiple sclerosis brain lesions.Neurol. Neuroimmunol. Neuroinflamm. 2018; 5: e466Crossref PubMed Scopus (13) Google Scholar]. Veroni [109Veroni C. et al.Transcriptional profile and Epstein–Barr virus infection status of laser-cut immune infiltrates from the brain of patients with progressive multiple sclerosis.J. Neuroinflammation. 2018; 15: 18Crossref PubMed Scopus (9) Google Scholar] identified widespread EBV infection in meninges of MS patients, and EBV-related gene expression profiles (associated with latent EBV infection) in both meningeal and white matter tissue. Of further interest was the reported detection of gene expression in EBV-infected cells associated with IFN-γ signaling, type I immunity effector functions, B cell differentiation, proliferation, lipid-antigen presentation, and T cell and myeloid cell recruitment. In another study, brain EBV was detected by PCR or EBV encoding region (EBER) in situ hybridization (ISH) in 90% of all MS cases compared with only 24% of non-MS samples [108Hassani A. et al.Epstein–Barr virus is present in the brain of most cases of multiple sclerosis and may engage more than just B cells.PLoS ONE. 2018; 13: e0192109Crossref PubMed Scopus (0) Google Scholar]. EBNA1 was detected by immunohistochemistry (IHC) in MS brain sections as was, to a lesser extent, the intermediate-early EBV transactivator gene, BZLF-1. Of note, this study also reported the detection of EBV in astrocytes and microglia. Viruses other than EBV (e.g., HSV-1, CMV, HHV-6) were not detected by PCR. A further study analyzed the expression of EBV latent proteins as well as proteins associated with lytic infection in archived brain samples [110Moreno M.A. et al.Signature of Epstein–Barr virus infection in multiple sclerosis brain lesions.Neurol. Neuroimmunol. Neuroinflamm. 2018; 5: e466Crossref PubMed Scopus (13) Google Scholar]. EBV-encoded protein and mRNA were detected by IHC and in situ hybridization in bo