Table 1.

EBV intersection with MS.

EBV features	Brief description	Compatibility with MS
EBV epidemiology	Ubiquitous DNA herpesvirus that infects about 90% of the global population. EBV infection is mostly asymptomatic in childhood but primary exposure during adolescence or adulthood frequently causes infectious mononucleosis (13).	Previous exposure to EBV is required, though not sufficient, to develop MS (14). Infectious mononucleosis (15) and high anti-EBNA-1 IgG titers (16–18) increase the risk of MS. There are similarities between the epidemiology of infectious mononucleosis and that of MS, including higher socioeconomic status, latitude gradient, earlier onset in women than in men (19, 20).
EBV biology	EBV is transmitted through saliva, infects mainly B cells and epithelial cells and establishes a life-long latent infection in memory B cells (11). The virus first establishes a lytic infection in the oropharyngeal epithelial cells and then switches to latent infection of B cells in the local lymphoid tissue (tonsils). Initially, EBV establishes a growth transforming latent infection of B cells (latency III or growth program) leading to proliferation of the infected cells; at this stage of infection, all EBV latent proteins [EBV nuclear antigen (EBNA) 1, 2, 3A, 3B, 3C, and -LP; latent membrane protein 1 (LMP1) and LMP2], several small noncoding RNAs and micro-RNAs, and the EBV-encoded small RNAs (EBER1, EBER2) are expressed. EBV infected B lymphoblasts receive survival and activating signals from LMP1 and LMP2A, that mimic B cell receptor stimulation by cognate antigen and T cell help through CD40 signalling, respectively. EBV then enters a more restricted form of latency where only EBNA1, LMP1 and LMP2 are expressed (latency II program), whereas only EBNA1, that is required for replication of the episomal EBV genome, is expressed in latency I. Viral persistence and avoidance of immune surveillance are achieved as the result of downregulation of all EBV gene products in EBV infected resting memory B cells that enter the blood circulation (latency 0). Occasional viral reactivations occur in the tonsils, where recirculating EBV infected B cells differentiate into plasma cells leading to production of viral particles that can infect new B cells locally and also result in viral shedding into saliva. The replicative viral cycle involves the sequential expression of a large array of immediate early, early and late lytic genes (>80) encoding proteins that are implicated in the production of new viral particles and (some of them) also in immune evasion.	The B-cell growth promoting properties of EBV could explain the expansion and differentiation of B cells in the CNS of MS patients throughout the disease.
		Persistent, treatment resistant intrathecal B-cell activation and immunoglobulin synthesis are a characteristic of MS (21, 22). More than 90% MS patients have oligoclonal IgG in the CSF. Oligoclonal IgG in the CSF are typically found in acute CNS infections, but their specificity in MS is unknown although recognition of EBV proteins has been reported (23–26). Polyspecific immunoglobulins recognizing common viruses (rubella, measles, varicella zoster, less commonly EBV) are synthesized in the CNS of nearly 80% of MS patients, and are believed to reflect non-specific bystander B-cell activation (22).
		B cells and plasmablasts/plasma cells, usually absent in the normal CSF, are found in the CSF of MS patients and their number correlates with inflammation, blood-brain barrier breakdown and intrathecal Ig synthesis (27–29). B cell receptor repertoire analysis in MS patients has revealed presence of clonally expanded B cells in CSF, brain tissue and meninges indicative of local antigenic stimulation (21, 30) as well as trafficking of activated B cells between the CNS, draining cervical lymph nodes and peripheral blood (29, 31, 32).
		B cells and plasma cells are found in CNS tissue in early and chronic MS stages (33–35), mainly within the perivascular space of intraparenchymal blood vessels and the subarachnoid space in the meninges. Large B-cell aggregates resembling B-cell follicles and containing stromal cells, proliferating B cells and plasma cells are present in the meninges of patients with progressive MS (36, 37). The pathogenic role of these structures is indicated by their association with increased cortical damage and MS disease severity (37–39).
EBV immunology	Continuous immune surveillance is essential to maintain virus-host homeostasis throughout the host’s life. Following primary infection, the rapid antibody response to EBNA2 and EBV lytic proteins, like the virus capsid antigen (VCA), is important to control the virus, and is followed by a slow increase of the neutralizing antibody response (mainly towards the major envelope EBV protein gp350) and a delayed EBNA1 IgG response. Most studies of the T-cell response to EBV during primary infection have been carried out in people with symptoms of infectious mononucleosis. Early control of EBV infection is associated with expansion of innate immune cells, mainly NK cells (40), IFNγ producing cytolytic CD8 T cells and, to a lesser extent, CD4 T cells. While CD4 T cells recognize a broad range of EBV latent and lytic proteins, CD8 T cells recognize mainly EBV lytic proteins (41). During persistent infection, memory CD4 T cells are present at low frequency, recognize mainly latent EBV proteins, do not express activation markers and belong to both the central memory and effector memory subsets. Compared to the CD4 T cell response, the EBV-specific CD8 T cell response in the blood of healthy carriers is much greater and skewed towards immunodominant EBV latent (EBNA3A, 3B, 3C) and immediate early (BZLF1, BRLF1) and early (BMRF1, BMLF1) lytic EBV proteins, the response to lytic proteins occurring at higher frequency. Circulating EBV-specific CD8 T cells are resting, antigen experienced T cells that exhibit potent effector functions, including cytotoxicity and cytokine (IFNγ, TNF) production, upon antigen challenge. Due to an efficient immune control, rare EBV infected memory B cells are present in the peripheral blood (1-50 in 106 B cells) and lymphoid tissue of healthy carriers. Cytotoxic lymphocytes, including NK cells and EBV-specific CD8 T cells, have a key role in limiting EBV lytic replication and preventing EBV-driven pathologies (40). The EBV-host balance is perturbed when genetic factors or other factors, such as immunosuppression, alter or abolish the cytotoxic control of EBV.	Altered humoral and cell-mediated immune responses to EBV in MS patients suggest EBV dysregulation/inadequate virus control. MS patients are 100% EBV seropositive and have higher serum titers of EBNA1 IgG and anti-VCA IgG compared to healthy individuals (10, 42). An increase in EBNA-1 IgG is detectable about 5 years up to 20 years before the first disease symptoms (17, 18). Higher EBNA-1 IgG titers have been associated with conversion to definite MS in patients with a clinically isolated syndrome (43) and with cortical atrophy and lesion burden in patients with MS (44). The association between anti-EBV antibodies and clinical or radiological MS disease activity is controversial (45–48). Regarding EBV-specific T cell responses, the CD4 T cell response to EBNA-1, but not CMV epitopes, is increased in the peripheral blood of MS patients compared to control subjects (49) and CD4 T cells recognizing EBNA-1 or EBV transformed B-cell lines have been detected in the CSF of MS patients (50–52). The EBV-specific CD8 T cell response is increased, reduced or unchanged in MS patients compared to control subjects depending on disease activity and duration, and on the T cell specificities investigated. Higher frequencies of EBV-specific T cells are found in the peripheral blood at disease onset and during relapses (53–56). The frequency and functionality of EBV-specific CD8 T cells decreases with increasing disease duration (54, 55, 57, 58), suggesting T cell exhaustion. EBV-specific CD8 T cells, recognizing mainly lytic EBV proteins, are enriched in the CSF (59–61) and brain tissue (62, 63) of MS patients. Defects in NK cell function are present in MS (64). Expansion of CD8+ NK cells in the blood of MS patients has been associated with a favourable clinical outcome (65), raising the possibility that this cell subset is involved in the control of EBV infection.
EBV pathogenic potential	EBV is etiologically linked to a wide range of human malignancies, including B-cell malignancies, like Hodgkin’s lymphoma, Burkitt’s lymphoma, diffuse large B cell lymphoma and post-transplant B-lymphoproliferative disease, NK/T cell lymphoma and nasopharyngeal carcinoma (12).	The mechanisms linking EBV infection to MS pathology remain elusive. Several hypotheses have been proposed, each calling for further studies:
	EBV is also the etiological agent of immunopathologic diseases that are caused by an excessive immune response towards uncontrolled EBV infection, like infectious mononucleosis, a self limiting lymphoproliferative disease, and chronic active EBV infection, a very serious condition with persistence of infectious mononucleosis-like symptoms and hemophagocytic lymphohistiocytosis. In infectious mononucleosis a large proportion of memory B cells are infected with EBV (up to 50% in the peripheral blood) causing the large expansions of NK cells and highly activated EBV-specific T cells, which are predominantly CD8 T cells specific for EBV lytic proteins, and release of pro-inflammatory cytokines, including high amounts of IFNγ. This aggressive cytotoxic immune response correlates with the severity of infectious mononucleosis symptoms, including fever, lymphadenopathy and prolonged fatigue (13). Several rare primary immunodeficiencies affecting NK and T cell function result in failure to control EBV infection predisposing to EBV-associated pathologies, like B-cell lymphomas, fulminant infectious mononucleosis and hemophagocytic lymphohistiocytosis (76).	▪ According to Pender (66), EBV infection may rescue autoreactive B cells producing antibodies to CNS proteins that migrate into the CNS and provide costimulatory signals for CNS autoreactive CD4 T cells. To date, MS-associated pathogenic autoantibodies remain undefined (67), and there is no evidence that EBV favors survival of autoreactive B cells (68). ▪ The molecular mimicry hypothesis is supported by several studies showing that antibodies (69–71) and CD4 T cells (72–75) from MS patients cross react with peptides from EBV proteins and peptides from myelin or other proteins expressed in the CNS. Crossreactive antibodies and CD4 T cells can be found also in healthy individuals, albeit at lower frequency. The pathogenicity of antibodies and T cells recognizing candidate MS autoantigens remains to be demonstrated. ▪ The mistaken self hypothesis proposes that EBV infection induces expression of the small heat shock protein alpha B-crystallin in B cells; HLA-DR-restricted presentation of alpha B-crystallin activates pathogenic CD4 T cells recognizing stress-induced alpha B-crystallin in glial cells in MS brain lesions (77). There is no evidence for increased T cell responses towards alpha B-crystallin in MS patients (78) or recognition of this protein by CSF-infiltrating T cells from MS patients (51, 61). ▪ EBV-driven immunopathology entails that bystander CNS tissue damage is caused by a cytotoxic T cell response towards a persistent, reactivated EBV infection in the CNS. In support of this hypothesis are the findings that CD8 T cells are activated and preferentially expand in the CNS of MS patients (79–83), and that EBV-specific CD8 T cells are selectively enriched in the CSF and brain tissue of MS patients (59–63). Presence of EBV latently infected B cells and EBV reactivation in the MS brain is highly debated (see text).