Elevated antibody titers to Epstein-Barr virus (EBV) have been found prior to the onset of multiple sclerosis (MS) and EBV has been found in serum of patients with MS exacerbations. We conducted a prospective, nested case-control study in the Nurses' Health Study and Nurses' Health Study II cohorts to determine whether plasma EBV viral load in healthy adults predicts the risk of MS. MS cases with blood collected before onset (n = 18) or diagnosis (n = 13) and 62 healthy controls were matched by age and time of blood collection. EBV viral load in plasma was measured by real time polymerase chain reaction. Presence of EBV in plasma was associated with an increased risk of MS (relative risk = 2.5, 95% CI 0.78-7.8, P = 0.12). Adjusting for smoking, ancestry, and latitude of residence at birth did not materially change this result. However, no association was found between the EBV viral load and risk of MS. These results support a role for EBV in the etiology of MS, but need to be confirmed in a larger study.