The temporal course of cellular pathology in virus-infected oligodendrocytes in vivo is not well defined. Here we study these events in the mouse brain using a novel system in which large numbers of oligodendrocytes can be reproducibly infected. In the mouse, following extraneural inoculation, the A7(74) strain of the alphavirus Semliki Forest virus (SFV) is efficiently neuroinvasive and central nervous system (CNS) infection leads to predominantly perivascular lesions of immune-mediated demyelination. This study demonstrates that direct intracerebral inoculation with SFV A7(74) or the SFV1 vector results in dramatic, selective and widespread infection of the major white matter tract of the brain, the corpus callosum. Mature oligodendrocytes are the predominant cell type infected. Subsequent events are complex; early virus-induced necrotic death of infected cells is followed by apoptotic death of adjacent apparently uninfected cells. A strong inflammatory response and considerable myelin loss are evident from 10 days and virus-positive cells are not observed after this time. In contrast, in athymic nu/nu mice, in the absence of T-cell responses, no inflammatory infiltrates are observed and virus-infected cells persist for over 30 days with extensive vacuolation but less demyelination. The change from an early destructive to a potentially persistent infection of oligodendrocytes is likely to reflect activation of innate immune responses. Activation of peripheral innate defences by inoculation of poly I: C prior to CNS virus infection abrogates the widespread corpus callosum infection. This widespread infection of the corpus callosum provides a novel in vivo system in which to study virus–oligodendrocyte interactions.