To evaluate the cause of MHV68.DHS attenuation, we first performed an infectious center assay to test whether the splenocytes from infected mice harbored latent virus capable of lytic reactivation. In this assay, equivalent numbers of splenocytes from infected mice are overlayed onto cultured mouse fibroblasts. Spontaneous lytic reactivation of the splenocytes leads to infection of the fibroblasts, which can be quantified by counting the resulting plaques [34] . Splenocytes from MHV68.DHS infected mice reactivated at a significantly lower frequency than those from MHV68.WT or MHV68.MR infections ( Figure 7B ). This reactivation frequency was independently assessed in a subset of the samples by a limiting dilution cytopathic effect assay, which yielded identical results (data not shown). Plaque assays on spleen homogenates confirmed that no preformed virus was present, indicating that the virus detected in these experiments originated from latently infected cells.