A clinical isolate of herpes simplex virus type 1 that is aciclovir resistant but neurovirulent in mice was described previously. The mutation in this virus is a double G insertion in a run of seven G residues that has been shown previously to be a mutational hotspot. Using a sensitive assay, it has been demonstrated that preparations of this virus are able to induce low but consistent levels of thymidine kinase (TK) activity. However, this activity results from a high frequency mutational event that inserts a further G into the 'G-string' motif and thus restores the TK open reading frame. Passage of this virus through the nervous system of mice results in the rapid selection of the TK-positive variant. Thus, this variant is the major component in virus reactivated from latently infected ganglia. Mutation frequency appears to be influenced by the genetic background of the virus.