High numbers of viral RNA copies in the central nervous system of mice during persistent infection with Theiler's virus

The low-neurovirulence Theiler's murine encephalomyelitis viruses (TMEV), s uch as BeAn virus, cause a persistent infection of the central nervous syst em (CNS) in susceptible mouse strains that results in inflammatory demyelin ation. The ability of TMEV to persist in the mouse CNS has traditionally be en demonstrated by recovering infectious virus from the spinal cord. Result s of infectivity assays led to the notion that TMEV persists at low levels. In the present study, we analyzed the copy number of TMEV genomes, plus- t o minus-strand ratios, and full-length species in the spinal cords of infec ted mice and infected tissue culture cells by using Northern hybridization. Considering the low levels of infectious virus in the spinal cord, a surpr isingly large number of viral genomes (mean of 3.0 X 10(9)) was detected in persistently infected mice. In the transition from the acute (approximatel y postinfection [p.i.] day 7) to the persistent (beginning on p.i. day 28) phase of infection, viral RNA copy numbers steadily increased, indicating t hat TMEV persistence involves active viral RNA replication. Further, BeAn v iral genomes were full-length in size; i.e., no subgenomic species were det ected and the ratio of BeAn virus plus- to minus-strand RNA indicated that viral RNA replication is unperturbed in the mouse spinal cord. Analysis of cultured macrophages and oligodendrocytes suggests that either of these cel l types can potentially synthesize high numbers of viral RNA copies if infe cted in the spinal cord and therefore account for the heavy viral load. A s cheme is presented for the direct isolation of both cell types directly fro m infected spinal cords for further viral analyses.