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Previous studies have shown that mouse adenovirus type-1 (MAV-1) caused a fatal hemorrhagic encephalitis in certain strains of mice. C57Bl/6 mice exhibited 100% mortality when given as little 103plaque-forming units (PFU) of MAV, in contrast to BALB/c mice which were resistant to as many as 106PFU. Susceptible animals died with a flaccid paralysis on the 3rd or 4th day after inoculation. The brains and spinal cords of these animals displayed numerous petechial hemorrhages that were found in virtually all areas of the brain, but were more numerous in white matter. In this paper, immunohistochemistry and electron microscopy were used to identify the viral target of replication within the CNS of susceptible mice. These studies showed that the CNS vascular endothelial cell was the primary site of viral replication within the CNS of mice infected with MAV-1. Characterization of cytokine mRNA levels and disease course in immunodeficient mice revealed that the host immune response played little, if any, role in the pathogenesis of MAV-1 disease in susceptible mice and was not responsible for the resistance of BALB/c mice. These results support the conclusion that disease course and outcome in susceptible and resistant strains of mice were determined primarily by the ability of the virus to replicate within the CNS vascular endothelium.

The pathogenesis of Venezuelan equine encephalitis virus (VEE) was examined in the mouse model using V3000, a virus derived from a molecular clone of the Trinidad donkey strain of VEE. These results were compared in parallel experiments with avirulent mutants of VEE derived by site-directed mutagenesis of the clone. Adult mice, inoculated subcutaneously in their left rear footpad with V3000, were followed in a time course study for 6 days in which 15 organs were tested for histopathological changes, for the presence of viral antigen by immunohistochemical staining, for the presence of viral nucleic acid by in situ hybridization analysis, and for content of viable virus. Virus was detected in the footpad inoculation site, but until 12 hr postinoculation (pi), the level of virus did not suggest early viral replication. By 4 hr pi, however, replication of V3000 was evident in the draining popliteal lymph node. At this early time point, no virus could be isolated from any other organ examined. At 12 hr, a significant serum viremia was observed, and virus was detected at a low level in a number of well vascularized organs, including spleen, heart, lung, liver, kidney, and adrenal gland. By 18 hr, high virus titers were present in serum and all the lymphoid organs examined, and these tissues appeared to be the major peripheral sites of V3000 replication. Virus in serum and peripheral organs was cleared by 3-4 days pi. In a second phase of the infection, V3000 invaded the central nervous system (CNS), replicated predominantly in neurons, and persisted in the brain until death by encephalitis. Pathologic findings as well as the results of immunocytochemical and in situ hybridization examination were generally coordinate with virus titration. A site-directed mutant of V3000, V3010, contained a mutation in the gene for the E2 glycoprotein at codon 76 (Glu to Lys) which rendered it avirulent after footpad inoculation. Detection of V3010 replication in the draining lymph node was sporadic and was sometimes delayed to as long as 3 days pi. Infrequent and/or delayed virus spread to other sites also was observed. Analogous experiments were performed with other mutants which were avirulent by the footpad inoculation route: V3014, a mutant differing from V3000 at three loci (E2 Lys 209, E1 Thr 272, and E2 Asn 239), as well as single-site mutants V3032 (E2 Lys 209) and V3034 (E1 Thr 272). The mutations in V3014 prevented spread beyond the draining lymph node. The single-site E2 Lys 209 mutation allowed spread to the draining lymph node and to other lymphoid organs without significant serum viremia or invasion of the CNS, and E1 Thr 272 was characterized by near normal peripheral replication, but only sporadic replication in the CNS. These results suggest that VEE-induced disease results from a sequence of events which can be defined by interdicting the pathogenic pathway with specific mutations in the VEE genome.