Rocio virus (ROCV) (Flavivirus genus, Flaviviridae family) was responsible for an outbreak of encephalitis in Sao Paulo State, Brazil, in 1975 (1-3). The objective of this study was to evaluate the C57BL/6 mice as model for ROCV infection. Immunocompetent C57BL/6 mice were infected via intraperitoneal with 2.76x108-2.76x104 PFU of ROCV and were then monitored for up to 20 days. Mice infected with the highest load viral (2.76x108 and 2.76x107 PFU) showed a dramatic decrease in weight, all of them dying after six days of infection. However, animals infected with lower viral loads (2.76x106, 2.76x105 and 2.76x104 PFU) showed a slow decrease in weight, the presence of encephalitis and a delay in death. Viral RNA was detected three and five days post-infection in serum, spleen, kidney, liver, brain, whole blood, lung, heart, spinal cord, and bone marrow. The highest viral load was found in the brain. The levels of hematocrit, platelets and the number of red blood cells in mice infected were not affected when compared to uninfected animals. However, leukopenia and lymphopenia were found in the infected animals as well as an increase in the number of monocytes. Hepatic enzyme levels were similar to the uninfected group. Histological analysis revealed an increase of polymorphonuclear cells in most tissues of infected mice but not in the spleen. Cytokine production in the sera of infected animals was investigated using a protein microarray. Interestingly, only the chemokine (C-C motif) ligand 1 (CCL-1) resulted in a significant increase. The lack of cytokine production in infected animals suggests that ROCV could be involved in inhibiting the immune response, similar to others viruses of the Japanese encephalitis virus complex, such as West Nile Virus (WNV)(4). This animal model will be an important tool for the study of the mechanisms of neurovirulence and neuroinvasiveness of ROCV.