Characterization of virus neuroinvasion, blood-brain barrier integrity and neuroinflammation following Powassan virus infection in mice.

Powassan virus (POWV) is a tick-borne Orthoflavivirus transmitted by Ixodes tick species. POWV causes fatal encephalitis in approximately 10-30% of neurological cases, and long-lasting neurological sequelae in approximately 50% of survivors. POWV entry into the central nervous system (CNS) is an important event in determining clinical outcome. In this study, we evaluated viral replication kinetics, neuropathology, as well as host immune response following POWV infection in C57BL/6J (WT) mice. Our data showed that infection with POWV by all inoculation routes, including the intravenous, intraperitoneal, intracranial and subcutaneous, led to severe neuroinvasive disease. We showed that POWV effectively replicates in WT mice, where replication and dissemination resulted in peripheral and neurotropic phases. Viral neuroinvasion correlated with severe neuropathological alterations as well as enhanced blood-brain barrier permeability. Next, we used transcriptomics to compare the induction of effector pathways in the brain during the acute and late stages of POWV infection in mice. At all examined time points, we found several dysregulated genes including genes associated with interferon signaling, neuroinflammation and cell death signaling. We detected significant increase in the protein levels of markers involved in neuroinflammation in POWV-infected brains. Immunofluorescence analyses further validated the transcriptomic findings and demonstrated increased activation of microglia (IBA1) and astrocytes (GFAP), infiltration of peripheral immune cells (CD45), and elevated neuronal cell death (TUNEL) in POWV-infected brains. Increased protein expression of caspase-3 and p16 further indicated activation of apoptotic and senescence-associated pathways. Interestingly, we detected viral RNA and found evidence of neuroinflammation persistence, albeit at lower levels, in mice that survived the acute POWV encephalitis phase. Overall, this study provides a comprehensive understanding of the pathogenic events that occur during the acute and late stages of POWV infection in mice.