Tick-borne flaviviruses (TBFVs), including Powassan virus and Tick-borne encephalitis virus cause encephalitis or hemorrhagic fevers in humans with case-fatality rates ranging from 1–30%. Despite severe disease in humans, TBFV infection of natural rodent hosts has little noticeable effect. Currently, the basis for resistance to disease is not known. We hypothesize that the coevolution of flaviviruses with their respective hosts has shaped the evolution of potent antiviral factors that suppress virus replication and protect the host from lethal infection. In the current study, we compared virus infection between reservoir host cells and related susceptible species. Infection of primary fibroblasts from the white-footed mouse (Peromyscus leucopus, a representative host) with a panel of vector-borne flaviviruses showed up to a 10,000-fold reduction in virus titer compared to control Mus musculus cells. Replication of vesicular stomatitis virus was equivalent in P. leucopus and M. musculus cells suggesting that restriction was flavivirus-specific. Step-wise comparison of the virus infection cycle revealed a significant block to viral RNA replication, but not virus entry, in P. leucopus cells. To understand the role of the type I interferon (IFN) response in virus restriction, we knocked down signal transducer and activator of transcription 1 (STAT1) or the type I IFN receptor (IFNAR1) by RNA interference. Loss of IFNAR1 or STAT1 significantly relieved the block in virus replication in P. leucopus cells. The major IFN antagonist encoded by TBFV, nonstructural protein 5, was functional in P. leucopus cells, thus ruling out ineffective viral antagonism of the host IFN response. Collectively, this work demonstrates that the IFN response of P. leucopus imparts a strong and virus-specific barrier to flavivirus replication. Future identification of the IFN-stimulated genes responsible for virus restriction specifically in P. leucopus will yield mechanistic insight into efficient control of virus replication and may inform the development of antiviral therapeutics.
Bibliographical noteFunding Information:
This work was primarily funded by a pilot grant from the Lyme Disease Association, Inc., N12484701 to RTT. This work was supported in part by the Intramural Research Program of the National Institutes of Health, National Institutes of Allergy and Infectious Diseases. ZIA-AI001125 to SMB. Research reported in this publication was also supported by the National Institute of General Medical Sciences of the National Institutes of Health. R15GM099055 to JMS. The content is solely the responsibility of the authors and does not represent the official views of the National Institutes of Health. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. We thank Dr. Saurabh Chattopadhyay for providing the IFIT antibodies and intellectual contributions. We also thank Drs. Malathi Krishnamurthy and R. Mark Wooten for their guidance in the virus entry assays and RT-qPCR respectively. We thank all the personnel at the flow cytometry and microscopy core facilities at the University of Toledo Health Science Campus.