Genbank file of the MNoV^CW3-HiBiT plasmid.

Caliciviruses are significant agricultural and human pathogens that are poorly understood due to the dearth of molecular tools, including reporter systems. We report the development of a robust luciferase-based reporter system for a model calicivirus, murine norovirus (MNoV). Genetic insertion of a HiBiT tag, an 11 amino acid fragment of nanolucifersase, at the junction of the nonstructural proteins NS4 and NS5 yields infectious virus. The resultant MNoV-HiBiT produces a robust signal that is detected early in infection and occurs only in cells susceptible to MNoV infection. The MNoV-HiBiT reporter is effective at monitoring acute infection in STAT1 deficient mice. Furthermore, we used this tool to characterize two unappreciated host directed anti-MNoV compounds. The use of the MNoV-HiBiT virus enables new mechanistic studies by a rapid and quantitative means of measuring MNoV replication. The HiBiT insertion strategy we describe may be useful for the generation of other calicivirus reporters.