A Recombinant Genotype I Japanese Encephalitis Virus Expressing a Gaussia Luciferase Gene for Antiviral Drug Screening Assay and Neutralizing Antibodies Detection

Japanese encephalitis virus (JEV) is the major cause of viral encephalitis in humans throughout Asia. In the past twenty years, the emergence of the genotype I (GI) JEV as the dominant genotype in Asian countries has raised a significant threat to public health security. However, no clinically approved drug is available for the specific treatment of JEV infection, and the commercial vaccines derived from the genotype III JEV strains merely provided partial protection against the GI JEV. Thus, an easy-to-perform platform in high-throughput is urgently needed for the antiviral drug screening and assessment of neutralizing antibodies specific against the GI JEV. In this study, we established a reverse genetics system for the GI JEV strain (YZ-1) using a homologous recombination strategy. Using this reverse genetic system, a gaussia luciferase (Gluc) expression cassette was inserted into the JEV genome to generate a reporter virus (rGI-Gluc). The reporter virus exhibited similar growth kinetics to the parental virus and remained genetically stable for at least ten passages in vitro. Of note, the bioluminescence signal strength of Gluc in the culture supernatants was well correlated with the viral progenies determined by viral titration. Taking advantage of this reporter virus, we established Gluc readout-based assays for antiviral drug screening and neutralizing antibody detection against the GI JEV. These Gluc readout-based assays exhibited comparable performance to the assays using an actual virus and are less time consuming and are applicable for a high-throughput format. Taken together, we generated a GI JEV reporter virus expressing a Gluc gene that could be a valuable tool for an antiviral drug screening assay and neutralization assay.