The inventors have created a reverse genetics system for RSV strain B1 of antigenic subgroup B encoding a replication-competent recombinant RSV that contains a codon-optimized G ORF and expresses enhanced green fluorescence protein (GFP). There are two antigenic subgroups of RSV, subgroups A and B, and most of the available information and reagents are for subgroup A. Immunity against either subgroup has reduced effectiveness in restricting the heterologous subgroup, suggesting that an effective RSV vaccine might need to contain both subgroups. The sequence of the wild type G gene was refractory to cloning into full-length antigenomic cDNA in E. coli, and so the inventors made and successfully used a codon optimized version. In addition, the inventors inserted an eGFP gene into the first gene position (promoter proximal). The resulting virus is replication-competent and efficiently expresses GFP in infected cells. This virus can be used as a tool to detect RSV-neutralizing antibodies to RSV subgroup B in a plaque-reduction assay. It also can be used to evaluate RSV infection in vitro and in vivo using GFP fluorescence to track infection. The antigenomic cDNA clone also provides the starting material for making live-attenuated subgroup B-specific RSV vaccine candidates containing defined mutations. These defined mutations can include ones that we previously developed for RSV subgroup A, and include stabilized point mutations, stabilized codon-deletions, and gene-deletions.
The present invention provides a reverse genetics system encoding strain B1 of RSV subgroup B containing a codon-optimized G ORF and encoding eGFP. This provides a tool for RSV subgroup B serology assays, for tracking RSV infection, and a starting point for making attenuated subgroup B strains for vaccine purposes.
This technology is available for licensing for commercial development in accordance with 35 U.S.C. § 209 and 37 CFR Part 404, as well as for further development and evaluation under a research collaboration.
- Viral diagnostics
- Vaccine research
- Serology assays
- Vaccine manufacture
- Ease of manufacture
- Unique research tool