Live attenuated antigenically marked classical swine fever vaccine

Inventors

Borca, Manuel V. • Risatti, Guillermo R. • Holinka-Patterson, Lauren G.

Assignees

University of Connecticut • US Department of Agriculture USDA

Abstract

Controlling Classical Swine Fever Virus (CSFV) involves either prophylactic vaccination or non-vaccination and elimination of infected herds depending on the epidemiological situation. Marker vaccines allowing distinction between naturally infected from vaccinated swine could complement “stamping out” measures. Previously, we reported the development of FlagT4v, a double antigenic marker live attenuated CSFV strain. FlagT4v was later shown as not to be completely stable in terms of its attenuation when assessed in a reversion to virulence protocol. We have developed a modified version of the FlagT4v where changes in the codon usage of genomic areas encoding for Flag and T4 were introduced to rectify the reversion to the virulent genotype. The new virus, FlagT4-mFT-Gv, possesses the same amino acid sequence as FlagT4v except for one substitution, Asparagine is replaced by Glycine at position 852 of the CSFV polypeptide. FlagT4-mFT-Gv protected swine against challenge with Brescia virulent virus at 21 days post vaccination.

Core Innovation

Classical swine fever (CSF) is a highly contagious disease of swine caused by CSF virus (CSFV), a Pestivirus with a single-stranded RNA genome encoding a polyprotein that produces essential structural glycoproteins E1 and E2. Current control strategies include prophylactic vaccination or non-vaccination with stamping out of exposed animals. However, existing live attenuated CSFV vaccines elicit humoral responses indistinguishable from infection, lacking DIVA (differentiating infected from vaccinated animals) capability, thus limiting their use especially in disease-free countries.

Previously developed FlagT4v, a double antigenic marker live attenuated CSFV strain, incorporated a FLAG epitope insertion in the E1 glycoprotein as a positive antigenic marker and abolished a CSFV-specific epitope recognized by monoclonal antibody WH303 in the E2 glycoprotein as a negative marker. Although FlagT4v provided rapid and solid protection and enabled serological differentiation from wild-type infections, it was not completely stable, showing some reversion to virulence upon successive passages in swine.

The invention addresses the need to improve stability while retaining dual antigenic markers and protective efficacy. It provides a modified FlagT4v virus, named FlagT4-mFT-Gv, which introduces changes in codon usage in genomic areas encoding the Flag epitope insertion in E1 and the T4 region in E2 to rectify reversion to virulence. FlagT4-mFT-Gv encodes the same amino acids as FlagT4v except one substitution: asparagine is replaced by glycine at position 852 in the CSFV polypeptide. FlagT4-mFT-Gv demonstrates stable attenuation in vivo and protects swine against challenge with the virulent Brescia strain at 21 days post-vaccination, while allowing serological distinction between vaccinated and infected animals.

Claims Coverage

The patent includes several claims, among which independent claims define the recombinant CSFV mutant, the vaccine composition, infected cells, and methods for protection and differentiation.

The claims cover the nucleic acid and protein composition of the modified double antigenic marker CSFV mutant, its use in vaccines, the infected host cells, and methods to immunize animals and distinguish vaccinated from infected animals based on serological markers.

Stated Advantages

Documented Applications