Live attenuated antigenically marked classical swine fever vaccine

Patent metadata

Field	Value
Publication number	US-9352032-B2
Application number	US-201414200802-A
Country	US
Kind code	B2
Filing date	Mar 7, 2014
Priority date	Mar 7, 2014
Publication date	May 31, 2016
Grant date	May 31, 2016

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.

First claim

We claim:

1. A recombinant classical swine fever virus (CSFV) mutant, FlagT4-mFT-Gv, comprising a cDNA sequence as set forth by SEQ ID NO: 1, wherein said recombinant CSFV mutant, FlagT4-mFT-Gv is a live attenuated CSFV capable of being recognized by monoclonal antibodies specifically binding to the FLAG™ epitope, which serves as a positive marker for said attenuated CSFV mutant, wherein said epitope is the eight amino acid peptide DYKDDDDK as set forth in positions 688-705 of SEQ ID NO: 2 and the nucleotide positions 2435-2488 Of SEQ ID NO: 1.

2. The recombinant CSFV mutant of claim 1 further comprising another mutation in CSFV E2 glycoprotein in addition to said positive mutation marker in CSFV E1 glycoprotein, wherein the mutant CSFV E2 glycoprotein of FlagT4-mFT-Gv comprises a substitution mutation in the wild-type WH303 immunogenic epitope, wherein the FlagT4-mFT-Gv substitution is TSFGMDTLR as set forth in positions 849-857 of SEQ ID NO: 2 thereby eliminating the immunodominant WH303 epitope of the wild-type CSFV, wherein said recombinant CSFV mutant is a live attenuated CSFV having both the positive marker as a result of the mutation in CSFV E1 glycoprotein of claim 1 and the negative marker as a result of the mutation in the wild-type WH303 epitope in CSFV E2 glycoprotein.

3. The recombinant CSFV mutant of claim 2 comprising a cDNA sequence as set forth by SEQ ID NO: 1.

4. The recombinant CSFV mutant according to claim 1 wherein the CSFV mutant comprises additional attenuating mutations.

5. A vaccine composition comprising the recombinant CSFV mutant according to any one of claims 1 , 2 , 3 and 4 .

6. An isolated cell infected with the CSFV mutant of any one of claims 1 , 2 , 3 and 4 .

7. A method for the protection of swine against CSF, comprising administering to swine a live attenuated CSF vaccine comprising a recombinant CSFV mutant according to claim 1 in an amount effective to protect said swine from clinical CSF.

8. A method of distinguishing animals naturally infected with CSFV from animals that have been vaccinated with a live attenuated CSF vaccine comprising the recombinant CSFV mutant according to claim 1 , comprising: analyzing serum from an animal under evaluation to determine if said serum binds specifically to the FLAG™ epitope.

9. A method for the protection of swine against CSF, comprising administering to swine a live attenuated CSF vaccine comprising a recombinant CSFV mutant according to any one of claims 2 and 3 in an amount effective to protect said swine from clinical CSF.

10. A method of distinguishing animals naturally infected with CSFV from animals that have been vaccinated with a live attenuated CSF vaccine comprising the recombinant CSFV mutant according to any one of claims 1 - 3 , comprising: analyzing serum from an animal under evaluation to determine if the positive marker, the FLAG™ epitope, is present and if the negative marker, the WH303 epitope, is absent.

Assignees

• Us Agriculture
• Univ Connecticut
• Us Agriculture
• Univ Conneticut

Inventors

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

Classifications

• C12N2770/24334

  Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein · CPC title

• C12N2770/24321

  Viruses as such, e.g. new isolates, mutants or their genomic sequences · CPC title

• G01N33/56983

  Viruses · CPC title

• G01N2333/183

  Flaviviridae, e.g. pestivirus, mucosal disease virus, bovine viral diarrhoea virus, classical swine fever virus (hog cholera virus) or border disease virus · CPC title

• C12N7/00

  Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof (preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00) · CPC title