Bovine viral diarrhea-mucosal virus with modified erns protein

A technology for bovine viral diarrhea and viruses, applied to cells modified by introducing foreign genetic material, antiviral agents, viruses/bacteriophages, etc., can solve problems such as inability to distinguish between inoculated and naturally infected animals

Inactive Publication Date: 2013-03-13
PFIZER INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, existing MLV vaccines cannot distinguish between vaccinated and naturally infected animals

Method used

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  • Bovine viral diarrhea-mucosal virus with modified erns protein
  • Bovine viral diarrhea-mucosal virus with modified erns protein
  • Bovine viral diarrhea-mucosal virus with modified erns protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0133] Example 1: Construction and serological characterization of chimeric pestiviruses

[0134]Escherichia coli K12 GM2163[F-ara-14,leuB6,thi-1,fhuA31,lacY1,tsx-78,galK2,galT22,supE44,hisG4,rpsL136,(Str r ), xyl-5, mtl-1, dam13::Tn9 (Cam r ), dcm-6, mcrB1, hsdR2 (rk - mk - ), mcrA] containing a plasmid containing the full-length genomic cDNA of bovine viral diarrhea virus strain NADL (BVDV-NADL), which was obtained from Dr. R. Donis of the University of Nebraska.

[0135] RD cells (bovine testicular cells transformed with SV40; obtained from Dr. R. Donis) were maintained in OptiMEM supplemented with 3% horse serum, 1% non-essential amino acids (NEAA) (in modified Eagle medium ( MEM), 2 mM GlutaMax and 10 μg / ml gentamicin. BK-6 cells were obtained from Pfizer Global Manufacturing (PGM). Cells were grown in Dulbecco's Modified Eagles Medium (DMEM) supplemented with 5% horse serum or donor bovine serum (PGM), 2 mM Glutamax, and 1% antibiotics and antifungals. All media co...

Embodiment 2

[0164] Example 2. Construction and serological characterization and efficacy testing of chimeric pestivirus vaccine candidates

[0165] Type 1 BVDV strain CM5960 and type 2 BVDV strain CM53637 were obtained from Pfizer Global Manufacturing. According to the manufacturer's instructions, use MagMax TM AI / ND Viral RNA Isolation Kit (Ambion) was used to extract viral RNA. According to the manufacturer's instructions, use ThermoScript TM RT-PCR System (Invitrogen) reverse transcribed RNA to generate cDNA. Using the overlapping PCR method, by combining the E of CM5960 and CM53637 rns Gene replacement for E of pronghorn pestivirus rns Gene (P-E rns ), resulting in chimeric pestiviruses. The oligonucleotide primers used for PCR are listed in Table 1.

[0166] To construct the chimeric CM5960 / P-E rns DNA, a fragment of CM5960 cDNA between the 5' UTR and the 3' end of the C gene was amplified from CM5960 cDNA by PCR using primers Oligo B-5 and Oligo 135. Using primers Oligo 1...

Embodiment 3

[0176] Example 3. Efficacy Testing of Chimeric Pestivirus Vaccine Candidates in a Bovine Respiratory Disease Model

[0177] Healthy cattle obtained negative for BVDV were randomized into study groups and maintained under the supervision of the attending veterinarian. The test vaccines were mixed with a sterile adjuvant and administered by intramuscular (IM) or subcutaneous (SC) injection or by intranasal (IN) inoculation. The vaccine is administered in 1 or 2 doses. Two doses of the vaccine were administered at intervals of 21-28 days. Animals were then challenged with type 1 or type 2 BVDV strains 21-28 days after the last inoculation. The challenge inoculum was administered intranasally in divided doses of 4 ml (2 ml / nostril). A control group consisting of non-vaccinated, non-challenged animals and / or non-vaccinated, challenged animals was also maintained throughout the study.

[0178] Clinical parameters including rectal temperature, depression, anorexia, and diarrhea w...

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PUM

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Abstract

The invention relates to a chimeric plague virus that can be used for an immunogenicity composition and a vaccine. The chimeric plague virus contains a bovine viral diarrhea-mucosal virus which is not expressing Erns protein with the same source, and expression of the chimeric plague virus is from allos Erns protein of another plague virus or a natrual, synthetic or genetic variant of the allos Erns protein. The invention also describes a method and a kit for treating or preventing transmission of infection of the bovine viral diarrhea-mucosal virus, and a method and a kit for differentiating vaccinal animals and wild infected animals.

Description

[0001] This application is a divisional application of the application with the same title filed on November 23, 2009 and with application number 200980148483.7. technical field [0002] The present invention relates to novel chimeric pestiviruses and their use in immunogenic compositions and vaccines. The invention also relates to methods and kits for treating or preventing the spread of bovine viral diarrhea virus infection. The invention also relates to the use of chimeric pestiviruses in methods and kits for differentiating vaccinated animals from animals infected with wild-type virus. Background technique [0003] Pestiviruses, including bovine viral diarrhea virus (BVD virus or BVDV), have been isolated from several animal species, both farmed and wild. Identified hosts of BVDV include buffalo, antelope, reindeer, and various deer species, and unique pestivirus species have been identified in giraffe and pronghorn. BVDV is a picornavirus of the Flaviviridae family. ...

Claims

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Application Information

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IPC IPC(8): C12N7/01C12N15/40C12N5/10C12N1/15C12N1/19C12N1/21C07K16/10A61K39/12A61K48/00A61P31/14G01N33/569
Inventor R・G・安肯鲍尔Y・罗S-K・W・韦尔奇Y・袁
Owner PFIZER INC
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