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Bovine viral diarrhea virus with a modified erns protein

A technology for bovine viral diarrhea and viruses, which is applied in the direction of positive-sense single-stranded RNA viruses, viruses, cells modified by introducing foreign genetic material, etc., and can solve the problem of not being able to distinguish between inoculated and naturally infected animals

Active Publication Date: 2011-11-09
ZOETIS SERVICE LLC
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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 virus with a modified erns protein
  • Bovine viral diarrhea virus with a modified erns protein
  • Bovine viral diarrhea virus with a modified erns protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0132] Example 1: Construction and serological characterization of chimeric pestiviruses

[0133]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.

[0134] 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), 2mM GlutaMax and 10ug / 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), 2mM Glutamax and 1% antibiotics and antifungals. All m...

Embodiment 2

[0163] Example 2. Construction and serological characterization and efficacy testing of chimeric pestivirus vaccine candidates

[0164] 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.

[0165] 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 ...

Embodiment 3

[0175] Example 3. Efficacy Testing of Chimeric Pestivirus Vaccine Candidates in a Bovine Respiratory Disease Model

[0176] 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.

[0177] Clinical parameters including rectal temperature, depression, anorexia, and diarrhea w...

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Abstract

The present invention relates to chimeric pestiviruses having utility as immunogenic compositions and vaccines wherein said chimeric pestivirus comprises a bovine viral diarrhea virus which does not express its homologous Ems protein, further wherein said chimeric pestivirus expresses a heterologous Ems protein derived from another pestivirus, or a natural, synthetic or genetic variant of said heterologous Erns protein. Also described herein are methods and kits for treating or preventing the spread of bovine viral diarrhea virus infection, as well as methods and kits for differentiating between vaccinated and wild-type infected animals.

Description

technical field [0001] 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 [0002] 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. It is closely related to other pestiviruses which are causative agents of border disease in sheep and classical swine fever in pigs. Recently, a distinct pestiviru...

Claims

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Application Information

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IPC IPC(8): C12N7/04C07K14/18C12N15/86A61K39/12C07K16/10G01N33/50A61K39/00
CPCA61K2039/5256C12N7/00A61K2039/525C12N2770/24322C07K14/005C12N2770/24361A61P1/04A61P1/12A61P31/14C12N5/10C12N7/04A61K39/12A61K2039/5252A61K2039/5254A61K2039/53A61K2039/552C07K16/1081C12N2770/24321G01N33/6854
Inventor R·G·安肯鲍尔Y·罗S-K·W·韦尔奇Y·袁
Owner ZOETIS SERVICE LLC
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