Pigeon-derived newcastle disease virus recombinant vaccine strain as well as construction method and application thereof

A technology of epidemic virus and attenuated vaccine, applied in application, vaccine, virus, etc., can solve the problem of lack of Newcastle disease vaccine products for pigeons

Active Publication Date: 2021-12-03
CHINA ANIMAL HEALTH & EPIDEMIOLOGY CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to construct and obtain a recombinant vaccine strain of pigeon-derived Newcastle disease virus and its construction method and application, thereby solving the problem of lack of pigeon Newcastle disease vaccine products in the prior art

Method used

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  • Pigeon-derived newcastle disease virus recombinant vaccine strain as well as construction method and application thereof
  • Pigeon-derived newcastle disease virus recombinant vaccine strain as well as construction method and application thereof
  • Pigeon-derived newcastle disease virus recombinant vaccine strain as well as construction method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1: Rescue of LaSota strain

[0044] 1. Virus purification

[0045] The purification of the virus is to obtain a single virus clone. The LaSota strain was purified by the limiting dilution method. The detailed steps are as follows: the virus liquid is diluted 10 times, and after dilution, the virus liquid of each titer is inoculated for 9 to 11 days. Age SPF chicken embryos (100 μL / piece), inoculate 5 pieces for each dilution. After 4 days, the allantoic fluid of chicken embryos was harvested to measure the activity of HA. The allantoic fluid with the highest dilution factor of HA activity was selected as the next-generation purified virus fluid. Doubling dilution and chicken embryo inoculation were carried out in the same way, the virus was continuously purified for 5 generations, and the 5th generation virus solution was subpackaged and preserved, and used as the original seed virus for the next test.

[0046] 2. Determination of virus sequence

[0047] Usi...

Embodiment 2

[0068] Embodiment 2 Rescue of vaccine candidate virus for recombinant pigeons

[0069] 1. Virus screening and purification

[0070] The Newcastle Disease Virus of Gene VI.2.1.2.2, which is mainly prevalent in the current Chinese pigeon population, was selected in the laboratory for reproduction and rejuvenation, and then the five strains were purified by the limiting dilution method. See Table 5 for hemagglutination titers. Finally, the QH1344 / 2017 strain purified for 5 generations was selected as the original seed virus for the construction of full-length cDNA clone.

[0071] Table 5: Hemagglutination titers of different strains of pigeon-derived Newcastle disease virus in each generation

[0072]

[0073] 2. Determination and analysis of the full-length genome sequence of the QH1344 / 2017 strain

[0074]Using the purified QH1344 / 2017 viral reverse-transcribed genomic cDNA as a template, PCR amplification was performed on the QH1344 / 2017 genome using the designed 13 pair...

Embodiment 3

[0093] Example 3 rLa-VI-QH17 vaccine strain to non-immune pigeon safety test and immune effect test

[0094] 1. Preparation of inactivated vaccine

[0095] The rLa-VI-QH17 vaccine strain was diluted 10,000 times with sterilized normal saline, inoculated with 9-11-day-old SPF chicken embryos, 0.1 mL per embryo, and incubated at 37°C. Discard the dead embryos within 24 hours after inoculation, put the dead embryos at 4°C in time between 24h and 120h, collect the mixed samples, and measure the HA for seedling toxicity to be 2 10.5 , the virus titer was 10 9.48 EID 50 / 0.1mL. Introduce the Newcastle disease virus solution with a determined titer into the inactivation tank, add formaldehyde solution with a final concentration of 0.1%, make it fully mixed, inactivate at 4°C for 48 hours, and shake it every 2 hours during this period. The oil adjuvant inactivated vaccine was prepared from the inactivated virus stock solution by a conventional method. Prepare the same dose of LaS...

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Abstract

The invention provides a pigeon-derived newcastle disease virus recombinant vaccine strain. A gene segment used in a process of constructing the pigeon-derived newcastle disease virus recombinant vaccine strain contains envelope glycoprotein F protein genes and HN protein genes of a pigeon-derived newcastle disease virus with attenuated toxicity. A preservation number of the specific pigeon-derived newcastle disease virus recombinant attenuated vaccine strain is CGMCC No. 21900. The pigeon-derived newcastle disease virus recombinant vaccine strain provided by the invention has the biological characteristics of high growth titer and low pathogenicity in a chick embryo, is stable in heredity, has a good immune protection effect on the pigeon-derived newcastle disease virus, can effectively inhibit and expel toxins and can be used for preventing and controlling the pigeon-derived newcastle disease virus which is popular at present, and the blank of lack of special vaccine products for prevention and control of the pigeon-derived newcastle disease in our country is filled.

Description

technical field [0001] The invention belongs to the field of veterinary biological products, and in particular relates to a pigeon-derived Newcastle disease virus recombinant vaccine strain and its construction method and application. Background technique [0002] Newcastle disease (ND) is an important infectious disease caused by a virulent strain of Newcastle disease virus (NDV) infecting poultry. The disease is a highly contagious, acute septic infectious disease characterized by damage to the respiratory tract and digestive tract. It spreads quickly and has a high fatality rate, causing huge economic losses to the world's poultry industry. The World Organization for Animal Health (OIE) lists Newcastle disease as a legally notifiable animal disease, and the Ministry of Agriculture and Rural Affairs of my country lists it as a first-class animal disease. [0003] Newcastle disease virus (NDV) is a single-stranded, negative-strand, non-segmented RNA virus belonging to the ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/45C12N7/04C12N15/85A61K39/17A61P31/14C12R1/93
CPCC07K14/005C12N7/00C12N15/85A61K39/12A61P31/14C12N2760/18121C12N2760/18122C12N2760/18134C12N2760/18162A61K2039/5252A61K2039/5254A61K2039/552
Inventor 于晓慧王静静刘华雷舒波李峥李阳蒋文明
Owner CHINA ANIMAL HEALTH & EPIDEMIOLOGY CENT
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