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A kind of attenuated strain of muscovy duck C subtype avian metapneumovirus and its preparation and application

A technology of avian metapneumovirus and attenuated strains, applied in the direction of viruses, antiviral agents, viruses/phages, etc.

Active Publication Date: 2021-07-30
WENS FOODSTUFF GRP CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Duck-derived subtype C avian metapneumovirus has only been reported in France and Canada, and attenuated vaccines for Muscovy duck-derived aMPV-C subtype have not yet been reported

Method used

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  • A kind of attenuated strain of muscovy duck C subtype avian metapneumovirus and its preparation and application
  • A kind of attenuated strain of muscovy duck C subtype avian metapneumovirus and its preparation and application
  • A kind of attenuated strain of muscovy duck C subtype avian metapneumovirus and its preparation and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] The subculture of embodiment 1 duck C subtype metapneumovirus

[0024] Use the duck subtype C avian metapneumovirus S-01 strain that is isolated in the laboratory and causes Muscovy ducks to be morbid in nature, inoculate the African green monkey kidney cell (Vero) T25 cell bottle that is covered with a single layer, and inoculate 100 times in each bottle Dilute 500ul of the virus solution, after 2 hours of adsorption, discard the virus solution, add 5ml of DMEM culture solution containing 2% fetal bovine serum, and cultivate for 24 to 96 hours until cytopathic changes appear, freeze and thaw 3 times repeatedly, and absorb the virus solution Inoculate the next generation. Continuously subcultured to 40 generations, and then continuously purified 5 times by limiting dilution method, and passed to 62 generations.

[0025] Subculture Results

[0026] F3 generation duck embryo yolk liquid was inoculated with Vero cells, F1-F3 generation had no obvious lesions, but F4 gene...

Embodiment 2

[0029] Embodiment 2C subtype avian metapneumovirus S-01 strain passage pathogenicity test

[0030] Select the above-mentioned S-01 strains for passage to generations S-01-5P, S-01-17P, S-01-28P, S-01-32P, S-01-40P, S-01-50P, S-01- 60P, Vero cell solution control group, a total of 9 groups, infected two-week-old ducklings with eye drops and nasal drops, and the infection dose was 10 4 TCID 50 / ml, observe the clinical symptoms every day after immunization and record the number and severity of diseased ducks in each group, collect the cloacal swabs and throat swabs of each group on the 5th day after immunization, and put the swabs of every 5 ducks in In a 15ml EP tube, carry out RT-PCR detection, a total of 6 samples to be tested in each group. By observing the clinical symptoms and lesions after necropsy, the pathogenicity of different generations to ducks was analyzed. (RT-PCR detection method refers to Ali A, Reynolds DL: A reverse transcription-polymerase chain reaction a...

Embodiment 3

[0035] Example 3C subtype avian metapneumovirus S-01 strain attenuated generation vaccine efficacy experiment

[0036] The weakened passages in the above results were subjected to immunogenicity and different titer immune challenge tests. F40, F50, and F60 were selected for the test, and the specific implementation was 200 1-day-old young muscovy ducks (maybe aMPV and The detection of aMPV was negative) were randomly divided into 10 groups, 20 in each group. The 40th generation, 50th generation, and 60th generation were divided into three gradients (10 2.5 、10 3.5 、10 4.5 TCID 50 / 0.1ml). Vero cell solution 0 is the blank group of the tenth group. The ducklings were immunized by eye drops and nasal drops at 2w. Observe the clinical symptoms every day after immunization, collect cloacal swabs and throat swabs on the 5th day, randomly collect 10 ducks in each group, and put them in 2ml EP tubes respectively. Add 300ul of normal saline containing 2000U double antibody to e...

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Abstract

The invention belongs to the field of microbial animal viruses, and in particular relates to an attenuated strain of muscovy duck subtype C avian metapneumovirus and its preparation and application. The attenuated Muscovy duck subtype C avian metapneumovirus strain of the present invention has a preservation number of CCTCC NO: V201823, a nucleotide sequence of SEQ ID NO: 1, and an amino acid sequence of SEQ ID NO: 2 to SEQ ID NO: 9. The attenuated muscovy duck subtype C avian metapneumovirus strain of the present invention is prepared by subculturing the C subtype avian metapneumovirus S-01 strain to 50 generations. The invention obtains the attenuated culture strain of Muscovy duck subtype C avian metapneumovirus S-01 strain for the first time, laying a foundation for the development of the attenuated vaccine.

Description

technical field [0001] The invention belongs to the field of microbial animal viruses, and in particular relates to an attenuated strain of muscovy duck subtype C avian metapneumovirus and its preparation and application. Background technique [0002] Avian metapneumovirus is one of the acute and highly contagious diseases of upper respiratory tract infection caused by avian metapneumovirus (aMPV). Avian lung disease was first isolated in young turkeys, accompanied by clinical symptoms such as sneezing, tracheal rales, sticky secretions in the nose and eyes, and conjunctivitis. Therefore it is called turkey rhinotracheitis. In South Africa in 1978, aMPV was first isolated and subsequently reported in Europe, Asia and South America. In 1998, my country first reported the isolation of avian pneumovirus A subtype in chickens with swollen head syndrome, and in 2012, B and C subtypes were successively isolated in chickens. In July 2010, in a duck farm in Guangdong Province, a ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00C12N7/08A61K39/155A61P31/14C12R1/93
CPCA61K39/12A61K2039/5254A61K2039/552A61P31/14C12N7/00C12N2760/18321C12N2760/18334C12N2760/18364
Inventor 刘佳佳陈峰王占新鲁俊鹏覃健萍操胜
Owner WENS FOODSTUFF GRP CO LTD
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