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Seneca valley virus vaccine and its preparation method and application

A virus vaccine, Seneca's technology, applied in the field of biotechnology and biological products, can solve problems such as loss of control and severe forms of prevention and control

Active Publication Date: 2019-02-19
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, so far, there is still no commercial SVV vaccine available in the world, the disease is still out of control, and the prevention and control situation is severe

Method used

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  • Seneca valley virus vaccine and its preparation method and application
  • Seneca valley virus vaccine and its preparation method and application
  • Seneca valley virus vaccine and its preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Embodiment 1: Isolation, cultivation and screening of SVV / FJ / 001 strain

[0060] 1.1 Virus isolation

[0061] Collect the vesicle skin, vesicle fluid and other diseased tissues of SVV-infected pigs, add PBS to grind, after repeated freezing and thawing 3 times, centrifuge at 4°C, 5000rpm for 10min, collect the supernatant, filter through a 0.22μm filter, extract RNA, and use RT-PCR And QRT-PCR and other methods to identify that the disease material does not contain other pathogens such as foot-and-mouth disease virus that can cause vesicular lesions in pigs except for SVV.

[0062] Inoculate the above-mentioned identified disease material into newly formed BHK-21 cells at 10% of the virus culture solution, incubate at 37°C for 1 hour, add DMEM culture solution, culture at 37°C for 5 days, harvest the virus solution, and pass through 3 times After repeated freezing and thawing, the second generation was inoculated, cultured for 4 days, and the toxic medium was harvested...

Embodiment 2

[0077] Example 2: Identification of Seneca Valley Virus

[0078] 2.1 Identification of virus by RT-PCR

[0079] The supernatant of cells such as BHK-21 cells, PK-15 cells, ST cells, SK-RST cells, IBRS-2 cells, H1299 cells or 293T cells infected with the stably passaged SVV / FJ / 001 strain was used with RNAeasy Mini Kit (Qiagen ) to extract total RNA, after reverse transcription, amplify the P1 gene, purify and recover it, and then send it for sequencing. The results show that the P1 gene can be amplified.

[0080] 2.2 Identification of viral antigens by indirect immunofluorescence

[0081] After the culture of BHK-21 cells infected with SVV / FJ / 001 strain was repeatedly frozen and thawed, it was inoculated into a six-well plate with slides placed at the bottom and grown with BHK-21 cells (monolayer cells grew to 60%-70 %), placed in 5% CO 2 In a 37°C incubator, indirect immunofluorescence was performed according to the conventional method after 24 hours. The primary antibody w...

Embodiment 3

[0082] Example 3: Establishment of SVV / FJ / 001 strain to pig challenge model

[0083] Six test pigs were screened from non-epidemic areas, and tested by neutralization experiments, the SVV neutralizing antibody titer was 9 TCID 50 , at the same time set up grinding tissue disease material virus as a control, the challenge dose is 2mL / head, observe continuously for 15 days, observe and record the disease situation, judge the disease situation of animals according to the symptoms such as hooves, nose mirror and lips blisters, and count Points (5-point system, the judgment standard is formulated according to the foot-and-mouth disease scoring standard, the higher the score, the more serious the clinical symptoms). The results of the challenge showed that the SVV / FJ / 001 strain showed typical clinical symptoms from the second day after the challenge, with blisters appearing on the hooves, and some blisters appeared on the nose mirror later. After the tissue virus was challenged, cl...

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Abstract

The invention relates to separated Seneca valley virus nucleic acid, a Seneca valley virus strain, a Seneca valley virus vaccine comprising the Seneca valley virus strain and a preparation method thereof, and application of the Seneca valley virus vaccine to preparation of a medicine for preventing and / or controlling related diseases caused by an animal Seneca valley virus.

Description

technical field [0001] The invention relates to a virus strain, in particular to a Seneca Valley virus strain, a Seneca Valley virus vaccine comprising the Seneca Valley virus strain, a preparation method and application thereof, and belongs to the field of biotechnology and biological products. Background technique [0002] Seneca valley virus (SVV), also known as Seneca virus A (SVA), is the only member of the genus Senecavirus in the Picornaviridae family and is a single-stranded positive-sense RNA. SVV-infected pigs can cause primary vesicular disease in pigs, causing vesicular lesions on the snout and crown of the hooves, accompanied by clinical manifestations such as lameness, anorexia, and lethargy. It is indistinguishable from clinical symptoms caused by foot-and-mouth disease, swine vesicular disease and vesicular stomatitis. SVV infection can cause vesicular lesions in weaned piglets, nursery pigs, fattening pigs and breeding pigs of all ages, accompanied by acute...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/41C12N7/00A61K39/125A61P31/14C12R1/93
CPCA61K39/12A61K2039/5252A61K2039/552C07K14/005C12N7/00C12N2770/32021C12N2770/32022C12N2770/32034
Inventor 郑海学杨帆朱紫祥刘华南张克山田宏曹伟军李丹马旭升靳野冯霞郭建宏何继军刘湘涛
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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