Attenuated strain of grass carp reovirus II and application of attenuated strain

A technology of reovirus and attenuated strains, applied in the directions of viruses, antiviral agents, viral antigen components, etc., can solve the problem that the protection power needs to be improved, etc.

Active Publication Date: 2020-08-28
广东富伦德生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Regarding the above-mentioned attenuated strains, there are still problems. First, the attenuated strain can cause about 10% mortality of the rare goner carp, which has a certain degree of virulence. Protection still needs to be improved

Method used

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  • Attenuated strain of grass carp reovirus II and application of attenuated strain
  • Attenuated strain of grass carp reovirus II and application of attenuated strain
  • Attenuated strain of grass carp reovirus II and application of attenuated strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Isolation and cultivation of grass carp reovirus type II:

[0029]The applicant purchased healthy grass carp for the test from a farm in Sanshui District, Foshan City, Guangdong Province. When using conventional PCR methods to exclude whether the test fish carried GCRV pathogens, GRCVⅡ virus was detected from one of the fish (this virus is present in this country). Invention or referred to as GCRV II-GD1207). Take the liver, kidney and spleen of GCRV-positive fish, wash them twice with PBS (pH7.2), weigh them, cut the tissues into small pieces, grind them with a tissue homogenizer, and use M199 medium to make 1: 10 tissue homogenate, after repeated freezing and thawing for 3 times, it was centrifuged at 3000rpm / min, 6000rpm / min and 10000rpm / min in turn, each time for 10min to remove sediment, and the supernatant was filtered through a microporous filter with a filter membrane pore size of 0.22μm Sterilized spare. After the CIK cells grown into a dense monolayer were r...

Embodiment 2

[0040] The safety of GCRV II-GD1207 in the preparation of grass carp hemorrhagic disease live vaccine:

[0041] After the prepared CIK cell virus liquid was filtered and sterilized, it was diluted to a GCRV II-GD1207 virus titer of 80 copies / μL, and then injected with grass carp with an average body length of 10-15 cm (weight of 20-25 g) and an average body length of 4 ±0.5cm (body weight: 5±0.5g) rare gobi carp, and a negative control group injected with sterile PBS, each grass carp intraperitoneally injected with 0.2ml, and rare gobi carp with 0.05ml intraperitoneally injected, each group of test fish Both are 20 tails. During the test, the water temperature was controlled at about 28°C, and the disease and death of the test fish in each group were observed every day for 3 consecutive weeks. At the same time, during the experiment, RT-PCR was used to detect the GCRV virus in the fish that did not die or became ill after 2 weeks of challenge.

[0042] The results showed tha...

Embodiment 3

[0044] Application of GCRV II-GD1207 in the preparation of grass carp hemorrhagic disease live vaccine:

[0045] Healthy grass carp (average body length 10-15cm, body weight 20-25g) not carrying GCRV virus were randomly divided into 4 groups, injection immunization group, immersion immunization group, oral immunization group and control group, 60 fish in each group. Dilute the attenuated vaccine virus content to 80copies / μL, inject 200μL per tail in the immunized group, soak the final concentration of vaccine virus in the immunized group to 10copies / μL, add 1% NaCl to soak for 1 hour, and oral vaccine with 1×10 3 Copies / μL GCRV II-GD1207 virus solution is sprayed on the surface of conventional fish feed by spraying, spraying 100mL of virus solution per kg of feed, drying at room temperature for 1-2 hours, and then spraying with appropriate amount of cod liver oil and virus Mix the feed with liquid, so that the cod-liver oil is wrapped on the outermost surface of the feed, so a...

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Abstract

The invention belongs to the technical field of vaccine preparation, and particularly relates to an attenuated strain of a grass carp reovirus II and an application of the attenuated strain. The preservation number of the attenuated strain is CCTCC NO:V201867. A live vaccine prepared from the attenuated strain can prevent diseases caused by the grass carp reovirus II; the immune protection effectfor the same genotype reaches 95% or above during injection use, reaches 65% or above during soaking immunization and reaches 40% or above during oral immunization; no stress reaction is generated toa body; the risk of virulence return is avoided; the vaccine can be applied to prevention of hemorrhagic diseases, especially viral hemorrhagic diseases of grass carp and black carp caused by the grass carp reovirus II; the morbidity of the hemorrhagic diseases of the grass carp and the black carp is reduced; and the survival rate of the grass carp and the black carp is increased.

Description

technical field [0001] The invention belongs to the technical field of vaccine preparation, in particular to an attenuated grass carp reovirus type II strain and application thereof. Background technique [0002] Grass carp (Ctenopharyngodon idella) is an important freshwater aquaculture fish in China. It has been cultured in my country for more than 60 years and is one of the most important freshwater fish species in the world. In 2018, the total output of freshwater fish was 29.5983 million tons, of which the output of grass carp was 5.345 million tons, accounting for 18.4% of the total output of freshwater aquaculture species, with an output value of more than 5 billion, ranking first among freshwater fish aquaculture species and having an important economic position. Grass carp hemorrhage disease (GCHD) caused by Grass Crap Reovirus (GCRV) is the most important pathogen causing the death of grass carp. Grass carp farming. GCRV belongs to the Reoviridae family (Reovirid...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00A61K39/15A61P31/14A61P7/04C12R1/93
CPCC12N7/00A61K39/12A61P31/14A61P7/04C12N2720/12221C12N2720/12234A61K2039/5254A61K2039/552A61K2039/542A61K2039/541Y02A50/30
Inventor 曾伟伟张同报王翠常丹丹
Owner 广东富伦德生物科技有限公司
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