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Largemouth bass iridovirus disease inactivated vaccine and preparation method thereof

A technology of inactivated vaccines and iridescent viruses, applied in biochemical equipment and methods, vaccines, viruses, etc., can solve the problems of largemouth bass economic losses, threats to the healthy development of the industry, and high lethality, and achieve good immune protection effects and production Low cost and good safety performance

Active Publication Date: 2020-05-12
ZHEJIANG INST OF FRESH WATER FISHERIES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the aquaculture output of largemouth bass has exceeded 500,000 tons. The disease occurs in all major breeding areas, and the incidence is high in summer. When the disease occurs, the water temperature is 25-30 ℃, which endangers adult fish and has a high mortality rate, which has caused major damage to the largemouth bass aquaculture industry. Economic loss, a serious threat to the healthy development of the industry

Method used

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  • Largemouth bass iridovirus disease inactivated vaccine and preparation method thereof
  • Largemouth bass iridovirus disease inactivated vaccine and preparation method thereof
  • Largemouth bass iridovirus disease inactivated vaccine and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] The present embodiment provides a kind of preparation method of largemouth bass iridovirus disease inactivated vaccine, comprises the following steps:

[0027]Step 1, EPC cell culture: Take 1 bottle of EPC that is full of monolayers, discard the old medium in a sterile ultra-clean bench, add 2ml of trypsin digestion solution with a concentration of 0.25% (V / V) to each bottle, and digest for 1min Quickly add 2ml of 199 medium containing 10% (V / V) calf serum with a pH of 7.4, gently blow the bottom of the cell bottle with a pipette, and then add 6ml of 199 medium containing 10% (V / V) calf serum medium to suspend cells. Take 2 T25 cell culture flasks, add 4ml of cell suspension to each flask, and culture them in a 25°C incubator. After the cells form a confluent monolayer and cover the culture dish, EPC cells are obtained for virus amplification.

[0028] Step 2, LMBV virus amplification: suck out the culture medium in the EPC cells, and then add the infected virus suspen...

Embodiment 2

[0034] Determination of LMBV inactivation conditions and safety test.

[0035] Obtain the amplified virus stock solution according to the method of Example 1, divide it into 3 equal parts, add BEI to the final concentration of BEI respectively 1%, 2%, and 2%, and inactivate it in a constant temperature shaker at 37°C at 120r / min for 24h, 48h After 72 hours, the inactivation was terminated with the same concentration of sodium thiosulfate solution and samples were taken. Take the prepared vaccine and inoculate EPC cells according to the above virus propagation method, and set up a negative control at the same time, observe for 7 to 10 days, if cytopathic effect appears, it indicates that the virus inactivation is not complete; if no cytopathic effect is seen, then blindly pass 2 Second, if cytopathy occurs in the blind transmission, it indicates that the virus inactivation is still incomplete. If no cytopathy occurs in the two blind transmissions, it indicates that the virus in...

Embodiment 3

[0041] Safety Test of Inactivated Vaccines

[0042] Sterility test: Take the vaccine prepared in Example 1, inoculate the brain infusion bacterial culture medium (BHI) plate, smear the plate with the streak method and incubate at 30°C for 15 days. If there are colonies growing, it indicates that the vaccine has bacterial contamination ; if no colonies grow, the vaccine is sterile.

[0043] Fish body safety test: Take the vaccine prepared above and inject 30 healthy largemouth bass of 50g to 70g, the injection dose is 0.1 to 0.2ml / tail, and the negative control is injected with the same dose of normal saline. After feeding for 15 to 30 days, if the vaccine group died or had clinical symptoms, but the negative control group did not show clinical symptoms or died, it indicated that the vaccine was unsafe; if neither the vaccine group nor the negative control group showed clinical symptoms or died, it indicated that the vaccine Safety.

[0044] Stress test and feeding effect aft...

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Abstract

The invention belongs to the technical field of biological medicines for veterinary use and in particular relates to a largemouth bass iridovirus disease inactivated vaccine and a preparation method thereof. The vaccine comprises EPC (epithelioma papulosum cyprinid) cells and largemouth bass ranavirus LMBV. The preparation method comprises the following steps: culturing EPC cells; performing LMBVvirus amplification on the cultured EPC cells so as to obtain an LMBV virus liquid; and performing inactivation treatment on the LMBV virus liquid. The vaccine prepared by the invention is good in immune protection effect and can be applied to prophylactic immunization of largemouth bass iridovirus, and the survival rate and the breeding benefits of largemouth bass can be increased.

Description

technical field [0001] The invention belongs to the technical field of veterinary biomedicine, and in particular relates to an inactivated vaccine for largemouth bass iridescent virus disease and a preparation method thereof. Background technique [0002] With the rapid development of my country's aquaculture industry, aquatic animal diseases, especially outbreaks of epidemics caused by viruses, have increased significantly, and the damage is extremely serious. Among the reported viruses, the iridescent virus is a more common virus. Iridoviruses mainly infect invertebrates and lower vertebrates. The viruses of the family Iridoviridae are all icosahedral viruses with a diameter of 120-300nm. 200kb. Including 5 genera, of which Ranavirus, Megalocytivirus and Lymphocystivirus can infect vertebrates, mainly fish, amphibians and reptiles, etc., while iridovirus (Iridovirus), Chloriridovirus infect invertebrates. A variety of iridescent viruses have been isolated and observed ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/12A61K39/39A61P31/20
CPCA61K39/12A61K39/39A61P31/20A61K2039/5252C12N2710/00034A61K2039/55566A61K2039/552Y02A50/30Y02A40/81
Inventor 沈锦玉姚嘉赟潘晓艺蔺凌云曹铮夏焱春尹文林刘忆瀚
Owner ZHEJIANG INST OF FRESH WATER FISHERIES
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