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Preparation method and application of attenuated goose astrovirus

An astrovirus and attenuated technology, which is applied in the field of goose astrovirus attenuated preparation, can solve the problems of potential safety hazards, short protection time, strong virulence, etc., and achieve short virus attenuation time, low cost and high virulence stable effect

Pending Publication Date: 2020-01-17
TIANJIN RINGPU BIO TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, the methods that can prevent goose gout are mainly vaccines, most of which are inactivated vaccines, but inactivated vaccines cannot induce a strong immune response, and the protection time is short, often requiring multiple immunizations
Although the attenuated live vaccine can induce a strong immune response and has a long acting time, there are safety hazards and high cost. In addition, the attenuated vaccine also has the risk of returning to strong virulence after immunization

Method used

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  • Preparation method and application of attenuated goose astrovirus
  • Preparation method and application of attenuated goose astrovirus
  • Preparation method and application of attenuated goose astrovirus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1: Isolation and Identification of Goose Astrovirus

[0029] 1. Isolation and cultivation of goose astrovirus

[0030] Take liver, spleen, kidney and other tissues of dead goslings, add sterile saline at a ratio of 1:4 for homogenization, place the homogenate in the refrigerator for 3 times, centrifuge at 6000r / min for 15min, and take the supernatant The bacteria were filtered through a 0.22 μm filter, and the virus solution was stored at -20°C.

[0031] 0.1 ml of virus liquid homogenate was continuously cultured on LMH cells for 4 generations with 0.1-2% serum, and the culture concentration of virus serum was 0.5-1%. With the development of 70%-80% cytopathic effect, cell cultures were frozen and stored at -80°C.

[0032] 2. Identification of Goose Astrovirus

[0033] Carry out viral RNA extraction to LMH cell culture, extraction method uses RNA / DNA virus extraction kit (Takara),

[0034] Utilizing the published goose Astrovirus sequence and the conserved O...

Embodiment 2

[0052] Embodiment 2. The preparation method of Goose Astrovirus Attenuated Virus

[0053] Use the same batch of isolated virus liquid to prepare attenuated virus, and adopt the following methods respectively:

[0054] 1. Conventional method

[0055] The isolated goose star virus liquid is inoculated with goose embryos and hatched. The hatching temperature is controlled at 35°C-38°C, and the allantoic fluid of goose embryos is collected after 6-7 days;

[0056] 2. High temperature treatment method

[0057] Treat the isolated goose star virus liquid in a water bath at 50°C-56°C for 55-65min, then inoculate the goose embryo and incubate it. The incubation temperature is controlled at 35°C-38°C, and the goose embryo allantois is harvested after 6-7 days liquid;

[0058] 3. Low temperature incubation method

[0059] Inoculate goose embryos with the isolated goose star virus liquid and incubate them. The incubation temperature is controlled at 29°C-30°C, and the allantoic fluid ...

Embodiment 3

[0076] Example 3. Monitoring of the immune effect of the inactivated vaccine prepared by goose Astrovirus

[0077] The virus processed by high-temperature method, low-temperature method, and high-low temperature combination method in Example 2, and untreated natural strength, and natural attenuated virus are prepared into inactivated vaccines, and the inactivated vaccines are necked at a dose of 2.0ml / only. 10 breeding geese were injected subcutaneously or intramuscularly, and 10 controls were set up. After the primary immunization, the blood was collected at the third, fourth, fifth, sixth, seventh, and eighth weeks to separate the serum, and the serum antibody titer was determined by the agar immunodiffusion test. The results showed that at the fifth week, after the high temperature in Example 2 Method, low temperature method, high and low temperature combination method The antibodies produced after the virus inactivated vaccine immunization can reach the level of antibodies...

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Abstract

The invention provides a preparation method of attenuated goose astrovirus and a live vaccine prepared by applying the attenuated goose astrovirus, and belongs to the technical field of waterfowl disease prevention and control. A method combining high-temperature treatment and low-temperature incubation is adopted to attenuate the virus. The method has the advantages of short virus attenuation time, low cost and long immune protection time. There is virulence return after multiple passages, thus effectively preventing goose gout disease caused by goose astrovirus.

Description

technical field [0001] The invention belongs to the field of virus preparation, and in particular relates to a preparation method and application of attenuated goose astrovirus. Background technique [0002] Goose gout is a disease caused by abnormal protein metabolism of goose. The main feature is that a large amount of urate is produced and deposited in the body. There is a large amount of urate deposition in the heart, liver, kidney, blood, and joints, which leads to goose standing. Unstable strength, less food, weight loss and even death. At the beginning of 2017, a large number of goose gout outbreaks occurred in Shandong, Anhui, Guangdong, Guangxi and other places in my country, causing huge economic losses to farmers. A new type of goose astrovirus was isolated from gooses, and it was found that goose astrovirus infection was the main factor inducing goose gout. [0003] At present, the methods that can prevent goose gout are mainly vaccines, most of which are inactiva...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/08A61K39/12A61P31/14C12R1/93
CPCC12N7/00A61K39/12A61P31/14C12N2770/12064C12N2770/12034A61K2039/5254A61K2039/552
Inventor 李守军刘冠星杨燚郁宏伟康亚男陈冰
Owner TIANJIN RINGPU BIO TECH
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