A kind of bovine rotavirus, bovine coronavirus double inactivated vaccine and preparation method thereof

A dual inactivated vaccine and bovine rotavirus technology, applied in biochemical equipment and methods, viruses, vaccines, etc., can solve problems such as the difficulty of virus separation, achieve safe and reliable vaccines, and prevent calves from diarrhea

Active Publication Date: 2022-07-26
QILU ANIMAL HEALTH PROD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the difficulty of BRV virus isolation, it needs to be treated with a certain concentration of proteolytic enzymes such as trypsin to effectively infect cells.

Method used

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  • A kind of bovine rotavirus, bovine coronavirus double inactivated vaccine and preparation method thereof
  • A kind of bovine rotavirus, bovine coronavirus double inactivated vaccine and preparation method thereof
  • A kind of bovine rotavirus, bovine coronavirus double inactivated vaccine and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] Embodiment 1---the preparation of bovine rotavirus, bovine coronavirus double inactivated vaccine (JY strain+YQ strain)

[0065] 1. Preparation of BRV-JY strain and BCV-YQ strain antigen by using roller bottle adherence

[0066] (1) Cultured cells: The seed cells MA-104 and HCT-8 were respectively recovered into cell flasks, grown for about 48 hours, digested with EDTA-trypsin cell dispersion, and passaged at a volume ratio of 1:4. Add serum-free DMEM medium (pH value is 7.0) for spinner flask culture, and when the cells are 90% to 100% full, they are used for continued passage or virus inoculation.

[0067] (2) Preparation of antigen (virus solution) for seedling preparation: Take the well-grown MA-104 cells, wash 2-3 times with PBS, and inoculate the BRV-JY strain at a dose of (V / V) 0.5%. inoculation. Serum-free DMEM medium (pH 7.0) was added for culture. After 5 to 7 days, when 80% of the cells appeared CPE, the spinner flask was placed at -20°C for 2 to 3 times o...

Embodiment 2

[0084] Embodiment 2---finished product inspection of bovine rotavirus, bovine coronavirus double inactivated vaccine (JY strain+YQ strain)

[0085] 1. Sterility test: According to the appendix of the current "Chinese Veterinary Pharmacopoeia", it should meet the regulations. The above test results are shown in Table 2.

[0086] Table 2 Physical properties, sterility test results

[0087]

[0088] 2. Safety inspection:

[0089] (1) Alternative animal test: 7 healthy rabbits of 1.5-2.0 kg were selected for the test, including 5 in the immunization group and 2 in the control group. The rabbits in the immunization group were injected subcutaneously with 1.0ml of vaccine (two-point injection, 0.5ml) / point), the control group rabbits were subcutaneously injected with 1.0ml of cell culture in each neck (two-point injection, 0.5ml / point), observed continuously for 7 days and measured the body temperature every afternoon at a fixed point. There is no abnormality to touch, no sym...

Embodiment 3

[0105] Example 3 - Antibody tracking test of newborn calves

[0106] Select 10 dry cows with BRV, BCV antigen and antibody double negative (antigen negative means RT-PCR detection is negative, antibody negative means serum neutralizing antibody titer ≤ 1:4), including 5 in the immunization group and 5 in the control group , 2.0ml of vaccine was injected into each neck muscle of cows in the immunization group, and 2.0ml of cell culture was injected into each neck muscle of cows in the control group. After 21 days, the immunization was boosted in the same way. After the test cows gave birth, blood was collected from the newborn calves that did not eat colostrum, and the neutralizing antibody titers of BRV and BCV were detected. Blood was collected from calves 60 days after birth to detect BRV and BCV neutralizing antibody titers. The 4 / 5BRV neutralizing antibody titers of immunized cows calving ≥1:64, and the 4 / 5BCV neutralizing antibody titers ≥1:64, the results are shown in T...

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Abstract

The invention relates to a double inactivated vaccine for preventing bovine rotavirus and bovine coronavirus and a preparation method thereof. The active components of the vaccine of the present invention include inactivated antigens of bovine rotavirus and bovine coronavirus, and the antigen components of the double inactivated vaccine can be suspended by passaged cell lines (for bovine rotavirus and bovine coronavirus) or microcarriers Cultivated (for bovine rotavirus) for virus propagation, and optimized the seedling production process. The results of vaccine safety, efficacy test and antibody tracking show that there is no adverse reaction after immunizing cattle with the dual inactivated vaccine of the present invention, and no abortion is caused after immunizing pregnant cattle. All the immunized cows produced immune protection, and the newborn calves could acquire high levels of maternal antibodies through breast milk. The results show that the vaccine of the present invention is safe and reliable, and can be used to prevent the occurrence of diarrhea in calves.

Description

technical field [0001] The invention relates to a dual inactivated vaccine of bovine rotavirus and bovine coronavirus and a preparation method thereof, belonging to the field of veterinary biological products. Background technique [0002] Calf diarrhea is a disease that causes high mortality in newborn calves and can cause serious economic losses. Its pathogens mainly include Bovine Rotavirus (BRV) and Bovine Coronavirus (BCV). BRV belongs to the family Reoviridae and Rotavirus. It is a double-stranded RNA virus that mostly infects calves within 1 month of age. The clinical symptoms are mainly watery diarrhea, mental depression, loss of appetite, severe dehydration and acidosis . BCV belongs to the family Coronaviridae and belongs to the genus Coronavirus. It is a single-stranded positive-stranded RNA virus that mainly encodes five proteins including S, M, N, E and HE. In addition to causing diarrhea in calves, it can also cause winter dysentery and hemorrhagic diarrhea in...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K39/295A61K39/215A61K39/15A61P31/14
CPCA61K39/12A61P31/14A61K2039/70A61K2039/552A61K2039/5252C12N2720/12334C12N2770/20034
Inventor 毕玉彧张娣高凤宋晓飞李营贾爱琴秦绪伟王蕾徐龙涛
Owner QILU ANIMAL HEALTH PROD
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