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Construction and screening of rabies virus Evelyn-Rokitnicki-Abelseth (ERA) attenuated live vaccine candidate strain

A technology of rabies virus and attenuated vaccines, which is applied in antiviral agents, inactivated/attenuated, medical preparations containing active ingredients, etc., can solve the problems of unpredictability and long cycle of live attenuated vaccines, and achieve Save operating time, have a good technical platform, and improve rescue efficiency

Active Publication Date: 2013-07-31
HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Another reason is that the development cycle of live attenuated vaccines using traditional methods is long and unpredictable. [5，6]

Method used

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  • Construction and screening of rabies virus Evelyn-Rokitnicki-Abelseth (ERA) attenuated live vaccine candidate strain
  • Construction and screening of rabies virus Evelyn-Rokitnicki-Abelseth (ERA) attenuated live vaccine candidate strain
  • Construction and screening of rabies virus Evelyn-Rokitnicki-Abelseth (ERA) attenuated live vaccine candidate strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 Construction and biological activity identification of rabies virus GP333 position R→E mutant strain

[0037] 1 Materials and methods

[0038] 1.1 Cells and viruses

[0039] Vero cells (African green monkey kidney cells, ATCC No.CCL-81): the culture medium is DMEM containing 10% fetal bovine serum; NA cells (neuroma cells, purchased from Changchun Academy of Military Medical Sciences): the culture medium is containing 10% MEM of fetal bovine serum; BHK-21 cells (suckling hamster kidney cells, ATCC No.CCL-10): the medium is DMEM containing 5% fetal bovine serum; the rabies virus vaccine strain used in the test is ERA strain vaccine (purchased from China Veterinary Drug Control Institute, AV61), the standard strain of rabies virus CVS used in the virus neutralization experiment was purchased from Changchun Academy of Military Medical Sciences.

[0040] 1.2 Main reagents and instruments

[0041] PrimerSTAR HS DNA Polymerase, T 4 DNA ligase and restriction en...

Embodiment 2

[0078] Example 2 Pathogenicity of rabies virus rRV-G333R / E mutant strain

[0079] Unless otherwise specified, the materials and methods of this example are the same as Example 1.

[0080] Pathogenicity detection of rRV-G333R / E mutant strain

[0081] Rabies virus ERA wild-type control strain (10 6 PFU / mL), rRV-G333R / E mutant strain (10 7 PFU / mL) stock solution was used to inoculate suckling mice and adult Balb / c mice (3 weeks old, 10-13g, purchased from Beijing Weitong Lihua Experimental Animal Technology Co., Ltd. 10 mice after inoculation were raised in negative pressure cages with high-efficiency filtration devices, observed daily until 28 days after inoculation, and recorded the number of survivors. The mice that died within 4 days after inoculation were considered non-specific deaths and were not counted.

[0082] Results: Intracerebral inoculation of rabies virus ERA wild-type control strain and rRV-G333R / E mutant strain in suckling mice were all fatal. All the suckli...

Embodiment 3

[0083] Example 3 Immunogenicity of rabies virus rRV-G333R / E mutant strain

[0084] Unless otherwise specified, the materials and methods of this example are the same as Example 1.

[0085] Detection of immunogenicity of rRV-G333R / E mutant strain

[0086] 1. Immunogenicity of rRV-G333R / E mutant strain to mice

[0087] Rabies virus ERA wild-type control strain (10 6 PFU / mL), rRV-G333R / E mutant strain (10 7 PFU / mL) stock solution was diluted to 10 6 PFU / mL and 2*10 6 PFU / mL, artificially immunize 5-week-old adult Balb / c mice (purchased from Beijing Weitong Lihua Experimental Animal Technology Co., Ltd.) with 100 μL / only by intramuscular injection. In the non-immune control group, 21 days after immunization, the virus of the same generation was used to boost immunization by intramuscular injection with the same dose. Three weeks after the booster immunization, blood was collected to determine the serum neutralizing antibody titer.

[0088] 2. Immunogenicity of rRV-G333R / E mu...

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Abstract

The invention relates to a rabies virus attenuated vaccine mutant strain. Arginine on the 333rd site of the glucoprotein of the rabies virus mutates into glutamic acid. The invention also relates to a preparation method and application of the rabies virus attenuated vaccine mutant strain.

Description

technical field [0001] The invention relates to the field of recombinant virus vaccines, more specifically, the invention relates to a rabies virus attenuated vaccine mutant strain, wherein the 333 position of the glycoprotein of the rabies virus is mutated from arginine to glutamic acid. The invention also relates to the preparation method and application of the rabies virus attenuated vaccine mutant strain. Background technique [0002] Rabies is a zoonotic disease with a high mortality rate caused by the rabies virus [1] . In the past two years, the incidence and death of human rabies in my country have exceeded 3,000 people per year, and the epidemic situation has shown a significant upward trend, posing a great threat to human health and social stability [2，3] . Epidemiological surveys show that almost all warm-blooded animals are susceptible to rabies virus. Wild animals are the natural storage hosts of rabies virus, and domestic animals such as dogs and cats are t...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/04A61K39/205A61P31/14
Inventor 步志高夏咸柱葛金英王喜军杨松涛冯娜
Owner HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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