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Eurasian avian like H1N1 swine influenza A virus cold-adapted attenuated virus strain and application thereof

A technology for H1N1 and swine flu, applied in veterinary vaccines, antisense single-stranded RNA viruses, medical preparations containing active ingredients, etc., can solve problems such as long residual time and lack of virus cross-protection

Active Publication Date: 2017-11-03
HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Generally speaking, inactivated vaccines have good immune protection and can protect against the same subtype of influenza virus. The disadvantage is that they cannot induce effective mucosal immune antibodies and cellular immune responses. Therefore, they lack protection against other subtypes or lineage viruses. Cross-protection, and the route of vaccination is usually through intramuscular injection, the vaccine needs to be added with corresponding adjuvants, and there will be a long residual time in the body

Method used

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  • Eurasian avian like H1N1 swine influenza A virus cold-adapted attenuated virus strain and application thereof
  • Eurasian avian like H1N1 swine influenza A virus cold-adapted attenuated virus strain and application thereof
  • Eurasian avian like H1N1 swine influenza A virus cold-adapted attenuated virus strain and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 Preparation of Eurasian avian H1N1 subtype swine influenza cold-adapted attenuated vaccine strain A / swine / Guangxi / 18 / 2011ca (H1N1)

[0027] 1. Take out one copy of the Eurasian avian H1N1 subtype swine influenza virus strain A / swine / Guangxi / 18 / 2011(H1N1) (GX18 for short) stored in our laboratory, dissolve it, and place it on ice to determine blood coagulation ( HA) price, and calculate 4HA unit / 0.2mL;

[0028] 2. 10-day-old SPF chicken embryo allantoic cavity was inoculated with 0.2 mL of GX18 virus diluent containing 4 HA units (diluted with phosphate buffer solution of pH 7.2 to 7.4);

[0029] 3. Incubate at 33°C for 72 to 96 hours and determine the HA value. When the HA value reaches 128 to 256, harvest the virus solution and use it as the next incubation temperature (1°C as the unit, gradient reduction) seed virus, continue to step 2 experiment.

[0030] In this way, 17 generations were uploaded on 10-day-old SPF chicken embryos, and a Eurasian avian H1N1 subtype...

Embodiment 2

[0035] Example 2 Determination of cold-adapted (ca) of swine influenza cold-adapted attenuated vaccine strain GX18ca

[0036] Cold adaptability (ca) is to compare the replication titers of virus strains on chicken embryos under 33°C suitable growth temperature conditions and 25°C low temperature growth conditions. Under the low temperature condition of 25℃, if the difference between the growth titer of the virus on the chicken embryo and the growth titer under the growth condition of 33℃ is within 100 times, the virus is considered to have cold adaptability characteristics, and vice versa .

[0037] Method: Dilute the virus GX18ca and the corresponding wild virus GX18 in a 10-fold ratio on ice, namely 100 μL virus + 900 μL PBS, and mix well. Pipette 100 μL of virus dilution into the next tube of 900 μL PBS, mix well, and then By analogy, dilute to 10 -9 . The diluted virus was inoculated with 4 10-day-old SPF chicken embryos for each dilution, and the inoculated chicken embryos w...

Embodiment 3

[0040] Example 3 Determination of temperature sensitive (ts) of swine influenza cold-adapted attenuated vaccine strain GX18ca

[0041] Temperature sensitivity (ts) is to compare the replication titer of virus strains on chicken embryos under suitable growth temperature of 33°C and high temperature of 39°C. If the growth titer of the virus on the chicken embryo under the high temperature condition of 39℃ is more than 100 times lower than its growth titer at the appropriate growth temperature of 33℃, then the virus is considered to have temperature sensitivity characteristics, and vice versa Of course.

[0042] Method: Dilute the virus 10 sequentially 2~7 To determine the EID of GX18ca and the corresponding wild virus GX18 at 33℃ or 39℃ 50 After 72 hours of incubation, the hemagglutination test was used to detect the replication of the virus at each temperature, and the Reed-Muench method was used to calculate the half of the chicken embryo infection dose (EID 50 ). The same metho...

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Abstract

The invention discloses a Eurasian avian like H1N1 swine influenza A virus cold-adapted attenuated virus strain and an application thereof. Eurasian avian like H1N1 swine influenza A viruses A / swine / Guangxi / 18 / 2011 (H1N1) are subjected to serial sub-culturing at the temperature of 33-25 DEG C to obtain the Eurasian avian like H1N1 swine influenza A virus cold-adapted attenuated virus strain named as a swine influenza cold-adapted attenuated virus strain A / swine / Guangxi / 18 / 2011ca (H1N1), the strain is collected in a China Center for Type Culture Collection, and the collection number of the strain is CCTCC NO. V201720. Researches show that an attenuated live vaccine prepared from the cold-adapted attenuated virus strain can resist attack of swine influenza A virus wild strains mainly epidemic in Chinese swine herds, and is of great significance for controlling prevalence and human infection of Eurasian avian like H1N1 and H1N2 swine influenza A viruses.

Description

Technical field [0001] The present invention relates to an attenuated influenza virus vaccine strain, in particular to a method that gradually reduces the Eurasian avian like (EA) H1N1 subtype swine influenza A virus (SIV) in SPF chicken embryos. Temperature cultivation to obtain a broad-spectrum cold-adapted attenuated vaccine strain is also related to the application of the vaccine strain in the prevention and control of EAH1N1 subtype swine influenza virus. The invention belongs to the technical field of veterinary medicine. Background technique [0002] Swine influenza is an acute and infectious respiratory infectious disease of pigs caused by swine influenza A virus (SIV). It is characterized by sudden onset, cough, dyspnea, fever, and rapid outcome. Under normal circumstances, swine flu has low mortality and high morbidity, and it is widespread worldwide, causing huge economic losses to the pig industry. [0003] There are three main subtypes of swine influenza virus: H1N1...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00A61K39/145A61P31/16
CPCA61K39/12A61K2039/5254A61K2039/552C12N7/00C12N2760/16134
Inventor 杨焕良陈化兰陈艳乔传玲
Owner HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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