Infectious bovine rhinotracheitis virus IBRV-JN03 isolate and application thereof

A technology of IBRV-JN03 and rhinotracheitis virus, applied in antiviral agents, viruses/bacteriophages, medical preparations containing active ingredients, etc., can solve the problems of poor virus protection and achieve good immunogenicity and broad coverage Market application prospect, highly targeted effect

Active Publication Date: 2015-09-23
SHANDONG NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no relevant vaccine product in my country, and the Ministry of Agriculture has not yet approved foreign IBRV vaccines to enter the Chinese market. More importantly, with the mutation of IBRV, even if foreign vaccines are allowed to be imported, their seed virus has poor protection against domestic epidemic strains

Method used

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  • Infectious bovine rhinotracheitis virus IBRV-JN03 isolate and application thereof
  • Infectious bovine rhinotracheitis virus IBRV-JN03 isolate and application thereof
  • Infectious bovine rhinotracheitis virus IBRV-JN03 isolate and application thereof

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Experimental program
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Effect test

Embodiment 1

[0029] The isolation and identification of embodiment 1 bovine infectious rhinotracheitis virus

[0030] 1.1 Isolation of bovine infectious rhinotracheitis virus

[0031] To observe the morbidity of cattle, the inventor collected nasal secretions, feces samples from cattle diarrhea, and tissue (liver, spleen, lymph node) samples from cattle that were preliminarily diagnosed as bovine infectious rhinotracheitis. For stool and bovine nose swab samples, dilute 1:5 with PBS (100U / mL penicillin, 100g / mL streptomycin). Centrifuge at 3000r / min for 10min, and the obtained supernatant is filtered and sterilized by a 0.22μm filter membrane to obtain a disease material treatment solution. The samples of infected bovine tissues (liver, spleen, and lymph nodes) were crushed with a tissue grinder, frozen and thawed three times, and diluted 1:3 with PBS (100 U / mL penicillin, 100 g / mL streptomycin). Centrifuge at 8000r / min for 10min, and the obtained supernatant is filtered and sterilized b...

Embodiment 2

[0058] Comparative analysis of the immunogenicity of embodiment 2 IBRV isolates

[0059] 2.1 Virus reproduction

[0060] Take MDBK cells in good growth state, wash them twice with PBS, and make the virus titer greater than 10 7.0 TCID 50 / mL, and IBRV isolates IBRV-JN03 and IBRV-LY9038 after 6 rounds of plaque purification, according to 100TCID 50 The amount of the virus was inoculated respectively, adding DMEM cell maintenance solution containing 2% (v / v) horse serum, pH value 7.0, 37 ℃ 5% CO 2 Cultivate, and when the cytopathy reaches 70%-80%, harvest the virus culture medium, freeze and thaw repeatedly 2-3 times, and centrifuge at 4°C at high speed to obtain the supernatant so as to obtain the IBRV antigen.

[0061] 2.2 Comparative analysis of virus immunogenicity

[0062] Add 0.2‰ formaldehyde to the supernatants of the above-mentioned amplified viruses IBRV-JN03 and IBRV-LY9038 and inactivate them at 37°C for 24 hours. Mix well to make inactivated antigen. Inject the ...

Embodiment 3

[0063] The development of embodiment 3 IBRV inactivated vaccine

[0064] 3.1 Virus reproduction

[0065] Take the MDBK monolayer cells in a good growth state, wash them twice with PBS, and prepare the cell culture fluid of the above-mentioned good immunogenic vaccine candidate strain (IBRV-JN03) by 100TCID 50 The amount of virus was inoculated, adding DMEM cell maintenance solution containing 2% (v / v) horse serum at 37°C 5% CO 2 Cultivate, and when the cell lesion is 70% to 80%, harvest the virus culture medium, freeze and thaw repeatedly 2 to 3 times, and centrifuge at 4°C at high speed to obtain the virus supernatant, take a sample for virus content determination, and the titer is 6.75×10 8.5 TCID 50 / mL, among the reported isolated strains of IBRV, this virus strain has a higher titer, which is more in line with the requirement of high antigenic mass for vaccine production.

[0066] 3.2 Preparation of virus inactivated vaccine

[0067] The above-mentioned vaccine candid...

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Abstract

The invention discloses an infectious bovine rhinotracheitis virus IBRV-JN03 isolate and application thereof. The preservation number of the infectious bovine rhinotracheitis virus is CGMCC No.10396 and the infectious bovine rhinotracheitis virus is preserved in the China General Microbiological Culture Collection Center of the China Committee for Culture Collection of Microorganisms, and has relatively high virus titer and good immunogenicity; after calves are immunized by an aluminium hydroxide adjuvant inactivated vaccine prepared through the infectious bovine rhinotracheitis virus, relatively high antibodies are generated, so that the infectious bovine rhinotracheitis virus IBRV-JN03 isolate is a good infectious bovine rhinotracheitis vaccine candidate virus strain. Therefore, an infectious bovine rhinotracheitis diagnostic reagent, inactivated vaccine, attenuated vaccine and genetic engineering vaccine prepared on the basis of the virus can be used for diagnosis and vaccine prevention and control of the endemic infectious bovine rhinotracheitis and a wide market application prospect is realized.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a bovine infectious rhinotracheitis virus IBRV-JN03 isolate and application thereof. Background technique [0002] Infectious bovine rhinotracheitis virus (Infectious bovine rhinotracheitis virus, IBRV) is an acute, febrile, contagious infectious disease of cattle. The disease can cause immunosuppression, secondary bacterial infection and lead to more severe respiratory disease. At present, the disease is widely prevalent in the world, and the disease seriously affects the international cattle production and trade, and has been listed as a B-type communicable disease by the World Organization for Animal Health (OIE). IBRV was detected in cows imported from New Zealand in the late 1970s in my country. Since there were no good immune protection measures, the virus was infected in cows in many provinces of my country, and the incidence rate was 10% to 90%. The fattening, milk productio...

Claims

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Application Information

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IPC IPC(8): C12N7/00A61K39/265A61P31/22C12Q1/70C12Q1/68G01N33/569C12R1/93
Inventor 王洪梅侯佩莉何洪彬
Owner SHANDONG NORMAL UNIV
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