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Bovine viral diarrhea virus low virulent strain and application thereof

A technology of bovine viral diarrhea and attenuated strains, applied in the direction of viruses, antiviral agents, viruses/bacteriophages, etc., can solve the problems of lack of safe and effective BVDV attenuated vaccines, and achieve good immune effect and high proliferation rate

Active Publication Date: 2021-07-27
SOUTHWEST UNIVERSITY FOR NATIONALITIES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Compared with inactivated vaccines, BVDV attenuated vaccine has the advantages of wider immune protection range, higher antibody level and longer antibody duration, but there is still a lack of safe and effective BVDV attenuated vaccine in the domestic market

Method used

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  • Bovine viral diarrhea virus low virulent strain and application thereof
  • Bovine viral diarrhea virus low virulent strain and application thereof
  • Bovine viral diarrhea virus low virulent strain and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1 Isolation and Sequence Analysis of Bovine Viral Diarrhea Virus Attenuated Strain

[0038] BVDV SMU-Z6 / 1a / SC / 2016 (Z6 for short, virus deposit number is CCTCC NO:V201813) strain, virus TCID50=10-8.11·0.1mL-1; MDBK (Madin-Darby bovine kidney, MDBK) cells.

[0039] (1) RT-PCR amplification primers were designed according to the Z6 (GenBank: MF693403.1) genome sequence published on the NCBI website. See Table 1 for information about the primers.

[0040] Table 1 Relevant information of primers for genome amplification of different generations of Z6 strains

[0041]

[0042]

[0043] (2) Determination of virus titer

[0044] Cell culture: resuscitate MDBK cells cryopreserved in a liquid nitrogen tank, pass 2 to 3 passages to restore cell viability, inoculate the virus, pass Z6 to the 100th passage, due to the continuous passaging, the adaptability of the virus to the cells is enhanced, and the fluorescence microscope The time for the first observation of cyt...

Embodiment 2

[0063] The pathogenicity analysis of different generation attenuated strains of embodiment 2

[0064] Infect BALB / c mice with different generations of viruses in the above-mentioned embodiment 1, and analyze the effect of different generations of viruses on small Pathogenicity of mice. The specific method is as follows:

[0065] (1) Infected mice: 63 BALB / c mice were randomly divided into 7 groups with 9 mice in each group, including 5 infection groups, 1 negative control group and 1 positive control group. The infection group was injected intraperitoneally with 1 mL of culture solution of Z6-P1, Z6-P25, Z6-P50, Z6-P75 and Z6-P100 (TCID 50 ≈1×10 -8.0 0.1mL -1 , P>0.05); the positive control group was intraperitoneally injected with Oregon C24V virus solution 1mL; the negative control group was intraperitoneally injected with cell culture medium DMEM 1mL. Continuously observe for 10 days after intraperitoneal injection, record the mental state, diet and excretion of the mi...

Embodiment 3

[0086] The immune effect of embodiment 3 attenuated strain Z6-P100

[0087] (1) Sterility test and safety test of Z6-P100

[0088] Take 0.2 mL of Z6-P100 culture solution, spread it evenly on LB solid medium by aseptic operation, and culture it in a constant temperature incubator at 37°C for 7 days, no bacterial growth was observed.

[0089] Three BALB / c mice were randomly selected and injected with Z6-P100 culture solution by intramuscular injection of fore and hind limbs, each 1 mL. The mice were observed within 7 days after the injection to see if there were side effects against immunity. The results showed that within 7 days after immunization, no obvious abnormalities in the appetite and spirit of the mice were observed, no obvious clinical symptoms (fever, diarrhea, etc.) appeared, and there was no significant difference in the weight of the mice. It shows that the Z6-P100 culture medium is sterile and safe, and will not cause immune side effects in mice.

[0090] (2)...

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Abstract

The invention discloses a bovine viral diarrhea virus low virulent strain and application thereof.The low virulent strain is obtained after 100 generations of continuous passage culture of an SMU-Z6 / 1a / SC / 2016 strain, the nucleotide sequence of the low virulent strain is shown as SEQ ID NO.1, and the microbial preservation number of the low virulent strain is V202131. After BABL / c mice are immunized with the bovine viral diarrhea virus attenuated strain, high serum neutralizing antibody, serum IgG antibody, lung and intestinal sIgA antibody and splenic lymphocyte proliferation rate can be generated, and a good immune effect is achieved. The bovine viral diarrhea virus attenuated strain has a very good application prospect in prevention, treatment and diagnosis of the bovine viral diarrhea virus.

Description

technical field [0001] The invention relates to the technical field of microbes and immunization, in particular to an attenuated strain of bovine viral diarrhea virus and its application. Background technique [0002] Bovine viral diarrhea virus (BVDV) belongs to the Flaviviridae family. Pestivirus can infect cattle, sheep, deer and other ruminants. The main host is cattle, which can cause bovine viral diarrhea. Diarrheal disease (Bovine viral diarrhea, BVD). The possible symptoms of BVDV infection include diarrhea, mucosal disease, respiratory disease, persistent infection and immune tolerance, and reproductive disorders in cows. BVDV spreads widely around the world and threatens the healthy development of animal husbandry. In my country, more than 5 million cattle die from BVDV infection every year. BVDV infection not only brings economic losses to the breeding industry, but also affects the quality and safety of bovine biological products such as serum and embryos, and ...

Claims

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Application Information

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IPC IPC(8): C12N7/04C12N7/08C12N15/40A61K39/12A61P31/14C12R1/93
CPCC12N7/00A61K39/12A61P31/14C12N2770/24321C12N2770/24322C12N2770/24334C12N2770/24364A61K2039/5254A61K2039/552
Inventor 张斌汤承岳华王丹
Owner SOUTHWEST UNIVERSITY FOR NATIONALITIES
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