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Duck BYD virus inactivated vaccine and preparation method thereof

A virus inactivation and virus technology, applied in biochemical equipment and methods, antiviral agents, viruses/bacteriophages, etc., can solve the problems of no effective treatment and immune prevention measures, lack of effective treatment drugs for flavivirus infection, etc. Achieve excellent immunogenicity, good immune effect, and good safety

Inactive Publication Date: 2012-07-04
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Since flavivirus infection has not been found in ducks before, this is the first outbreak and epidemic of duck BYD virus infection. However, there is still no effective treatment for flavivirus infection at home and abroad, and there is no vaccine to protect it. Therefore, there is no effective treatment and immunoprophylaxis

Method used

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  • Duck BYD virus inactivated vaccine and preparation method thereof
  • Duck BYD virus inactivated vaccine and preparation method thereof
  • Duck BYD virus inactivated vaccine and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1 Virus isolation and pathogenicity

[0029] 1. Isolation and identification The virus was isolated from the brain tissue of infected ducks in a duck farm in Baiyangdian, Hebei Province, China. Make a 10% homogenate of the diseased duck brain tissue sample with sterile PBS, centrifuge at 3000r / min for 20min, filter and sterilize, inoculate 9-12 day old duck embryos without maternal antibody through the allantoic cavity, and incubate at 37°C for 5 On the first day, the allantoic fluid was harvested and passaged in duck embryos. Inoculate the second-generation allantoic fluid on the BHK-21 cell monolayer, culture it statically in a 5% carbon dioxide incubator, and observe it for 7 days. When the cell changes appear, the culture is harvested and stored at -70°C.

[0030] Prepare the ultrathin sections of infected cells and observe under the transmission electron microscope. A large number of spherical enveloped virus particles with a diameter of about 50 nm can be...

Embodiment 2

[0034] Example 2 Preparation of duck BYD virus JXSP strain virus solution

[0035] 1. Propagation of virus seeds Dilute BYDV-JXSP 2nd generation allantoic duck embryo venom 5 times and inoculate it into BHK-21 cell culture, place it in a 5% carbon dioxide incubator at 37°C for static culture until 75% of the cells have lesions Harvest the cell fluid, freeze and thaw twice, take the supernatant and dilute it 100 times, then infect and inoculate BHK-21 cells, pass it to the 8th passage, harvest the cell fluid and store it at -70°C as virus seeds, the virus content is ≥10 5.5 TCID 50 / mL.

[0036] 2. Cell seed propagation Take the BHK-21 cells stored in liquid nitrogen, immediately place them in a 37°C water bath to melt, centrifuge at 800r / min for 5min, discard the supernatant, add DMEM cell growth medium containing 10% fetal bovine serum to suspend the cell pellet, Transferred to a cell culture bottle, cultured statically at 37°C in a 5% carbon dioxide incubator, and subcultu...

Embodiment 3

[0038] Example 3 Preparation of duck BYD virus inactivated vaccine

[0039] 1. Inactivation of virus fluid

[0040] Add the virus liquid prepared in Example 2 to the formaldehyde solution at a final concentration of 0.2% (volume ratio), mix well, shake well, and then act at 37° C. for 24 hours to inactivate the virus.

[0041] 2. Inactivation effect test

[0042] BHK-21 cells were cultured to a monolayer on a 24-well plate, and the 10-fold diluted BYDV-JXSP virus solution treated with formaldehyde inactivation in step 1 was inoculated on the BHK-21 cell monolayer at 0.2 mL / well, and each sample Inoculate 3 wells, discard the sample solution after incubation for 1 hour, continue to maintain the culture after washing with maintenance solution, and set positive control and blank control wells at the same time; observe the positive control wells after 48 hours. Cytopathies can be seen, and the samples and blank control wells are negative; Blind passage for 2 generations, the sam...

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Abstract

The invention provides a vaccine for preventing duck egg reduction syndrome caused by BYD virus and a preparation method thereof. The invention provides a duck BYD virus (Family Flaviviridae, Genus Flavivirus, Ntaya virus group, Duck BYD virus) JXSP strain, and the preservation number is CGMCC No.5266. The virulent strain provided by the invention is duck BYD virus virulent strain with excellent immunogenicity. The virulent strain is vaccinated onto a sensitive cell to obtain cell sap, which is emulsified after being inactivated so as to obtain the safe, effective and controllable duck BYD virus inactivated vaccine, so that the prevention and the control of the duck egg reduction syndrome can be favored.

Description

technical field [0001] The invention relates to the technical field of veterinary biological products, in particular to a duck BYD virus vaccine and a preparation method thereof. Background technique [0002] Since April 2010, a severe disease has been prevalent in duck farms in some southeastern provinces of China. The disease has spread rapidly to all major duck-growing provinces and cities in China, including Zhejiang, Fujian, Jiangxi, Shandong, Hebei, Jiangsu and Beijing. The disease affects a variety of laying ducks, including Peking ducks and shelducks. The most prominent clinical symptoms of infected ducks are a sudden drop in feed intake and a sudden drop in egg production rate. The egg production rate can drop to 10 within 5 days. %, necropsy showed serious lesions such as hemorrhage, atrophy, and rupture of the ovary. In the early stage, some follicles can be seen to have bleeding points or spots. With the development of the disease, the follicles are severely hem...

Claims

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Application Information

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IPC IPC(8): C12N7/00A61K39/12A61P31/14C12R1/93
Inventor 苏敬良田克恭陈瑞爱苏文良遇秀玲练炳洲周智
Owner CHINA AGRI UNIV
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