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A kind of poultry adenovirus type 4 strain, vaccine composition and application thereof

A vaccine composition, avian adenovirus technology, applied in the directions of viruses, vaccines, antiviral agents, etc., to achieve the effects of high challenge protection rate, good specificity and immunogenicity, and strong immunity

Active Publication Date: 2020-09-08
HENAN AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In terms of diagnosis, the current diagnosis is mainly based on clinical symptoms, histopathological changes, PCR amplification, and virus isolation; in terms of disease prevention and control, although there have been research reports on the use of chicken embryos and poultry-derived cells to culture avian adenoviruses, However, the serotypes of most of the reported strains are inconsistent with the current market strains. Therefore, the current market urgently needs to develop a poultry gland with a serotype consistent with the current market strains, high titer, and good immunogenicity. virus vaccine

Method used

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  • A kind of poultry adenovirus type 4 strain, vaccine composition and application thereof
  • A kind of poultry adenovirus type 4 strain, vaccine composition and application thereof
  • A kind of poultry adenovirus type 4 strain, vaccine composition and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1: Isolation and identification of avian adenovirus type 4 HNJZ strain

[0030] 1.1 Isolation of virus

[0031] Take the liver of chicken suspected of chicken hepatitis-pericardial effusion syndrome in Henan area, cut it into pieces with sterile scissors, add sterile PBS (pH7. Grind completely, collect the homogenate after grinding, transfer it to a sterile centrifuge tube, centrifuge at 10000g for 10min, filter the supernatant after centrifugation with a 0.22μm filter membrane, and use the filtrate to inoculate 11-day-old SPF chicken embryos Monolayer of primary chicken embryo liver cells at 37 °C 5% CO 2 After culturing, the cells were observed for lesions every day, and the cells showed typical adenovirus grape cluster lesions. Harvest the virus when the cell lesion reaches 90%. After repeated freezing and thawing three times, centrifuge at 10,000 g for 10 minutes.

[0032] 1.2 Virus identification

[0033] Primers were designed according to the Hexon gen...

Embodiment 2

[0045] Example 2: Sequence determination and analysis of avian adenovirus type 4 HNJZ strain

[0046] 2.1 Sequence determination of avian adenovirus type 4 HNJZ strain

[0047] Utilize the QIAamp DNA Blood Mini kit of Qiagen Company to extract its viral DNA from the virus isolated from Example 1 step 1.1 as a template, according to the genome sequence of the avian adenovirus type 4 ON1 strain (GenBank accession number GU188428) published in GenBank 26 pairs of specific primers (Table 1) were designed to perform PCR amplification reaction on the genome of avian adenovirus type 4 HNJZ strain. PCR amplification reaction system (50 μL) is: 5×PCR Buffer 10uL, template 10ng, 10mM deoxyribonucleoside triphosphate (dNTP) 1μl, upstream and downstream primers (25μM) each 1μl, high-fidelity thermostable DNA polymerase 1 unit , ddH 2 O make up. The PCR amplification reaction conditions were: pre-denaturation at 98°C for 2 min, 1 cycle of amplification; denaturation at 98°C for 10 s, an...

Embodiment 3

[0064] Embodiment 3: avian adenovirus type 4 HNJZ strain virus titer TCID 50 Determination of

[0065] Take 9-day-old SPF chicken embryos, after the eggshells are sterilized by 75% alcohol, open the eggshells from the air chamber side, take out the embryo body, remove the head and limbs with sterile ophthalmic scissors, take the liver, cut it into pieces, and wash it with sterile PBS Three times, add 0.25% trypsin, digest in a water bath at 37°C for 10 minutes; then add medium to blow the digested tissue into single cells, centrifuge at 200g for 5 minutes, and collect the cells; reweight in DMEM medium containing 10% fetal bovine serum Suspend the cells to get the hepatocyte suspension, count the hepatocyte suspension, and then count the number according to 1×10 5 The amount of inoculum of cells / well inoculated the hepatocyte suspension into a 96-well cell culture plate; the virus liquid obtained in 1.1 separation in Example 1 was diluted 10 times, and 10 -5 、10 -6 、10 -7 ...

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Abstract

The invention discloses a newly isolated avian adenovirus 4-type HNJZ strain. The microbial preservation number of the newly isolated avian adenovirus 4-type HNJZ strain is CGMCC NO.13385, and the strain has good specificity and immunogenicity, can serve as an inactivated vaccine production strain and an inspection strain and is used for preventing hepatitis-hydropericardium syndromes of poultry. The invention further discloses a vaccine composition prepared from the avian adenovirus 4 type HNJZ strain. The vaccine composition contains the inactivated avian adenovirus 4-type HNJZ strain and pharmaceutically acceptable adjuvant, the vaccine composition has good immunogenicity, high immunity can be generated after immunization, the protection rate is high, prevalence and spread of avian adenoviruses can be effectively prevented, and application prospects are wide.

Description

technical field [0001] The invention belongs to the field of microbial viruses and relates to a virus strain, in particular to an avian adenovirus type 4 strain, a vaccine composition and an application thereof. Background technique [0002] Avian adenovirus (Avian adenovirus) is divided into three groups according to group-specific antigens, and each has its own group-specific antigens. Group I adenoviruses include traditional fowl adenovirus (Fowl adenovirus, FAdV) and other poultry isolates, with a total of 12 serotypes. The clinical diseases caused by avian adenovirus mainly include inclusion body hepatitis (Inclusion body hepatitis, IBH), hepatitis-hydropericardium syndrome (HHS), gizzard erosion and some other diseases. Of these diseases, HHS is the most detrimental to the poultry industry. HHS was first reported in the Ankara region of Pakistan in 1987, and subsequently reported in India, Kuwait, Iran, Japan, South Korea, Russia, Mexico, South America and North Amer...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K39/235A61P31/20
CPCA61K39/12A61K2039/5252A61K2039/552C12N7/00C12N2710/10221C12N2710/10231C12N2710/10234
Inventor 赵军王川庆杨霞常洪涛陈陆王新卫李永涛刘红英姚慧霞高冬生刘延珂
Owner HENAN AGRICULTURAL UNIVERSITY
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