Avian pathogenic escherichia coli strain and application thereof in vaccine preparation

A technology of avian pathogenic Escherichia coli, which is applied in the field of avian pathogenic Escherichia coli strains and its application in vaccine preparation, can solve problems such as drug residues in poultry products and affect the safety of poultry products, and achieve good public health significance, The effect of reduced use

Inactive Publication Date: 2016-08-17
SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the emergence of drug resistance, a large number of antibiotics are currently used in the prevention and control of the disease, which can easily cause drug residues in poultry products, which will directly affect the food safety of poultry products. This is not only related to animal husbandry production and animal husbandry. The development of industrial economy is also related to human health and living environment

Method used

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  • Avian pathogenic escherichia coli strain and application thereof in vaccine preparation
  • Avian pathogenic escherichia coli strain and application thereof in vaccine preparation
  • Avian pathogenic escherichia coli strain and application thereof in vaccine preparation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1 Construction of avian pathogenic Escherichia coli luxS and aroA deletion strains

[0024] Using the genome of avian pathogenic Escherichia coli DE17 as a template and luxS-UF / luxS-UR as primers, the upstream homology arm fragment of the luxS gene (775bp) was amplified, and luxS-DF / luxS-DR was used as primers to amplify Amplification of the homology arm fragment downstream of the luxS gene (1023bp). Use aroA-UF / aroA-UR as primers to amplify the upstream homology arm fragment of aroA gene (886bp), and use aroA-DF / aroA-DR as primers to amplify the downstream homology arm fragment of aroA gene (957bp ).

[0025] The upper and lower homology arm fragments of the luxS and aroA genes were cloned into the pUC19 vector to construct pUC19-luxS-up-down and pUC19-aroA-up-down recombinant vectors, respectively.

[0026] The pKD3 plasmid was used as a template and pkD3-F / pkD3-R were used as primers to amplify the chloramphenicol resistance gene (cat). After the PCR produc...

Embodiment 2

[0035] Embodiment 2 median lethal dose (LD 50 ) determination

[0036] In this example, the wild strain DE17, the deletion strains DE17ΔaroA and DE17ΔaroAΔluxS were cultured to the logarithmic phase, the bacteria were collected, washed three times with sterile PBS, the number of bacteria was adjusted, and the ratio was diluted. Each strain of bacteria is divided into five groups, with 8 cherry valley ducks in each group, so that the number of bacteria in each strain of bacteria and each group is 10 7 CFU / piece, 10 6 CFU / piece, 10 5 CFU / piece, 10 4 CFU / piece, 10 3 CFU / only and 10 2 CFU / only was injected into the thigh muscle, observed for 7 days after the challenge, and recorded the death of Cherry Valley duck. Data statistics using Reed-Muench method to calculate LD 50 .

[0037] The detection results showed that the LD of DE17, the deletion strain DE17ΔaroA and the double deletion strain DE17ΔluxSΔaroA 50 1.2×10 4 CFU, 3.8×10 5 CFU and 2.4×10 6 CFU. After the del...

Embodiment 3

[0038] Example 3 Effect of aroA and luxS deletion on APEC motility

[0039] In this example, the wild strain DE17, the deletion strain DE17ΔaroA and the double deletion strain DE17ΔluxSΔaroA were cultivated to the logarithmic phase, the bacteria were washed three times with sterile PBS, and the OD 600 Adjust to 1.0. In the middle of the motility medium, add 10 μl of bacteria, and culture overnight in a 37°C incubator. Use a vernier caliper to record the size of the bacterial circle for comparative analysis. The results of motility analysis showed that the deletion of aroA gene alone did not affect the motility of APEC, while the simultaneous deletion of luxS and aroA genes could significantly reduce the motility of APEC (pfigure 1 shown.

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Abstract

The invention belongs to the field of biotechnology, and provides an avian pathogenic escherichia coli strain, which is an avian pathogenic escherichia coli luxS and aroA double-gene-deleted strain with the preservation numbr of CGMCC10601. The avian pathogenic escherichia coli strain provided by the invention can be used for preparing avian pathogenic escherichia coli inactivation and attenuated vaccine and bacterial ghost vaccine, and preventing avian pathogenic escherichia coli infection.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular, the invention relates to an avian pathogenic Escherichia coli strain and its application in vaccine preparation. Background technique [0002] Avian colibacillosis is one of the infectious diseases that seriously affect the poultry industry caused by avian pathogenic Escherichia coli (APEC). APEC infection of poultry can cause symptoms such as pneumonia, air sacculitis, pericarditis, perihepatitis, peritonitis, and salpingitis. In severe cases, it causes systemic infection characterized by sepsis, leading to acute death. At present, the complex serotypes and broad-spectrum drug resistance of avian pathogenic Escherichia coli pose new challenges to the prevention and control of avian colibacillosis. APEC not only endangers the health of poultry, but also affects the egg production and the time to market of meat poultry, and increases the cost of feeding, causing serious economic losse...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/21A61K39/108A61P31/04C12R1/19
CPCA61K39/0258A61K2039/552C12N9/1092C12N9/88C12Y205/01019C12Y404/01021
Inventor 韩先干于圣青王少辉杨立军陈兆国黄燕米荣升贾海燕
Owner SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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