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Pullorum disease salmonella spiC-rfc double-gene knockout attenuated strain and DIVA vaccine application thereof

A technology for Salmonella pullorum and Salmonella pullorum is applied in the direction of recombinant DNA technology, bacteria, antibacterial drugs, etc., which can solve the problems of enhancement and drug resistance of bacteria, and achieve the effect of reducing virulence, solid immune protection, The effect of lethal dose increase

Inactive Publication Date: 2016-04-13
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The traditional way to prevent and treat this disease is to use antibiotics. Long-term use is likely to cause many effects such as drug residues and bacterial drug resistance. Therefore, the research on preventing pullorum with vaccines has attracted much attention.

Method used

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  • Pullorum disease salmonella spiC-rfc double-gene knockout attenuated strain and DIVA vaccine application thereof
  • Pullorum disease salmonella spiC-rfc double-gene knockout attenuated strain and DIVA vaccine application thereof
  • Pullorum disease salmonella spiC-rfc double-gene knockout attenuated strain and DIVA vaccine application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Example 1 Construction of Salmonella pullorum spiC-rfc double gene knockout attenuated strain S06004ΔspiCΔrfc

[0046] 1. Materials and Methods

[0047] 1.1 Strains and plasmids

[0048] The Salmonella pullorum S06004ΔspiC mutant strain was constructed and preserved by the inventor's laboratory using a recombinant suicide plasmid method to knock out the spiC gene without trace (the Salmonella pullorum S06004ΔspiC mutant strain is a prior art, and its specific construction method is detailed in " Construction and identification of Salmonella pullorum S06004ΔspiC mutant strain, Chinese Veterinary Science, 2014, Issue 04").

[0049] Salmonella pullorum S06004 (Nal R ), E.coliDH5α, E.coliSpy372 (λpir) and plasmid pKD46 (Amp R ), pKD3 (Cm R ), pCP20 (Amp R 、Cm R ) are preserved by the inventor's laboratory, and these bacteria and plasmids are all prior art. For details, see the construction and biological characteristics of the pathogenic island-2 deletion strain of Sa...

Embodiment 2

[0083] Example 2 DIVA vaccine application of Salmonella pullorum spiC-rfc double gene knockout attenuated strain S06004ΔspiCΔrfc

[0084] The test material is the double-gene knockout S06004ΔspiCΔrfc attenuated strain of Salmonella pullorum constructed in Example 1.

[0085] Pick a single colony of bacteria and streak the whole plate on the LB culture plate, culture at 37°C for 20 hours, use sterile PBS to carefully scrape off the bacteria on the LB plate, wash 3 times with sterile PBS, and adjust the bacterial concentration to 10 9 CFU / mL, that is to make attenuated Salmonella pullorum vaccine.

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Abstract

The invention belongs to the field of fowl salmonellosis prevention and control and particularly relates to a pullorum disease salmonella spiC-rfc double-gene knockout attenuated strain and preparation and application thereof. The attenuated strain is pullorum disease salmonella in which a spiC gene or an rfc gene is not expressed. It is found through animal experiments that virulence of the mutant strain is remarkably reduced, and colonization time in a host body is shortened obviously. The attenuated strain is adopted for preparing pullorum disease salmonella live vaccine, after chickens are inoculated with the vaccine, no adverse reaction is generated, and strong virulence strain infection can be effectively eliminated. The host does not generate an anti-body aiming at O-antigen, by means of pullorum disease fowl typhoid multi-valence detection antigen, the attenuated stain can be effectively applied to glass plate agglutination experiments to distinguish immunized chickens and naturally infected chickens, and the characteristic of the DIVA vaccine is achieved.

Description

technical field [0001] The invention belongs to the field of prevention and treatment of poultry salmonellosis, and in particular relates to a spiC-rfc double gene knockout attenuated strain of Salmonella pullorum and application of the DIVA vaccine thereof. Background technique [0002] Pullorum is a multiple infectious disease caused by Salmonella Pullorum, which is very harmful to the chicken industry in my country. In addition to being able to spread horizontally, the pathogen can also spread vertically through eggs, resulting in a decrease in hatching rate and hatching rate. It usually infects chicks less than 20 days old. It is characterized by white diarrhea, showing the course of acute sepsis, and the morbidity and mortality are quite high. ; Adult chickens are mostly chronic or latent infection, generally without obvious clinical symptoms. The traditional way to prevent and treat this disease is to use antibiotics. Long-term use can easily cause many effects such a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N15/74A61K39/112A61P31/04C12R1/42
CPCC07K14/255A61K39/0275A61K2039/522A61K2039/54A61K2039/545
Inventor 焦新安郭荣显耿士忠焦扬朱姗姗潘志明孙林陈祥李求春
Owner YANGZHOU UNIV
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