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Attenuated Salmonella pullorum and application thereof

A technology for salmonella and avian salmonella, applied in the direction of bacteria, antibacterial drugs, medical preparations containing active ingredients, etc., can solve problems such as environmental and food safety impacts, and achieve good immunogenicity

Inactive Publication Date: 2011-08-17
HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] These existing vaccine strains are all capable of colonizing the host, which still has some impact on the environment and food safety

Method used

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  • Attenuated Salmonella pullorum and application thereof
  • Attenuated Salmonella pullorum and application thereof
  • Attenuated Salmonella pullorum and application thereof

Examples

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Effect test

Embodiment 1

[0023] Example 1 Construction of Double Gene Deletion Attenuated Salmonella (Salmonella pullorum) SM091-DED Strain

[0024] 1. Test materials and methods

[0025] 1.1 Strains and plasmids

[0026] The Salmonella avian strain was isolated and identified by the inventor's laboratory and named SM091. Plasmid pKD46 (temperature-sensitive, containing gam, bet and exo genes regulated by the arabinose promoter, Ampr), pKD3 (containing the chloramphenicol resistance gene with FRT sites on both sides, Cmr), pCP20 (temperature-sensitive , encoding FLP recombinase, which can specifically recognize the FLP site).

[0027] 1.2 Reagents

[0028] dNTP and Taq DNA polymerase were purchased from MBI Company, plasmid extraction kit was purchased from QIAGEN Company, DNA gel recovery kit was purchased from Shanghai Huasun Company, L-arabinose was purchased from Sigma Company, and primer synthesis was completed by Nanjing Boya Company.

[0029] 1.3 Construction method of double-gene deletion ...

Embodiment 2

[0049] The preparation of embodiment 2 attenuated salmonella vaccine

[0050] 1. Test material: the attenuated Salmonella SM091-DED strain constructed in Example 1;

[0051] 2. Test method

[0052] The frozen attenuated Salmonella strains were diluted with sterilized saline, streaked on LB agar plates, and cultured overnight at 37°C. On the next day, pick a single colony and inoculate it into non-resistant LB liquid medium, and culture it at 37°C for 18-20 hours. Centrifuge at 3000rpm, wash 3 times with sterilized PBS, remove the medium components, and then count the bacteria to 10 7 CFU, divided into cryopreservation tubes, freeze-dried and stored to make attenuated Salmonella vaccine.

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Abstract

The invention discloses an attenuated Salmonella pullorum and application thereof. By using gene knockout technology, the in vivo colonization gene and the main virulence gene of Salmonella pullorum are deleted, thus obtaining an dual-gene-deleted attenuated Salmonella pullorum SM091-DED strain; and the microbiological preservation number of the strain is CGMCC NO.4604. The virulence of the attenuated Salmonella pullorum disclosed by the invention is obviously reduced, and the in vivo colonization time is short after the attenuated Salmonella pullorum is inoculated into a host; the herd infection test shows that the attenuated Salmonella pullorum has no horizontal transmission capability; the attenuated Salmonella pullorum provides full cross protection for homotype and allotype high-virulence Salmonella pullorum; and after SPF (Specific Pathogen Free) chicks are immunized, the infection of the high-virulence strain can be effectively eliminated, and the herd infection can not be caused. Thus, the invention provides a safe, fine-immunogenicity and low-virulence Salmonellosis avium attenuated live vaccine for preventing Salmonellosis avium.

Description

technical field [0001] The present invention relates to a strain of Salmonella, in particular to an attenuated Salmonella pullorum (Salmonella pullorum) SM091-DED obtained after gene knockout and its construction method. The present invention also relates to the use of the attenuated Salmonella as a medicine for preventing salmonellosis The invention belongs to the field of prevention and treatment of poultry salmonellosis. Background technique [0002] The safety of animal food is not only related to the health of consumers, but also affects the safety of public health in society. Salmonellosis is one of the zoonotic diseases of great significance in public health. The pathogen Salmonella belongs to the Enterobacteriaceae family. Salmonella not only causes poultry infections, but also causes bacterial infections in humans, leading to food poisoning and acute gastroenteritis and other diseases. Salmonella exists in various animal meats such as chicken, duck, pig, cattle, s...

Claims

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Application Information

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IPC IPC(8): C12N1/20A61K39/112A61P31/04C12R1/42
Inventor 刘思国刘慧芳王秀梅于申业王牟平司薇杨志
Owner HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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