Cross-genus virulent bacteriophage, preparation process and application of cross-genus virulent bacteriophage in prevention and treatment of swine paratyphoid fever and pullorum disease

A porcine paratyphoid, preparation technology, applied in the field of microbiology, can solve the problems of limited application range, no therapeutic effect, weak cracking effect, etc., achieve the effects of low production cost, broaden the scope of sterilization, and restore appetite

Active Publication Date: 2021-05-14
山东仙普爱瑞科技股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

(1) Phages are highly specific, so therapeutic phages are often only effective against a single strain or subtype of bacteria, and have weak or no therapeutic effect on other strains, which greatly limits the scope of their application

Method used

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  • Cross-genus virulent bacteriophage, preparation process and application of cross-genus virulent bacteriophage in prevention and treatment of swine paratyphoid fever and pullorum disease
  • Cross-genus virulent bacteriophage, preparation process and application of cross-genus virulent bacteriophage in prevention and treatment of swine paratyphoid fever and pullorum disease
  • Cross-genus virulent bacteriophage, preparation process and application of cross-genus virulent bacteriophage in prevention and treatment of swine paratyphoid fever and pullorum disease

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1 Isolation of a virulent phage XPAR_CPS01 that cross-genus infects Salmonella pullorum and Salmonella choleraesuis

[0045] Sample treatment: Collect feces from pigs infected with paratyphoid fever from a pig farm in Yantai City, Shandong Province, soak in sterile water overnight in a 37 °C incubator, filter out excess impurities with gauze, filter and sterilize at 0.22 μM, and then use it for processing sample.

[0046] Preparation of host bacteria: Pick a single colony of Salmonella choleraesuis virulent strain C78-2 and inoculate it in 100 mL of LB broth, culture with shaking at 180 r / min at 37°C for 16-18 h to obtain a suspension of the host bacteria, and set aside.

[0047] Bacteriophage proliferation: Take 10 mL of the above-mentioned treated sample and add it to the above-mentioned Erlenmeyer flask containing the host bacterial solution. After mixing, shake and cultivate overnight in a 37°C air bath shaker at 160rpm to obtain a phage proliferation soluti...

Embodiment 2

[0053] Example 2 Determination of cleavage profile of bacteriophage XPAR_CPS01

[0054] 5 mL of LB semi-solid medium was mixed with the host bacteria in the logarithmic phase of Salmonella choleraesuis virulent strain C78-2, attenuated vaccine strain C500, Salmonella pullorum C79-13, S06004, RKS5078, and CDC1983-67 (100 μl, OD =1.0) and mix well, pour it into a solid LB plate to form a double-layer plate, after standing still, take 200 μL of the above-mentioned purified phage and spread it on the plate, after it dries, culture it upside down at 37°C overnight, and observe whether there are lysis spots . The results showed that the Salmonella choleraesuis attenuated vaccine strain C500 (with a plaque diameter of 0.43 mm), Salmonella pullorum C79-13 (with a plaque diameter of 0.50 mm), S06004 (with a plaque diameter of 0.05 mm), The plaques on the plates of RKS5078 (0.16 mm in diameter of plaque) and CDC1983-67 (0.12 mm in diameter of plaque) were extremely bright, indicating t...

Embodiment 3

[0056] Example 3 Determination of the titer of bacteriophage XPAR_CPS01

[0057] Using the Salmonella choleraesuis virus vaccine strain C500 as the host bacterium, 1 mL of the preserved phage XPAR_CPS01 stock solution was diluted 2 to 13 times by a 10-fold gradient to make dilutions of different gradients for use.

[0058] Pour 10 mL of sterilized NA medium into an empty Petri dish, and make NA agar plates after solidification. Add 0.1mL of diluents of different gradients and 1mL of host bacteria to a 15mL vial, mix with 5mL of NA medium dissolved and cooled to 60°C, and immediately pour it into the pre-prepared NA plate. As a blank control, set up three parallels. After cooling and solidifying, place it upside down in a constant temperature incubator at 37°C for 16-24 hours, read the phage titer, and determine the number of phages. Phage titer (pfu / mL) = number of plaques × dilution factor × 10. The titer of phage XPAR_CPS01 stock solution was 1.05×10 10 pfu / mL.

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Abstract

The invention provides a strain of cross-genus virulent bacteriophage, and the preservation number of the bacteriophage is CCTCC NO: M 2020838. According to the invention, a strain of cross-genus infected salmonella pullorum and salmonella choleraesuis bacteriophage is separated for the first time, and compared with a single lytic salmonella bacteriophage, the bactericidal range of the bacteriophage is greatly widened, and the bacteriophage has good lytic ability for a salmonella choleraesuis virulent strain C78-2, a attenuated vaccine strain C500, salmonella pullorum C79-13, S06004, RKS5078 and CDC1983-67. The bacteriophage disclosed by the invention has the characteristics of high sterilization speed and high yield. Meanwhile, the cross-genus virulent bacteriophage can be drunk with water, can eliminate animal gastrointestinal tract salmonella, is used for preventing and treating swine paratyphoid fever and pullorum disease, and is convenient to use.

Description

technical field [0001] The invention relates to the technical field of microbes, in particular to the preparation of a transgenus potent phage and its combination with Bacillus subtilis in the prevention and treatment of swine paratyphoid and pullorum. Background technique [0002] Salmonella is one of the important zoonotic pathogens in public health. Food poisoning caused by salmonella often ranks first. Different serotypes of Salmonella can cause various diseases such as pullorum, chicken typhoid, swine paratyphoid, and animal abortion. Some serotypes of Salmonella can also parasitize animals in a carrier state, seriously affecting the healthy development of the breeding industry. At present, antibiotics are still considered to be the main method to treat Salmonella infection. However, with the long-term use and abuse of veterinary antibiotics, drug-resistant Salmonella not only makes the prevention and control of animal diseases more and more difficult, but also serious...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00A61K35/76A61P31/04A61K35/742C12R1/92
CPCC12N7/00A61K35/76A61K35/742A61P31/04C12N2795/10321C12N2795/10332C12N2795/10352A61K2300/00Y02A50/30
Inventor 韩国英李琦王兴业刘刚王晓冉王茂超郭海岩
Owner 山东仙普爱瑞科技股份有限公司
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