Preparation method of drug-resistant pseudomonas aeruginosa broad-host range phage

A technology of Pseudomonas aeruginosa and bacteria, which is applied in the direction of viruses/phages, biochemical equipment and methods, microorganisms, etc., can solve the problems of poor separation and purification effects, achieve simple preparation methods, avoid drug-resistant Pseudomonas aeruginosa, and isolate Good purification effect

Inactive Publication Date: 2011-02-02
INST OF MICROBIOLOGY HEILONGJIANG ACADEMY OF SCI
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  • Abstract
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  • Application Information

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Problems solved by technology

[0006] The purpose of the present invention is to solve the problem of poor separation and purification effect in the existing separation method of Pseudomonas aeruginosa phage, and to provide a preparation method of drug-resistant Pseudomonas aeruginosa wide phage

Method used

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  • Preparation method of drug-resistant pseudomonas aeruginosa broad-host range phage

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specific Embodiment approach 1

[0011] Specific embodiment one: the preparation method of drug-resistant Pseudomonas aeruginosa wide phage in this embodiment is carried out according to the following steps: one, take 1L hospital sewage, add CaCl after gauze filtration 2 To a concentration of 1mmol / L, centrifuge after stirring and get the supernatant to obtain a sewage sample; 2. Add 100ml three times concentrated beef extract peptone liquid medium, 200ml sewage sample and 2ml Pseudomonas aeruginosa suspension to the 500ml Erlenmeyer flask lysate, then 160r / min shaker culture at 37°C for 12-24h, centrifuge, take the supernatant and filter, and then culture at 37°C overnight, after passing the sterility test, the lysate was obtained; 3. Take a loop The lysate was inoculated in LB liquid medium, and then 0.1ml was added at a concentration of 10 7 ~10 8 Pfu / ml Pseudomonas aeruginosa host bacterial suspension, mixed at room temperature for 15 minutes to obtain a mixed solution; 4. Take LB medium to dissolve and ...

specific Embodiment approach 2

[0016] Embodiment 2: This embodiment differs from Embodiment 1 in that the centrifugation in step 1 is carried out at a speed of 5000 r / min for 15 minutes. Other steps and parameters are the same as those in Embodiment 1.

specific Embodiment approach 3

[0017] Specific embodiment three: the difference between this embodiment and specific embodiment one or two is the preparation of Pseudomonas aeruginosa bacterial suspension in step 2: Pseudomonas aeruginosa is inoculated on LB solid medium, cultivated at 37°C for 17~26h, take Inoculation of a single colony in 5ml sterile water is complete. Other steps and parameters are the same as those in Embodiment 1 or Embodiment 2.

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Abstract

The invention discloses a preparation method of drug-resistant pseudomonas aeruginosa broad-host range phage, which relates to a preparation method of phage and solves the problem of poor separation and purification effect in the traditional separation method of the pseudomonas aeruginosa phage. The method comprises the following steps: 1, selecting a sewage sample; 2, preparing a lysis solution; 3, carrying out mixed cultivation on the lysis solution and a pseudomonas aeruginosa host cell suspension to obtain a mixed solution; 4, preparing a single phage plaque; and 5, inoculating the phage into a pseudomonas aeruginosa-containing culture medium, culturing until the pseudomonas aeruginosa is completely dissolved, and finally filtering. The preparation method of pseudomonas aeruginosa phage has the advantages of simple operation, good separation and purification effect and no phage breakage. The prepared drug-resistant pseudomonas aeruginosa broad-host range phage can be used for manufacturing autoimmune (vaccines) biological agents for treating diseases caused by the infection of drug-resistant pseudomonas aeruginosa, can be especially used as a topical spray for treating epidermal infectious diseases, and can avoid the drug-resistant pseudomonas aeruginosa produced by the use of antibiotics.

Description

technical field [0001] The invention relates to a preparation method of phage. Background technique [0002] In clinical practice, the abuse of antibiotics has led to the continuous growth of bacterial drug resistance, making bacterial infectious diseases, especially the infection of drug-resistant bacteria, a global problem in clinical treatment. Pseudomonas aeruginosa (Pseudomonas aeruginosa.PA), also known as Pseudomonas aeruginosa, is one of the important opportunistic pathogens in clinically acquired infections. According to the data of my country's National Nosocomial Pathogen Resistance Surveillance (NPRS), Pseudomonas aeruginosa ranks first among all nosocomial Gram-negative bacteria. Drug-resistant PA strains have caused great difficulties in the clinical treatment of PA infections. [0003] Bacteriophages are obligate parasites, so wherever there are bacteria in nature, you can find the existence of specific phages, that is, phages are distributed along with the d...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00
Inventor 孙建华曲晓军于冲夏海华王金英齐桂云佟忠山郭薇媛
Owner INST OF MICROBIOLOGY HEILONGJIANG ACADEMY OF SCI
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