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High-cracking pseudomonas aeruginosa bacteriophage RDP-PA-20001 and application thereof

A technology of RDP-PA-20001 and Pseudomonas aeruginosa, applied in the direction of phage, virus/phage, application, etc., can solve the problems of untimely treatment and ineffective medication, and achieve wide acid-base tolerance range and strong lytic activity , the effect of high temperature tolerance

Active Publication Date: 2021-03-19
RECOM QINGDAO BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to provide a highly lytic Pseudomonas aeruginosa phage RDP-PA-20001, which aims to solve the problem of acute onset of Pseudomonas aeruginosa infection in poultry farms, untimely treatment and drug resistance of pathogenic bacteria. Difficulties such as drug ineffectiveness due to sex

Method used

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  • High-cracking pseudomonas aeruginosa bacteriophage RDP-PA-20001 and application thereof
  • High-cracking pseudomonas aeruginosa bacteriophage RDP-PA-20001 and application thereof
  • High-cracking pseudomonas aeruginosa bacteriophage RDP-PA-20001 and application thereof

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Experimental program
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Effect test

Embodiment 1

[0029] The isolation and identification of embodiment 1 Pseudomonas aeruginosa BP-20001

[0030] Sampling from the feces of chicken farms with Pseudomonas aeruginosa bacterial disease, using the method of aseptic operation, carry out 1:10 dilution with sterilized LB broth, take 10ml of 1:10 sample dilution and add to In 90ml common broth medium, culture at 37°C for 18-24 hours. A thin film of bacteria grows on the surface of the culture medium, and the culture medium is yellow-green.

[0031] Pick the culture from the thin film of the enrichment solution, inoculate it on the hexadecyltrimethylammonium bromide agar plate by streaking, and culture it at 37°C for 18 to 24 hours, and it will appear off-white, flat and wet on the surface. And the surrounding agar showed yellow-green colonies with a diameter of about 2-4mm. Pick a typical colony and streak it on a hexadecyltrimethylammonium bromide agar plate for three consecutive purifications, then pick a single colony and inocu...

Embodiment 2

[0032] Example 2 Isolation and identification of Pseudomonas aeruginosa phage RDP-PA-20001

[0033] The feces sample used in the experiment of the present invention was collected from a manure pond in a broiler chicken farm in Jimo, Qingdao in 2020, as a sample for phage isolation.

[0034] (1) Isolation of phage RDP-PA-20001:

[0035] Take 10㎎ of feces and put them in 10ml LB broth, soak overnight at 4°C, centrifuge at 10,000rpm for 5min, and filter the supernatant with a 0.22μm microporous membrane to sterilize. Take 3ml of the filtrate and 3ml of the host bacterium culture solution in the logarithmic growth phase, add them together to 20ml of autoclaved LB broth, put them in a 37°C incubator, and culture them overnight to multiply the possible phages. Take 3ml of the sample culture solution, centrifuge at 10000rpm for 5min, and take the supernatant to filter and sterilize with a 0.22μm microporous membrane. Assume that the above filtrate contains phages. The LB broth was...

Embodiment 3

[0041] Morphological observation of embodiment 3 phage

[0042] Using phosphotungstic acid negative staining method: 20 μL of phage proliferation solution (titer 10 9 pfu / mL) was dropped on a paraffin sheet, and the copper grid was placed on it for 10 minutes and then removed to absorb phage. Dry at natural room temperature for 2-3 minutes, stain with 2% phosphotungstic acid aqueous solution, blot the water after 2 minutes, dry naturally for 5 minutes, and observe the phage morphology with an electron microscope. Photo by electron microscope ( figure 2 ) It can be seen that the head of bacteriophage RDP-PA-20001 is icosahedral and stereosymmetric, the length of the head is 60.865nm, and the length of the phage tail is 183.254nm. According to the latest classification and naming of phages, it belongs to the long-tailed phage family .

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Abstract

The invention discloses high-cracking pseudomonas aeruginosa bacteriophage RDP-PA-20001 and application thereof in poultry pathogenic pseudomonas aeruginosa infection. The bacteriophage is separated from excrement of a Qingdao Jimo broiler chicken farm by using a double-layer plate method, is named as the pseudomonas aeruginosa bacteriophage RDP-PA-20001, and has a preservation number of CGMCC No.21410. The pseudomonas aeruginosa bacteriophage RDP-PA-20001 provided by the invention has relatively high temperature tolerance and a relatively wide acid-base tolerance range, and is beneficial to industrial production and preparation; and the bacteriophage has a strong cracking effect on pseudomonas aeruginosa BP-20001, can effectively prevent and control the generation and propagation of pseudomonas aeruginosa, and can be used for efficiently and reliably treating poultry animal diseases caused by the pseudomonas aeruginosa.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a highly lytic Pseudomonas aeruginosa phage RDP-PA-20001 and its application in poultry animals infected by Pseudomonas aeruginosa. Background technique [0002] Pseudomonas aeruginosa is a common Gram-negative bacillus, referred to as Pseudomonas aeruginosa. , mainly causing nosocomial infections, often causing infections of the respiratory tract, urinary system and burn wounds. It is stipulated in the hygienic standard of cosmetics that this bacterium should not be detected, and Pseudomonas aeruginosa in swimming pool water can also cause human upper respiratory tract infection. Human consumption of food contaminated by Pseudomonas aeruginosa can cause food poisoning, and foodborne Pseudomonas aeruginosa contamination has become one of the risk factors that endanger human health. Pseudomonas aeruginosa has asymptomatic carrier phenomenon in poultry, which can cause septicemia in ...

Claims

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Application Information

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IPC IPC(8): C12N7/00A01N63/40A01P1/00
CPCA01N63/40C12N7/00C12N2795/10321
Inventor 杜新永李先胜罗成盛赵丹丹王晓铃张庆王立坤刘玉庆盖春云马如霞刘爽
Owner RECOM QINGDAO BIOTECH CO LTD
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