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Salmonella bacteriophage capable of simultaneously splitting escherichia coli, and application of salmonella bacteriophage

A technology for Salmonella and Escherichia coli, which is applied to phages, viruses/phages, and medical raw materials derived from viruses/phages, etc. It can solve the problems of bacterial resistance, ineffective medication, untimely treatment, etc., and achieve the effect of strong cracking ability

Active Publication Date: 2021-11-02
RECOM QINGDAO BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0011] The purpose of the present invention is to provide a Salmonella phage that can lyse Escherichia coli at the same time and its application, aiming to solve the problem of acute onset and untimely treatment of poultry infected with Salmonella in farms, and ineffective medication due to drug resistance of pathogenic bacteria. question

Method used

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  • Salmonella bacteriophage capable of simultaneously splitting escherichia coli, and application of salmonella bacteriophage
  • Salmonella bacteriophage capable of simultaneously splitting escherichia coli, and application of salmonella bacteriophage
  • Salmonella bacteriophage capable of simultaneously splitting escherichia coli, and application of salmonella bacteriophage

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] The isolation and identification of embodiment 1 pathogenic Salmonella BS20078

[0042] Sampling from the diseased farm, aseptically take the liver of diseased poultry, streak it on the selective medium (SS agar), culture it at 37°C for 18-24 hours, and form a round, flat, neat edge and smooth surface on the medium Wet red colonies, pick typical colonies and continue to streak and purify 3 times, then pick a single colony and inoculate in 5mL LB broth, shake and culture at 200rpm at 37°C for 8h to obtain a uniform turbid bacterial suspension. Through 16sRNA molecular identification and serotype identification, it was determined to be pathogenic Salmonella, and one of the strains was named BS20078 and stored in a -80°C refrigerator.

Embodiment 2

[0043] Example 2 Isolation and Identification of Phage RDP-SA-21003

[0044] Manure treatment: Take manure from the farm, weigh 5g of chicken manure and add it to 10mL of sterile water to soak overnight, then centrifuge the overnight leaching solution at 10,000rpm for 5min, take the supernatant and pass it through a 0.22μm filter, and use the filtrate for later use.

[0045] Prepare mixed bacterial suspension: Take 0.2mL bacterial suspension and 0.1mL filtrate and add it to 5mL LB broth, culture overnight at 37°C with shaking at 200rpm, then centrifuge at 10000rpm for 5min, pass the supernatant through a 0.22μm filter, and filter out liquid for use.

[0046] Phage Isolation: Use the double-plate method to separate the phage. Take 0.1mL of the filtrate of the mixed bacterial suspension and 0.2mL of the Salmonella suspension, mix evenly, bathe in 37°C water for 10min, spread double plates, and place them in a 37°C incubator for incubation After 6-8h, observe the result. If the...

Embodiment 3

[0049] Morphological observation of embodiment 3 phage

[0050] Take 20 μL of the liquid containing crude phage particles and drop them on the copper grid, let it settle naturally for 15 minutes, and absorb the excess liquid from the side with filter paper, add a drop of 2% phosphotungstic acid (PTA) on the copper grid to stain the phage for 10 minutes, and then use the filter paper to remove the Aspirate the staining solution from the side, and observe the phage morphology with an electron microscope after the sample is dry, as shown in figure 1 shown.

[0051] Depend on figure 1 It can be seen that the phage RDP-SA-21003 has a polyhedral stereosymmetric head, which wraps the nucleic acid, has a diameter of about 60 nm, and has a tail about 30 nm in length, and the neck connects the head and the tail. According to the Ninth Report of the International Virus Taxonomy Organization Virus Classification, the phage can be classified as Brachyviridae of the order Brachyviridae. ...

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Abstract

The invention discloses a salmonella bacteriophage RDP-SA-21003 capable of simultaneously splitting escherichia coli, and the preservation number is CGMCC No. 22494. After whole genes are sequenced, the salmonella bacteriophage RDP-SA-21003 does not have virulence genes and lyogenic genes; the cracking efficiency of the obtained bacteriophage on host bacteria BS20078 is 98.7% or above, when the bacteriophage is independently used, the cracking capacity on salmonella is relatively high, and the cracking spectrum is wide; and the bacteriophage RDP-SA-21003 has the capability of cracking escherichia coli causing chicken air sacculitis and peritonitis.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a Salmonella phage capable of simultaneously lysing Escherichia coli and its application. Background technique [0002] Salmonella pullorum (Salmonella pullorum) belongs to the Enterobacteriaceae family, Gram-negative enterobacteria, and is the pathogen of salmonellosis. It exists widely in nature, and about 1,000 strains have been discovered so far. Salmonella can be isolated from most vertebrates, and it mainly lives in the intestinal tract of humans and animals. The size of the bacteria is (0.6-0.9)×(1-3) μm, generally without spores and capsules, most of them have flagella, and the antigenic structure and biochemical characteristics are relatively complex. [0003] Salmonella species are diverse and more than 2500 Salmonella enterica serotypes have been described based on antigenic properties defined by antigens on the surface of the bacteria. Poultry infection caused by Salmo...

Claims

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Application Information

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IPC IPC(8): C12N7/00A61K35/76A61K45/06A61P31/04A23K10/18A23K50/75A01N63/40A01P1/00C11D3/38C11D3/48C12R1/92
CPCC12N7/00A61K35/76A61K45/06A61P31/04A23K10/18A23K50/75A01N63/40C11D3/381C11D3/48C12N2795/10221C12N2795/10232C12N2795/10231
Inventor 杜新永张得彦李先胜马如霞王立坤
Owner RECOM QINGDAO BIOTECH CO LTD
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