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A strong lytic Staphylococcus aureus phage rdp-sr-20001 and its application

A technology of RDP-SR-20001 and Staphylococcus, which is applied in the direction of bacteriophage, virus/phage, medical raw materials derived from virus/phage, etc., can solve the problems of ineffective medication and untimely treatment, and achieve the effect of treating pollution

Active Publication Date: 2022-03-22
RECOM QINGDAO BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The purpose of the present invention is to provide a strong lytic Staphylococcus aureus phage RDP-SR-20001, aiming to solve the problem of acute onset of Staphylococcus aureus infection in poultry farms and untimely treatment, as well as the problem caused by the drug resistance of pathogenic bacteria. Difficulties such as ineffectiveness of medication

Method used

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  • A strong lytic Staphylococcus aureus phage rdp-sr-20001 and its application
  • A strong lytic Staphylococcus aureus phage rdp-sr-20001 and its application
  • A strong lytic Staphylococcus aureus phage rdp-sr-20001 and its application

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Effect test

Embodiment 1

[0031] Embodiment 1 Isolation and Identification of Pathogenic Staphylococcus aureus BS-20001

[0032] Sampling from diseased farms, aseptically take joint exudate, liver and spleen of diseased chickens, streak on BHI medium, culture at 37°C for 24-48 hours, then perform Gram staining and microscope observation on the bacteria growing on the plate , for Gram-positive cocci colonies were streaked and purified again, and the above steps were repeated 3 times until colonies with uniform size and shape were grown, then a single colony was picked and inoculated in 5mL LB broth, 37°C 200rpm Shake culture for 10 h to obtain a uniform turbid bacterial suspension. Through biochemical identification and 16sRNA molecular identification, it was determined to be pathogenic Staphylococcus aureus, and one of them was named BS-20001 (refer to figure 1 shown), and stored in a -80°C refrigerator.

Embodiment 2

[0033] Example 2 Isolation and Identification of Staphylococcus aureus Phage RDP-SR-20001

[0034] (1) Sewage treatment: Take sewage from the farm, filter the collected sewage samples with multi-layer gauze, then place the filtrate at 4°C for 10,000×g centrifugation for 15 minutes, and use a disposable filter with a pore size of 0.22 μm on the top The clear liquid was filtered, and the filtrate was placed at 4°C for later use;

[0035] (2) Preparation of mixed bacterial suspension: Take 0.2 mL of bacterial suspension and 1 mL of filtrate and add it to 5 mL of LB broth, culture overnight at 37°C with shaking at 200 rpm, then centrifuge at 10,000 rpm for 5 min, and pass the supernatant through a 0.22 μm filter. The filtrate is used for later use.

[0036](3) Separation of bacteriophages: Separation of phages was carried out using the double-plate method. After mixing 0.1mL of the filtrate of the mixed bacterial suspension and 0.2mL of Staphylococcus aureus BS-20001, they were p...

Embodiment 3

[0039] Electron microscope observation of embodiment 3 phage

[0040] Phages were treated with phosphotungstic acid negative staining method: take 100 μL of purified and proliferated phages, drop them on paraffin sheets, place the film side of the copper mesh on the phage droplet, and remove the copper mesh from the droplet after 10 minutes Put it on a clean filter paper, let it dry for 3 minutes, add a drop of 2% phosphotungstic acid (PTA) on the copper grid to stain the phage for 10 minutes, then use the filter paper to absorb the staining solution from the side, and observe the shape of the phage with an electron microscope after the sample is dry .

[0041] Depend on image 3 It can be seen that the head of phage RDP-SR-20001 is a regular polyhedron structure with a diameter of about 41nm and no tail; according to the ninth report of the International Virus Taxonomy Organization on the Classification of Viruses, the morphology of the phage conforms to the characteristics ...

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Abstract

The invention discloses a strongly lytic Staphylococcus aureus phage RDP-SR-20001 and its application in poultry pathogenic Staphylococcus aureus infection. The present invention utilizes the double-layer plate method to obtain a Staphylococcus aureus BS‑20001 strain from diseased broiler lesion tissue of Qingdao Jimo broiler farm, and then uses Staphylococcus aureus BS‑20001 as the host to isolate gold from the farm sewage. Staphylococcus aureus phage RDP‑SR‑20001, the deposit number is CGMCCNo.21409. The Staphylococcus aureus phage RDP‑SR‑20001 provided by the present invention has a strong cracking effect on pathogenic Staphylococcus aureus BS‑20001 in the poultry breeding environment, and is used for industrial production of phages for pathogenic Staphylococcus aureus in the poultry breeding environment The control provides a phage source.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a strongly lytic Staphylococcus aureus phage RDP-SR-20001 and its application. Background technique [0002] Staphylococcus aureus (Staphylococcus aureus) is an important human pathogenic bacteria, belonging to the genus Staphylococcus (Staphylococcus). Staphylococcus aureus is ubiquitous in nature and can be found in air, water, dust, and human and animal waste. Therefore, there are many opportunities for food to be contaminated by it. Staphylococcus aureus is pathogenic to both animals and humans. Among them, air pollution is the main route of transmission in animal populations. Rabbits are sensitive to Staphylococcus aureus and can cause sepsis in rabbits when they are infected through damaged skin surfaces. Mice infected with Staphylococcus aureus can develop soybean-sized blotches on the skin or sepsis. Broiler chickens and other poultry infected with Staphylococcus aureus ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A23K50/75A23K10/16A23L33/10A61K35/76A61P31/04C11D3/38A01N63/40A01P3/00A01P1/00C12R1/92
CPCC12N7/00A23K50/75A23K10/16A23L33/10A61K35/76A61P31/04C11D3/381A01N63/40C12N2795/00021C12N2795/00031C12N2795/00032A23V2002/00A23V2200/30
Inventor 杜新永李先胜罗成盛刘爽赵丹丹张庆刘玉庆盖春云王立坤马如霞
Owner RECOM QINGDAO BIOTECH CO LTD
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