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Isolation and application of a Salmonella phage rdp-sa-17118

A technology of RDP-SA-17118 and Salmonella, applied in the field of microorganisms, can solve the problems of drug failure and lack of antibiotics, and achieve the effect of good acid-base tolerance

Active Publication Date: 2020-08-28
QINGDAO RUNDA BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in actual farming, due to the lack of relevant knowledge of antibiotic use by farmers, broad-spectrum antibiotics are often used or a large number of drugs are used, resulting in the gradual increase of bacterial resistance to antibiotics and the gradual ineffectiveness of drugs.

Method used

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  • Isolation and application of a Salmonella phage rdp-sa-17118
  • Isolation and application of a Salmonella phage rdp-sa-17118
  • Isolation and application of a Salmonella phage rdp-sa-17118

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1 Isolation and Identification of Pathogenic Salmonella Avian S6

[0038] 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, named S6, and stored in a -80°C refrigerator.

Embodiment 2

[0039] Example 2 Isolation and identification of phage RDP-SA-17118:

[0040](1) Manure treatment: Weigh 5 g of chicken manure and add it to 10 mL of sterile water to soak overnight, then centrifuge the overnight leachate at 10,000 rpm for 5 minutes, take the supernatant and pass it through a 0.22 μm filter, and use the filtrate for later use;

[0041] (2) Preparation of mixed bacterial suspension: Take 0.2mL of bacterial suspension and 0.1mL of filtrate and add it to 5mL of LB broth, culture at 37°C, shake at 200rpm overnight, then centrifuge at 10,000rpm for 5min, take the supernatant and pass it through a 0.22μm filter. Reserve the filtrate;

[0042] (3) Separation of bacteriophages: Separation of phages was carried out using the double-plate method. After mixing 0.1 mL of the filtrate of the mixed bacterial suspension and 0.2 mL of the host Salmonella suspension, they were bathed in water at 37°C for 10 minutes, and then double-plates were spread and placed at 37°C. After...

Embodiment 3

[0045] Electron microscope observation of embodiment 3 phage

[0046] 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.

[0047] The bacteriophage RDP-SA-17118 has a polyhedral three-dimensional symmetrical head wrapped around nucleic acid, with a diameter of about 70nm, a tail about 120nm in length, a tail sheath, and a neck connecting the head and tail. According to the Ninth Report of the International Virus Taxonomy Organization Virus Classification, the bacteriophage is classified as Myoviridae of the order Cauviridae.

[0048] Whol...

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Abstract

The present invention discloses the isolation and application of a Salmonella phage RDP-SA-17118. Its host is Salmonella S6. The phage can form phage plaques with a diameter of 2mm-4mm on a double plate; through electron microscope observation, the phage has A polyhedral three-dimensional symmetrical head, wrapped in nucleic acid, with a diameter of 70nm, a tail with a length of 120nm, a tail sheath, and a neck connecting the head and tail. The phage titer is high, up to 10 12 pfu / mL, it has good tolerance to temperature and pH, after 29 consecutive passages, its titer is still maintained at 10 12 Above pfu / mL, dilute 10 6 After doubling, it still has a good cracking effect on the host. After feeding, the existence of the phage can be detected in the heart, liver, lung, kidney, thymus and serum species. Through animal experiments, it has the potential to treat chicken salmonellosis. better effect.

Description

technical field [0001] The invention relates to the field of microorganisms, in particular to the isolation and application of a strain of Salmonella phage RDP-SA-17118. Background technique [0002] The problem of increased bacterial resistance has long received international attention. The European Union has banned the use of large amounts of antibiotics in feed since 1999, and the European Union, the United States and other regions and countries have begun to monitor the problem of drug resistance in their regions. The results not only show that Salmonella has high resistance to a variety of antibiotics, but also the problem of multi-drug resistance is becoming more and more serious. In my country, the problem of bacterial drug resistance has been paid more attention in recent years, and related drug resistance and drug sensitivity tests are also being carried out continuously. Current research on drug resistance mainly focuses on the molecular level analysis of drug res...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K35/76A61P31/04C12R1/92
CPCA61K35/76A61P31/04C12N7/00C12N2795/10121C12N2795/10132C12N2795/10151
Inventor 杜新永李先胜马如霞
Owner QINGDAO RUNDA BIOTECH
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