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Cracking vibrio parahaemolyticus bacteriophage RDP-VP-21007 and application thereof

A technology of RDP-VP-21007 and Vibrio hemolyticus, which is applied in the direction of phage, virus/phage, and medical raw materials derived from virus/phage, and can solve problems such as ineffective medication, acute onset of aquatic animals, and drug resistance of pathogenic bacteria , to achieve strong cracking activity and good application prospects

Inactive 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

[0006] The purpose of the present invention is to provide a specific lytic Vibrio parahaemolyticus phage RDP-VP-21007, to replace the application of antibiotics in the prevention and treatment of aquaculture, and to solve the problem of acute onset of aquatic animals and untimely treatment, and the Antibiotic resistance of bacteria leads to problems such as ineffective medication

Method used

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  • Cracking vibrio parahaemolyticus bacteriophage RDP-VP-21007 and application thereof
  • Cracking vibrio parahaemolyticus bacteriophage RDP-VP-21007 and application thereof
  • Cracking vibrio parahaemolyticus bacteriophage RDP-VP-21007 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Embodiment 1 Isolation and identification of pathogenic bacteria and bacteriophage

[0036] (1) Isolation and identification of pathogenic Vibrio parahaemolyticus BV-21001

[0037] Sampling from the diseased sea cucumber lesions, using aseptic technique, streaking on TCBS medium, culturing at 37°C for 18-24 hours, showing round green colonies with neat edges on TCBS agar plates, with a diameter of about 2-4mm ; Then pick typical colonies and streak on the Chromagar Vibrio chromogenic medium. After culturing at 37°C for 18-24 hours, purple colonies appear on the Chromagar Vibrio chromogenic medium. Pick typical colonies and continue to streak After purification for 3 times, a single colony was picked and inoculated in 5 mL of 2216E liquid medium, and cultured with shaking at 200 rpm at 37°C for 8 hours to obtain a uniform turbid bacterial suspension. Through 16sRNA molecular identification, it was identified as Vibrio parahaemolyticus, and one of them was named BV-21001...

Embodiment 2

[0046] Electron microscope observation of embodiment 2 phage

[0047] Drop phage liquid (titer 10 10 pfu / m L) 100 μL, place the membrane side of the copper mesh on the phage droplet, take it off after 10 minutes, and let it dry naturally in the air for 2-3 minutes. Then drop a drop of 2% phosphotungstic acid (PTA) aqueous solution on the copper grid for staining, take it off after 10 minutes, dry it in the air for 10-15 minutes, observe with an electron microscope, and select a clear phage image to take pictures.

[0048] photo by electron microscope figure 1 It can be seen that the phage has a regular hexagonal head with a diameter of about 55 nm and a tail.

Embodiment 3

[0049] Example 3 Phage Genome Sequencing

[0050] After the enrichment culture of a single phage, centrifuge at 8000g for 15 minutes at 4°C, add 10% PEG8000 and 0.5M NaCl and let it stand overnight, then add an equal amount of chloroform to mix well, and centrifuge at 5000g for 10 minutes after standing for stratification , after removing the chloroform layer and PEG layer, add restriction endonuclease for digestion treatment, and use gradient density cesium chloride to suspend phage, and then use TM buffer to dialyze 3 times, each time for 30 minutes, and send it to Huada Gene for biological sequencing The company performs whole genome sequencing. The results are shown in the sequence listing.

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Abstract

The invention discloses a cracking vibrio parahaemolyticus bacteriophage RDP-VP-21007 and an application thereof. The host of the bacteriophage is vibrio parahaemolyticus. Through electron microscope observation, the head of the bacteriophage is in a regular hexagon shape, the diameter of the head is about 55 nm, the head is provided with a tail, and the preservation number is CGMCC No.22492. The vibrio parahaemolyticus bacteriophage RDP-VP-21007 has a strong cracking effect on vibrio parahaemolyticus in a water body, has relatively high temperature tolerance and a relatively wide acid-base tolerance range, can be used for efficiently treating vibrio parahaemolyticus diseases caused by the pathogenic bacteria in aquaculture, and can be used as a disinfectant. A bacteriophage source is provided for industrial, large-scale and programmed production of bacteriophages and treatment of aquaculture bacterial diseases caused by vibrio parahaemolyticus.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a lytic Vibrio parahaemolyticus phage and its application in aquaculture Vibrio parahaemolyticus infection. Background technique [0002] Vibrio parahaemolyticus, also known as pathogenic halophilic bacteria, is a marine pathogenic bacteria and is a zoonotic pathogen. It was isolated and discovered in Japan in 1950 in an outbreak of food poisoning. Vibrio parahaemolyticus is widely distributed, mainly in seawater and aquatic products. According to statistics, the detection rate of Vibrio parahaemolyticus in coastal seawater in East China is 47.5% to 66.5%. 45.6% to 48.7%, up to 90% or more in summer. In addition to seafood, the presence of Vibrio parahaemolyticus has been found in livestock and poultry meat, pickled vegetables, salted eggs, and freshwater fish. [0003] In recent years, my country's aquaculture industry has developed rapidly. With the increasing scale of marine a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00A01N63/40A01P1/00A61K35/76A61P31/04A23L33/135A23K10/18C11D3/38C11D3/48C12R1/92
CPCC12N7/00A01N63/40A61K35/76A61P31/04A23L33/135A23K10/18C11D3/381C11D3/48C12N2795/10021C12N2795/10031C12N2795/10032A23V2002/00A01N63/20A23V2200/30
Inventor 杜新永马如霞崔耐李先胜王晓铃
Owner RECOM QINGDAO BIOTECH CO LTD
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