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Lytic vibrio parahaemolyticus bacteriophage RDP-VP-19003 and application thereof

A technology of RDP-VP-19003, Vibrio hemolyticus, applied in the direction of bacteriophage, virus/phage, medical raw materials derived from virus/phage, etc. Effect

Active Publication Date: 2021-01-08
RECOM QINGDAO BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The invention is based on the order of the corresponding country to completely ban antibiotics, solves the problems of bacterial drug resistance and antibiotic abuse, and provides a safe and reliable lytic proteus mirabilis phage for treating poultry proteus mirabilis disease

Method used

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  • Lytic vibrio parahaemolyticus bacteriophage RDP-VP-19003 and application thereof
  • Lytic vibrio parahaemolyticus bacteriophage RDP-VP-19003 and application thereof
  • Lytic vibrio parahaemolyticus bacteriophage RDP-VP-19003 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Example 1 Isolation and identification of Vibrio parahaemolyticus BV-19004

[0032] Samples were taken from the diseased sea cucumber lesions, using aseptic operation, streaked on TCBS medium, and cultured at 37°C for 18-24 hours, on the TCBS agar plate appeared round green colonies with neat edges and a diameter of about 2-4. mm; then pick a typical colony and streak it 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 Line purification was performed 3 times, and then 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-19004.

Embodiment 2

[0033] Example 2 Isolation and identification of Vibrio parahaemolyticus phage RDP-VP-19003

[0034] The sewage sample used in the test of the present invention was collected from the surrounding sewage of farmers in Jimo, Qingdao in 2018, and was used as a water sample for the separation of bacteriophage.

[0035] Take 10 ml of sewage, centrifuge at 10,000rpm for 5 minutes, remove larger impurities and most of the bacteria, and then use a 0.22 micron microporous membrane to filter and sterilize. Take 3 milliliters of the filtrate and 3 milliliters of the host bacterial suspension, add them together into 20 milliliters of autoclaved LB broth, and then place them in a 37°C incubator for overnight culture. After culturing, take 5 ml, centrifuge at 10,000 rpm for 5 minutes, and then filter and sterilize with a 0.22 micron microporous filter. The filtrate is the stock solution intended to contain phage. Then, use the double-plate method to identify whether there are phages; pick ...

Embodiment 3

[0037] Example 3 Morphological observation of bacteriophage

[0038] Drop phage liquid (titer 10 10 pfu / m L) 100 μL, put the membrane side of the copper mesh on the phage droplet, take it off after 10 min, and let it dry naturally in the air for 2-3 min. Then drop a drop of 2% phosphotungstic acid (PTA) aqueous solution on the copper grid for staining, take it off after 10 min, dry it in the air for 10-15 min, observe with an electron microscope, and select a clear phage image to take pictures. Photo by electron microscope ( figure 2 ) It can be seen that the phage has no tail, is a regular hexagon, and the diameter of the head is about 55nm.

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Abstract

The invention discloses a lytic vibrio parahaemolyticus bacteriophage RDP-VP-19003 and an application thereof to pathogenic vibrio parahaemolyticus infection of aquatic animals. According to the invention, the bacteriophage is separated from wastewater of Qingdao Jimo aquaculture farmers by using a double-layer plate method, is named as vibrio parahaemolyticus bacteriophage RDP-VP-19003, and has apreservation number of CGMCC No. 19385. The vibrio parahaemolyticus bacteriophage RDP-VP-19003 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. The bacteriophage has a strong cracking effect on the vibrio parahaemolyticus BV-19004, can effectively prevent and control generation and propagation of the vibrio parahaemolyticus, and can be used for efficiently and reliably treating aquatic animal diseases caused by the vibrio parahaemolyticus and improving the water environment.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a lytic Vibrio parahaemolyticus phage RDP-VP-19003 and its application in pathogenic Vibrio parahaemolyticus infection of aquatic animals. Background technique [0002] In recent years, my country's aquaculture industry has developed rapidly. With the increasing scale of marine aquaculture and the continuous promotion of intensive aquaculture methods, the frequent occurrence of fish and shrimp diseases has posed a serious threat to the healthy development of large-scale aquaculture of aquatic products. The pathogens of aquatic animals mainly include bacteria, viruses, parasites, etc. Among them, bacterial diseases are more serious diseases to aquatic animals. [0003] Vibrio parahaemolyticus is an important zoonotic pathogen, which not only causes severe marine animal vibriosis, but also causes human infection and even death. Every summer and autumn is the outbreak period of the di...

Claims

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Application Information

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IPC IPC(8): C12N7/00A23K50/80A23K10/18C11D3/38A01N63/40A01P1/00A61K35/76A61P31/04C12R1/92
CPCA61K35/76A23K10/18A23K50/80A61P31/04A01N63/40C11D3/381C12N7/00C12N2795/00021C12N2795/00031C12N2795/00032
Inventor 杜新永李霞马如霞李先胜赵丹丹罗成盛张庆刘玉庆盖春云衣蕾刘爽
Owner RECOM QINGDAO BIOTECH CO LTD
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