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Edwardsiella catfish bacteriophage EIP-1 and application thereof

An EIP-1, bacteriophage technology, applied in virus/bacteriophage, medical raw materials derived from virus/bacteriophage, application, etc., can solve the problem of phage treatment with few phage species, and achieve the effect of improving the therapeutic effect and expanding the scope of application

Pending Publication Date: 2022-07-22
UNIVERSITY OF CHINESE ACADEMY OF SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the small number of isolated phage species is an important factor limiting the further development of phage therapy.

Method used

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  • Edwardsiella catfish bacteriophage EIP-1 and application thereof
  • Edwardsiella catfish bacteriophage EIP-1 and application thereof
  • Edwardsiella catfish bacteriophage EIP-1 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Example 1 Isolation and purification of phage

[0020] Take 38.5 g of brain-heart infusion broth medium and put it in 1 L of ultrapure water to obtain brain-heart infusion liquid medium.

[0021] Take 38.5 g of brain-heart infusion broth medium, put it in 1 L of ultrapure water, and add 7.5 g of agar powder to obtain brain-heart infusion semi-solid medium.

[0022] Take 52 g of brain heart leachate agar medium and put it in 1 L of ultrapure water to obtain brain heart leachate solid medium.

[0023] After mixing the above medium, autoclave at 121 °C for 15 min.

[0024] The test water sample of the present invention was collected in a fish farm in Beijing in September 2020, and the phage was isolated from the water sample as a matrix.

[0025] 10 mL of environmental water samples were filtered and sterilized with a 0.22 μm microporous membrane, and Edwardella catfish MCCC 1K00244 was cultured overnight to logarithmic growth phase. Take 5 mL of filtered water sample, ...

Embodiment 2

[0027] Example 2 Morphological observation of bacteriophage

[0028] Take 20 μL of the phage suspension and drop it on the copper mesh. After it settles naturally for 10 minutes, it is blotted dry from the side with dry filter paper. After drying for about 1 minute, add 1 drop of 1% uranyl acetate on the copper mesh, stain for 2 minutes, and then use it carefully to dry. The excess dye was blotted dry from the side of the filter paper, and after 30 minutes of natural drying in the dark, observation was performed with a transmission electron microscope (JEM-1400).

[0029] Transmission electron microscopy results such as figure 2 As shown, phage EIP-1 ​​has an icosahedral head and a retractable tail, the head diameter is about 45.74 nm, the tail length is about 96.61 nm, and the tail diameter is about 24.49 nm. According to the eighth report on the classification of viruses by the International Organization for Taxonomy of Viruses (ICTV), the phage EIP-1 ​​can be preliminaril...

Embodiment 3

[0030] Example 3 Phage genome analysis and identification

[0031] Phage DNA was extracted by the phenol-chloroform method, and the DNA precipitate was dissolved in TE buffer for whole-genome sequencing. The sequencing platform was IlluminaNovaSeq. Before sequencing on the machine, the sample DNA was randomly broken into small fragments of about 350bp, and then the NEB standard library building kit was used to select specific adapters, the sample was prepared for library preparation and the size of the library fragments was detected by aglent2100, and then the molar concentration of the library was detected by qPCR , Illumina NovaSeq was used for PE 2×150 sequencing after the library was qualified. Then use spades v.3.11.1 splicing software to splicing the optimized sequence to get the best assembly result; use CARD (the Comprehensive Antibiotic Research Database) and VFDB (Virulence Factors Database) database to predict online whether the phage EPP-1 genome contains antibioti...

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Abstract

The invention discloses an edwardsiella catfish bacteriophage EIP-1 and application thereof, and belongs to the technical field of biology. The invention discloses a strain of edwardsiella catfish bacteriophage EIP-1, the preservation number of which is CGMCC (China General Microbiological Culture Collection Center) No.45077. The separation and identification of the bacteriophage not only enriches a bacteriophage germplasm resource library, but also provides a selectable potential scheme for solving the problem of fish death caused by Edwardsiella catfish infection in aquaculture; through whole genome sequencing of the bacteriophage EIP-1, functional genes and functional proteins of the bacteriophage EIP-1 can be further deeply excavated, and one or more enzymes with a wide-spectrum splitting effect are provided for bacteriophage treatment and biological prevention and treatment of Edwardsiella catfish infection in aquaculture, so that the application universality and effectiveness of the bacteriophage are improved; the bacteriophage EIP-1 can effectively split edwardsiella catfish and is used for preventing and treating fish infection death caused by edwardsiella catfish infection in aquaculture.

Description

technical field [0001] The present invention relates to the field of biotechnology, in particular to a catfish Edwardsiella phage EIP-1 ​​and its application. Background technique [0002] E. catfish is a common fish pathogen that can infect a variety of aquaculture fish including turbot, eel, catfish, zebrafish, yellow catfish, channel catfish, rainbow trout, and tilapia. E. catfish primarily infects the digestive tract and nervous system of fish. The specific forms of digestive tract infection are: catfish Edwardella enter the blood of fish through the digestive tract, and then use the blood circulatory system to infect various tissues, resulting in symptoms such as inflammation, congestion, bleeding and ulceration in various organs; the specific forms of nervous system infection are: Catfish Edwardella enter the olfactory bulb through the nasal cavity of the fish, then enter the brain through the olfactory bulb, and finally infect the meninges, skull and head skin of the...

Claims

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Application Information

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IPC IPC(8): C12N7/00A61K35/76A61P31/04A01N63/40A01P1/00C02F3/34C12R1/92C02F103/20
CPCC12N7/00A61K35/76A61P31/04A01N63/40C02F3/34C12N2795/10121C12N2795/10131C12N2795/10132C02F2103/20
Inventor 刘如铟寸淑娟韩岗华刘新春余志晟
Owner UNIVERSITY OF CHINESE ACADEMY OF SCIENCES
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