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Vibrio parahaemolyticus bacteriophage, bdellovibrio bacteriovorus and application thereof

A technology of Vibrio hemolyticus and bacteriophage, applied in the direction of bacteriophage, virus/phage, application, etc., can solve the problems of long reproduction time and resistance mutation of leech vibrio, and achieve strong host specificity, long-lasting curative effect, and wide lysis spectrum. Effect

Active Publication Date: 2020-08-18
QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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  • Abstract
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  • Application Information

AI Technical Summary

Problems solved by technology

[0008] Aiming at the current disadvantages of single phage infection that easily produces resistance mutations and long breeding time of Bdellovibrio, the purpose of the present invention is to provide a combination of Vibrio parahaemolyticus phage VP-HYP MCS-1 and Bdellovibrio Halobacteriovorax sp.MCS-1 Drugs and their application in the prevention and treatment of pathogenic Vibrio parahaemolyticus infection in prawns

Method used

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  • Vibrio parahaemolyticus bacteriophage, bdellovibrio bacteriovorus and application thereof
  • Vibrio parahaemolyticus bacteriophage, bdellovibrio bacteriovorus and application thereof
  • Vibrio parahaemolyticus bacteriophage, bdellovibrio bacteriovorus and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0046] Example 1 Separation and purification of Vibrio parahaemolyticus phage VP-HYP MCS-1

[0047] 1) The host bacteria obtain:

[0048] Water samples were collected from prawn breeding ponds suffering from acute hepatopancreatic necrosis, and were spread on RO solid plates (yeast powder 1g, peptone 1g, sodium acetate 1g, trace elements 10mL, agar 15g, deionized water 1000mL, pH 7.8) -8.0), cultured at 28°C for 24h. Randomly pick a single colony, and repeatedly streak and purify the single colony 5 times. The purified single colony is MCS-1, which is stored in a glycerol tube at -80°C.

[0049] To clarify the taxonomic status of strain MCS-1, the 16S rDNA sequence was amplified using universal primers 27F (5'-AGAGTTTGATCCTGGCTCAG-3') and 1492R (5'-GGTTACCTTGTTACGACTT-3'). The sequence that was successfully amplified by PCR was cloned and sequenced to obtain the full-length sequence of 16S rDNA (GenBank accession number MN901166), and the full-length sequence was analyzed an...

Embodiment 2

[0064] Example 2 Separation and purification of Vibrio parahaemolyticus Bdellovibrio Halobacteriovorax sp.MCS-1

[0065] Seawater from the coast of Qingdao was collected, filtered through a sterile filter with a pore size of 0.22 μm, and then added to the logarithmic growth phase of Vibrio parahaemolyticus MCS-1 bacteria liquid, and cultured in a shaker (28°C, 150rpm). Samples were taken on the 1d, 3d, 5d, and 7d, and the RO liquid medium (yeast powder 1g, peptone 1g, sodium acetate 1g, trace elements 10mL, deionized water 100mL, pH 7.8-8.0) was used for gradient dilution, and the dilution gradient was 10 -1 , 10 -2 , 10 -3, 10 -4 , 10 -5 , 10 -6 , 10 -7 , 10 -8 Take 1mL of each dilution gradient and mix with 1mL of host bacteria, and then place them in a shaker (28°C, 150rpm) and incubate for 30min. After the incubation, add 5mL RO semi-solid medium, mix well and pour on each RO solid plate, shake well until the semi-solid solidifies. Seal with parafilm and place in ...

Embodiment 3

[0078] Example 3 Detection of host range of Vibrio parahaemolyticus phage VP-HYP MCS-1

[0079] Select the existing 13 Vibrio strains in the laboratory to detect the range of phage hosts. Among them, 13 Vibrio strains are Vibrio parahaemolyticus (Vibrio parahaemolyticus), Vibrio owesii (Vibrio Owensii), Vibrioneocaledonicus (Vibrio Subvibrio), Vibrio plantisponsor (plant probiotic Vibrio), Vibriohannami, Vibrio hyugaensis, Vibrio azureus (Vibrio aspirinus), Vibrio natriegens (Na Vibrio), Vibrio alginolyticus (Vibrio alginolyticus), Vibrio harveyi (Ha Vibrio vibrio), Vibrio variabilis (Vibrio variant), Vibrio galatheae, Vibrio gangliei.

[0080] Take 1 mL of the above-mentioned different bacteria in the logarithmic growth phase and mix them evenly with 5 mL of RO semi-solid (about 48 ° C), pour them into a double-layer plate, and then add 10 μL of the above-mentioned examples dropwise to the solidified RO semi-solid plate to obtain purified Phages were cultured in a constant t...

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Abstract

The invention belongs to the technical field of microorganism prevention and treatment, and particularly relates to a vibrio parahaemolyticus bacteriophage VP-HYP MCS-1 and bdellovibrio bacteriovorusHalobacterovoraxsp.MCS-1 composition and application thereof to prevention and treatment of prawn pathogenic vibrio parahaemolyticus infection. The vibrio parahaemolyticus bacteriophage is vibrio parahaemolyticus bacteriophage VP-HYPMCS-1, is preserved in China General Microbiological Culture Collection Center, and has a preservation number of CGMCC No.199693. The vibrio parahaemolyticus bacteriophage is a vibrio parahaemolyticus bacteriophage VP-HYPMCS-1. The bdellovibrio parahaemolyticus is bdellovibrio parahaemolyticus Halobacterovoraxsp.MCS-1, is preserved in the China General Microbiological Culture Collection Center (CGMCC), and has a preservation number of CGMCC No.199694. The invention also discloses a preparation method of the bdellovibrio parahaemolyticus strain. The virulent bacteriophage VP-HYP MCS-1 obtained through separation is high in host specificity and has cracking and killing effects on vibrio parahaemolyticus; the bdellovibrio bacteriovorus Halobacterium ovalax sp.MCS-1 is obtained through separation, the host spectrum of the bdellovibrio bacteriovorus Halobacterium ovalax sp.MCS-1 is wide, and the bdellovibrio bacteriovorus Halobacterium ovalax sp.MCS-1 has cracking and killing effects on various vibrios including vibrio parahaemolyticus; the vibrio parahaemolyticus bacteriophage and bdellovibrio bacteriovorus provided by the invention have good application prospects in prevention and control of prawn pathogenic vibrio parahaemolyticus infection.

Description

technical field [0001] The invention belongs to the technical field of microbial control, and in particular relates to a composition of Vibrio parahaemolyticus phage VP-HYP MCS-1, Bdellovibrio Halobacteriovorax sp. application. Background technique [0002] In recent years, my country's aquaculture industry has developed rapidly. With the increasing scale of marine aquaculture and the promotion of intensive farming methods, economic benefits have gradually increased. However, the frequent occurrence of bacterial diseases in aquaculture animals has caused great harm to the aquaculture industry. Among them, Vibrio anguillarum, Vibrio alginolyticus, Vibrio harveyi, Vibrio salmonicida, Vibrio parahaemolyticus and other Vibrio bacteria caused vibriosis is the most serious. [0003] As one of the main aquaculture animals in my country, prawn occupies an important position in the fishery economy. However, the acute hepatopancreas necrosis disease (AHPND) caused by Vibrio parah...

Claims

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Application Information

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IPC IPC(8): C12N7/00A61K35/76A61P31/04A61P1/16A61P1/18A01K61/13
CPCC12N7/00A61K35/76A61P31/04A61P1/16A61P1/18A01K61/13C12N2795/00021A61K2300/00Y02A40/81
Inventor 张永雨王增猛孙越超
Owner QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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