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Vibrio harveyi phage vB_KaS_PK22, Vibrio harveyi phage-containing phage composition and application of Vibrio harveyi phage

A technology of Vibrio harveii and bacteriophage, applied in the field of microorganisms, can solve problems such as antibiotic residues, and achieve the effects of avoiding antibiotic residues, reducing costs, and reducing morbidity and mortality.

Active Publication Date: 2020-09-25
QINGDAO PHAGEPHARM BIO TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The phage and its phage composition are safe to use and have no side effects, and are used to solve the infection caused by Vibrio harveyi and the water problem caused by the large proliferation of Vibrio harveyi in water, avoiding the antibiotics caused by the use of antibiotics. Residues, and the problem of induction of drug-resistant Vibrio harveii

Method used

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  • Vibrio harveyi phage vB_KaS_PK22, Vibrio harveyi phage-containing phage composition and application of Vibrio harveyi phage
  • Vibrio harveyi phage vB_KaS_PK22, Vibrio harveyi phage-containing phage composition and application of Vibrio harveyi phage
  • Vibrio harveyi phage vB_KaS_PK22, Vibrio harveyi phage-containing phage composition and application of Vibrio harveyi phage

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] The preparation of embodiment 1 phage

[0034] (1) Preparation of host bacterial suspension

[0035] The host bacterium VK02 was divided and marked on the TCBS plate, and a single colony was picked and inoculated into 5ml 2216E liquid medium, cultured at 170rpm and 37°C for 16-18h to obtain a fresh bacterial solution.

[0036] (2) Isolation and purification of phage

[0037] (1) Isolation of phage

[0038] Sewage collected from a seafood market in Binzhou City, Shandong Province was centrifuged at 11,000 r / min for 5 minutes, and the supernatant obtained by centrifugation was filtered with a 0.22 μm filter membrane, and then 2216E broth medium and 100 μl of host bacterial suspension were added. After culturing for 16 hours at 170 rpm and 37° C., centrifuge at 11,000 r / min for 5 minutes, and finally filter with a 0.22 μm filter membrane to obtain the phage stock solution.

[0039] Dilute the phage stock solution 10 times with 1×PBS solution, take 100 μl of the above-me...

Embodiment 2

[0048] The biological characteristic determination of embodiment 2 bacteriophage

[0049] (1) Electron microscope detection of phage

[0050] Use the above-mentioned phage suspension for electron microscope detection, take 20 μl of phage suspension and drop it on the copper grid, settle for 15 minutes, absorb excess liquid with filter paper, stain with 2% phosphotungstic acid for 30 minutes, and observe with electron microscope after drying.

[0051] The result is as figure 1 As shown, the length of the phage head is 75nm, the width of the head is 70nm, the length of the stretchable tail is 140nm, and the length of the stretchable tail is 140nm. According to the classification standard identification of the ninth report of the International Committee on Taxonomy of Viruses (ICTV), it can be determined that the phage belongs to the family Myoviridae, and it is named vB_KaS_PK22.

[0052] (2) Determination of lysis profile of phage

[0053] 1. Experimental method: In this exa...

Embodiment 3

[0081] Example 3 Determination of the effect of phage vB_KaS_PK22 on preventing Vibrio harveyi from infecting Penaeus vannamei 1. Experimental method:

[0082] Healthy Penaeus vannamei with a body weight of about 5g were randomly divided into 4 groups, 20 in each group. The test group was set up in parallel, and the bacteriophage vB_KaS_PK22 was evenly mixed with the prawn feed according to the volume-to-mass ratio of 5%, and dried in the shade. Feed at a dose of 3% of body weight with a phage content of 1 x 10 8 PFU / g. The control group (that is, the challenge group) was fed with 2216E medium soaked in the same dose. After 1 hour, all the prawns in the test group 1, test group 2 and control group were injected with Vibrio harveii at a dose of 4.05×10 6 CFU / only; at the same time, a blank control group was set up, and the prawns in this group were not treated in any way. The number of dead prawns in each group was recorded within 48 hours, and the protection rate of phage w...

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Abstract

The invention discloses a Vibrio harveyi phage vB_KaS_PK22, a Vibrio harveyi phage-containing phage composition and application of the Vibrio harveyi phage. The phage vB_KaS_PK22 is deposited in the China General Microbiological Culture Collection Center on May 15, 2020, and has a collection number of CGMCC No. 19974. The phage composition includes the Vibrio harveyi phage vB_KaS_PK22 and other matched phage. The phage has a wide lysis spectrum and high specificity, and is capable of effectively preventing and controlling Vibrio harveyi in aquaculture farms and greatly reducing the incidence of multiple diseases which are caused by Vibrio harveyi; and the Vibrio harveyi phage can also be applied widely to steps with easy losses which are caused by infection of Vibrio harveyi, daily disinfection of breeding environments and bacteriostasis of fresh aquatic products in the process of aquaculture, so that healthy development of the aquaculture industry is facilitated.

Description

technical field [0001] The invention relates to the technical field of microorganisms, in particular to a Vibrio harveyi phage vB_KaS_PK22, a composition containing the phage and applications thereof. Background technique [0002] Vibrio harveyi is a Gram-negative bacterium with luminescent properties. It is a normal seawater microbial population and an opportunistic pathogen commonly found in seawater. It mainly causes infections in prawns, fish and shellfish. In Thailand, India, Australia, Ecuador, Venezuela and China, Vibrio harveyi is one of the main pathogenic bacteria in cultured prawns. Studies have shown that Vibrio harveyi will multiply in large numbers when the aquaculture environment is poor and the immunity of prawns is reduced, which will cause the outbreak and rapid spread of prawn vibriosis. [0003] Due to the great impact of vibriosis on the aquaculture industry, predecessors have made a lot of research on vibriosis. At present, antibiotics have played a ma...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00A61K35/76A61P31/04A23K10/18A23K50/80C02F3/34A01N63/40A01P1/00A23L3/3571A23L5/20A23B4/22C12R1/92C02F103/20
CPCA23B4/22A23L3/3571A61K35/76A23K10/18A23K50/80A23L5/28A61P31/04A01N63/40C02F3/34C02F2103/20C02F2303/04C12N7/00C12N2795/10121C12N2795/10132
Inventor 潘强任慧英孙虎芝闫艳新戚铭芮
Owner QINGDAO PHAGEPHARM BIO TECH CO LTD
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