Vibrio alginolyticus bacteriophage GVA-P21 with high fermentation rate and lasting bacteriostasis and application thereof

A technology of GVA-P21 and Vibrio alginolyticus, which is applied in the field of microbial technology application, can solve the problems of drug resistance of pathogenic bacteria, and achieve the effect of strong specificity, intuitive antibacterial effect and excellent antibacterial performance

Pending Publication Date: 2022-07-05
武汉格瑞农生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, antibiotics are mainly used for the treatment of vibriosis related to aquaculture. However, with the abuse of antibiotics, more

Method used

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  • Vibrio alginolyticus bacteriophage GVA-P21 with high fermentation rate and lasting bacteriostasis and application thereof
  • Vibrio alginolyticus bacteriophage GVA-P21 with high fermentation rate and lasting bacteriostasis and application thereof
  • Vibrio alginolyticus bacteriophage GVA-P21 with high fermentation rate and lasting bacteriostasis and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Isolation and Purification of Vibrio alginolyticus Phage GVA-P21

[0025] Three water samples were collected from an aquaculture pond in Zhanjiang, Guangdong, with 50 mL each, centrifuged at 6000 rpm / min for 10 min, the supernatant was taken, and sterilized with a 0.22 μm filter, and 5 mL of the treated supernatant and 0.5 mL of Vibrio alginolyticus ( The concentration is about 5×108CFU / mL) and mixed evenly, after static adsorption for 15min, placed in a 28°C incubator for 150rpm / min shaking fermentation overnight; centrifuge the fermented liquid at 6000rpm / min for 10min, take the supernatant, and use a 0.22μm filter Sterilize to obtain seed filtrate, take 100 μL of the filtrate and mix it with 300 μL of Vibrio alginolyticus (concentration is about 5×108 CFU / mL), and after static adsorption for 15min, mix the mixture with 2% sodium chloride TSB semi-solid (containing 0.65% agar) ) mixed evenly, poured on a 2% sodium chloride TSA bottom plate, and after solidification, t...

Embodiment 2

[0028] Experiment on the lysis range of Vibrio alginolyticus by phage GVA-P21

[0029] Phage lysis profiles were determined using a double-layer drop method. Prepare enough EP tubes containing 3 mL of 2% sodium chloride TSB, inoculate 355 strains of Vibrio alginolyticus in the medium, and shake at a constant temperature of 200 rpm / min at 37°C for 7 hours to obtain 355 copies of Vibrio alginolyticus bacteria solution ; Prepare a sufficient amount of TSA bottom plate, mix 0.9ml of Vibrio alginolyticus with 6ml of TSB semi-solid medium (containing 0.65% agar) at about 50°C, and slowly spread it on the TSA bottom plate; wait for the plate medium After solidification to room temperature, take 7 μL of Vibrio alginolyticus bacteriophage GVA-P21 solution (titer 1×109PFU / ml) and drop it in the center of the double-layer plate. After the solution is naturally air-dried, it is placed in a constant temperature biochemical incubator at 37°C for about 7h. ,Observation results. Each group ...

Embodiment 3

[0040] Determination of the titer of Vibrio alginolyticus bacteriophage GVA-P21 at different multiplicity of infection and different infection time

[0041] Prepare the host strain of Vibrio alginolyticus phage GVA-P21, Vibrio alginolyticus GVA1, prepare an EP tube filled with 3 mL of 2% sodium chloride TSB, pick a single colony of Vibrio alginolyticus VAJS1 and inoculate it in the medium at 37°C Under the condition of shaking culture for 8h, the host bacteria liquid was obtained. 1% of the inoculum was inoculated into 100 mL of TSB medium, and when Vibrio alginolyticus grew to the initial logarithmic phase (concentration 1×10 CFU / mL), the Vibrio alginolyticus phage GVA-P21 was then inoculated into the culture In the base, according to different multiplicity of infection (MOI; MOI=number of phages / number of bacteria), each group of experiments was performed in parallel. Incubate with shaking at 150 rpm in a shaker at 28°C for 12 h. A part of the culture medium was taken out ...

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Abstract

The invention provides a vibrio alginolyticus bacteriophage GVA-P21 with high fermentation rate and lasting bacteriostasis and application thereof, the preservation number of the vibrio alginolyticus bacteriophage is CCTCC NO: M20211480, and the vibrio alginolyticus bacteriophage is named as GVA-P21, the vibrio alginolyticus bacteriophage GVA-P21 is high in fermentation efficiency, when MOI is 0.01, the titer reaches 8.24 * 10 < 10 > PFU/mL after 6 h of culture, the cracking rate of 355 strains of pathogenic vibrio alginolyticus is as high as 95.5%, and the vibrio alginolyticus bacteriophage GVA-P21 can be used for preparing the vibrio The vibrio alginolyticus bacteriophage GVA-P21 has a long-time antibacterial effect on vibrio alginolyticus, and is more obvious than that of other bacteriophages on the vibrio alginolyticus, so that the vibrio alginolyticus bacteriophage GVA-P21 has good prevention and treatment effects on diseases caused by the vibrio alginolyticus in aquaculture.

Description

technical field [0001] The invention belongs to the application field of microorganism technology, and particularly relates to a high fermentation rate and lasting bacteriostasis vibrio phage GVA-P21 and its application. Background technique [0002] With the rapid development of the marine aquaculture industry and the continuous expansion of the scale of intensive aquaculture, the incidence of bacterial diseases in the marine aquaculture industry is getting higher and higher. Among the pathogenic bacteria that cause the infection of aquaculture animals, the pathogenic Vibrio species occupies a relatively high proportion, which not only brings high risks to the health of aquatic animals, but also causes serious economic losses to the marine aquaculture industry. [0003] The common pathogenic Vibrio species of aquatic animals mainly include Vibrio eel, Vibrio harvey, Vibrio parahaemolyticus and Vibrio alginolyticus, among which, Vibrio alginolyticus ranks first in the number...

Claims

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Application Information

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IPC IPC(8): C12N7/00A61K35/76A61P31/04C12R1/92
CPCC12N7/00A61K35/76A61P31/04C12N2795/10121C12N2795/10132
Inventor 王喜亮黄金梅田甲张秀玲徐岳张晓东李越胡雄
Owner 武汉格瑞农生物科技有限公司
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