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Attenuated eel vibrio strain and its application

A technology of vibrio eel and strains, applied in the field of attenuated strains of vibrio eel, to achieve high-efficiency immune protection, significant immune effect, and good control effect

Inactive Publication Date: 2005-07-13
EAST CHINA UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In view of this, this type of attenuated vaccine has been identified as a biological product with high environmental safety risks by the approval regulations of biological product safety inspection and management agencies in various countries, including China, and it is difficult to enter the commercial development process

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Preparation of Live Attenuated Vaccine of Vibrio anguillarum

[0023] Composition of culture medium and seawater saline:

[0024] 1) LB slant medium: soybean peptone (Difco) 10g / L, yeast extract (Merck) 5g / L, NaCl 25g / L, agar 18g / L, pH 7.5;

[0025] 2) Seed medium: soybean peptone (Difco) 10g / L, yeast extract (Merck) 5g / L, NaCl 25g / L, phenylalanine 20mg / L, tyrosine 20mg / L, tryptophan 20mg / L , p-hydroxybenzoic acid 20mg / L, p-aminobenzoic acid 20mg / L, pH 7.5;

[0026] 3) Fermentation medium: soybean peptone (Difco) 10g / L, yeast extract (Merck) 5g / L, NaCl 25g / L, ferric ammonium citrate 0.2-0.5mmol, phenylalanine 20mg / L, tyrosine 20mg / L, tryptophan 20mg / L, p-hydroxybenzoic acid 20mg / L, p-aminobenzoic acid 20mg / L, pH6.8;

[0027] 4) Seawater saline: NaCl 20, KCl 0.7, MgCl 2 ·6H 2 O 4.8, NaHCO 3 0.11, MgSO 4 ·7H 2 O 3.5, CaCl 2 2H 2 O 1.6, g / L, pH7.2, filter sterilized.

[0028] Vaccine preparation:

[0029] Take the seeds of the attenuated vaccine strain (Vibri...

Embodiment 2

[0031] The median lethal dose LD of flounder as experimental animal 50 Determination:

[0032] The fish used in the test were first placed in the SPF (Specific Pathogen Free) laboratory to adapt to breeding for 1 week to eliminate abnormal individuals. Before the infection test, the SPF test fish were stocked in the 20L infection test tank in the infection laboratory (ChallengeLab), and continued to be fed for 1 week, with 20 fish (average body length 12-15cm) in each tank. The test tank replaced 2 / 3 volume of aquaculture water with sterile old sea water every day, and the water temperature was 20°C, with a fluctuation of 2°C.

[0033] The fish used in the test were randomly divided into groups, and two tanks were tested in parallel in each group. In the infection test, each group of test fish was dosed with a certain concentration gradient (10 2 ~10 8 CFU / tail) wild strain and attenuated vaccine strain were artificially infected by intramuscular injection. Record the num...

Embodiment 3

[0043] Injection-administered immune protection test with flounder as experimental animal:

[0044] The experimental flounder was randomly divided into 12 groups, each group had 3 parallel water tanks, 10 fish / tank. The prepared attenuated live vaccine was immunized by intramuscular injection. The immunization dose is 10 6 CFU / tail and 10 8 CFU / tail, intramuscular injection of flounder. The control group was injected with sterile normal saline. After 4 weeks and 8 weeks, each group was immunized with live bacteria of Vibrio anguillaris wild strain (intramuscular injection for 10 days). 8 CFU / tail) for artificial infection challenge. Observe and count the number of deaths in the control group and the immune group within 15 days, and calculate the immune protection rate of each group (see Table 2, Table 3).

[0045] Calculate the immune protection rate according to the following formula:

[0046] Immune protection rate %=(control group mortality rate-immune group mortalit...

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Abstract

The present invention discloses two attenuated gene null deficiency strains, Vibrio anguillarum MVAV6203 (delta aroC) and Vibrio anguillarum MVAV6204 (delta aroC delta angE), obtained from eel vibrio wild strain Vibrio anguillarum MVM425, and their application in vaccine for preventing and treating vibriosis of raised fishes. The strains of the present invention contains no foreign gene segment and antibiotic resistance label, and has large segment deletion of toxicity and toxicity relevant gene frame, irrecoverable toxicity and technological safety. Experiment shows that the attenuated live vaccine prepared with the strain of the present invention has excellent effect of preventing and treating vibriosis of raised fishes.

Description

technical field [0001] The invention relates to an attenuated strain of Vibrio anguillarum and its application. Background technique [0002] Aquaculture is becoming increasingly important in view of the growing world population and the depletion of natural fisheries. Since the 1990s, the world's mariculture production has increased at a rate of 5%. In 2001, the global mariculture production reached 15.1 million tons. It is estimated that by 2010, my country's marine fish farming output will reach 500,000 tons in order to meet people's demand for marine fish products. Due to the limited water and soil resources, it is difficult to continue to develop by increasing the production area by increasing the breeding area. Therefore, large-scale, intensive, and high-density farming models have gradually become the mainstream of the development of my country's marine fish farming industry. However, with the continuous and steady development of the marine aquaculture industry, var...

Claims

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Application Information

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IPC IPC(8): A61K39/106C12N1/21
Inventor 马悦赵东玲张元兴
Owner EAST CHINA UNIV OF SCI & TECH
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