Attenuated live vaccine of vibrion from eel

A live attenuated vaccine, Vibrio eel technology, applied in bacteria, antibacterial drugs, antibody medical ingredients, etc., can solve problems such as low level, inconvenient administration, unstable immune response, etc., and achieve good control effects, The effect of low toxicity and low immune protection rate

Inactive Publication Date: 2006-11-08
上海浩思海洋生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Inactivated vaccines often have the following defects: incomplete protective antigens, unstable and low levels of immune responses, even unable to stimulate correct immune responses and cause side effects, inconvenient administration, etc.
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

[0024] Embodiment 1: the preparation of Vibrio anguillaris attenuated live vaccine and inactivated vaccine

[0025] Preparation of live attenuated vaccine preparation: get 1 inoculation loop and store it in LB slant medium (soybean peptone (Difco) 10g / L, yeast extract (Merck) 5g / L, NaCl 30g / L, agar 18g / L, pH 7.5 ) on MVAV6202 seeds, inoculated in a 500ml shake flask containing 100ml liquid LB seed medium (soybean peptone (Difco) 10g / L, yeast extract (Merck) 5g / L, NaCl 30g / L, pH 7.5), in Shake culture at 28°C (rotating speed 200 rpm). After 12 hours, take 5ml of vigorously growing bacterial solution (O.D=4.0 or so) and inoculate 100ml of fresh iron-rich LB medium (soybean peptone (Difco) 10g / L, yeast extract (Merck) 5g / L, NaCl 30g / L , ferric ammonium citrate 0.2-0.5mmol, pH6.8), cultivated at 28°C for 12 hours. Wash with sterile saline (0.85% NaCl) and dilute to a certain concentration suspension (10 6 -10 8 pcs / ml) for spare use.

[0026] Preparation of inactivated vaccin...

Embodiment 2

[0027] Example 2: Infection test and median lethal dose LD with grouper as object 50 Determination of pathogenicity test:

[0028] The fish used in the experiment was the grouper provided by Guangdong Zhanjiang 863 Breeding Base, with an average weight of 13g, and those who had no abnormality in the aquarium for more than one week were used as experimental fish. The test fish were divided into random groups, 15 fish in each group, and 1 / 3 of the breeding water was changed every day to provide sufficient oxygen, and the temperature of the breeding water was 28°C. No food is fed during the entire process. Abnormal individuals were excluded before the experiment.

[0029] Every tail experimental fish abdominal injection 0.3ml embodiment 1 prepared attenuated live vaccine preparation and wild vibrio anguillae virulence strain MVM425 preparation with medical injection gun, control group injected 0.85% sterile normal saline, recorded the cumulative death number in 10 days and mor...

Embodiment 3

[0037] Embodiment 3: to the immune protection test of grouper:

[0038] Experimental groupers were divided into random groups, 30 fish / group. The grouper was immunized with the attenuated live vaccine preparation prepared in Example 1 by intraperitoneal injection and immersion. In the injection immunization, the concentration of the prepared vaccine preparation was adjusted to 10 8 Individual / ml, 0.3ml / tail, intraperitoneal injection. During the immersion immunization, the concentration of the vaccine preparation soaking solution was adjusted to 10 6 pcs / ml, the soaking time is controlled at about 1 minute. The control group was injected with sterile physiological saline, 0.3ml / tail. After 15 days, a booster immunization was performed by repeating the above-mentioned doses. After another 15 days, each group was challenged with Vibrio anguillarum wild strain MVM425 live bacteria: 9×10 8 / ml physiological saline preparation, 0.3ml / tail, intraperitoneal injection. Then obs...

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PUM

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Abstract

The invention provides an attenuated live vaccine of Vibrio anguillarum. Compared with the wild strain MVM425, the vaccine has obvious low toxicity and immune protection rate, and does not contain any antibiotic resistance markers. Tests have proved that the live attenuated vaccine can effectively protect susceptible fish from the infection of the wild strain of vibrio anguillarum with strong vibriosis pathogenicity.

Description

technical field [0001] The invention relates to an attenuated live vaccine for aquaculture fish, in particular to an attenuated live vaccine of Vibiro anguillarum. Background technique [0002] With the gradual development of the marine aquaculture industry, a large-scale outbreak of various diseases followed, and the disease problem became increasingly prominent, restricting the further development of the marine aquaculture industry. The prevention and control of diseases has become the primary problem that mariculture should solve. [0003] Vibrio eelium is an important bacterial pathogen that seriously harms aquaculture fish in my country and the world. It mainly causes vibriosis such as hemorrhagic septicemia in farmed fish. The main virulence factor contained in Vibrio eelium is An iron uptake system encoded by a pJM1-like plasmid (Crosa, J.H., Walsh, C.T. Genetics and assembly line enzymology of siderophore biosynthesis in bacteria. Microbiol ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K39/104A61P31/04C12N1/21
Inventor 马悦张元兴赵东玲刘琴邵明非何建国黄志坚
Owner 上海浩思海洋生物科技有限公司
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