Attenuated live vaccine of eel vibrio

An attenuated live vaccine, Vibrio anguillarum technology, applied in the direction of bacteria, antibacterial drugs, bacterial antigen components, etc., can solve the problems of low level, unstable immune response, inconvenient administration, etc., and achieve high immune protection rate , good vibriosis prevention and control effect, excellent immune effect

Inactive Publication Date: 2005-03-30
EAST CHINA UNIV OF SCI & TECH
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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.

Method used

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  • Attenuated live vaccine of eel vibrio
  • Attenuated live vaccine of eel vibrio
  • Attenuated live vaccine of eel vibrio

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Embodiment 1: the preparation of vibrio anguillarum live attenuated vaccine

[0028] Take 1 inoculation loop and save the seeds on LB slant medium (soybean peptone (Difco) 10g / L, yeast extract (Merck) 5g / L, NaCl 30g / L, agar 18g / L, pH7.5), inoculate in A test tube containing 1 ml of liquid LB seed medium (soy peptone (Difco) 10 g / L, yeast extract (Merck) 5 g / L, NaCl 30 g / L, pH 7.5) was incubated at 28°C. After 12 hours, take 0.01ml of vigorously growing bacterial solution (O.D=4.0 or so) and inoculate 1ml of fresh iron-rich LB medium (soybean peptone (Difco) 10g / L, yeast extract (Merck) 5g / L, NaCl 30g / L L, FeCl 3 0.2-0.5mmol, pH6.8), cultured at 37°C for 16 hours. Take 0.01ml bacterial liquid, transfer to 1ml fresh iron-rich LB medium (containing 0.2mM Fe 3+ ), and cultured at 28°C for 12 hours. As mentioned above, after repeating 2 to 3 cycles, the obtained bacterial liquid is the bacterial liquid to be screened containing candidate strains.

[0029] Repeat the ab...

Embodiment 2

[0038] Example 2: Infection and immunization with flounder as object

[0039] Infectious experiment:

[0040] Vibrio anguillarum wild strain V.anguillarumMVM425 and pEIB1 plasmid deletion strain V.anguillarumMVAV6201 were cultured in iron-containing LB medium (the aforementioned LB seed medium+0.5mM Fe 3+ ) in a shaker (200r / min) at 28°C for 24h. The bacterial cells were collected by centrifugation (10,000 r / min, 10 min), washed with sterilized physiological saline (pH 7.4) for 3 times, and prepared into a bacterial suspension for later use.

[0041] The fish used in the experiment was the flounder (Paralichthysolivaceus Temminck et Schlegel) purchased from the Experimental Base of Damai Island of the Yellow Sea Fisheries Research Institute, which is sensitive to Vibrio anguillarum, with an average body length of 10±1cm and a weight of 50±2.5g.

[0042] The test fish were divided into random groups, 30 fish in each group, and 1 / 3 of the breeding water was changed every day t...

Embodiment 3

[0054] Embodiment 3: Infection and immunity with American redfish as object

[0055] The fish used in the test were American redfish provided by the 863 breeding base in Zhanjiang, Guangdong, with an average weight of 40 g. Those who had no abnormalities in the aquarium for more than one week were used as experimental fish. Inoculate the wild strain MVM425 and the attenuated strain on the aforementioned LB slant medium, culture at 28°C for 18-24 hours, wash with sterile PBS solution (pH7.2) to make a suspension, and adjust the concentration of the bacterial solution by referring to the McFarland turbidimetric tube into MCF3 tubes (9×10 8 / ml), then intramuscularly inject American red fish, 15 tails / group, dosage is 0.4ml / tail, and the control group injects aseptic PBS liquid. After the injection, the American redfish was kept in an aquarium, oxygenated, and the water temperature was controlled at 27.3-32.4°C. Daily observations were made to record the morbidity and mortalit...

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Abstract

An attenuated live vaccine of eel vibrio for preventing the sensitive fish from being infected by wild vibrio and its preparing process are disclosed.

Description

technical field [0001] The invention relates to an attenuated live vaccine, in particular to an attenuated live vaccine of Vibrio 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 a kind of important bacterial pathogen that seriously harms marine fish cultured 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 Mol Biol Rev, 2002, 66(2): 223-49). ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/02A61P31/04C12N1/20
Inventor 马悦张元兴邵明非孙修勤何建国叶巧真黄志坚刘允坤
Owner EAST CHINA UNIV OF SCI & TECH
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