Bdellovibrio preparation for preventing frog wryneck diseases and application thereof

A technology for phagocytosis of Bdellovibrio and Bdellovibrio, which is applied in the field of freshwater aquaculture, can solve the problems of introducing pathogenic bacteria, lack of function of Bdellovibrio, and potential safety hazards, and achieve safe and efficient use, conducive to growth and reproduction, and low cost of use Effect

Pending Publication Date: 2019-08-27
XIAMEN HUIYING ANIMAL TECH +1
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Through research on Bdellovibrio-related patent documents, it was found that most bacteriophage Bdellovibrio preparations are generally cultured with live host bacteria Escherichia coli, which has several defects. On the one hand, it is necessary to separate unlysed E. coli after the cultivation, which is cumbersome to operate; on the one hand If the live host bacteria are not cleanly separated, there will be safety hazards, which may damage the environment and introduce pathogenic bacteria, which is not conducive to the large-scale use of Bdellovibrio preparations; Decreased or lost ability to infect other pathogenic bacteria

Method used

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  • Bdellovibrio preparation for preventing frog wryneck diseases and application thereof

Examples

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Effect test

Embodiment 1

[0038] Embodiment 1 effective phage Bdellovibrio isolation test

[0039] This example is used to illustrate the source of Bdellovibrio and its acquisition method of the present invention.

[0040] Sewage samples were obtained from the drainage ditch of a bullfrog farm, centrifuged at 3000r / min for 5min, the supernatant was taken, and 100mL was divided into 250mL triangular flasks, and 4 small bottles were added with a bacterial concentration of 10 8 CFU / mL Flavobacterium meningitidis, Elisabeth mil, Pseudomonas aeruginosa, Aeromonas hydrophila enriched culture, 30 ℃, 100r / min aerobic culture for 48h. After the enrichment culture, the tap water agar double-layer plate was used for separation and detection, and Flavobacterium meningitidis, Elisabeth mil, Pseudomonas aeruginosa, and Aeromonas hydrophila were used as host bacteria for testing, and the plates were cultured at 30°C for 2 days , multiple transparent plaques appeared. Pick multiple upper layer of phage plaque cultur...

Embodiment 2

[0041] Embodiment 2 preparation of phage Bdellovibrio bacterial liquid

[0042] This example is used to illustrate the bacteriophage Bdellovibrio liquid provided by the present invention and its preparation method.

[0043] The Bdellovibrio phage described in this example is provided by Example 1.

[0044] 1. Preparation of inactivated host bacteria concentrate: Inoculate Flavobacterium meningitidis, Elizabeth mil, Pseudomonas aeruginosa, Aeromonas hydrophila respectively in 10g / L peptone, 3g / L beef extract, 5g / L L sodium chloride, pH7.0-7.5 culture medium, after aerobic culture at 30°C for 24 hours, collect the sludge by centrifugation, mix and place in a water bath at 60°C for 30 minutes, and cool for later use.

[0045] 2. Preparation of bacteriophage Bdellovibrio bacteria liquid: Take the uninactivated mixed host bacteria in the above step 1 and dilute them with sterilized physiological saline to make the bacterial concentration at 10 9 CFU / mL was inoculated with Bdellov...

Embodiment 3

[0046] Preparation of Example 3 Bdellovibrio Phage Preparation 1

[0047] This example is used to illustrate a phage Bdellovibrio preparation and its preparation method for preventing bullfrog tilted head disease provided by the present invention.

[0048] The phage Bdellovibrio bacterial liquid obtained in Example 2 was inserted into the inactivated host bacterium, and the bacterial concentration was about 10 6 CFU / mL, after aerobic culture for 2 hours, add 2% (V / V) glycerol, stir evenly, and then mix with freeze-dried carrier (sucrose: skimmed milk powder = 6%: 94%) at a liquid-solid mass ratio of 1:0.5 Proportionally mixed to obtain a mixture to be freeze-dried.

[0049] The above-mentioned mixed solution to be freeze-dried was divided into stainless steel shallow pans, placed in a vacuum freeze dryer at -40°C, pre-frozen for 3 hours, and then started to vacuumize. First raise the temperature to 0°C within 24 hours, then gradually raise the temperature to 10°C in the next...

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Abstract

The invention discloses a bdellovibrio preparation for preventing frog wryneck diseases. Pathogenic bacteria capable of causing the frog wryneck diseases are taken as various mixed living host bacteria to culture bdellovibrio; then the uncracked host pathogenic bacteria are removed by filtration to obtain a bdellovibrio seed solution; then co-culture is carried out through various mixed inactivated host bacteria; the obtained bdellovibrio seed solution is firstly added with a freeze-drying protecting agent to be uniformly mixed, and then is added with a freeze-drying carrier to be mixed to obtain a preparation to be freeze-dried; finally, the bdellovibrio preparation is prepared by freeze-drying. The invention further discloses a preparation method and application of the bdellovibrio preparation. The bdellovibrio preparation can effectively prevent the frog wryneck diseases, obviously reduces the incidence rate, and has the advantages of environment-friendly effects and safe and efficient use.

Description

technical field [0001] The invention relates to the technical field of freshwater aquaculture, in particular to a Bdellovibrio bacteriophage preparation for preventing head tilt of frogs and its application, which is mainly used in the breeding process of frogs to prevent head tilt. Background technique [0002] Frogs belong to the order Anura. The body of this species is short and wide, and the limbs are relatively long. The larva has a tail and the adult has no tail. There are about 2,500 species of each, and there are about 130 species of frogs in China. The more common species in breeding are bullfrogs, frogs, and black-spotted frogs. Frogs generally have the advantages of fast growth, delicious taste, rich nutrition, and high protein content, and are valuable foods. Its skin can also be used to make high-grade leather, the internal organs of frogs can be used for medicine, and frog oil can be used to make high-grade lubricating oil. Therefore, it has high economic va...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K35/74A61K9/19A61K47/46A61P31/04A61P25/00C12N1/20C12N1/04C12R1/01
CPCA61K9/19A61K35/74A61K47/46A61P25/00A61P31/04C12N1/04C12N1/20
Inventor 邹文政林章秀林松泉乔欣君李惠静
Owner XIAMEN HUIYING ANIMAL TECH
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