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A kind of Bacillus subtilis 7k and its application

A technology of Bacillus subtilis and microbial strains, applied in the fields of application, bacteria, antibacterial drugs, etc., can solve the problem that it is difficult to distinguish virus infection and bacterial infection in time, lack of effective weapons for virus infection, and it is difficult to reduce farmers' concerns about antibiotics and illegal drugs. Substance Abuse and Other Issues

Active Publication Date: 2020-07-28
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Similar to the abuse of antibiotics in the human population, people in the aquatic industry also lack effective weapons to deal with viral infections, and it is difficult to distinguish viral infections from bacterial infections in the actual breeding process, which has also become an important reason for the abuse of antibiotics
Therefore, if the outbreak of viral-derived diseases in aquatic animals cannot be controlled, it will be difficult to reduce the abuse of antibiotics and prohibited drugs by farmers

Method used

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  • A kind of Bacillus subtilis 7k and its application
  • A kind of Bacillus subtilis 7k and its application
  • A kind of Bacillus subtilis 7k and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0034] Isolation and identification of Bacillus subtilis 7K

[0035] 1) Sampling: The cultured grouper was purchased from a farm in Hainan; the marine sediment was sourced from the mangrove forests along the coast of Guangdong, the South China Sea, and the deep sea of ​​the Indian Ocean.

[0036] 2) Medium:

[0037] Liquid medium for enrichment (LB medium): peptone 10g, yeast powder 5g, NaCl 10g, distilled water 1000ml, pH 7.0-7.2.

[0038] Separation medium: 1.5%-2% agar was added to LB medium.

[0039] Salt tolerance test medium: change the amount of NaCl added in the LB medium to make medium with different salinity (NaCl 2%, 5%, 7%, 10%).

[0040] Sugar fermentation test medium: (NH 4 ) 2 HPO 4 1g, MgSO 4 ·7H 2 O 0.2g, KCl 0.2g, yeast extract 0.2g, sugar or alcohol 10g, distilled water 1000mL, 0.2% bromothymol blue solution 1.2ml, adjust pH 7.0-7.2, divide into test tubes, each tube contains 10mL medium, Sterilize at 115°C for 20 minutes.

[0041] Gelatin medium: ...

experiment example 1

[0061] Experimental example 1 Bacillus subtilis (Bacillus subtilis) 7K characteristic experiment

[0062] (1) Preparation of spores: Pick 7K colonies of single clone Bacillus subtilis and inoculate them in 5ml LB liquid medium, culture on a shaking table at 37°C overnight, then dilute 1:100, add to DSM medium, Cultivate on a shaker at 250 r / min at 37°C. At 0h, 12h, and 24h of spore culture, 100 μL of culture solution smears were taken for spore staining and microscopic examination to observe the formation of spores. Spore collection and washing methods: centrifuge at 8000r / min for 15min to collect the bacteria, remove the supernatant, add lysozyme to a final concentration of 4mg / mL, and let stand at room temperature for 15min. Wash the spores with 1mol / L NaCl, add PMSF to a final solubility of 1μmol / L, mix thoroughly, and centrifuge at 8000r / min for 15min to collect the bacteria. Then wash once with 1mol / L KCl, and wash twice with distilled water. Dissolve the spores in a c...

experiment example 2

[0073] Experimental example 2 The probiotic effect of Bacillus subtilis (Bacillus subtilis) 7K on grouper

[0074] 1) Preparation of Bacillus subtilis (Bacillus subtilis) 7K added feed

[0075] Bacillus subtilis (Bacillus subtilis) 7K isolated from the intestinal tract of the pearl gentian grouper was inoculated in LB liquid medium, cultured at 37°C for 24 hours, and then the culture solution was mixed into the spore-forming medium at a ratio of 1:100 ( Soybean powder 10g / l, yeast powder 2.5g / L, glucose 10g / L, (NH 4 ) 2 SO 4 2.5g / L, inorganic salts and trace elements) were cultured at 37°C for 48h in a 5L automatic fermenter. Then the fermentation broth was centrifuged at 8000rpm for 15min and the supernatant was discarded. In order to remove residual vegetative bodies, all spores were transferred to lysozyme solution (4mg / ml) and treated at room temperature for 15min. The treated spores were washed with sterile water and centrifuged twice (8000rpm, 15min). The washed sp...

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Abstract

The invention discloses a Bacillus subtilis 7K and application thereof. Bacillus subtilis (Bacillus subtilis) 7K of the present invention is preserved in the Guangdong Microbial Culture Collection Center, the preservation number is GDMCC No: 60226, and the preservation address is 5th Floor, Building 59, Compound, No. 100 Xianlie Middle Road, Guangzhou City, and the preservation date is 2017 August 31st. Bacillus subtilis (Bacillus subtilis) 7K of the present invention has the characteristics of extreme environments such as resistance to 100°C high temperature, low pH resistance, bile salt resistance, and gastrointestinal digestive enzyme resistance, and it is confirmed by experiments that the Bacillus subtilis obtained by screening in the present invention ( Bacillus subtilis) 7K new strain can promote the growth of aquatic animals and regulate the expression of various immune genes, and can also inhibit the growth of various aquatic animal pathogenic bacteria, thereby improving the ability of aquatic animals to resist pathogenic infection, and has a good application prospect and market value.

Description

technical field [0001] The invention belongs to the technical field of fisheries and microbiology, more specifically, the invention relates to the isolation and identification of a new bacterial strain of Bacillus subtilis (Bacillus subtilis) 7K with the effect of disease-resistant probiotics, which can promote the growth of aquatic animals and regulate The expression of various immune genes can also inhibit the growth of various aquatic animal pathogenic bacteria, thereby improving the ability of aquatic animals to resist pathogenic infection. Background technique [0002] According to the UN Food and Agriculture Organization's survey on the status of the global aquatic industry, in 2014 the global fishery produced a total of 167,200,000 tons of fish and generated a trade value of nearly 148 billion US dollars. This also means that people's consumption of aquatic products has reached an unprecedented high value (Fisheries F A O. The state of world fisheries and aquaculture2...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20A23K10/18A23K50/80C02F3/34A61K35/742A61P31/04C12R1/125
CPCA61K2035/115A23K10/18A23K50/80C02F3/34C12N1/205C12R2001/125
Inventor 秦启伟周胜黄晓红黄友华魏京广
Owner SOUTH CHINA AGRI UNIV
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