Bacillus subtilis and application thereof

A technology of Bacillus subtilis and microbial strains, applied in the field of Bacillus subtilis, can solve problems such as antibiotic pollution and increased drug resistance, and achieve strong protease and amylase production, good tolerance, and strong cell adhesion Effect

Active Publication Date: 2020-05-22
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, with the long-term use of antibiotics, the increase in the resistance of pathogenic bacteria, the residues of antibiotics in livestock and poultry products, and the pollution of the environment caused by excessive excretion have brought new challenges to human health.

Method used

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  • Bacillus subtilis and application thereof
  • Bacillus subtilis and application thereof
  • Bacillus subtilis and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Isolation and identification of embodiment one Bacillus subtilis

[0024] 1.1 Isolation of strains

[0025] The Bacillus subtilis BSC16a (Bacillus subtilis BSC16a) of the present invention is isolated and screened from fresh feces of healthy pigs in Sichuan Province, with a total of 86 samples. The specific separation and purification method is as follows: weigh 10 g of stool sample, dilute and mix with 90 mL sterile PBS, filter, take 1 mL of the filtrate, centrifuge, discard the supernatant; Cultivate for 48 hours, heat-treat at 80°C for 10 minutes, centrifuge, and discard the supernatant; add 1 mL of nutrient broth culture solution again, vortex, and shake at 37°C for 24 hours; the mixture is properly diluted and spread on a nutrient agar (NA) plate for 37 Cultivate at ℃ for 10h.

[0026] The aseptic PBS formula of the above-mentioned use of the present invention is as follows:

[0027] Weigh NaCl 8.0g, KCl 0.2g, NaCl 2 HPO 4 1.42 g, KH 2 PO 4 Add 0.27 g into 8...

Embodiment 2

[0036] The screening of embodiment Bacillus subtilis

[0037] 2.1 Screening of tolerant Bacillus subtilis

[0038] Take 1 mL of the fresh culture solution of Bacillus subtilis and inoculate it into sterile PBS with pH = 2.5 and containing 0.3% porcine bile salt. After culturing for 3 hours at 37°C, take samples and serially dilute them to 10 -4 、10 -5 , draw 0.1mL of the diluted solution and spread it on the NA plate, count and measure the number of viable bacteria after incubating at 37°C for 12 hours, and calculate the survival rate with the number of viable bacteria at 0h as the control.

[0039] The results of the tolerance test showed that only 18% of the tested Bacillus subtilis were extremely sensitive to acids and bile salts, and the survival rate was lower than 30%; the rest could tolerate both pH=2.5 and In the environment of 0.3% bile salts, the survival rate is higher than 80%.

[0040] 2.2 Screening of antibacterial Bacillus subtilis

[0041] Centrifuge the ac...

Embodiment 3

[0055] The biological characteristic research of embodiment Bacillus subtilis BSC16a

[0056] 3.1 Activation of Bacillus subtilis BSC16a and preparation of its seed solution

[0057] The frozen Bacillus subtilis BSC16a was taken out from the -20°C refrigerator, melted rapidly in a 37°C water bath, and streaked on the NA plate. After culturing at 37°C for 10 hours, a single colony was picked and inoculated in 10 mL of nutrient broth medium. Cultivate with shaking at 220rpm / min for 18h. After two consecutive subcultures, the seed solution was prepared and set aside.

[0058] 3.2 Determination of the growth curve of Bacillus subtilis BSC16a

[0059] Take the seed solution of Bacillus subtilis BSC16a in 3.1, inoculate it in a nutrient broth medium with a 2% inoculum amount, and culture it with shaking at 37°C for 24 hours. Taking the uninoculated liquid medium as the control, samples were taken every 2 hours after inoculation to measure the OD of the bacterial solution 600 val...

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Abstract

The invention discloses a Bacillus subtilis BSC16a and application thereof. The Bacillus subtilis is preserved in China General Microbiological Culture Collection Center on September 6, 2019, and thepreservation number is CGMCC NO: 18473. The strain has a good in-vitro probiotic property. The strain disclosed by the invention plays a good probiotic role in the aspects of improving the growth performance and body immunity of growing and fattening pigs and reducing enteropathogenic bacteria. In addition, the Bacillus subtilis BSC16a disclosed by the invention is used as a probiotic preparationfor feeding diarrhea young animals. The strain has a good prevention and treatment effect on diarrhea animals. The strain can also be used as a feed additive for preventing and treating diarrhea of piglets, lambs and calves.

Description

technical field [0001] The invention relates to a strain of Bacillus subtilis and its application. Background technique [0002] Since the discovery of antibiotics, they have brought great benefits to human medical treatment and animal husbandry. However, with the long-term use of antibiotics, the increase in the resistance of pathogenic bacteria, the residue of antibiotics in livestock and poultry products, and the environmental pollution caused by excessive excretion have brought new challenges to human health. Subsequently, countries successively promulgated bans to restrict and prohibit the application of antibiotics in livestock breeding. In 1986, Sweden became the first country to ban antibiotics in livestock and poultry feed, and then the European Union, South Korea, and the United States successively proposed to ban antibiotics from being used as feed additives. The Ministry of Agriculture and Rural Affairs of my country announced in 2019 that all pharmaceutical fe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20A61K35/742A61P1/12A23K50/30A23K50/10A23K50/60A23K10/18C12R1/125
CPCA61K35/742A23K10/18A23K50/10A23K50/30A23K50/60A61P1/12C12N1/205C12R2001/125
Inventor 李有全贾丹王锦明关贵全孟林明李贺海王佳慧刘军龙刘爱红殷宏罗建勋刘志杰白小明独军政高闪电郝建刚刘光远任巧云杨吉飞李东平
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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