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Bacillus cereus phage composition and application thereof

A technology of Bacillus cereus and bacteriophage, which is applied in the directions of viruses/bacteriophages, applications, and microorganisms, and can solve problems such as food poisoning, human health, and ecological security threats, and achieve the effects of easy storage, low preparation, and environmental friendliness

Active Publication Date: 2022-04-26
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The main defect that existing technology exists is: the spore of bacillus cereus is easy to spread in soil, is attached to vegetables, and eats this kind of improperly handled food and it easily causes food poisoning, endangers human health and ecological environment. Security poses extremely serious potential threats
There are few reports on the removal of antibiotic-resistant Bacillus cereus in soil-vegetable systems. Therefore, there is an urgent need for biologically targeted inactivation to reduce and eliminate the risk of antibiotic-resistant Bacillus cereus accumulation in soil-vegetable systems Technology research and development work

Method used

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  • Bacillus cereus phage composition and application thereof
  • Bacillus cereus phage composition and application thereof
  • Bacillus cereus phage composition and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] 1. Isolation and purification of Bacillus cereus phage

[0029] The soil samples for the test were collected from the polluted soil around the excrement accumulation pool of the Hengliang Dairy Farm in Nanjing City, Jiangsu Province. Basic physical and chemical properties of soil: 23.8% sand, 45.4% soil, 31.8% clay, pH 7.7, total nitrogen 1.7g kg -1 , water-soluble nitrogen 1.7g·kg -1 , total phosphorus 1.3g·kg -1 , total potassium 17.5g·kg -1 , CEC 19.4cmol·kg -1 .

[0030] Take 10g of fresh soil sample, add it into 100mL sterile water, culture at 30℃, 250rpm for 5h, centrifuge at 10 000rpm for 5min, filter the supernatant through a 0.22μm membrane, take 9mL of the filtrate and 1mL of cereus spores grown to the logarithmic phase Add 40mL of 3×LB liquid medium to the bacillus suspension, add solid calcium chloride to a final concentration of 1mmol·L -1 , 30°C, 250rpm shaking table for 12h, the obtained culture solution was centrifuged at 10 000rpm for 5min, and th...

Embodiment 2

[0038] To verify the antibacterial effect of single phage and its composition on Bacillus cereus in aqueous phase.

[0039] Set up 4 groups of experimental treatments. Control group (treatment 1): Take 100 μL logarithmic phase Bacillus cereus (tetW) and add it to 100 mL LB liquid medium, inoculate 100 μL according to the optimal MOI value Inoculate a single phage Bacteriostatic effect on host bacteria (treatment 2): On the basis of the control group, inoculate 100 μL of Shake to mix; inoculate a single phage (Treatment 3): On the basis of the control group, inoculate 100 μL of phage according to the optimal MOI value Shake and mix; inoculate 1 / 2 phage with 1 / 2 phage (Treatment 4): On the basis of the control group, according to the optimal MOI value, inoculate 50 μL and 50 μL After shaking and mixing each group of treatments, shake culture at 37°C and 150rpm, take samples every 2h, and count the bacteria of Bacillus cereus (tetW), and draw the antibacterial eff...

Embodiment 3

[0041] The potting soil for the test was collected from the polluted soil around the manure accumulation pool of the Hengliang Dairy Farm in Nanjing City, Jiangsu Province. The planted crop is potato (Solanum tuberosum), Jiangsu Academy of Agricultural Sciences. Basic physical and chemical properties of soil: 23.8% sand, 45.4% soil, 31.8% clay, pH 7.7, total nitrogen 1.7g kg -1 , water-soluble nitrogen 1.7g·kg -1 , total phosphorus 1.3g·kg -1 , total potassium 17.5g·kg -1 , CEC 19.4cmol kg -1 .

[0042] Four groups of treatments were set up in the experiment: ①Control group (CK): plant 1 potato in each pot, potato tuber 2-4cm, overlying soil 2-3cm, compacted overburden, room temperature 15±2℃; ②bacteriophage Treatment (P1): On the basis of the control group, inoculate 100mL with a concentration of 10 according to the optimal MOI 6 pfu mL -1 Phage ③ Phage Treatment (P2): On the basis of the control group, inoculate 100 mL with a concentration of 10 5 pfu mL -1 Pha...

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Abstract

Bacillus cereus phage composition and its application, including two strains of phage, respectively phage φYSZBA1, the preservation number is: CCTCC M 2018517, and the classification is named Bacillus cereus phage φYSZBA1; bacteriophage φYSZBA2, the preservation number is: CCTCC M 2018518, and the classification is named Bacillus cereus phage φYSZBA2. The present invention uses the mixed "cocktail" therapy to inoculate the phage mixture into the polluted soil-vegetable system, infects and inactivates the resistant Bacillus cereus in the system in a collaborative way to remove the resistance gene. After the restoration, the resistant wax Bacillus-like bacteria and their resistance genes were greatly reduced, and at the same time, the functional diversity and stability of the soil ecological environment could be maintained.

Description

technical field [0001] The invention belongs to the technical field of remediation of soil contaminated by antibiotic-resistant pathogenic bacteria, and in particular relates to a bacillus cereus phage composition and an application thereof. Background technique [0002] In recent years, due to the abuse of medicine and veterinary antibiotics, the lack of safe treatment technology for sewage treatment plants and livestock manure, and the lack of environmental management, the farmland soil-vegetable system in the suburbs of China and many countries around the world has often become residual and breeding High-risk hotspot sources of Antibiotic Resistance Bacteria (ARB) and Antibiotic Resistance Genes (ARGs), especially horizontal transfer of mobile genetic elements (plasmids, integrons, transposons) in a large number of settings Under the promotion of vertical transduction or vertical transduction, the risk of the spread of some antibiotic-resistant pathogenic bacteria will be...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A01N63/40A01P1/00
Inventor 赵远超孙明明胡锋晁会珍郑晓璇
Owner NANJING AGRICULTURAL UNIVERSITY
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