Bacteriophage and application of same to soil remediation

一种噬菌体、土壤的技术，应用在噬菌体领域，能够解决微生物群落功能稳定性和多样性生态风险评估没有受到重视、微生物群落功能稳定性和多样性的相关生态风险评估匮乏、缺少抗生素抗性致病细菌同步灭活等问题，达到改良土壤理化性质、抗逆能力强、孔隙度发达的效果

Active Publication Date: 2019-01-29
INST OF SOIL SCI CHINESE ACAD OF SCI
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AI Technical Summary

Problems solved by technology

[0006] These methods will all point out that both biochar and phage can block or even inactivate pathogenic bacteria, and can also simultaneously prevent crop diseases, but generally lack the overall restoration effect on soil-crop (vegetable) system synergistic removal of ARB and ARGs
In addition, the existing patents have never involved the introduction of the combination of biochar and phages to inactivate pathogenic bacteria carrying antibiotic resistance genes in soil-vegetable systems
[0007] The main defects in the existing technology are: most of the existing researches on biochar patents are used as adsorption materials to remove organic, non-polar, heavy metals and other pollutants in polluted soil, or are used in improving soil properties, adjusting soil fertility, improving crop quality and Yield, but there are few studies on broad-spectrum adsorptive removal of antibiotic-resistant pathogenic bacteria in soil. At the same time, the ecological risk assessment related to the functional stability and diversity of microbial communities in soil-vegetable systems after inoculation is relatively scarce; and now Most of the phage therapy applications are phages with strong selectivity and high specificity as the initial material, and there is a lack of biological control technology for synchronous inactivation of multiple antibiotic-resistant pathogenic bacteria in the soil-vegetable system. Ecological risk assessment associated with functional stability and diversity of microbial communities in vegetable systems has received little attention
[0008] The main reasons for the defects are as follows: In recent years, the academic community has gradually recognized that the soil-vegetable system is the "source" and "sink" for the accumulation and occurrence of various antibiotic-resistant bacteria and resistance genes, and this type of new resistance causes Bacteria and genes will seriously threaten human health and ecological environment safety through the transmission of food chains; and most of the existing restoration research is on the fire-fighting and restoration of E. Bacteria research, lack of attention to bacteria with high risk of pathogenicity in soil-vegetables (such as: Klebsiella pneumoniae, Pseudomonas aeruginosa, etc.)

Method used

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  • Bacteriophage and application of same to soil remediation
  • Bacteriophage and application of same to soil remediation
  • Bacteriophage and application of same to soil remediation

Examples

Experimental program
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Effect test

Embodiment 1

[0034] 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 basic physical and chemical properties of the soil: sand 23.8%, soil 45.4%, clay 31.8%, pH 7.7, total nitrogen 1.7 g·kg -1 , water-soluble nitrogen 1.7 g·kg -1 , total phosphorus 1.3 g·kg -1 , total potassium 17.5 g·kg -1 , CEC 19.4 cmol kg -1 .

[0035] Take 5 g of fresh soil samples, add 50 mL of sterile water, culture at 28 °C, 150 rpm for 5 h, centrifuge at 10 000 rpm for 5 min, filter the supernatant through a 0.22 μm membrane, take 9 mL of the filtrate and 1 mL of the growth medium Pseudomonas aeruginosa PAO1 suspension in the logarithmic phase was added to 40 mL LB liquid medium, and calcium chloride solid was added to the final concentration of the solution to 1 mmol L -1 , 30°C, 150 rpm shaker culture for 12 h, the resulting culture solution was centrifuged at 10 000 rpm for 5 min, and the...

Embodiment 2

[0038] 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. Growing Vegetables for Carrots Seoul Six Inches ( Daucus L .), Beijing Zhongnong Tianteng Vegetable Seed Company. Basic physical and chemical properties of soil: 23.8% sand, 45.4% soil, 31.8% clay, pH 7.7, total nitrogen 1.7 g·kg -1 , water-soluble nitrogen 1.7 g·kg -1 , total phosphorus 1.3 g·kg -1 , total potassium 17.5 g·kg -1 , CEC 19.4 cmol kg -1 .

[0039] Four groups of treatments were set up in the experiment: ① Control group (CK): 3 carrots were planted in each pot (cover soil 0.5-1 cm above the seeds, room temperature 20±2°C); ② Biochar treatment (B): On the basis of the control group Apply biochar (1 g / kg); ③ Phage φYSZPK treatment (P): Inoculate 100 mL on the basis of the control group with a concentration of 10 6 pfu mL -1 phage φYSZPK; ④ Combined treatment of biochar and phage φYSZPK...

Embodiment 3

[0042] 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. Plant vegetables as Hong Pin No. 1 Chaotian Pepper ( Capsicum frutescens var ), Qianshu Baihua Seed Industry. Basic physical and chemical properties of soil: 23.8% sand, 45.4% soil, 31.8% clay, pH 7.7, total nitrogen 1.7 g·kg -1 , water-soluble nitrogen 1.7 g·kg -1 , total phosphorus 1.3 g·kg -1 , total potassium 17.5 g·kg -1 , CEC 19.4 cmol kg -1 .

[0043] Four groups of treatments were set up in the experiment: ① Control group (CK): Plant 3 chili peppers in each pot (cover soil 0.5-1 cm above the seeds, room temperature 25±2°C); ② biochar treatment (B): On the basis of the control group, biochar (1 g / kg) was applied; ③ Phage φYSZPK treatment (P): On the basis of the control group, 100 mL was inoculated with a concentration of 10 6 pfu mL -1 phage φYSZPK; ④ Combined treatment of biochar and phag...

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Abstract

The invention provides a bacteriophage and application of the same to soil remediation. The bacteriophage Phi YSZPK is preserved in China Center for Type Culture Collection on August 01, 2018, under the accession number of CCTCC M 2018516, and is classified and named as Pseudomonas aeruginosa and Klebsiella phage Phi YSZPK. Biomass charcoal and the screened bacteriophage are used in combination and returned back to contaminated soil to synergistically prevent and control and deeply track the propagation and spreading of resistant pathogenic bacteria and resistant genes in an inactivated soil-vegetable system; the combined use of the biomass charcoal and the bacteriophage has positive promotion effect on the functional stability of microbial communities in the soil-vegetable system, effectively alleviates the spreading of resistant pathogenic bacteria in the soil-vegetable system and prevents secondary pollution; so a novel solution is provided for bioremediation and treatment of farmland soil contaminated by antibiotic resistant pathogenic bacteria in China.

Description

technical field [0001] The invention belongs to the technical field of remediation of multi-antibiotic-resistant pathogenic bacteria and their related resistant genes, and particularly relates to a strain of bacteriophage φYSZPK and its use in resistance-controlling inactivation of antibiotic-resistant pathogenic bacteria in soil-vegetable systems. application. Background technique [0002] In recent years, due to the abuse of antibiotic veterinary drugs, the lack of safe treatment technology for livestock and poultry manure, and the lack of environmental management, the farmland soil-vegetable system around the suburban animal husbandry farms in China and many countries around the world has often become a source of residual and breeding antibiotics. High-risk hotspot "sources" and "sinks" of Antibiotic Resistance Bacteria (ARB) and Antibiotic Resistance Genes (ARGs), especially mobile genetic elements (plasmids, integrons, transgenic Under the promotion of horizontal tra...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00B09C1/10B09C1/08C12R1/92
CPCB09C1/08B09C1/10C12N7/00C12N2795/00021C12N2795/00032C12N2795/00011
Inventor 叶茂赵远超孙明明张忠云黄丹卞永荣胡锋蒋新
Owner INST OF SOIL SCI CHINESE ACAD OF SCI
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