CN104560836B - One bacillus licheniformis and its application in degraded effective cypermethrin - Google Patents
One bacillus licheniformis and its application in degraded effective cypermethrin Download PDFInfo
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- CN104560836B CN104560836B CN201510053324.6A CN201510053324A CN104560836B CN 104560836 B CN104560836 B CN 104560836B CN 201510053324 A CN201510053324 A CN 201510053324A CN 104560836 B CN104560836 B CN 104560836B
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- bacillus licheniformis
- effective cypermethrin
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/07—Bacillus
- C12R2001/10—Bacillus licheniformis
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B09—DISPOSAL OF SOLID WASTE; RECLAMATION OF CONTAMINATED SOIL
- B09C—RECLAMATION OF CONTAMINATED SOIL
- B09C1/00—Reclamation of contaminated soil
- B09C1/10—Reclamation of contaminated soil microbiologically, biologically or by using enzymes
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Abstract
The invention discloses a bacillus licheniformis, the entitled BBCY 014 of bacterial strain, Classification And Nomenclature is：Bacillus licheniformis Bacillus licheniformis, are preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center on October 24th, 2014, and deposit number is CGMCC NO.9839.The bacillus licheniformis of the present invention, the beta-cypermethrin degrading rate to 60mg/L concentration under the optimal conditions of laboratory is 95.10%；Field test shows, to the residual of field effective cypermethrin, and the clearance in 7d reaches more than 90%.The bacillus licheniformis of the present invention has the function of degrading pesticide effective cypermethrin, bacillus licheniformis is probiotics, therefore the vegetable soil that its bacteria suspension can be directly used in by pollution by pesticides is biological prosthetic, with preferable repairing effect, and with it is sustainable, cost is low the characteristics of.
Description
Technical field
The present invention relates to a bacillus licheniformis, and its application in degraded effective cypermethrin.
Background technology
Pyrethroid pesticide (Pyrethroid Insecticide) is an analoglike natural pyrethrum structure people
The high-efficiency low-toxicity biomimetic type insecticide of work synthesis, using neural sodium-ion channel as action target, with consumption is low, drug effect is high, kill
The advantages of worm spectrum is wide.Effective cypermethrin (Beta-cypermethrin) be in pyrethroid insecticides application compared with
One of wide agricultural chemicals, by two couple of cypermethrin, efficiently raceme mixture is constituted, and its insecticidal activity is far above cypermethrin,
The food leaf being currently used primarily on the crops such as preventing and treating fruits and vegetables, tealeaves, cereal or food fruit insect, and preventing and treating mosquitos and flies, livestock are parasitic
Worm etc..Although pyrethroid pesticide low toxicity, dosage is low, with frequently using, about the residue problem of the agricultural chemicals
And its also exposed day by day comes out harmfulness.Existing studies have shown that pyrethroid pesticide is to some beneficial insects and aquatic life
Thing has very high toxicity, has neurotoxicity, cumulative toxicity and genotoxicity to human body, effective cypermethrin is by U.S.'s ring
Border protection office is classified as carcinogen, therefore it is a big hot issue to eliminate or reduce residues of pesticides.Biodegradation is to solve agricultural chemicals
One of new way of residual, it is limited for the bacillus research report for effective cypermethrin of degrading at present, and due to soil ring
Border is complicated, and strain degradation ability can not reach laboratory level, therefore this research can not only enrich beta-cypermethrin degrading
Flora, and the application for being it in biological prosthetic provides theoretical foundation.
The content of the invention
For above-mentioned prior art, the present invention has obtained one plant of lichens gemma by enrichment culture, the screening of gradient method for domesticating
Bacillus, its degradation rate to effective cypermethrin is up to 95.10%.
The present invention is achieved by the following technical solutions：
One bacillus licheniformis, the entitled BBCY-014 of bacterial strain, Classification And Nomenclature is：Bacillus licheniformis Bacillus
Licheniformis, is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms on October 24th, 2014
Center, deposit number is CGMCC NO.9839.
The bacillus licheniformis BBCY-014 bacterial strains of the present invention, are tamed on March 10th, 2014 using enrichment culture, gradient
Method is located away from Sichuan Chengdu Pixian County vegetable soil, bacillus licheniformis is accredited as through 16S rDNA, through to effective cypermethrin
Biodegradable experimental study, the bacterium is to the beta-cypermethrin degrading rate of 60mg/L concentration under the optimal conditions of laboratory
95.10%.
The bacillus licheniformis BBCY-014 bacterial strains that the present invention is screened have very strong vitality, in the effect of environment
The lower ability with the various enzymes of generation, has the characteristics such as high temperature resistant, acid and alkali-resistance concurrently.Enzyme-specific and efficient chlorine cyanogen chrysanthemum produced by it
Ester is acted on, and makes its ester linkage breaking, generates corresponding intermediate product, to the last mineralising.Field test shows, bacillus licheniformis
BBCY-014 bacterial strains are to the residual of field effective cypermethrin, and the clearance in 7d reaches more than 90%.
The bacillus licheniformis BBCY-014 new strains that the present invention is screened have the function of degrading pesticide effective cypermethrin,
Bacillus licheniformis is probiotics, therefore the vegetable soil that its bacteria suspension can be directly used in by pollution by pesticides is biological prosthetic, has
Preferable repairing effect, and there is the characteristics of sustainable, cost is low.
Brief description of the drawings
One bacillus licheniformis, the entitled BBCY-014 of bacterial strain, Classification And Nomenclature is：Bacillus licheniformis Bacillus
Licheniformis, is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms on October 24th, 2014
Center, deposit number is CGMCC NO.9839.Preservation address is：Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Chinese science
Institute of microbiology of institute, postcode：100101.
Embodiment
With reference to embodiment, the present invention is further illustrated.
Test the degradeds of 1 bacillus licheniformis BBCY-014 in laboratory conditions to effective cypermethrin
1. bacterial strain storage medium：Conventional method makes improvement LB inclined-plane [(NH4)2SO41.5g, KH2PO40.5g,
K2HPO41.5g, MgSO40.2g, NaCl 10.5g, tryptone 10g, yeast extract powder 5.0g, ultra-pure water 1000mL, agar
2% (w/v), pH7.5, add 0.2% Tween 80 (w/v)] after 121 DEG C of sterilizing 20min, add certain volume effective cypermethrin
Mother liquor 100mg/L concentration for needed for bacterial strain preservation, to maintain strains for degrading ability.Finally the inclined-plane lawn of culture is placed in
Preservation at -80 DEG C in 20% glycerol tube.
2. the preparation of seed liquor/bacteria suspension：By inoculation in the improvement LB nutrient solutions of the effective cypermethrin containing 100mg/L
In [(NH4)2SO41.5g, KH2PO40.5g, K2HPO41.5g, MgSO40.2g, NaCl10.5g, tryptone 10g, yeast are leached
Powder 5.0g, ultra-pure water 1000mL, pH7.5, add 0.2% Tween 80 (w/v)], 37 DEG C of shaking table cultures (180r/min) to logarithm
Phase, with sterile saline centrifuge washing (8000r/min, 4 DEG C) three times, it is resuspended, adjustment cell optical density OD600About 1.0,
Seed liquor/bacteria suspension is made.
3. fluid nutrient medium：(NH4)2SO41.5g, KH2PO40.5g, K2HPO41.5g, MgSO40.2g, NaCl 10.5g, pancreas
Peptone 10g, yeast extract powder 5.0g, ultra-pure water 1000mL, pH7.5, add 0.2% Tween 80 (w/v), effective cypermethrin
Concentration 60mg/L, 121 DEG C of sterilizing 20min.
Prepare such as 3. fluid nutrient medium, effective cypermethrin concentration 60mg/L, aseptically addition is by 2. seed liquor
Preparation method prepare bacillus licheniformis BBCY-014 bacteria suspensions, inoculum concentration 7.0% (v/v), the initial pH7.5 of culture medium,
Fluid nutrient medium conical flask liquid amount is 20mL/100mL, 37 DEG C of shaking table culture (180r/min) 66h.
As a result：After 66h, effective cypermethrin concentration is reduced to 2.94mg/L, bacillus licheniformis BBCY-014 bacteria suspension
Degradation rate to effective cypermethrin is 95.10%.
Test degradeds of the 2 bacillus licheniformis BBCY-014 in vegetable soil to effective cypermethrin
Test in Chengdu Pixian County feux rouges town vegetable garden in November, 2014, soil crop is pakchoi.Soil physical chemistry
Matter is determined to：Dauk, soil pH 6.7, organic matter 17.9g/kg, total nitrogen content 2.3g/kg, total phosphorus content 0.38g/
kg.It is about 10m that 4 pieces of areas are divided in vegetable garden2Soil, wherein two blocks of soil sprinkling empirically 2. preparation of seed liquor in 1
Bacteria suspension prepared by method, the bacteria suspension amount of spraying is 4.5 × 108cfu/g.Soil sample is regularly adopted daily, by 5 point sampling methods, with special
With the instrument of fetching earth, 0~10cm ground surface soils are taken, control group (not spraying Bacillus licheniformis suspension) and spray bacterium group are determined after one week
The residual quantity of the effective cypermethrin of (spraying Bacillus licheniformis suspension), the results are shown in Table 1, spray high in the soil of bacteria suspension
Cypermethrin Residues concentration is imitated well below the control group for not spraying bacterium, illustrates that bacillus licheniformis BBCY-014 can effectively degrade dish
Effective cypermethrin in Orchard Soil, biological prosthetic effect can be played to vegetable soil.
In the vegetable soil repair process of table 1 in soil the residual quantity of effective cypermethrin change
Claims (1)
1. application of the bacillus licheniformis in degraded effective cypermethrin；
The entitled BBCY-014 of bacterial strain of the bacillus licheniformis, Classification And Nomenclature is：Bacillus licheniformis Bacillus
Licheniformis, is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms on October 24th, 2014
Center, deposit number is CGMCC NO.9839；
Bacillus licheniformis is cultivated, bacteria suspension is made；The bacteria suspension, which is prepared by the following method, to be obtained：
Bacillus licheniformis is inoculated in the improvement LB nutrient solutions of the effective cypermethrin containing 100mg/L, 37 DEG C, 180r/min shakes
Bed culture is to logarithmic phase, and centrifuge washing three times under the conditions of 8000r/min, 4 DEG C with sterile saline are resuspended, and adjust cell
Optical density OD600For 1.0, produce；
The improvement LB nutrient solutions of the effective cypermethrin containing 100mg/L, which are prepared by the following method, to be obtained：Take (NH4)2SO41.5g, KH2PO40.5g, K2HPO41.5g, MgSO40.2g, NaCl 10.5g, tryptone 10g, yeast extract powder
5.0g, ultra-pure water 1000mL, mixing are mixed, adjustment pH to 7.5,0.2% Tween 80 of addition, 100mg/L effective cypermethrins, i.e.,
.
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Families Citing this family (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108085281A (en) * | 2018-01-08 | 2018-05-29 | 四川大学 | One plant of β-Cypermethrin in Secticides-degrading Strains and its application |
| CN109439555B (en) * | 2018-11-27 | 2020-10-30 | 四川农业大学 | Saccharomyces cerevisiae capable of effectively adsorbing cypermethrin |
| CN110791456B (en) * | 2019-11-26 | 2022-03-04 | 黄河三角洲京博化工研究院有限公司 | Salt-tolerant growth-promoting bacillus and application thereof |
Citations (3)
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| CN101591623A (en) * | 2009-04-15 | 2009-12-02 | 洪永聪 | A kind of bacillus cereus strain and with the degrading enzyme and preparation method thereof of the pyrethroid pesticide of its preparation |
| CN102604859A (en) * | 2012-01-17 | 2012-07-25 | 华南农业大学 | Pyrethroid pesticide residue degradation bacteria as well as bacteria agent and application |
| CN103776943A (en) * | 2013-09-24 | 2014-05-07 | 四川农业大学 | Method for simultaneously detecting cypermethrin and 3-PBA in microbial degradation system |
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2015
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101591623A (en) * | 2009-04-15 | 2009-12-02 | 洪永聪 | A kind of bacillus cereus strain and with the degrading enzyme and preparation method thereof of the pyrethroid pesticide of its preparation |
| CN102604859A (en) * | 2012-01-17 | 2012-07-25 | 华南农业大学 | Pyrethroid pesticide residue degradation bacteria as well as bacteria agent and application |
| CN103776943A (en) * | 2013-09-24 | 2014-05-07 | 四川农业大学 | Method for simultaneously detecting cypermethrin and 3-PBA in microbial degradation system |
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| Effects of Two Surfactants and Beta-Cyclodextrin on Beta-Cypermethrin Degradation by Bacillus licheniformis B‑1;Jiayuan Zhao等;《J.Agric.Food Chem.》;20151129;第63卷;10729-10735 * |
| Simultaneous Degradation of Cypermethrin and Its Metabolite,3‑Phenoxybenzoic Acid, by the Cooperation of Bacillus licheniformis B‑1 and Sphingomonas sp. SC‑1;Fangfang Liu等;《J.Agric.Food Chem.》;20140728;第62卷;8256-8262 * |
| 拟除虫菊酯类农药残留降解菌的筛选及其生理特性研究;丁海涛;《土壤学报》;20030131;第40卷(第1期);123-129 * |
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