CN106119168B - One plant has rich phosphorus, degrading organic phosphor and the Serratieae for inhibiting plant pathogenic fungi - Google Patents
One plant has rich phosphorus, degrading organic phosphor and the Serratieae for inhibiting plant pathogenic fungi Download PDFInfo
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- CN106119168B CN106119168B CN201610580077.XA CN201610580077A CN106119168B CN 106119168 B CN106119168 B CN 106119168B CN 201610580077 A CN201610580077 A CN 201610580077A CN 106119168 B CN106119168 B CN 106119168B
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- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 title claims abstract description 50
- 229910052698 phosphorus Inorganic materials 0.000 title claims abstract description 41
- 239000011574 phosphorus Substances 0.000 title claims abstract description 41
- 244000000004 fungal plant pathogen Species 0.000 title claims abstract description 12
- 230000000593 degrading effect Effects 0.000 title claims abstract description 11
- 230000002401 inhibitory effect Effects 0.000 title claims abstract description 10
- MCWXGJITAZMZEV-UHFFFAOYSA-N dimethoate Chemical compound CNC(=O)CSP(=S)(OC)OC MCWXGJITAZMZEV-UHFFFAOYSA-N 0.000 claims abstract description 20
- 239000005562 Glyphosate Substances 0.000 claims abstract description 18
- XDDAORKBJWWYJS-UHFFFAOYSA-N glyphosate Chemical compound OC(=O)CNCP(O)(O)=O XDDAORKBJWWYJS-UHFFFAOYSA-N 0.000 claims abstract description 18
- 229940097068 glyphosate Drugs 0.000 claims abstract description 18
- 230000002265 prevention Effects 0.000 claims description 3
- 238000004321 preservation Methods 0.000 claims description 2
- 230000001580 bacterial effect Effects 0.000 abstract description 24
- 230000015556 catabolic process Effects 0.000 abstract description 23
- 238000006731 degradation reaction Methods 0.000 abstract description 23
- 239000003987 organophosphate pesticide Substances 0.000 abstract description 10
- 201000010099 disease Diseases 0.000 abstract description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 abstract description 3
- 230000000813 microbial effect Effects 0.000 abstract description 3
- 244000005700 microbiome Species 0.000 abstract description 3
- 241000233866 Fungi Species 0.000 abstract description 2
- 241000894006 Bacteria Species 0.000 description 26
- 241000607714 Serratia sp. Species 0.000 description 20
- 239000000575 pesticide Substances 0.000 description 16
- 239000000243 solution Substances 0.000 description 14
- 238000000034 method Methods 0.000 description 13
- 241000123650 Botrytis cinerea Species 0.000 description 8
- 238000000855 fermentation Methods 0.000 description 8
- 230000004151 fermentation Effects 0.000 description 8
- 239000001963 growth medium Substances 0.000 description 8
- 239000002609 medium Substances 0.000 description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 8
- 108020004465 16S ribosomal RNA Proteins 0.000 description 6
- 241000607715 Serratia marcescens Species 0.000 description 6
- 230000005764 inhibitory process Effects 0.000 description 6
- 239000002351 wastewater Substances 0.000 description 6
- 241000196324 Embryophyta Species 0.000 description 5
- 230000000694 effects Effects 0.000 description 5
- 239000002054 inoculum Substances 0.000 description 5
- 238000004519 manufacturing process Methods 0.000 description 5
- 244000052769 pathogen Species 0.000 description 5
- 230000001717 pathogenic effect Effects 0.000 description 5
- 239000000126 substance Substances 0.000 description 5
- 235000017060 Arachis glabrata Nutrition 0.000 description 4
- 244000105624 Arachis hypogaea Species 0.000 description 4
- 235000010777 Arachis hypogaea Nutrition 0.000 description 4
- 235000018262 Arachis monticola Nutrition 0.000 description 4
- XYFCBTPGUUZFHI-UHFFFAOYSA-N Phosphine Chemical compound P XYFCBTPGUUZFHI-UHFFFAOYSA-N 0.000 description 4
- 241000832823 Serratia nematodiphila Species 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 238000005259 measurement Methods 0.000 description 4
- 235000020232 peanut Nutrition 0.000 description 4
- 238000012545 processing Methods 0.000 description 4
- 238000005070 sampling Methods 0.000 description 4
- 238000010186 staining Methods 0.000 description 4
- 238000013456 study Methods 0.000 description 4
- 241000726221 Gemma Species 0.000 description 3
- 241000813090 Rhizoctonia solani Species 0.000 description 3
- 241000607720 Serratia Species 0.000 description 3
- 238000004737 colorimetric analysis Methods 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 238000012216 screening Methods 0.000 description 3
- 239000002689 soil Substances 0.000 description 3
- 239000007787 solid Substances 0.000 description 3
- 230000001629 suppression Effects 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- 235000016623 Fragaria vesca Nutrition 0.000 description 2
- 240000009088 Fragaria x ananassa Species 0.000 description 2
- 235000011363 Fragaria x ananassa Nutrition 0.000 description 2
- ZOKXTWBITQBERF-UHFFFAOYSA-N Molybdenum Chemical compound [Mo] ZOKXTWBITQBERF-UHFFFAOYSA-N 0.000 description 2
- 241000244206 Nematoda Species 0.000 description 2
- 240000007594 Oryza sativa Species 0.000 description 2
- 235000007164 Oryza sativa Nutrition 0.000 description 2
- 238000012408 PCR amplification Methods 0.000 description 2
- 239000001888 Peptone Substances 0.000 description 2
- 108010080698 Peptones Proteins 0.000 description 2
- 241000607479 Yersinia pestis Species 0.000 description 2
- 229960004373 acetylcholine Drugs 0.000 description 2
- 239000003905 agrochemical Substances 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 229940041514 candida albicans extract Drugs 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 235000009508 confectionery Nutrition 0.000 description 2
- 239000003344 environmental pollutant Substances 0.000 description 2
- 229940088598 enzyme Drugs 0.000 description 2
- 210000003495 flagella Anatomy 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- 230000002538 fungal effect Effects 0.000 description 2
- 230000028644 hyphal growth Effects 0.000 description 2
- 230000001788 irregular Effects 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 229910052750 molybdenum Inorganic materials 0.000 description 2
- 239000011733 molybdenum Substances 0.000 description 2
- 235000019319 peptone Nutrition 0.000 description 2
- 229910000073 phosphorus hydride Inorganic materials 0.000 description 2
- 231100000719 pollutant Toxicity 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 230000000452 restraining effect Effects 0.000 description 2
- 235000009566 rice Nutrition 0.000 description 2
- 239000004576 sand Substances 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 239000012138 yeast extract Substances 0.000 description 2
- QCVGEOXPDFCNHA-UHFFFAOYSA-N 5,5-dimethyl-2,4-dioxo-1,3-oxazolidine-3-carboxamide Chemical compound CC1(C)OC(=O)N(C(N)=O)C1=O QCVGEOXPDFCNHA-UHFFFAOYSA-N 0.000 description 1
- 241000588624 Acinetobacter calcoaceticus Species 0.000 description 1
- 241001530056 Athelia rolfsii Species 0.000 description 1
- 241000193830 Bacillus <bacterium> Species 0.000 description 1
- 241000186146 Brevibacterium Species 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- 244000025254 Cannabis sativa Species 0.000 description 1
- 102000012286 Chitinases Human genes 0.000 description 1
- 108010022172 Chitinases Proteins 0.000 description 1
- 102000003914 Cholinesterases Human genes 0.000 description 1
- 108090000322 Cholinesterases Proteins 0.000 description 1
- 108020004414 DNA Proteins 0.000 description 1
- 102000002322 Egg Proteins Human genes 0.000 description 1
- 108010000912 Egg Proteins Proteins 0.000 description 1
- 108090000371 Esterases Proteins 0.000 description 1
- 239000007836 KH2PO4 Substances 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 241000588772 Morganella morganii Species 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 102000052812 Ornithine decarboxylases Human genes 0.000 description 1
- 108700005126 Ornithine decarboxylases Proteins 0.000 description 1
- 241000589517 Pseudomonas aeruginosa Species 0.000 description 1
- 238000012300 Sequence Analysis Methods 0.000 description 1
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 1
- 230000000895 acaricidal effect Effects 0.000 description 1
- OIPILFWXSMYKGL-UHFFFAOYSA-N acetylcholine Chemical compound CC(=O)OCC[N+](C)(C)C OIPILFWXSMYKGL-UHFFFAOYSA-N 0.000 description 1
- -1 acetylcholine ester Chemical class 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
- PYMYPHUHKUWMLA-WDCZJNDASA-N arabinose Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)C=O PYMYPHUHKUWMLA-WDCZJNDASA-N 0.000 description 1
- PYMYPHUHKUWMLA-UHFFFAOYSA-N arabinose Natural products OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 description 1
- SRBFZHDQGSBBOR-UHFFFAOYSA-N beta-D-Pyranose-Lyxose Natural products OC1COC(O)C(O)C1O SRBFZHDQGSBBOR-UHFFFAOYSA-N 0.000 description 1
- 238000010170 biological method Methods 0.000 description 1
- 230000033558 biomineral tissue development Effects 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 239000002775 capsule Substances 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 229940048961 cholinesterase Drugs 0.000 description 1
- 239000000356 contaminant Substances 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000004064 dysfunction Effects 0.000 description 1
- 235000014103 egg white Nutrition 0.000 description 1
- 210000000969 egg white Anatomy 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 229910052564 epsomite Inorganic materials 0.000 description 1
- 238000012851 eutrophication Methods 0.000 description 1
- 238000009313 farming Methods 0.000 description 1
- 239000003337 fertilizer Substances 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 230000002363 herbicidal effect Effects 0.000 description 1
- 239000004009 herbicide Substances 0.000 description 1
- BHEPBYXIRTUNPN-UHFFFAOYSA-N hydridophosphorus(.) (triplet) Chemical compound [PH] BHEPBYXIRTUNPN-UHFFFAOYSA-N 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 230000000749 insecticidal effect Effects 0.000 description 1
- 239000002917 insecticide Substances 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
- 230000005012 migration Effects 0.000 description 1
- 238000013508 migration Methods 0.000 description 1
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 229940076266 morganella morganii Drugs 0.000 description 1
- 231100000252 nontoxic Toxicity 0.000 description 1
- 230000003000 nontoxic effect Effects 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 239000003986 organophosphate insecticide Substances 0.000 description 1
- 230000000361 pesticidal effect Effects 0.000 description 1
- 150000003014 phosphoric acid esters Chemical class 0.000 description 1
- 238000013081 phylogenetic analysis Methods 0.000 description 1
- 238000000053 physical method Methods 0.000 description 1
- 231100000614 poison Toxicity 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000009418 renovation Methods 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 238000004062 sedimentation Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- RYYWUUFWQRZTIU-UHFFFAOYSA-K thiophosphate Chemical compound [O-]P([O-])([O-])=S RYYWUUFWQRZTIU-UHFFFAOYSA-K 0.000 description 1
- 239000003440 toxic substance Substances 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000033 toxigenic Toxicity 0.000 description 1
- 230000001551 toxigenic effect Effects 0.000 description 1
- 230000032258 transport Effects 0.000 description 1
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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
-
- 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
- C12R2001/425—Serratia
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- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
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- Genetics & Genomics (AREA)
- Zoology (AREA)
- Bioinformatics & Cheminformatics (AREA)
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- General Health & Medical Sciences (AREA)
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Abstract
The invention discloses one plant to have rich phosphorus, degrading organic phosphor and the Serratieae for inhibiting plant pathogenic fungi, belongs to microorganisms technical field.The bacterial strainSerratiaSp.MEW06 is preserved in China typical culture collection center, deposit number is CCTCC NO.M 2015779, Serratieae of the invention has the rich phosphorus ability of transitory efficient, it can be used for preparing bio-feritlizer, also there is inhibitory effect to the disease fungus of crop, can be applied to microbial control, simultaneously, the bacterial strain has good degradation to organophosphorus pesticide glyphosate and Rogor, can effectively remove organophosphorus pesticide pollution, Serratieae of the invention is with a wide range of applications.
Description
Technical field
The invention belongs to microorganisms technical fields, and in particular to Serratieae MEW06 and its bacterial strain in rich phosphorus, degradation
Application in terms of organic phosphorus and inhibition plant pathogenic fungi.
Background technique
Organophosphorus pesticide is the big classification in current pesticide, accounts for 80% of Pesticidal products or so, because it is to prevention and treatment agricultural
Disease pest and weed has the characteristics that economical, efficient, quick, at home and abroad mass production and is widely used always.But due to organic
The expanding production of phosphorus pesticide chemical factory, the wastewater flow rate being discharged in production process is big, and COD content is high, and pollutant component is complicated,
Toxicity is big, and difficult for biological degradation causes great harm environment.And can be spread around from applying area after the applications of pesticide,
Migration is accumulated in soil, and into water body or plant etc., and by the enrichment of food chain, hidden danger is generated to human security.It is organic phosphorus
Pesticide is mostly phosphoric acid ester or group thiophosphate, can inhibit cholinesterase activity, makes it that can not decompose phatidylcholine, causes mind
Through dysfunction, serious person may cause death；Secondly, part organophosphorus pesticide is by microbial mineralization in environment at CO2And nothing
After machine phosphorus, causes content of inorganic phosphorus in water body to increase, cause the eutrophication of water body.The uncontrolled production of pesticide chemical factory and
Pesticide is unreasonable to be widely used, and increasingly severe consequence is caused to the mankind and ecological environment, the pollution problem of pesticide is
Hot spot as global concern.Domestic and international scientist has carried out many correlative studys work, to the degradation technique of pesticide have compared with
Great development, including various physics, chemistry, biological method etc..It is high with traditional physical method and chemical method processing cost, it may
New pollutant is generated to compare, using microorganism and its biological product come the biological renovation method of degradation of contaminant have it is nontoxic,
Noresidue, it is without secondary pollution the advantages that, be to eliminate and a kind of method safely, effectively, inexpensively of removing toxic substances agricultural chemicals waste water.
Rogor is the organic phosphorus Insecticidal and acaricidal agent of absorbability, is suitable for prevention and treatment sucking pest, and purposes is wide, yield is big,
But Rogor is volatile to be assembled in the soil to air, water body and sedimentation, pollutes farming animals fishing byproduct, and pass through food chain transport
To human body, harm is generated to people.Glyphosate is the steriland herbicide of a kind of high-efficiency low-toxicity, inner sucting conduction type, due to not having
Selectivity, application range gradually expand.But chemical residual is big in glyphosate production wastewater, and environmental pollution is extremely tight
Weight.Therefore, the bioremediation for studying Rogor and glyphosate seems extremely necessary.
The Rogor degradation bacteria being separated at present mainly has acinetobacter calcoaceticus, Bacillus cercus, pseudomonas aeruginosa, gemma bar
Bacterium etc.；Glyphosate degradation bacterium mainly has pseudomonad, morganella morganii etc.；Existing pesticide degradation bacteria is to organophosphorus insecticide Rogor
Low with the degradation efficiency of glyphosate, bacterial strain has a single function, and practical application is limited.Serratieae of the invention can be with efficient degradation grass
Sweet phosphine and Rogor, and inorganic phosphorus in wastewater can be adsorbed, the improvement for being applied to pollution environment has important practical significance
And application prospect.Currently, about serratia marcescens high yield chitinase and its red non-dispersive pigment-Bacterium prodigiosum of secretion
Element has antibacterial, adjusts immune function, the suppression functions such as cancer report both at home and abroad it is more, but with Rogor and glyphosate degradation effect
Serratieae has not been reported.
Summary of the invention
The purpose of the present invention is the practical problems and needs according to current organophosphorus pesticide pollution, and providing one plant has richness
Phosphorus, degrading organic phosphor pesticides and the Serratieae for inhibiting plant pathogenic fungi.
The present invention has rich phosphorus, degrading organic phosphor pesticides and the Serratieae Serratia for inhibiting plant pathogenic fungi
For sp.MEW06 in China typical culture collection center preservation (Wuhan, China Wuhan University), deposit number is CCTCC NO.M
2015779, the deposit date is on December 25th, 2015.
The bacterium colony and thallus feature of bacterial strain of the present invention are as follows: in LB solid medium culture, bacterium colony is in cerise, surface light
It is sliding, it moistens, opaque, edge is irregular, and size is 1.5-2.0 μm of 0.9-1.2 μ m, and bacterium solution presentation is scarlet when 28 DEG C of cultures
Color, whens 37 DEG C of cultures, do not develop the color.By SEM observation and spore staining, flagella staining, capsule stain and Gram's staining, hair
Existing MEW06 bacterial strain is subsphaeroidal brevibacterium, does not produce gemma, peritrichous, without pod membrane, Gram-negative.
The molecular biological variety identification method of bacterial strain of the present invention are as follows: using Serratia sp.MEW06 genome as template, use
Universal primer (27F and 1492R) expands 16S rDNA sequence (see sequence table), MEW06 16S rDNA PCR amplification sequence warp
NCBI Blast carries out homologous comparison and analyzes, discovery bacterial strain MEW06 and Serratia marcescens (GenBank:
KT894728 sequence similarity) is 99%, the sequence with Serratia nematodiphila (GenBank:KC172028)
Similitude is also 99%.Serratia sp.MEW06 16S rDNA sequence is in the upper and serratia marcescens (Serratia that evolves
Marcescens) close with thermophilic nematode Serratieae (Serratia nematodiphila).
It is provided by the invention that there are rich phosphorus, degrading organic phosphor pesticides and plant pathogenic fungi Serratieae is inhibited to pass through domestication
Method obtains, comprising the following steps: acquisition Sand lake and its water sample on periphery take 5-10mL to be inoculated in LB culture solution, be enriched with
Culture；Then bacteria suspension is inoculated in the fermentation medium containing pesticide by the inoculum concentration (V/V) of 5%-10%, passage and attenuation 5 times
Afterwards, LB solid medium is lined, according to the generation situation of bacterium colony on plate, the well-grown bacterium colony of picking is inoculated in containing agriculture
The fermentation medium of medicine measures organophosphorus pesticide content using acetylcholine ester enzyme inhibition rate method (GB/T5009.199-2003),
Bacterial strain Serratia sp.MEW06 higher to degradation of pesticide efficiency is filtered out, and passes through molybdenum blue colorimetric method, tablet face-off method point
It Ce Ding not rich phosphorus ability and inhibition plant pathogenic fungi ability.
The bacterial strain Serratia sp.MEW06 of separation screening is tested for the property:
A. the measurement of strains for degrading organophosphorus pesticide ability
To inquire into aimed strain Serratia sp.MEW06 to the degradation capability of organophosphorus pesticide, it is raw to choose agricultural at present
There are commonly machine phosphorus insecticide Rogor and glyphosate as target pesticide on producing.By the Serratia sp.MEW06 of screening after purification
Bacterial strain is activated overnight, and adjusts bacterium solution OD600=1.0, by 4% inoculum concentration be forwarded to containing 50mg/L, 100mg/L, 150mg/L,
Degradation rate is measured by sampling in 1d, 3d, 5d, 7d in the fermentation culture of 200mg/L glyphosate or Rogor, 150rpm, 28 DEG C of cultures.
B. the measurement of bacterial strain richness phosphorus ability
By the Serratia sp.MEW06 bacterial strain of screening after purification, being inoculated in phosphorus content respectively by 2% inoculum concentration is
In the PAM culture medium of 10mg/L, 20mg/L, in 0h, 6h, 12h, poly- phosphorus rate is measured by sampling for 24 hours.10min is centrifuged through 6000r/min
Afterwards, supernatant is taken to measure content of inorganic phosphorus (GB 11893-89- molybdenum antimony resistance colorimetric method), with the enrichment culture liquid before access bacterial strain
Inorganic phosphorus concentration compare, calculate poly- phosphorus rate.
The measurement of c.MEW06 bacterial strain restraining epiphyte ability
Taking diameter is the eugonic sharp knife Fusariumsp of 5mm, apple wheel line bacterium, peanut leaf mold, Rhizoctonia solani, strawberry
Grey mold and the pathogen of Botrytis cinerea fungus block are inoculated in the center of PDA plate, 28 DEG C of culture 2d.It is put in the position apart from mycelia edge 2cm
Sterilized filter paper (D=6mm) is set, the bacterium solution of 60 μ L and PBS (blank control) are dripped on filter paper respectively.28 DEG C of continuation
After culture 2 days, inhibition situation of the observation test sample to Fungal hyphal growth.
Compared with prior art, present invention has the advantage that
1, bacterial strain Serratia sp.MEW06 of the invention is strong to the degradation capability of Rogor, glyphosate, and does not cause solution
Middle phosphorus content increases, and can grow under extremely low nutrient environment, and resistance ability is high, therefore can be applied to the biology of agricultural chemicals waste water
Processing, while the bacterial strain has suppression disease fungus ability, to sharp knife Fusariumsp, apple wheel line bacterium, peanut leaf mold, rice banded sclerotial blight
Bacterium, Botrytis cinerea and the pathogen of Botrytis cinerea have and preferably have an inhibitory effect, and Serratia sp.MEW06 can be in biological control, micro-
The application of the fields such as bio-feritlizer, water harnessing, has a vast market foreground.
2, bacterial strain Serratia sp.MEW06 expand cultural method it is simple, toxigenic capacity is low, can with large-scale culture,
Being prepared into remains of pesticide or removal in bio-feritlizer or microbial bacterial agent degradation soil, phosphorous chemical industry enterprise is organic phosphorus and Phos
Pollution.
Detailed description of the invention
Fig. 1 is Serratia sp.MEW06 colonial morphology (A), spore staining (B) and scanning electron microscope (C) figure.
Fig. 2 is that the organic phosphorus degrading rate of Serratia sp.MEW06 changes with time ((A): Rogor, (B): careless sweet
Phosphine).
Fig. 3 is Serratia sp.MEW06 restraining epiphyte lab diagram, (A) sharp knife Fusariumsp；(B) apple wheel line bacterium；(C) flower
Leave is mould；(D) Rhizoctonia solani；(E) Botrytis cinerea；(F) the pathogen of Botrytis cinerea；(a), (c), (e), (g), (i) and (k) blank pair
According to PBS；(b), (d), (f), (h), (j) and (l) MEW06 fermentation liquid.
Case is embodied
The following is specific embodiments of the present invention, is described further to technical solution of the present invention, but of the invention interior
Appearance is not limited solely to range described in embodiment, all to be included in this without departing substantially from the change of present inventive concept or equivalent substitute
Within the protection scope of invention.
Embodiment 1
Acquisition Sand lake and its water sample on periphery are added to LB culture medium, and 28 DEG C of 150r/min enrichment cultures are for 24 hours.It will be upper
State the fermentation medium that bacteria suspension is inoculated in the 200mg/L containing glyphosate by the inoculum concentration of 5%-10%, 28 DEG C of 150r/min enrichments
After cultivating 3d, it is forwarded to same medium, continues to cultivate under the same conditions, passage and attenuation process glyphosate content gradually mentions
It is 1000mg/L that height, which arrives,；The bacterium solution for passing on 5 times is lined into LB solid medium, 28 DEG C, 24-48h is cultivated, according on culture medium
The generation situation of bacterium colony, the well-grown colony inoculation of picking is in the LB culture solution (dilution 5 times) of the 50mg/L containing glyphosate, and 28 DEG C
150r/min cultivates 72h, measures organophosphorus pesticide content using acetylcholine enzyme inhibition rate method (GB/T5009.199-2003),
Bacterial strain is calculated to the degradation rate of organophosphorus pesticide；And by molybdenum blue colorimetric method, tablet face-off method measures rich phosphorus ability and suppression respectively
Plant pathogenic fungi ability processed, obtains bacterial strain MEW06.Using MEW06 genome as template, using universal primer (27F and
1492R), expand 16S rDNA sequence (see sequence table).MEW0616S rDNA PCR amplification sequence carries out same through NCBI Blast
Source compares analysis, and the sequence similarity of discovery bacterial strain MEW06 and Serratia marcescens (GenBank:KT894728) is
99%, the sequence similarity with Serratia nematodiphila (GenBank:KC172028) is also 99%.Use MEGA6
Phylogenetic analysis is carried out it is found that Serratia sp.MEW06 16S rDNA sequence is in upper and serratia marcescens of evolving
(Serratia marcescens) is close with thermophilic nematode Serratieae (Serratia nematodiphila).
Bacterial strain Serratia sp.MEW06 of the present invention is G-, amphimicrobian, be in subsphaeroidal rod-short, do not produce gemma, Zhousheng
Flagellum, without pod membrane.On LB plate, for 24 hours, bacterium colony is in cerise, and convex, surface is smooth for 28 DEG C of cultures, is moistened, opaque, side
Edge is irregular, and size is 1.5-2.0 μm of 0.9-1.2 μ m.Cerise, 37 DEG C of culture Shi Buxian are presented in bacterium solution when 28 DEG C of cultures
Color, the result is shown in Figure 1.Physiological biochemical property are as follows: ornithine decarboxylase, esterase and DNA enzymatic are positive, and arabinose fermentation is negative.
Combining form, physiological and biochemical index and 16S rDNA sequence analysis (see sequence table), are initially identified as Serratia
Bacterium.
Case study on implementation 2
By the Serratia sp.MEW06 bacterial strain of purifying, after being activated overnight, bacterium solution OD is adjusted600=1.0, it is connect by 4%
Kind amount, which is forwarded in the fermentation culture of the glyphosate of content containing a certain concentration or Rogor, cultivates, and is measured by sampling and degrades in 1d, 3d, 5d
Rate.Fermentative medium formula is as follows: peptone 2g/L, yeast extract 1g/L, NaCl 2g/L, pH7.0-7.2.The result shows that
Serratia sp.MEW06 is fabulous to the degradation effect of Rogor, when handling low concentration Rogor (50mg/L), cultivates 5d, supernatant
It is substantially not detectable Rogor concentration, degradation rate nearly reaches 100%.It handles high concentration Rogor (200mg/L), cultivates 7d
Afterwards, measurement degradation rate reaches 90.12%.MEW06 processing glyphosate ability is weaker than Rogor, but under 50mg/L glyphosate concentration,
7d is cultivated, degradation rate is 95% or so.To sum up, for the Rogor of various concentration, the Serratieae of present embodiment has preferably
Degradation effect, for low concentration glyphosate, the Serratieae of present embodiment has preferable degradation effect, as a result sees Fig. 2.
Case study on implementation 3
Serratieae single colonie is inoculated in LB culture medium and is activated overnight in Example 1, and LB culture medium prescription is as follows: egg
White peptone 10g/L, yeast extract 5g/L, NaCl 10g/L, pH7.0-7.2, is inoculated in phosphorus content by 2% inoculum concentration respectively
(in terms of P) is that poly- phosphorus rate, PAM culture medium is measured by sampling in 0,12,24,48,72h in the PAM culture medium of 10mg/L, 20mg/L
Formula are as follows: CH3COONa 5g/L, NH4Cl2g/L, KH2PO40.022~0.0878g/L, MgSO4·7H2O 0.5g/L, CaCl2
0.2g/L, deionized water 1000mL, pH7.0~7.2.The results are shown in Table 1, when phosphorus content is 10mg/L in solution, by for 24 hours
Enrichment, be substantially not detectable content of inorganic phosphorus in solution, rich phosphorus rate reaches 99.94%, and phosphorus content is 20mg/L in solution,
By enrichment for 24 hours, rich phosphorus rate reaches 63.79%, and concrete outcome is shown in Table 1.The Serratieae of present embodiment can be high in short-term
Poly- phosphorus is imitated, the preparation applied to biologic phosphorus fertilizer and the processing of phosphorus-containing wastewater are expected to.
1 Serratia sp.MEW06 of table poly- phosphorus rate (%) in PAM culture medium culture changes over time
Case study on implementation 4
Taking diameter is the eugonic sharp knife Fusariumsp of 5mm, apple wheel line bacterium, peanut leaf mold, Rhizoctonia solani, strawberry
Grey mold and the pathogen of Botrytis cinerea fungus block are inoculated in the center of PDA plate, 28 DEG C of culture 2d.It is put in the position apart from mycelia edge 2cm
The filter paper (D=6mm) for setting sterilizing, the bacterium solution of 60 μ L and PBS (blank control) are dripped on filter paper respectively.28 DEG C are continued to train
After supporting 2 days, inhibition situation of the observation test sample to Fungal hyphal growth.From figure 3, it can be seen that Serratia sp.MEW06
Fermentation liquid can obviously inhibit the growth of sharp knife Fusariumsp, apple wheel line bacterium, peanut leaf mold and Botrytis cinerea mycelia, to rice
The withered bacterium of line and the pathogen of Botrytis cinerea have certain inhibiting effect.
<110>Hubei University
<120>one plants have rich phosphorus, degrading organic phosphor and the Serratieae for inhibiting plant pathogenic fungi
<160>1
<210>1
<211>1400
<212>DNA
<213>Serratieae (Serratia sp.)
<400>1
gtcgagcggt agcacagggg gagcttgctc cccgggtgac gagcggcgga cgggtgagta 60
atgtctggga aactgcctga tggaggggga taactactgg aaacggtagc taataccgca 120
taacgtcgca agaccaaaga gggggacctt cgggcctctt gccatcagat gtgcccagat 180
gggattagct agtaggtggg gtaatggctc acctaggcga cgatccctag ctggtctgag 240
aggatgacca gccacactgg aactgagaca cggtccagac tcctacggga ggcagcagtg 300
gggaatattg cacaatgggc gcaagcctga tgcagccatg ccgcgtgtgt gaagaaggcc 360
ttcgggttgt aaagcacttt cagcgaggag gaaggtggtg agcttaatac gttcatcaat 420
tgacgttact cgcagaagaa gcaccggcta actccgtgcc agcagccgcg gtaatacgga 480
gggtgcaagc gttaatcgga attactgggc gtaaagcgca cgcaggcggt ttgttaagtc 540
agatgtgaaa tccccgggct aacctgggaa ctgcatttga aactggcaag ctagagtctc 600
gtagaggggg gtagaattcc aggtgtagcg gtgaaatgcg tagagatctg gaggaatacc 660
ggtggcgaag gcggccccct ggacgaagac tgacgctcag gtgcgaaagc gtggggagca 720
aacaggatta gataccctgg tagtccacgc tgtaaacgat gtcgatttgg aggttgtgcc 780
cttgaggcgt ggcttccgga gctaacgcgt taaatcgacc gcctggggag tacggccgca 840
aggttaaaac tcaaatgaat tgacgggggc ccgcacaagc ggtggagcat gtggtttaat 900
tcgatgcaac gcgaagaacc ttacctactc ttgacatcca gagaactttc cagagatgga 960
ttggtgcctt cgggaactct gagacaggtg ctgcatggct gtcgtcagct cgtgttgtga 1020
aatgttgggt taagtcccgc aacgagcgca acccttatcc tttgttgcca gcggttcggc 1080
cgggaactca aaggagactg ccagtgataa actggaggaa ggtggggatg acgtcaagtc 1140
atcatggccc ttacgagtag ggctacacac gtgctacaat ggcgtataca aagagaagcg 1200
acctcgcgag agcaagcgga cctcataaag tacgtcgtag tccggattgg agtctgcaac 1260
tcgactccat gaagtcggaa tcgctagtaa tcgtagatca gaatgctacg gtgaatacgt 1320
tcccgggcct tgtacacacc gcccgtcaca ccatgggagt gggttgcaaa agaagtaggt 1380
agcttaacct tcgggagggc 1400
Claims (1)
1. one plant has rich phosphorus, degrading organic phosphor and the application for the Serratieae for inhibiting plant pathogenic fungi, it is characterised in that: should
Serratieae can be used for being enriched with Phos and/or degrading organic phosphor and/or prevention and treatment plant pathogenic fungi；
It is described organic phosphorus for Rogor, glyphosate；
The Serratieae is SerratiaSerratiaSp.MEW06 is preserved in Chinese allusion quotation on December 25th, 2015
Type culture collection preservation, deposit number are CCTCC NO.M 2015779.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102676439A (en) * | 2012-06-26 | 2012-09-19 | 四川大学 | Serratia strain and method for disposing methidathion pesticide residues on surfaces of soil and fruits/vegetables |
| CN103122330A (en) * | 2012-12-31 | 2013-05-29 | 陈秀蓉 | Alpine grassland pasture endogenous serratia plymuthica strain GH010 and application thereof as well as microbial agent prepared from same and preparation method |
| CN104087544A (en) * | 2014-06-19 | 2014-10-08 | 徐州工程学院 | Engineering bacterium capable of degrading organophosphorus pesticides, and construction method and application thereof |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102676439A (en) * | 2012-06-26 | 2012-09-19 | 四川大学 | Serratia strain and method for disposing methidathion pesticide residues on surfaces of soil and fruits/vegetables |
| CN103122330A (en) * | 2012-12-31 | 2013-05-29 | 陈秀蓉 | Alpine grassland pasture endogenous serratia plymuthica strain GH010 and application thereof as well as microbial agent prepared from same and preparation method |
| CN104087544A (en) * | 2014-06-19 | 2014-10-08 | 徐州工程学院 | Engineering bacterium capable of degrading organophosphorus pesticides, and construction method and application thereof |
Non-Patent Citations (1)
| Title |
|---|
| 2株沙雷氏菌Serratia spp.部分生物学功能测定及发酵工艺参数的初探;高晓星;《中国硕士学位论文 农业科技辑》;20130131;D043-101 * |
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