CN104745510B - Lactobacillus fermentum and application thereof in plant fungal disease control - Google Patents
Lactobacillus fermentum and application thereof in plant fungal disease control Download PDFInfo
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- 241000186840 Lactobacillus fermentum Species 0.000 title claims abstract description 33
- 229940012969 lactobacillus fermentum Drugs 0.000 title claims abstract description 6
- 208000031888 Mycoses Diseases 0.000 title abstract description 11
- 238000000855 fermentation Methods 0.000 claims abstract description 28
- 230000004151 fermentation Effects 0.000 claims abstract description 28
- 241000196324 Embryophyta Species 0.000 claims abstract description 11
- 235000007688 Lycopersicon esculentum Nutrition 0.000 claims abstract description 8
- 244000005700 microbiome Species 0.000 claims abstract description 6
- 238000004321 preservation Methods 0.000 claims abstract description 5
- 240000003768 Solanum lycopersicum Species 0.000 claims abstract 2
- 239000012530 fluid Substances 0.000 claims description 27
- 239000001963 growth medium Substances 0.000 claims description 26
- 239000002609 medium Substances 0.000 claims description 26
- 241000894006 Bacteria Species 0.000 claims description 20
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 20
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 claims description 15
- 229940041514 candida albicans extract Drugs 0.000 claims description 15
- 239000001632 sodium acetate Substances 0.000 claims description 15
- 235000017281 sodium acetate Nutrition 0.000 claims description 15
- 239000007787 solid Substances 0.000 claims description 15
- 239000012138 yeast extract Substances 0.000 claims description 15
- CEYULKASIQJZGP-UHFFFAOYSA-L disodium;2-(carboxymethyl)-2-hydroxybutanedioate Chemical compound [Na+].[Na+].[O-]C(=O)CC(O)(C(=O)O)CC([O-])=O CEYULKASIQJZGP-UHFFFAOYSA-L 0.000 claims description 14
- 238000009630 liquid culture Methods 0.000 claims description 13
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 claims description 12
- 235000019262 disodium citrate Nutrition 0.000 claims description 12
- 239000002526 disodium citrate Substances 0.000 claims description 12
- 229940079896 disodium hydrogen citrate Drugs 0.000 claims description 12
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 11
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- 108010080698 Peptones Proteins 0.000 claims description 10
- 235000015278 beef Nutrition 0.000 claims description 10
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- 235000019319 peptone Nutrition 0.000 claims description 10
- 229920000742 Cotton Polymers 0.000 claims description 9
- 241000082085 Verticillium <Phyllachorales> Species 0.000 claims description 9
- 239000000284 extract Substances 0.000 claims description 9
- 229940099596 manganese sulfate Drugs 0.000 claims description 9
- 235000007079 manganese sulphate Nutrition 0.000 claims description 9
- 239000011702 manganese sulphate Substances 0.000 claims description 9
- SQQMAOCOWKFBNP-UHFFFAOYSA-L manganese(II) sulfate Chemical compound [Mn+2].[O-]S([O-])(=O)=O SQQMAOCOWKFBNP-UHFFFAOYSA-L 0.000 claims description 9
- 239000000306 component Substances 0.000 claims description 7
- 239000012533 medium component Substances 0.000 claims description 7
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- 235000009566 rice Nutrition 0.000 claims description 6
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 claims description 5
- 229920001817 Agar Polymers 0.000 claims description 5
- 229930006000 Sucrose Natural products 0.000 claims description 5
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 5
- 239000008272 agar Substances 0.000 claims description 5
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- 241001530056 Athelia rolfsii Species 0.000 claims description 4
- 201000010099 disease Diseases 0.000 claims description 4
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 4
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 claims 1
- 239000005864 Sulphur Substances 0.000 claims 1
- 239000006071 cream Substances 0.000 claims 1
- WPBNNNQJVZRUHP-UHFFFAOYSA-L manganese(2+);methyl n-[[2-(methoxycarbonylcarbamothioylamino)phenyl]carbamothioyl]carbamate;n-[2-(sulfidocarbothioylamino)ethyl]carbamodithioate Chemical compound [Mn+2].[S-]C(=S)NCCNC([S-])=S.COC(=O)NC(=S)NC1=CC=CC=C1NC(=S)NC(=O)OC WPBNNNQJVZRUHP-UHFFFAOYSA-L 0.000 claims 1
- 239000002689 soil Substances 0.000 abstract description 3
- 239000000126 substance Substances 0.000 abstract description 3
- 241000186660 Lactobacillus Species 0.000 abstract description 2
- 229940039696 lactobacillus Drugs 0.000 abstract description 2
- 238000002360 preparation method Methods 0.000 abstract description 2
- 241000606750 Actinobacillus Species 0.000 abstract 1
- 230000000443 biocontrol Effects 0.000 abstract 1
- 244000052769 pathogen Species 0.000 description 20
- 230000001717 pathogenic effect Effects 0.000 description 20
- 239000012141 concentrate Substances 0.000 description 18
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 18
- 239000007788 liquid Substances 0.000 description 15
- 239000006228 supernatant Substances 0.000 description 15
- 230000002401 inhibitory effect Effects 0.000 description 12
- 239000000243 solution Substances 0.000 description 12
- 230000000844 anti-bacterial effect Effects 0.000 description 9
- 239000004310 lactic acid Substances 0.000 description 9
- 235000014655 lactic acid Nutrition 0.000 description 9
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 8
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 8
- 235000011130 ammonium sulphate Nutrition 0.000 description 8
- 239000007979 citrate buffer Substances 0.000 description 8
- 235000021186 dishes Nutrition 0.000 description 8
- 230000009514 concussion Effects 0.000 description 7
- 241000227653 Lycopersicon Species 0.000 description 6
- 230000001629 suppression Effects 0.000 description 6
- 244000052616 bacterial pathogen Species 0.000 description 5
- 239000000575 pesticide Substances 0.000 description 5
- 241000896292 Odontothrips loti Species 0.000 description 4
- 239000003905 agrochemical Substances 0.000 description 4
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- 230000003385 bacteriostatic effect Effects 0.000 description 4
- 229910052739 hydrogen Inorganic materials 0.000 description 4
- 239000001257 hydrogen Substances 0.000 description 4
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- 238000001556 precipitation Methods 0.000 description 4
- 239000013049 sediment Substances 0.000 description 4
- TWFZGCMQGLPBSX-UHFFFAOYSA-N Carbendazim Natural products C1=CC=C2NC(NC(=O)OC)=NC2=C1 TWFZGCMQGLPBSX-UHFFFAOYSA-N 0.000 description 3
- 241001465754 Metazoa Species 0.000 description 3
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 3
- 239000006013 carbendazim Substances 0.000 description 3
- JNPZQRQPIHJYNM-UHFFFAOYSA-N carbendazim Chemical compound C1=C[CH]C2=NC(NC(=O)OC)=NC2=C1 JNPZQRQPIHJYNM-UHFFFAOYSA-N 0.000 description 3
- 238000012258 culturing Methods 0.000 description 3
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- 238000000034 method Methods 0.000 description 3
- -1 phosphoric acid hydrogen Chemical class 0.000 description 3
- QXJKBPAVAHBARF-BETUJISGSA-N procymidone Chemical compound O=C([C@]1(C)C[C@@]1(C1=O)C)N1C1=CC(Cl)=CC(Cl)=C1 QXJKBPAVAHBARF-BETUJISGSA-N 0.000 description 3
- 238000011218 seed culture Methods 0.000 description 3
- 241000208340 Araliaceae Species 0.000 description 2
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 2
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 2
- 235000005035 Panax pseudoginseng ssp. pseudoginseng Nutrition 0.000 description 2
- 235000003140 Panax quinquefolius Nutrition 0.000 description 2
- KCIDZIIHRGYJAE-YGFYJFDDSA-L dipotassium;[(2r,3r,4s,5r,6r)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl] phosphate Chemical compound [K+].[K+].OC[C@H]1O[C@H](OP([O-])([O-])=O)[C@H](O)[C@@H](O)[C@H]1O KCIDZIIHRGYJAE-YGFYJFDDSA-L 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- 235000008434 ginseng Nutrition 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 241000193830 Bacillus <bacterium> Species 0.000 description 1
- 208000031968 Cadaver Diseases 0.000 description 1
- 102000016938 Catalase Human genes 0.000 description 1
- 108010053835 Catalase Proteins 0.000 description 1
- QSJXEFYPDANLFS-UHFFFAOYSA-N Diacetyl Chemical group CC(=O)C(C)=O QSJXEFYPDANLFS-UHFFFAOYSA-N 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 241000192125 Firmicutes Species 0.000 description 1
- 206010017533 Fungal infection Diseases 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- 241000726221 Gemma Species 0.000 description 1
- 241000238631 Hexapoda Species 0.000 description 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- JVTAAEKCZFNVCJ-UHFFFAOYSA-M Lactate Chemical compound CC(O)C([O-])=O JVTAAEKCZFNVCJ-UHFFFAOYSA-M 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 229910002651 NO3 Inorganic materials 0.000 description 1
- NHNBFGGVMKEFGY-UHFFFAOYSA-N Nitrate Chemical compound [O-][N+]([O-])=O NHNBFGGVMKEFGY-UHFFFAOYSA-N 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- 241001052560 Thallis Species 0.000 description 1
- 241000251539 Vertebrata <Metazoa> Species 0.000 description 1
- 235000009754 Vitis X bourquina Nutrition 0.000 description 1
- 235000012333 Vitis X labruscana Nutrition 0.000 description 1
- 240000006365 Vitis vinifera Species 0.000 description 1
- 235000014787 Vitis vinifera Nutrition 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- 238000012271 agricultural production Methods 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 235000013405 beer Nutrition 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000003115 biocidal effect Effects 0.000 description 1
- 230000023852 carbohydrate metabolic process Effects 0.000 description 1
- 239000001569 carbon dioxide Substances 0.000 description 1
- 229910002092 carbon dioxide Inorganic materials 0.000 description 1
- 239000007795 chemical reaction product Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 244000038559 crop plants Species 0.000 description 1
- 101150038956 cup-4 gene Proteins 0.000 description 1
- 235000013365 dairy product Nutrition 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
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- 230000000694 effects Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000009313 farming Methods 0.000 description 1
- 210000003608 fece Anatomy 0.000 description 1
- 239000003337 fertilizer Substances 0.000 description 1
- 235000015203 fruit juice Nutrition 0.000 description 1
- 208000024386 fungal infectious disease Diseases 0.000 description 1
- 239000008273 gelatin Substances 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 235000011852 gelatine desserts Nutrition 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 235000003869 genetically modified organism Nutrition 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 235000013622 meat product Nutrition 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 238000009629 microbiological culture Methods 0.000 description 1
- 239000006916 nutrient agar Substances 0.000 description 1
- 235000008935 nutritious Nutrition 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 230000001590 oxidative effect Effects 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 239000000049 pigment Substances 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 239000010865 sewage Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 210000003708 urethra Anatomy 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
Classifications
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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
- C12R2001/225—Lactobacillus
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/10—Animals; Substances produced thereby or obtained therefrom
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P1/00—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes
- C12P1/04—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes by using bacteria
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- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Zoology (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Wood Science & Technology (AREA)
- General Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- Genetics & Genomics (AREA)
- General Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- Virology (AREA)
- Pest Control & Pesticides (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Mycology (AREA)
- Environmental Sciences (AREA)
- Dentistry (AREA)
- Plant Pathology (AREA)
- Agronomy & Crop Science (AREA)
- Medicinal Chemistry (AREA)
- Tropical Medicine & Parasitology (AREA)
- Biomedical Technology (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
the invention discloses a lactobacillus fermentum and application thereof in plant fungal disease control, belonging to the field of biocontrol preparations, wherein the lactobacillus strains are obtained by separating soil planted under tomatoes and are classified and named as lactobacillus fermentum, the Latin chemical name of the strains is L actinobacillus fermentum, the microorganism of the strain is NJHYHWGY20130877, the preservation date is 2014, 01, 17, the registration number of the preservation center is CGMCC No.8735, the lactobacillus fermentum is fermented and cultured to obtain fermentation liquor, the fermentation liquor is concentrated to prepare a lactein concentrated solution, and the lactein concentrated solution is diluted and sprayed on the leaf surfaces of crops.
Description
Technical field
The invention belongs to biological prevention and control agent field, and in particular to a kind of lactic acid bacteria and its answering in fungal diseases of plants preventing and treating
With.
Technical background
Today's society, agricultural sustainable development have turned into world's most countries and have dominated trend.Agricultural problem is as economical
Social development matter of utmost importance, various countries' agenda is put on.China is used as a large agricultural country, and secure agricultural production is concerning national economy
It is stable with social harmony.Largely although agricultural product can be made significantly to increase production using agricultural chemicals such as chemical fertilizer, agricultural chemicals, also band
Many adverse effects are carried out.For this problem, there is the friendly biological pesticide of green ecological in succession in China, is used with Traditional Agricultural
Chemicals is compared, and is had the advantages that low toxicity, selectively strong, efficient, low-residual, is easily decomposed.Mainly include Natural Enemies of Insects, plant
Source pesticide, microbial pesticide, genetically modified organism, biochemical pesticides and antibiotic agricultural chemicals.
Lactobacillus is also known as Bacillus acidi lactici, is a category for lactic acid bacteria.Cell is generally in regular elongated rod shape, 0.5~1.2 μ m
1.0~10.0 μm, sometimes to be spherical, into short chain.Gram-positive, spore is not given birth to, cell is rare to be moved with peritrichous.It is facultative to detest
Oxygen, sometimes micro- aerobic, bacterium colony projection, full edge, colourless, 2~5mm of diameter on nutrient agar.Chemoheterotrophic bacteria is, it is necessary to nutritious
Culture medium.Glycometabolism is decomposed in fermentation, and more than 50% is lactic acid in end-product.Azymic lactate, nitrate is not reduced, not liquid
Change gelatin, catalase and oxidizing ferment are all negative, rare to cause a disease.30~40 DEG C of optimum temperature, acid resistance is strong, and optimal pH 5.5~
Remain to survive under the conditions of 6.0, pH3.0~4.5.
Lactic acid bacteria is distributed widely in environment, particularly in animal, vegetables and food, generally inhabits bird and vertebrate
Alimentary canal, and in the urethra of mammal.Plant surface, soil, dairy products, meat products, beer, grape wine, fruit juice,
In brewer's wort, sewage and human and animal excreta, separate.Because lactic acid bacteria can produce a large amount of bateriostatics in fermentating metabolism
Matter, including organic acid, enzyme, lactein, hydrogen peroxide, carbon dioxide, biacetyl etc., thus can effectively suppress spoilage organisms and
The growth of pathogenic bacteria, and there is growth promoting function.In addition, lactic acid bacteria has abundant species diversity, be not only research classification,
The ideal material of biochemical, heredity, molecular biology and genetic engineering, and have in fields such as farming and animal husbandry, industry, food and medicines
There is high application value.
The content of the invention
The purpose of the present invention is to propose to one plant of lactobacillus fermenti, it is a further object of the present invention to provide above-mentioned fermentation side
Method, further object of the present invention are to provide application of the lactobacillus fermenti in terms of fungal diseases of plants suppression.
Technical scheme is such as：One strains of lactic acid bacteria, its Classification And Nomenclature are lactobacillus fermenti, the Classification system of strain
It is：Lactobacillus fermentum, join the microorganism of evidence：NJHYHWGY20130877, preservation date are 01 month 2014
17, collection numbering of registering on the books was CGMCC No.8735.
Present invention also offers the fermentation process of above-mentioned lactobacillus fermenti, and it is comprised the following steps that：By lactobacillus fermenti
CGMCC No.8735 are inoculated in solid medium, 30~40 DEG C of 12~24h of culture；A bacterium is chosen in solid medium
Fall, picking thalline, in conical flask of the access equipped with seed liquid culture medium, it is 100~180r/min to control rotating speed, 30~40 DEG C of bars
10~22h is cultivated under part and obtains lactobacillus fermenti seed liquor；Take in conical flask of the seed liquor access equipped with fermentation medium, control turns
Speed is 120~160r/min, and cultivating 24~36h under the conditions of 30~40 DEG C obtains zymotic fluid.
It is preferred that the component of above-mentioned solid medium is：9~11g/L of peptone, 9~11g/L of beef extract, yeast extract 4.5~
5.5g/L, 1.5~2.5g/L of DisodiumHydrogen Citrate, 18~22g/L of glucose, 4.5~5.5g/L of sodium acetate, dipotassium hydrogen phosphate
1.5~2.5g/L, 0.5~0.6g/L of magnesium sulfate, 0.2~0.3g/L of manganese sulfate, 15~25g/L of agar.
It is preferred that the component of above-mentioned seed liquid culture medium is：9~12g/L of peptone, 8~12g/L of beef extract, yeast extract 4~
6g/L, 16~24g/L of glucose, 1.5~2.5g/L of DisodiumHydrogen Citrate, 4~6g/L of sodium acetate, dipotassium hydrogen phosphate 1.5~
2.5g/L, 0.5~0.6g/L of magnesium sulfate, 0.2~0.3g/L of manganese sulfate.
It is preferred that the component of above-mentioned fermentation medium is：20~30g/L of sucrose, yeast extract 15~25g/L, KCl0.6~
1.0g/L, 1.5~2.5g/L of DisodiumHydrogen Citrate, 4~6g/L of sodium acetate, dipotassium hydrogen phosphate 1.5~2.5g/L, pH 5.5~
6.5。
It is preferred that the liquid amount of above-mentioned seed liquid culture medium, which is seed liquor culture volume, accounts for the 16~40% of conical flask capacity；
The liquid amount of fermentation medium is that fermentation medium volume accounts for the 16~40% of conical flask capacity, seed liquor and fermentation medium
Volume ratio is 1：(10~100).
Present invention also offers application of the above-mentioned lactobacillus fermenti in fungal diseases of plants preventing and treating, wherein plant mycosis
Evil bacterial strain preferred cotton verticillium wilt, graw mold of tomato or rice sheath blight disease；Its application process, will to prepare lactein concentrate
The lactein concentrate dilutes 1000~1500 times, is sprayed on corps leaf surface；
Wherein, the lactein concentration liquid and preparation method thereof concretely comprises the following steps, by zymotic fluid obtained by above-mentioned fermentation 8000
Under the conditions of~12000r/min, 20~40min is centrifuged, lower floor's bacterium mud is discarded, it is standby to obtain fermented supernatant fluid；Fermented supernatant fluid is existed
Concentrate is concentrated to give under vacuum condition, wherein volume concentration multiple is 10~20 times；Ammonium sulfate is added into concentrate,
Whirlpool shakes 1~2h, and ammonium sulfate quality saturation degree is 40~50% wherein in solution；Under the conditions of 18000~20000r/min from
25~40min of the heart, supernatant is removed, take lower sediment to be dissolved in the citrate buffer solution that pH is 4.0~5.0, obtain lactic acid bacteria
The concentrate of element, wherein precipitation and the ratio of citrate buffer solution are 1：(100~1000).
The antibacterial bacteriostatic diameter for being verified as, lactein concentrate suppression pathogen being surveyed using Odontothrips loti, is calculated
Bacteriostasis rate；With oese picking pathogen mycelium in centrifuge tube, whirlpool concussion crushes to obtain pathogen hypha fluid, pipettes bacterium
Filament liquid, is spread evenly across in PDA culture medium, and its middle plateform specification is diameter 90mm, high 16mm；In the flat board containing pathogen
On be equidistantly placed Oxford cup 4, after standing 5~10min, the concentration of 100~200 μ L lacteins is added into each Oxford cup
Liquid, wherein Oxford cup specification are 6.0 ± 0.1mm of internal diameter, external diameter 8.0 ± 0.1mm, high 10.0 ± 0.1mm；After 28 DEG C of culture 24h
Survey antibacterial circle diameter.Pathogen inhibiting rate is calculated, wherein bacteriostasis rate calculation formula is：
Growth of pathogenic bacteria inhibiting rate (%)=(antibacterial circle diameter-cups and dishes diameter)/(control colony diameter-cups and dishes diameter)
× 100%
The microorganism of above-mentioned lactobacillus fermenti Lactobacillus fermentum ginsengs evidence：NJHYHWGY20130877, it is
By experimental group from plantation tomato soil in it is isolated.
CGMCC No.8735 bacterial strains have following properties：
1st, form and cultural characteristic
Cultivated on MRS culture mediums, bacterium colony is flat, circular or irregularly arrive coarse, usually transparent, non-pigment；Gram
Positive bacteria, it is shaft-like, gemma is not produced, size is 0.4~1.0 × 2.5~3.0 μm, paired or chaining, is not moved；Heterofermentation, from
Glucose produces sour aerogenesis；Lactobacillus fermenti amphimicrobian, its thalli growth and metabolite generation will appropraite conditions：Growth temperature
Spend for 15~45 DEG C, optimum temperature is 34~40 DEG C, likes slant acidity, pH5.5~7.0, optimal pH 6.0~6.8.
2nd, physiological and biochemical property：
The major physiological biochemical character of lactobacillus fermenti CGMCC No.8735 bacterial strains is shown in Table 1：
The physiological and biochemical property of the bacterial strain of table 1
Note：+：Positive or growth；—：Feminine gender does not grow
Beneficial effect：
The invention discloses one plant of lactobacillus fermenti CGMCC No.8735 and its in crop plants fungal disease preventing and treating side
The application in face, wherein have obvious inhibiting effect to cotton verticillium wilt, it is inhibited to graw mold of tomato or rice sheath blight disease.
Preservation information
The microorganism of above-mentioned lactobacillus fermenti Lactobacillus fermentum ginsengs evidence：NJHYHWGY20130877 is
By this laboratory seed selection and it is preserved in common micro-organisms center (court of Beijing of China General Microbiological culture presevation administration committee
The positive institute 3 of area's North Star West Road 1, Institute of Microorganism, Academia Sinica), it is referred to as CGMCC, and the numbering registered on the books is
CGMCC No.8735, preservation date are：On 01 17th, 2014.
Embodiment
The present invention provides a kind of lactobacillus fermenti Lactobacillus fermentum CGMCC No.8735 and its true
Application in terms of fungus diseases suppression, example is set forth below and is further described.
Example one：
(1) strain culturing
Lactobacillus fermenti CGMCC No.8735 are inoculated in solid medium, 30 DEG C of culture 12h.From solid medium
One bacterium colony of upper selection, picking thalline, access are equipped with the 250mL conical flasks of seed liquid culture medium, wherein seed liquid culture medium
Volume is 16% (in 250mL conical flask load 40mL seeds liquid culture medium) of conical flask capacity, rotating speed 100r/min, 30 DEG C of bars
10h is cultivated under part and obtains seed liquor；Take in 250mL conical flasks of the seed liquor access equipped with fermentation medium, wherein fermentation medium
Volume is 16% (in 250mL conical flask load 40mL seeds liquid culture medium) of conical flask capacity, seed liquor and fermentation medium
Volume ratio be 1：100, rotating speed 120r/min, culture 28h obtains zymotic fluid under the conditions of 30 DEG C.
Above-mentioned solid medium component is (g/L)：Peptone 9, beef extract 9, yeast extract 4.5, glucose 18, hydrogen citrate
Disodium 1.5, sodium acetate 4.5, dipotassium hydrogen phosphate 1.5, magnesium sulfate 0.5, manganese sulfate 0.2, agar 15.
Seed culture medium component is (g/L)：Peptone 9, beef extract 8, yeast extract 4, glucose 16, DisodiumHydrogen Citrate
1.5, sodium acetate 4, dipotassium hydrogen phosphate 1.5, magnesium sulfate 0.5, manganese sulfate 0.2.
Fermentation medium component is (g/L)：Yeast extract 15, sucrose 20, DisodiumHydrogen Citrate 1.5, sodium acetate 4, phosphoric acid hydrogen
Dipotassium 1.5, KCl 0.6, pH5.5.
(2) prepared by fermented supernatant fluid
By zymotic fluid under the conditions of 8000r/min, 20min is centrifuged, lower floor's bacterium mud is discarded, takes fermented supernatant fluid standby.
(3) prepared by lactein concentrate
Fermented supernatant fluid is concentrated to give concentrate under vacuum, wherein volume concentration multiple is 10 times；To concentration
Ammonium sulfate is added in liquid, whirlpool concussion 1h, ammonium sulfate saturation degree is 40% wherein in solution；Under the conditions of 18000r/min from
Heart 25min, supernatant is removed, take lower sediment to be dissolved in the citrate buffer solution that pH is 4.0, obtain the concentration of lactein
Liquid, wherein precipitation and the ratio of citrate buffer solution are 1：100.
(4) bacteriostasis rate calculates
The bacteriostatic diameter of lactein concentrate suppression pathogen is surveyed using Odontothrips loti, calculates bacteriostasis rate.Use oese
Picking pathogen mycelium the broken pathogen hypha fluid of whirlpool concussion, pipettes hypha fluid, in 1.5mL centrifuge tubes
It is even to be coated in PDA culture medium；4 Oxford cups are equidistantly placed on the flat board containing pathogen, after standing 5min, to each ox
100 μ L lactein concentrates are added in the cup of Tianjin；Antibacterial circle diameter is surveyed after 28 DEG C of culture 24h.Calculate pathogen inhibiting rate：
Above-mentioned growth of pathogenic bacteria inhibiting rate (%)=(antibacterial circle diameter-cups and dishes diameter)/(control colony diameter-cups and dishes
Diameter) × 100%
Flat board specification is (mm)：Diameter 90 is high by 16.
The amount that PDA culture medium is poured into flat board is (mL)：25.
Oxford cup specification is (mm)：Internal diameter 6.0 ± 0.1, external diameter 8.0 ± 0.1 are high by 10.0 ± 0.1.
Concrete outcome is shown in Table 1：
Bacteriostasis rate of the table 1 to different fungal diseases
Note：Carbendazim and procymidone concentration are 10 μ g/mL
Lactobacillus fermenti CGMCC No.8735 fermented liquids can effectively suppress cotton verticillium wilt, tomato ash as shown in Table 1
Mildew or rice banded sclerotial blight pathogen.It is wherein best to the inhibition of cotton verticillium wilt, inhibiting rate 59.14%.
Example two：
(1) strain culturing
Lactobacillus fermenti CGMCC No.8735 are inoculated in solid medium, 36 DEG C of culture 18h.From solid medium
One bacterium colony of upper selection, picking thalline, access are equipped with the 250mL conical flasks of seed liquid culture medium, wherein seed liquid culture medium
Volume is the 32% of conical flask capacity, rotating speed 140r/min, and culture 16h obtains seed liquor under the conditions of 36 DEG C；Take seed liquor access dress
Have in the 250mL conical flasks of fermentation medium, wherein fermentation medium volume is the 32% of conical flask capacity, seed liquor and fermentation
The volume ratio of culture medium is 1：20, rotating speed 140r/min, culture 28h obtains zymotic fluid under the conditions of 36 DEG C.
Above-mentioned solid medium component is (g/L)：Peptone 10, beef extract 10, yeast extract 5, glucose 20, hydrogen citrate
Disodium 2, sodium acetate 5, dipotassium hydrogen phosphate 2, magnesium sulfate 0.55, manganese sulfate 0.25, agar 18.
Seed culture medium component is (g/L)：Peptone 10, beef extract 10, yeast extract 5, glucose 20, DisodiumHydrogen Citrate
2, sodium acetate 5, dipotassium hydrogen phosphate 2, magnesium sulfate 0.55, manganese sulfate 0.25.
Fermentation medium component is (g/L)：Yeast extract 18, sucrose 23, DisodiumHydrogen Citrate 2, sodium acetate 5, phosphoric acid hydrogen two
Potassium 2, KCl 0.85, pH6.0.
(2) prepared by fermented supernatant fluid
By zymotic fluid under the conditions of 10000r/min, 30min is centrifuged, lower floor's bacterium mud is discarded, takes fermented supernatant fluid standby.
(3) prepared by lactein concentrate
Fermented supernatant fluid is concentrated to give concentrate under vacuum, wherein volume concentration multiple is 15 times；To concentration
Ammonium sulfate is added in liquid, whirlpool concussion 1.5h, ammonium sulfate saturation degree is 45% wherein in solution；Under the conditions of 19000r/min
32min is centrifuged, removes supernatant, takes lower sediment to be dissolved in the citrate buffer solution that pH is 4.5, obtains the concentration of lactein
Liquid, wherein precipitation and the ratio of citrate buffer solution are 1：1000.
(4) bacteriostasis rate calculates
The bacteriostatic diameter of lactein concentrate suppression pathogen is surveyed using Odontothrips loti, calculates bacteriostasis rate.Use oese
Picking pathogen mycelium the broken pathogen hypha fluid of whirlpool concussion, pipettes hypha fluid, in 1.5mL centrifuge tubes
It is even to be coated in PDA culture medium；4 Oxford cups are equidistantly placed on the flat board containing pathogen, after standing 5min, to each ox
150 μ L lactein concentrates are added in the cup of Tianjin；Antibacterial circle diameter is surveyed after 28 DEG C of culture 24h.Calculate pathogen inhibiting rate：
Above-mentioned growth of pathogenic bacteria inhibiting rate (%)=(antibacterial circle diameter-cups and dishes diameter)/(control colony diameter-cups and dishes
Diameter) × 100%
Flat board specification is (mm)：Diameter 90 is high by 16.
The amount that PDA culture medium is poured into flat board is (mL)：25.
Oxford cup specification is (mm)：Internal diameter 6.0 ± 0.1, external diameter 8.0 ± 0.1 are high by 10.0 ± 0.1.
Concrete outcome is shown in Table 2：
Bacteriostasis rate of the table 2 to different fungal diseases
Note：Carbendazim and procymidone concentration are 10 μ g/mL
Lactobacillus fermenti CGMCC No.8735 fermented liquids can effectively suppress cotton verticillium wilt, tomato ash as shown in Table 2
Mildew or rice banded sclerotial blight pathogen.It is wherein best to the inhibition of cotton verticillium wilt, inhibiting rate 71.41%.
Example three：
(1) strain culturing
Lactobacillus fermenti CGMCC No.8735 are inoculated in solid medium, 40 DEG C of culture 24h.From solid medium
One bacterium colony of upper selection, picking thalline, access are equipped with the 250mL conical flasks of seed liquid culture medium, wherein seed liquid culture medium
It is the 40% of conical flask capacity, rotating speed 180r/min, culture 22h obtains seed liquor under the conditions of 40 DEG C；Seed liquor access is taken equipped with hair
In the 250mL conical flasks of ferment culture medium, wherein fermentation medium volume is the 40% of conical flask capacity, seed liquor and fermented and cultured
The volume ratio of base is 1：10, rotating speed 160r/min, culture 36h obtains zymotic fluid under the conditions of 40 DEG C.
Above-mentioned solid medium component is (g/L)：Peptone 11, beef extract 11, yeast extract 5.5, glucose 22, citric acid
Disodium hydrogen 2.5, sodium acetate 5.5, dipotassium hydrogen phosphate 2.5, magnesium sulfate 0.6, manganese sulfate 0.3, agar 20.
Seed culture medium component is (g/L)：Peptone 12, beef extract 12, yeast extract 6, glucose 24, DisodiumHydrogen Citrate
2.5, sodium acetate 6, dipotassium hydrogen phosphate 2.5, magnesium sulfate 0.6, manganese sulfate 0.3.
Fermentation medium component is (g/L)：Yeast extract 25, sucrose 30, DisodiumHydrogen Citrate 2.5, sodium acetate 6, phosphoric acid hydrogen
Dipotassium 2.5, KCl 1.0, pH6.5.
(2) prepared by fermented supernatant fluid
By zymotic fluid under the conditions of 12000r/min, 40min is centrifuged, lower floor's bacterium mud is discarded, takes fermented supernatant fluid standby.
(3) prepared by lactein concentrate
Fermented supernatant fluid is concentrated to give concentrate under vacuum, wherein volume concentration multiple is 20 times；To concentration
Ammonium sulfate is added in liquid, whirlpool concussion 2h, ammonium sulfate saturation degree is 50% wherein in solution；Under the conditions of 20000r/min from
Heart 40min, supernatant is removed, take lower sediment to be dissolved in the citrate buffer solution that pH is 5.0, obtain the concentration of lactein
Liquid, wherein precipitation and the ratio of citrate buffer solution are 1：500.
(4) bacteriostasis rate calculates
The bacteriostatic diameter of lactein concentrate suppression pathogen is surveyed using Odontothrips loti, calculates bacteriostasis rate.Use oese
Picking pathogen mycelium the broken pathogen hypha fluid of whirlpool concussion, pipettes hypha fluid, in 1.5mL centrifuge tubes
It is even to be coated in PDA culture medium；4 Oxford cups are equidistantly placed on the flat board containing pathogen, after standing 10min, to each
200 μ L lactein concentrates are added in Oxford cup；Antibacterial circle diameter is surveyed after 28 DEG C of culture 24h.Calculate pathogen inhibiting rate：
Above-mentioned growth of pathogenic bacteria inhibiting rate (%)=(antibacterial circle diameter-cups and dishes diameter)/(control colony diameter-cups and dishes
Diameter) × 100%
Flat board specification is (mm)：Diameter 90 is high by 16.
The amount that PDA culture medium is poured into flat board is (mL)：25.
Oxford cup specification is (mm)：Internal diameter 6.0 ± 0.1, external diameter 8.0 ± 0.1 are high by 10.0 ± 0.1.
Concrete outcome is shown in Table 3：
Bacteriostasis rate of the table 3 to different fungal diseases
Note：Carbendazim and procymidone concentration are 10 μ g/mL
Lactobacillus fermenti CGMCC No.8735 fermented liquids can effectively suppress cotton verticillium wilt, tomato ash as shown in Table 3
Mildew or rice banded sclerotial blight pathogen.It is wherein best to the inhibition of cotton verticillium wilt, inhibiting rate 61.32%.
The present invention is used for green bio and prevented and treated, and is had broad application prospects in agriculture field.By to lactobacillus fermenti
The process exploitation of lactobacillus fermenti Lactobacillus fermentum CGMCC No.8735 fermentation lactic acid producing rhzomorphs, is reduced
Chemical pesticide usage amount, agricultural product security is improved, be advantageous to environmental protection and agricultural sustainable development.
Claims (4)
1. one plant of lactobacillus fermenti, the Classification system of strain are：Lactobacillus fermentum, join the microorganism of evidence：
NJHYHWGY20130877, preservation date are on 01 17th, 2014, and collection numbering of registering on the books is CGMCC
No.8735。
2. a kind of fermentation process using lactobacillus fermenti as claimed in claim 1, it is comprised the following steps that：By lactobacillus fermenti
CGMCC No.8735 are inoculated in solid medium, 30~40 DEG C of 12~24h of culture；A bacterium is chosen in solid medium
Fall, picking thalline, in conical flask of the access equipped with seed liquid culture medium, it is 100~180r/min to control rotating speed, 30~40 DEG C of bars
10~22h is cultivated under part and obtains lactobacillus fermenti seed liquor；Take in conical flask of the seed liquor access equipped with fermentation medium, control turns
Speed is 120~160r/min, and cultivating 24~36h under the conditions of 30~40 DEG C obtains zymotic fluid；The group of wherein described solid medium
It is divided into：9~11g/L of peptone, 9~11g/L of beef extract, 4.5~5.5g/L of yeast extract, 1.5~2.5g/L of DisodiumHydrogen Citrate,
18~22g/L of glucose, 4.5~5.5g/L of sodium acetate, 1.5~2.5g/L of dipotassium hydrogen phosphate, 0.5~0.6g/L of magnesium sulfate, sulphur
Sour 0.2~0.3g/L of manganese, 15~25g/L of agar；The component of described seed liquid culture medium is：9~12g/L of peptone, beef
8~12g/L of cream, 4~6g/L of yeast extract, 16~24g/L of glucose, 1.5~2.5g/L of DisodiumHydrogen Citrate, 4~6g/ of sodium acetate
L, 1.5~2.5g/L of dipotassium hydrogen phosphate, 0.5~0.6g/L of magnesium sulfate, 0.2~0.3g/L of manganese sulfate；Described fermentation medium
Component be：20~30g/L of sucrose, yeast extract 15~25g/L, KCl 0.6~1.0g/L, 1.5~2.5g/ of DisodiumHydrogen Citrate
L, 4~6g/L of sodium acetate, dipotassium hydrogen phosphate 1.5~2.5g/L, pH 5.5~6.5.
3. fermentation process as claimed in claim 2, it is characterised in that seed liquor culture volume for conical flask capacity 16~
40%；Fermentation medium volume is the 16~40% of conical flask capacity, and the volume ratio of seed liquor and fermentation medium is 1：(10~
100)。
4. a kind of lactobacillus fermenti as claimed in claim 1 is prevented in cotton verticillium wilt, graw mold of tomato or rice banded sclerotial blight disease
Control the application of aspect.
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| CN102226164A (en) * | 2011-06-22 | 2011-10-26 | 天津科技大学 | High density culture method of lactobacillus fermentium |
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| CN102226164A (en) * | 2011-06-22 | 2011-10-26 | 天津科技大学 | High density culture method of lactobacillus fermentium |
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| 一株活性细菌的抑菌活性研究;朱红霞等;《河南科技学院学报》;20131231;第41卷(第6期);21-24 * |
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