CN103232940B - Bionectria ochroleuca Bo-1 strain, cultures thereof, and applications of the strain and the cultures thereof in resisting pathogenic bacteria - Google Patents
Bionectria ochroleuca Bo-1 strain, cultures thereof, and applications of the strain and the cultures thereof in resisting pathogenic bacteria Download PDFInfo
- Publication number
- CN103232940B CN103232940B CN201310125494.1A CN201310125494A CN103232940B CN 103232940 B CN103232940 B CN 103232940B CN 201310125494 A CN201310125494 A CN 201310125494A CN 103232940 B CN103232940 B CN 103232940B
- Authority
- CN
- China
- Prior art keywords
- bacterial strain
- light color
- red shell
- shell bacterium
- raw red
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 241001149472 Clonostachys rosea Species 0.000 title claims abstract description 12
- 244000052616 bacterial pathogen Species 0.000 title abstract description 7
- 235000014469 Bacillus subtilis Nutrition 0.000 claims abstract description 10
- 241000588724 Escherichia coli Species 0.000 claims abstract description 10
- 241000191967 Staphylococcus aureus Species 0.000 claims abstract description 9
- 241001272684 Xanthomonas campestris pv. oryzae Species 0.000 claims abstract description 8
- 244000063299 Bacillus subtilis Species 0.000 claims abstract description 7
- 241001363490 Monilia Species 0.000 claims abstract description 6
- 238000004321 preservation Methods 0.000 claims abstract description 5
- 241000894006 Bacteria Species 0.000 claims description 85
- 230000001580 bacterial effect Effects 0.000 claims description 62
- 239000000287 crude extract Substances 0.000 claims description 27
- 238000000855 fermentation Methods 0.000 claims description 23
- 230000004151 fermentation Effects 0.000 claims description 23
- 239000007788 liquid Substances 0.000 claims description 23
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 18
- 241000209094 Oryza Species 0.000 claims description 15
- 235000007164 Oryza sativa Nutrition 0.000 claims description 15
- 201000010099 disease Diseases 0.000 claims description 15
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 15
- 235000009566 rice Nutrition 0.000 claims description 15
- 238000000034 method Methods 0.000 claims description 13
- 241000607142 Salmonella Species 0.000 claims description 12
- 230000001717 pathogenic effect Effects 0.000 claims description 9
- 241000222122 Candida albicans Species 0.000 claims description 8
- 241000186781 Listeria Species 0.000 claims description 8
- 229940095731 candida albicans Drugs 0.000 claims description 8
- 230000000968 intestinal effect Effects 0.000 claims description 8
- 230000012010 growth Effects 0.000 claims description 7
- 239000001888 Peptone Substances 0.000 claims description 6
- 108010080698 Peptones Proteins 0.000 claims description 6
- 241000607768 Shigella Species 0.000 claims description 6
- 235000019319 peptone Nutrition 0.000 claims description 6
- 238000002360 preparation method Methods 0.000 claims description 6
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 5
- 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 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
- 229930006000 Sucrose Natural products 0.000 claims description 5
- 229910052799 carbon Inorganic materials 0.000 claims description 5
- 238000001704 evaporation Methods 0.000 claims description 5
- 230000008020 evaporation Effects 0.000 claims description 5
- 239000000284 extract Substances 0.000 claims description 5
- 239000008103 glucose Substances 0.000 claims description 5
- 239000005720 sucrose Substances 0.000 claims description 5
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 4
- 241000186779 Listeria monocytogenes Species 0.000 claims description 4
- 239000000706 filtrate Substances 0.000 claims description 4
- QJGQUHMNIGDVPM-UHFFFAOYSA-N nitrogen group Chemical group [N] QJGQUHMNIGDVPM-UHFFFAOYSA-N 0.000 claims description 4
- 235000015097 nutrients Nutrition 0.000 claims description 4
- 229920002472 Starch Polymers 0.000 claims description 3
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 claims description 3
- 229910052921 ammonium sulfate Inorganic materials 0.000 claims description 3
- 235000011130 ammonium sulphate Nutrition 0.000 claims description 3
- 235000015278 beef Nutrition 0.000 claims description 3
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 claims description 3
- 239000008107 starch Substances 0.000 claims description 3
- 235000019698 starch Nutrition 0.000 claims description 3
- 108010046845 tryptones Proteins 0.000 claims description 3
- 238000003745 diagnosis Methods 0.000 claims description 2
- 238000000605 extraction Methods 0.000 claims description 2
- 238000001914 filtration Methods 0.000 claims description 2
- 229910052757 nitrogen Inorganic materials 0.000 claims description 2
- 241000196324 Embryophyta Species 0.000 abstract description 15
- 241000111332 Molanna albicans Species 0.000 abstract description 3
- 230000002401 inhibitory effect Effects 0.000 abstract description 2
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 24
- 230000008485 antagonism Effects 0.000 description 15
- 230000005764 inhibitory process Effects 0.000 description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 8
- 239000002609 medium Substances 0.000 description 6
- 239000001965 potato dextrose agar Substances 0.000 description 6
- 229920001817 Agar Polymers 0.000 description 5
- 244000061456 Solanum tuberosum Species 0.000 description 5
- 235000002595 Solanum tuberosum Nutrition 0.000 description 5
- 239000008272 agar Substances 0.000 description 5
- 230000000694 effects Effects 0.000 description 5
- 239000000463 material Substances 0.000 description 5
- 241000235349 Ascomycota Species 0.000 description 4
- 230000000844 anti-bacterial effect Effects 0.000 description 4
- WIIZWVCIJKGZOK-RKDXNWHRSA-N chloramphenicol Chemical compound ClC(Cl)C(=O)N[C@H](CO)[C@H](O)C1=CC=C([N+]([O-])=O)C=C1 WIIZWVCIJKGZOK-RKDXNWHRSA-N 0.000 description 4
- 229940049547 paraxin Drugs 0.000 description 4
- 241000142531 Clonostachys Species 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- 241000233866 Fungi Species 0.000 description 3
- 241000607479 Yersinia pestis Species 0.000 description 3
- 230000003042 antagnostic effect Effects 0.000 description 3
- 238000009792 diffusion process Methods 0.000 description 3
- 230000006872 improvement Effects 0.000 description 3
- -1 pyrrolidine alkaloid pyrroles Chemical class 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 230000000452 restraining effect Effects 0.000 description 3
- 238000000926 separation method Methods 0.000 description 3
- 239000007787 solid Substances 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 241000221775 Hypocreales Species 0.000 description 2
- 241000147158 Nectriaceae Species 0.000 description 2
- PCNDJXKNXGMECE-UHFFFAOYSA-N Phenazine Natural products C1=CC=CC2=NC3=CC=CC=C3N=C21 PCNDJXKNXGMECE-UHFFFAOYSA-N 0.000 description 2
- KYQCOXFCLRTKLS-UHFFFAOYSA-N Pyrazine Chemical compound C1=CN=CC=N1 KYQCOXFCLRTKLS-UHFFFAOYSA-N 0.000 description 2
- 241000589634 Xanthomonas Species 0.000 description 2
- 238000013459 approach Methods 0.000 description 2
- 244000000005 bacterial plant pathogen Species 0.000 description 2
- 230000004071 biological effect Effects 0.000 description 2
- 239000011248 coating agent Substances 0.000 description 2
- 238000000576 coating method Methods 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 230000001408 fungistatic effect Effects 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 238000011081 inoculation Methods 0.000 description 2
- 238000009630 liquid culture Methods 0.000 description 2
- 230000000813 microbial effect Effects 0.000 description 2
- 239000000575 pesticide Substances 0.000 description 2
- 239000003375 plant hormone Substances 0.000 description 2
- 239000013641 positive control Substances 0.000 description 2
- 230000002265 prevention Effects 0.000 description 2
- 238000012797 qualification Methods 0.000 description 2
- 238000012216 screening Methods 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- OWEGMIWEEQEYGQ-UHFFFAOYSA-N 100676-05-9 Natural products OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(OC2C(OC(O)C(O)C2O)CO)O1 OWEGMIWEEQEYGQ-UHFFFAOYSA-N 0.000 description 1
- VEPOHXYIFQMVHW-XOZOLZJESA-N 2,3-dihydroxybutanedioic acid (2S,3S)-3,4-dimethyl-2-phenylmorpholine Chemical compound OC(C(O)C(O)=O)C(O)=O.C[C@H]1[C@@H](OCCN1C)c1ccccc1 VEPOHXYIFQMVHW-XOZOLZJESA-N 0.000 description 1
- UPLPHRJJTCUQAY-WIRWPRASSA-N 2,3-thioepoxy madol Chemical compound C([C@@H]1CC2)[C@@H]3S[C@@H]3C[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@](C)(O)[C@@]2(C)CC1 UPLPHRJJTCUQAY-WIRWPRASSA-N 0.000 description 1
- UHPMCKVQTMMPCG-UHFFFAOYSA-N 5,8-dihydroxy-2-methoxy-6-methyl-7-(2-oxopropyl)naphthalene-1,4-dione Chemical compound CC1=C(CC(C)=O)C(O)=C2C(=O)C(OC)=CC(=O)C2=C1O UHPMCKVQTMMPCG-UHFFFAOYSA-N 0.000 description 1
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 1
- 241000193830 Bacillus <bacterium> Species 0.000 description 1
- 241001488339 Bionectriaceae Species 0.000 description 1
- 238000009631 Broth culture Methods 0.000 description 1
- 241000050051 Chelone glabra Species 0.000 description 1
- 241000223218 Fusarium Species 0.000 description 1
- 229930191978 Gibberellin Natural products 0.000 description 1
- 241000238631 Hexapoda Species 0.000 description 1
- 240000007171 Imperata cylindrica Species 0.000 description 1
- GUBGYTABKSRVRQ-PICCSMPSSA-N Maltose Natural products O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-PICCSMPSSA-N 0.000 description 1
- 229930182555 Penicillin Natural products 0.000 description 1
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 1
- 241000371966 Penicillus <bivalve> Species 0.000 description 1
- 241001480064 Quercus variabilis Species 0.000 description 1
- 108010093894 Xanthine oxidase Proteins 0.000 description 1
- 102100033220 Xanthine oxidase Human genes 0.000 description 1
- 241000589652 Xanthomonas oryzae Species 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 239000011149 active material Substances 0.000 description 1
- 238000012271 agricultural production Methods 0.000 description 1
- GZCGUPFRVQAUEE-SLPGGIOYSA-N aldehydo-D-glucose Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C=O GZCGUPFRVQAUEE-SLPGGIOYSA-N 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 229940124350 antibacterial drug Drugs 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 230000003385 bacteriostatic effect Effects 0.000 description 1
- 230000004888 barrier function Effects 0.000 description 1
- GUBGYTABKSRVRQ-QUYVBRFLSA-N beta-maltose Chemical compound OC[C@H]1O[C@H](O[C@H]2[C@H](O)[C@@H](O)[C@H](O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@@H]1O GUBGYTABKSRVRQ-QUYVBRFLSA-N 0.000 description 1
- 239000012620 biological material Substances 0.000 description 1
- 229930183167 cerebroside Natural products 0.000 description 1
- RIZIAUKTHDLMQX-UHFFFAOYSA-N cerebroside D Natural products CCCCCCCCCCCCCCCCC(O)C(=O)NC(C(O)C=CCCC=C(C)CCCCCCCCC)COC1OC(CO)C(O)C(O)C1O RIZIAUKTHDLMQX-UHFFFAOYSA-N 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 210000001072 colon Anatomy 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 229930004069 diterpene Natural products 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000009509 drug development Methods 0.000 description 1
- 238000007877 drug screening Methods 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 210000002615 epidermis Anatomy 0.000 description 1
- 230000003628 erosive effect Effects 0.000 description 1
- 238000003810 ethyl acetate extraction Methods 0.000 description 1
- 239000003448 gibberellin Substances 0.000 description 1
- IXORZMNAPKEEDV-OBDJNFEBSA-N gibberellin A3 Chemical class C([C@@]1(O)C(=C)C[C@@]2(C1)[C@H]1C(O)=O)C[C@H]2[C@]2(C=C[C@@H]3O)[C@H]1[C@]3(C)C(=O)O2 IXORZMNAPKEEDV-OBDJNFEBSA-N 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- SEOVTRFCIGRIMH-UHFFFAOYSA-N indole-3-acetic acid Chemical compound C1=CC=C2C(CC(=O)O)=CNC2=C1 SEOVTRFCIGRIMH-UHFFFAOYSA-N 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 244000000010 microbial pathogen Species 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 230000000877 morphologic effect Effects 0.000 description 1
- 239000004570 mortar (masonry) Substances 0.000 description 1
- 239000005645 nematicide Substances 0.000 description 1
- 229940049954 penicillin Drugs 0.000 description 1
- 230000035479 physiological effects, processes and functions Effects 0.000 description 1
- 239000000049 pigment Substances 0.000 description 1
- 230000008635 plant growth Effects 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 229930002352 pyrrolidine alkaloid Natural products 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 230000002441 reversible effect Effects 0.000 description 1
- 229930000044 secondary metabolite Natural products 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 230000001568 sexual effect Effects 0.000 description 1
- 239000002002 slurry Substances 0.000 description 1
- SUKJFIGYRHOWBL-UHFFFAOYSA-N sodium hypochlorite Chemical compound [Na+].Cl[O-] SUKJFIGYRHOWBL-UHFFFAOYSA-N 0.000 description 1
- 239000008399 tap water Substances 0.000 description 1
- 235000020679 tap water Nutrition 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 239000003643 water by type Substances 0.000 description 1
Landscapes
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
The present invention relates to a plant endophyte Bionectria ochroleuca Bo-1 strain, cultures thereof, and applications of the strain and the cultures thereof in resisting pathogenic bacteria. The strain is preserved in China Center for Type Culture Collection (CCTCC), the address of the center is Wuhan University, Wuhan, China, the accession number is CCTCCNO: M2013027, and the preservation date is January 18, 2013. The Bionectria ochroleuca Bo-1 strain of the invention has high resisting performance for Xanthomonas campestris pv.oryzae, and a strong inhibitory effect on Monilia albicans (M.albican), Escherichia coli (E. coli), Staphylococcus aureus (S. aureus ), Bacillus subtilis (B. subtilis) and other pathogenic bacteria.
Description
Technical field
The invention belongs to microbial technique agricultural production, relate in particular to the raw red shell bacterium of plant endogenesis epiphyte light color (
bionectria ochroleuca) Bo-1 bacterial strain and culture and the application in antagonism pathogenetic bacteria.
Background technology
Bacterial blight of rice is one of Major Diseases in China's Rice Production, can cause paddy rice to subtract 20%-30%, and when serious, even No kernels or seeds are gathered, as in a year of scarcity.Bacterial blight of rice be called again white leaf pest, cogongrass pest, burn etc., be by Xanthomonas oryzae pv oryzae (
xanthomonas oryzaepv.
oryzae, Xoo) cause, all there is generation in Ge Dao district of China, be the Major Diseases of paddy rice.The main controlling way of present stage is chemical prevention, but the excessive medicament such as phenazine, MBAMT of using can cause pathogenic bacteria resistance to strengthen and the havoc of ecotope.
In recent years, the application of plant endogenesis epiphyte in agricultural received much concern.Endophyte of plant refers to and colonizes in health plant stem or blade and root, completes its all or part of life cycle, but does not form a class fungi of obviously infecting.Research is found, the endogenetic fungus colonizing in plant materials can produce various active material, as pyrrolidine alkaloid pyrroles's pyrazine, ergotine, indoles diterpene isoreactivity material, can improve the Anti-bacterium of plant, the ability of nematicide, and phytophage insect is had to stronger toxicity, as Shu etc. from Quercus variabilis endogenous fusanin (
fusariumspp.) in, be separated to 2 kinds and there is the active cerebroside compound with suppressing xanthine oxidase activity of antagonistic bacterium.Also discovery of the research such as Rice, plant endogenesis epiphyte not only can be secreted plant hormone as the growth of Plant hormones regulators,gibberellins, indolylacetic acid isoreactivity material promotion plant, but also can improve the resistance of plant.In addition, the secondary metabolite that plant endogenesis epiphyte produces enriches, chemical structure is special, biological activity is various and structure type is novel, opens up new approach for finding new biological activity material and natural drug development and application.Also will become development green agriculture, developing green agricultural-food effective means simultaneously.Therefore, plant endogenesis epiphyte has the great potential as novel antibacterial drug screening source.
Bionectria (
bionectria) be Rossman in 1999 etc. according to morphology and molecular biological characteristics by a genus of the independent foundation of a part of Nectriaceae (Nectriaceae) bacterial classification, the classified Chi Ke section (Bionectriaceae) of making a living, Hypocreales (Hypocreales), Ascomycetes (Ascomycetes), Ascomycota (Ascomycota).According to research reports, some raw red shell bacterium (
bionectriaspp.) the antibiotics material producing can play specific antagonistic action to some pathogenic bacteria, can suppress the Growth and reproduction of pathogenic micro-organism, as Cao Jinzhong from the raw red shell bacterium N.SBA35 of isolated light color in the root of large-flowered skullcap to subtilis (
bacillus subtilis), colon bacillus (
escherichia coli), sickle-like bacteria (
fusarium) etc. there is better fungistatic effect.But about the plant endogenesis epiphyte-Bionectria of antagonism bacterial leaf spot pathogenic bacteria activity (
bionectria) the rarely seen report of bacterium.
Summary of the invention
For chemical pesticide dangerous with the less problem of microbial pesticide kind, first object of the present invention be to provide the raw red shell bacterium of a kind of light color (
bionectria ochroleuca) Bo-1 bacterial strain, second object of the present invention is to provide the raw red shell bacterium Bo-1 bacterial strain fermentation liquor of light color and crude extract thereof, the 3rd object of the present invention is to provide the culture of the raw red shell bacterium Bo-1 bacterial strain of light color or this bacterial strain for the method for disease-resistant indigenous bacteria, and the 4th object of the present invention is to provide a kind of biological disease-resistant indigenous bacteria preparation.
In order to realize first above-mentioned object, the present invention has adopted following technical scheme:
The raw red shell bacterium of light color (
bionectria ochroleuca) Bo-1 bacterial strain, this bacterial strain is preserved in Chinese Typical Representative culture collection center (CCTCC), address: China. Wuhan. and Wuhan University, deposit number is CCTCC NO:M 2013027, preservation date: on January 18th, 2013.
In order to realize second above-mentioned object, the present invention has adopted following technical scheme:
The raw red shell bacterium Bo-1 bacterial strain fermentation liquor of light color, this fermented liquid is by the raw red shell bacterium Bo-1 bacterial strain of described light color by utilizing organic carbon source and nitrogen source fermentation to prepare, and described strain growth temperature range is 10 DEG C-35 DEG C, and pH scope is 5-9.
As preferably, described strain growth temperature is 30 DEG C, and pH is 6.0.
As preferably, described organic carbon source is selected from one or more in glucose, sucrose, Fructus Hordei Germinatus sugar and starch; Nitrogenous source is selected from one or more in Tryptones, peptone, beef extract and ammonium sulfate.
The raw red shell bacterium Bo-1 bacterial strain fermentation liquor crude extract of light color, it is characterized in that this fermented liquid crude extract by described fermented liquid through filtering, extraction, then evaporation concentration makes.
As preferably, this fermented liquid crude extract is prepared by following method:
1) picking the raw red shell bacterium Bo-1 bacterial strain mycelium of purifying light color be inoculated into and be equipped with in PD liquid nutrient medium, on shaking table, 200 ~ 250 r/min, 25 ~ 30 DEG C, shaking culture;
2) cultivate after 6 ~ 8 days, obtain fermented liquid;
3) gained fermented liquid at room temperature obtains filtrate after filtered through gauze, and filtrate repeatedly extracts through ethyl acetate, until upper strata ethyl acetate layer is colourless;
4) the ethyl acetate layer solution obtaining is merged, on Rotary Evaporators, evaporation concentration is to doing and obtain crude extract.
In order to realize the 3rd above-mentioned object, the present invention has adopted following technical scheme:
The culture of the raw red shell bacterium Bo-1 bacterial strain of light color or this bacterial strain is for the method for disease-resistant indigenous bacteria, but present method forecloses for diagnosis and the treatment of disease.
As preferably, described pathogenetic bacteria be rice leaf spot bacteria (
xanthomonas campestris pv.oryzae), intestinal bacteria (
escherichia coli), Candida albicans (
monilia albican), subtilis (
bacillus subtilis), Salmonellas (
salmonella), Shigellae (
shigella), listeria spp (
listeria monocytogenesor streptococcus aureus (
staphyloccocus aureus).
In order to realize the 4th above-mentioned object, the present invention has adopted following technical scheme:
A kind of biological disease-resistant indigenous bacteria preparation, the disease-resistant indigenous bacteria preparation of this biology comprises the culture of the raw red shell bacterium Bo-1 bacterial strain of described light color or this bacterial strain.
As preferably, described pathogenetic bacteria be rice leaf spot bacteria (
xanthomonas campestris pv.oryzae), intestinal bacteria (
escherichia coli), Candida albicans (
monilia albican), subtilis (
bacillus subtilis), Salmonellas (
salmonella), Shigellae (
shigella), listeria spp (
listeria monocytogenes) or streptococcus aureus (
staphyloccocus aureus).
The raw red shell bacterium Bo-1 bacterial strain of light color of the present invention has the performance of efficient antagonism rice leaf spot bacteria, simultaneously to Candida albicans (
m. albican), intestinal bacteria (
e. coli), streptococcus aureus (
s. aureus), subtilis (
b. subtilis) etc. pathogenetic bacteria all have stronger restraining effect.Therefore Bo-1 bacterial strain and culture thereof having broad application prospects in antagonism pathogenetic bacteria.Be expected to for the research and development of novel microorganism biological prevention and control agent increase new approach, lay the foundation for the biological and ecological methods to prevent plant disease, pests, and erosion work of bacterial blight of rice etc. advances.
Brief description of the drawings
Fig. 1 be the raw red shell bacterium of light color (
bionectria ochroleuca) cultural characteristic of Bo-1 bacterial strain, a left side: bacterium colony front; Right: bacterium colony reverse side.
Fig. 2 is the micro-morphology of the raw red shell bacterium Bo-1 bacterial strain of light color; Left: conidiophore; Right: conidium.
Fig. 3 is the raw red shell bacterium Bo-1 bacterial strain phylogenetic tree of light color of setting up based on ITS rDNA gene order.
Fig. 4 is the raw red shell bacterium Bo-1 bacterial strain fermentation liquor crude extract antagonism Xoo P6 physiological strain inhibition zone of light color; Upper: 0.2% acetone contrast, under left and right: 100 mg/L crude extract.
Fig. 5 is paraxin antagonism Xoo P6 physiological strain inhibition zone; Left: 0.2% acetone contrast, the right side: 100 mg/L paraxin.
Fig. 6 is the raw red shell bacterium Bo-1 bacterial strain fermentation liquor crude extract antagonism Candida albicans inhibition zone of light color; Left: 0.2% acetone contrast, the right side: 100 mg/L crude extract.
Fig. 7 is the raw red shell bacterium Bo-1 bacterial strain fermentation liquor crude extract antagonism subtilis inhibition zone of light color; (left side: 0.2% acetone contrast, the right side: 100 mg/L crude extract).
Fig. 8 is the raw red shell bacterium Bo-1 bacterial strain fermentation liquor crude extract antagonism intestinal bacteria inhibition zone of light color; Left: 0.2% acetone contrast, the right side: 100 mg/L crude extract.
Fig. 9 is the raw red shell bacterium Bo-1 bacterial strain fermentation liquor crude extract antagonism streptococcus aureus inhibition zone of light color; (left side: 0.2% acetone contrast, the right side: 100 mg/L crude extract).
biomaterial preservation explanation
The raw red shell bacterium of light color (
bionectria ochroleuca) Bo-1 bacterial strain, this bacterial strain is preserved in Chinese Typical Representative culture collection center (CCTCC), address: China. Wuhan. and Wuhan University, deposit number is CCTCC NO:M 2013027, preservation date: on January 18th, 2013.
Embodiment
embodiment 1
From the healthy paddy rice blade interior of rice terrace separation screening obtain the raw red shell bacterium of a strain antagonism rice leaf spot bacteria light color (
b.ochroleuca) Bo-1 bacterial strain.
(1) test plant pathogenic bacteria
Rice leaf spot bacteria (
xanthomonas oryzaepv.
oryzae, Xoo) P6 physiological strain.
(2) substratum
Potato dextrose agar (PDA): (200 g), and (20 g), agar (15-20 g), water (1000 mL), pH 6.0 ~ 6.5 for glucose for potato.
Potato glucose liquid state fermentation substratum (PD): (200 g), and (20 g), water (1000 mL), pH 6.0 ~ 6.5 for glucose for potato.
Improvement PDA solid medium: (300 g), and (5.0 g), and (15.0 g), Ca (NO for sucrose for peptone for potato
3)
24H
2o(0.5 g), NaHPO
412H
2g), (15.0 g), water (1000 mL), pH 6.0 ~ 6.5 for agar for O(2.0.
Improvement PD liquid nutrient medium: (300 g), and (5.0 g), and (15.0 g), Ca (NO for sucrose for peptone for potato
3)
24H
2o(0.5 g), NaHPO
412H
2o(2.0 g), water (1000 mL), pH 6.0 ~ 6.5.
(3) experimental technique
1. the separation of paddy rice endogenetic fungus
First with tap water, rice leaf is rinsed well, and with 75% Ethanol Treatment 4 min, aseptic water washing 3 times, process 3 min with 2.5% clorox again, sterilized water washing 3 times, removes blade epidermis, getting 0.01 g puts into aseptic mortar and adds 10 mL sterilized waters to grind to form homogenate, draw 0.5 mL slurries in being added with the PDA culture dish of penicillin (100 μ g/mL), coating is even, 28 DEG C of constant temperature culture.With sterilized water in contrast.After 3 d ~ 5 d, observe, after thalline grows, the required colony edge mycelia of picking purifying is cultivated, and the pure strain number of gained (Bo-1 bacterial strain) is stored in-20 DEG C of refrigerators, for subsequent use.
antagonism
xoothe mensuration of P6 physiological strain activity
Picking purifying Bo-1 bacterial strain mycelium inoculation, to being equipped with in 250 mL Erlenmeyer flasks of approximately 100 mL PD liquid nutrient mediums, after (220 r/min, 28 DEG C) shaking culture 7 d, obtains Bo-1 bacterial strain fermentation liquor on shaking table.Gained fermented liquid to dry, obtains Bo-1 strain fermentation crude extract through filtered through gauze, ethyl acetate extraction, Rotary Evaporators evaporation concentration.
Adopt Oxford agar diffusion method to measure fermented liquid crude extract pair
xoo the antagonistic activity of P6 physiological strain, concrete operations are as follows: (1)-80 DEG C of refrigerators of going bail for and be hidden in
xoo p6 physiological strain is inoculated in drawing together of improvement PD liquid culture medium shaking table and cultivates (28 DEG C, 120 r/min), in the time that bacteria suspension concentration reaches OD value 0.6, gets 100 μ L bacterium liquid and is evenly applied on PDA solid medium, stand-by.(2) become concentration to be the liquid of 100 μ g/mL by 2% acetone diluted respectively Bo-1 bacterial strain fermentation liquor crude extract and paraxin (positive control), stand-by.(3) adopt Oxford agar diffusion method to measure fungistatic effect, and measure the size of inhibition zone by right-angled intersection method.
the qualification of bacterial strain
Bo-1 bacterial strain is carried out to morphology, Physiology and biochemistry and ITS rNDA Sequence Identification, determine its kind.
(4) experimental result
1. high antagonism
xoothe screening of P6 physiological strain Bo-1 bacterial strain
Through flat board coating, separation, purifying and antagonism
xoothe mensuration of P6 physiological strain activity, obtains a strain and is numbered Bo-1 bacterial strain plant endogenesis epiphyte, its fermented liquid crude extract pair
xoop6 physiological strain has stronger bacteriostatic activity, and maximum antibacterial circle diameter is 22.4 mm(accompanying drawings 4), and the maximum antibacterial circle diameter of positive controls paraxin is 23.1 mm(accompanying drawings 5).
the qualification of bacterial strain
Morphological specificity: Bo-1 inoculation, on PDA substratum, is cultivated after 7 d for 30 DEG C, and its colony diameter is about 45.5 mm, and bacterium colony is rounded, and edge is more smooth, and it is cotton-shaped or velvet-like, dense that mycelia is; Initial stage of culture, the positive color of bacterium colony be white, after gradually become pink colour, be finally orange; Bacterium colony back side color is changed gradually into orange and is had a small amount of radial rill by colourless; After cultivating 7 d, bacterial strain is secreted pigment, makes substratum become yellow-green colour (accompanying drawing 1).Micro-Microscopic observation: Bo-1 bacterial strain mycelia is thicker, and branch is less, colourless, has barrier film; Conidiophore betides matrix or surperficial mycelia, falx stem (160 μ m-400 μ m) × (3.5 μ m-4 μ m), wall is smooth or be bordering on smoothly, and some isolates are coarse; Most of is the raw penicillus of single-wheel, a small amount of two-wheel raw (accompanying drawing 2); Conidium is wide ellipse or ellipse, size be (3.0 μ m-4.5 μ m) × (spherical or subsphaeroidal, thecaspore is spherical or subsphaeroidal, unicellular for m) (accompanying drawing 2) of 2.5 μ m-3 μ, sexual generation cleistothecium.
Physiological and biochemical property: strain growth temperature range is 10 DEG C-35 DEG C, 30 DEG C of optimum temperutures; Appropriate pH scope is 5-9, and the most suitable growth pH is 6.0; The carbon source that bacterial strain can utilize has glucose, sucrose, maltose, starch etc.; The nitrogenous source that can utilize has Tryptones, peptone, beef extract and ammonium sulfate etc.
ITS rDNA electrophoresis and sequencing analysis result show, its length is 542 bp, with the raw red shell bacterium of light color (
bionectria ochroleuca) homology reaches 99%(accompanying drawing 3).
embodiment 2
The raw red shell bacterium of light color (
b. ochroleuca) restraining effect of Bo-1 bacterial strain fermentation liquor crude extract to encountered pathogenic bacterium.
(1) test plant pathogenic bacteria:intestinal bacteria (
escherichia coli), Candida albicans (
monilia albican), subtilis (
bacillus subtilis), Salmonellas (
salmonella), Shigellae (
shigella), listeria spp (
listeria monocytogenes), streptococcus aureus (
staphyloccocus aureus).
(2) substratum:beef-protein medium, Half Fraser broth culture, buffered peptone water substratum, GN enrichment liquid substratum.
(3) experimental technique:above-mentioned pathogenetic bacteria to be measured is inoculated in respectively to corresponding liquid culture medium shaking table (120 r/min) activation culture, in the time that bacteria suspension concentration reaches OD value 0.6, gets 100 μ L bacterium liquid and be evenly applied to corresponding solid medium, stand-by; Bo-1 bacterial strain fermentation liquor crude extract is made into 100 μ g/mL solution with 2% acetone, stand-by; Measure respectively inhibitory potency with Oxford agar diffusion method, and measure the size of inhibition zone by right-angled intersection method.
(4) experimental result:bo-1 bacterial strain fermentation liquor crude extract to Candida albicans (
m. albican), intestinal bacteria (
e. coli), streptococcus aureus (
s. aureus), subtilis (
b. subtilis) all have stronger restraining effect, antibacterial circle diameter to be respectively 38.9 mm, 26.4 mm, 28.1 mm and 27.6 mm(accompanying drawings 6,7,8,9); To listeria spp (
l. monocytogenes), Salmonellas (
salmonella), Shigellae (
shigella) effect slightly a little less than, antibacterial circle diameter is respectively 15.1 mm, 15.9 mm, 14.5 mm.
Claims (11)
- The raw red shell bacterium of light color ( bionectria ochroleuca) Bo-1 bacterial strain, this bacterial strain is preserved in Chinese Typical Representative culture collection center (CCTCC), address: China. Wuhan. and Wuhan University, deposit number is CCTCC NO:M 2013027, preservation date: on January 18th, 2013.
- 2. the raw red shell bacterium Bo-1 bacterial strain fermentation liquor of light color, it is characterized in that: this fermented liquid is given birth to red shell bacterium Bo-1 bacterial strain by utilizing organic carbon source and nitrogen source fermentation to prepare by the light color described in claim 1, described strain growth temperature range is 10 DEG C-35 DEG C, and pH scope is 5-9.
- 3. the raw red shell bacterium Bo-1 bacterial strain fermentation liquor of light color according to claim 2, is characterized in that: described strain growth temperature is 30 DEG C, and pH is 6.0.
- 4. the raw red shell bacterium Bo-1 bacterial strain fermentation liquor of light color according to claim 2, is characterized in that: organic carbon source is selected from one or more in glucose, sucrose, Fructus Hordei Germinatus sugar and starch; Nitrogenous source is selected from one or more in peptone, beef extract and ammonium sulfate.
- 5. the raw red shell bacterium Bo-1 bacterial strain fermentation liquor of light color according to claim 4, is characterized in that: nitrogenous source is selected from Tryptones.
- 6. the raw red shell bacterium Bo-1 bacterial strain fermentation liquor crude extract of light color, it is characterized in that this fermented liquid crude extract by the fermented liquid described in claim 2 ~ 5 any one claim through filtering, extraction, then evaporation concentration makes.
- 7. the raw red shell bacterium Bo-1 bacterial strain fermentation liquor crude extract of light color according to claim 6, is characterized in that this fermented liquid crude extract is prepared by following method:1) picking the raw red shell bacterium Bo-1 bacterial strain mycelium of purifying light color be inoculated into and be equipped with in PD liquid nutrient medium, on shaking table, 200 ~ 250 r/min, 25 ~ 30 DEG C, shaking culture;2) cultivate after 6 ~ 8 days, obtain fermented liquid;3) gained fermented liquid at room temperature obtains filtrate after filtered through gauze, and filtrate repeatedly extracts through ethyl acetate, until upper strata ethyl acetate layer is colourless;4) the ethyl acetate layer solution obtaining is merged, on Rotary Evaporators, evaporation concentration is to doing and obtain crude extract.
- 8. the culture of the raw red shell bacterium Bo-1 bacterial strain of light color according to claim 1 or this bacterial strain, for the method for disease-resistant indigenous bacteria, forecloses but the method is used for diagnosis and the treatment of disease.
- 9. the culture of the raw red shell bacterium Bo-1 bacterial strain of light color according to claim 8 or this bacterial strain, for the method for disease-resistant indigenous bacteria, is characterized in that: described pathogenetic bacteria be rice leaf spot bacteria ( xanthomonas campestris pv.oryzae), intestinal bacteria ( escherichia coli), Candida albicans ( monilia albican), subtilis ( bacillus subtilis), Salmonellas ( salmonella), Shigellae ( shigella), listeria spp ( listeria monocytogenesor streptococcus aureus ( staphyloccocus aureus).
- 10. the disease-resistant indigenous bacteria preparation of biology, is characterized in that: the disease-resistant indigenous bacteria preparation of this biology comprises the culture of the raw red shell bacterium Bo-1 bacterial strain of light color described in claim 1 or this bacterial strain.
- The biological disease-resistant indigenous bacteria preparation of 11. one according to claim 10, is characterized in that: described pathogenetic bacteria is rice leaf spot bacteria (Xanthomonas campestris pv.oryzae), intestinal bacteria (Escherichia coli), Candida albicans (Monilia albican), subtilis (Bacillus subtilis), Salmonellas (Salmonella), Shigellae (Shigella), listeria spp (Listeria monocytogenes) or streptococcus aureus (Staphyloccocus aureus).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310125494.1A CN103232940B (en) | 2013-04-10 | 2013-04-10 | Bionectria ochroleuca Bo-1 strain, cultures thereof, and applications of the strain and the cultures thereof in resisting pathogenic bacteria |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310125494.1A CN103232940B (en) | 2013-04-10 | 2013-04-10 | Bionectria ochroleuca Bo-1 strain, cultures thereof, and applications of the strain and the cultures thereof in resisting pathogenic bacteria |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103232940A CN103232940A (en) | 2013-08-07 |
| CN103232940B true CN103232940B (en) | 2014-08-06 |
Family
ID=48881010
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310125494.1A Active CN103232940B (en) | 2013-04-10 | 2013-04-10 | Bionectria ochroleuca Bo-1 strain, cultures thereof, and applications of the strain and the cultures thereof in resisting pathogenic bacteria |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103232940B (en) |
Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104774768B (en) * | 2014-12-30 | 2018-07-31 | 四川农业大学 | A kind of watt cloth fritillaria endogenetic fungus WBS003 and its application |
| CN107794229B (en) * | 2017-11-24 | 2020-12-04 | 浙江师范大学 | Aspergillus sclerotiorum As-75 strain for antagonizing rice bacterial blight and fermentation culture solution and application thereof |
| CN108191663B (en) * | 2017-12-13 | 2021-01-26 | 浙江师范大学 | Antagonistic rice bacterial leaf blight active monomer compound and preparation method thereof |
| CN116751690A (en) * | 2023-07-05 | 2023-09-15 | 黑龙江省农业科学院经济作物研究所 | New strain of Myxomycetes and its application |
-
2013
- 2013-04-10 CN CN201310125494.1A patent/CN103232940B/en active Active
Non-Patent Citations (4)
| Title |
|---|
| 4个黄岑内生真菌菌株的鉴定及抗菌活性研究;曹晋忠等;《安徽农业大学学报》;20110419;第38卷(第3期);393-399 * |
| 刘琴英等.抗水稻白叶枯病菌植物内生真菌的筛选、鉴定.《微生物学杂志》.2013,第33卷(第4期),4-8. * |
| 曹晋忠等.4个黄岑内生真菌菌株的鉴定及抗菌活性研究.《安徽农业大学学报》.2011,第38卷(第3期),393-399. |
| 陆铮铮等.烟草青枯菌土壤拮抗真菌的筛选机鉴定.《贵州农业科学》.2012,第40卷(第1期),86-89. * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN103232940A (en) | 2013-08-07 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Fouda et al. | Biotechnological applications of fungal endophytes associated with medicinal plant Asclepias sinaica (Bioss.) | |
| Saravanan et al. | Development of integrated approach to manage the fusarial wilt of banana | |
| CN104498386B (en) | The preparation method and application of raw Bacillus amyloliquefaciens strain SZ23 and zymotic fluid in wild jujube | |
| Corrado et al. | Antimicrobial evaluation of fungal extracts produced by endophytic strains of Phomopsis sp. | |
| CN107736379B (en) | Application of bacillus amyloliquefaciens in preventing and treating plant fungal diseases | |
| CN101525587B (en) | Streptomyces strain and application thereof | |
| CN102643760B (en) | Antagonistic bacterium capable of generating siderophore for controlling plant diseases | |
| CN105886428A (en) | Streptomyces albidoflavus and applications thereof in microbial fertilizers | |
| CN101851597B (en) | Streptomyces griseoflavus for resisting alfalfa diseases and screening method thereof | |
| CN113801808B (en) | Streptomyces albocongensis and application thereof | |
| CN103232940B (en) | Bionectria ochroleuca Bo-1 strain, cultures thereof, and applications of the strain and the cultures thereof in resisting pathogenic bacteria | |
| Duan et al. | Fermentation optimization and disease suppression ability of a Streptomyces ma. FS-4 from banana rhizosphere soil | |
| CN101928673B (en) | Trichoderma harzianum | |
| CN103087924A (en) | Entomogenous fungus Nomuraearileyi, as well as culture method and application thereof | |
| CN107629985B (en) | Plant endophytic bacterium with antagonistic effect on plant pathogenic fungi | |
| CN105695344B (en) | The cultural method of Isaria Microsclerotia and its application | |
| CN105062897B (en) | The Trichoderma viride of one plant height production chlamydospore and its application | |
| CN102618455B (en) | Stain of streptomyces fradiae and application thereof | |
| CN102363753B (en) | Streptomyces termitum ACT-2 strain, culture and application thereof | |
| CN110317747A (en) | A kind of bacillus amyloliquefaciens JT68 and its application in prevention and treatment tea anthracnose | |
| CN104195064A (en) | Paddy rice endophytic actinomycete realizing in-vitro efficient antagonism on rice blast pathogen | |
| CN103834585A (en) | Rhizospheric pseudomonad capable of largely producing phenazine-1-carboxylic acid and phenazine-1-amide | |
| CN103382447A (en) | Streptomyces viridocyaneus Sv-29 and application thereof | |
| CN113604376A (en) | Sugarcane endophytic bacillus subtilis and application thereof | |
| CN106520572B (en) | A kind of Chinese toon endogenetic fungus 56-50 and its secondary metabolite, preparation method and application |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20191210 Address after: 313000 No.5-8, Changhe Road, Huaxi street, Changxing County, Huzhou City, Zhejiang Province Patentee after: Zhejiang Changxing green battery technology Co.,Ltd. Address before: Room 1,020, Nanxun Science and Technology Pioneering Park, No. 666 Chaoyang Road, Nanxun District, Huzhou City, Zhejiang Province, 313000 Patentee before: Huzhou You Yan Intellectual Property Service Co.,Ltd. Effective date of registration: 20191210 Address after: Room 1,020, Nanxun Science and Technology Pioneering Park, No. 666 Chaoyang Road, Nanxun District, Huzhou City, Zhejiang Province, 313000 Patentee after: Huzhou You Yan Intellectual Property Service Co.,Ltd. Address before: 321004 No. 688 Yingbin Road, Zhejiang, Jinhua Patentee before: Zhejiang Normal University |
|
| TR01 | Transfer of patent right | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20200916 Address after: Room 501, Office Building of Market Supervision Bureau of Langchuan Avenue, Jianping Town, Langxi County, Xuancheng City, Anhui Province Patentee after: Langxi pinxu Technology Development Co.,Ltd. Address before: 313000 No.5-8, Changhe Road, Huaxi street, Changxing County, Huzhou City, Zhejiang Province Patentee before: Zhejiang Changxing green battery technology Co.,Ltd. |
|
| TR01 | Transfer of patent right | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20230921 Address after: Room 1101-7, Yuanlong Building, No. 111 Jianshe North Road, High tech Industrial Park, Tangshan City, Hebei Province, 063000 Patentee after: Hebei Peptide Medical Technology Co.,Ltd. Address before: Room 501, office building, Langchuan Avenue, Jianping Town, Langxi County, Xuancheng City, Anhui Province Patentee before: Langxi pinxu Technology Development Co.,Ltd. |