CN116445357A - 一株贝莱斯芽孢杆菌及其应用 - Google Patents
一株贝莱斯芽孢杆菌及其应用 Download PDFInfo
- Publication number
- CN116445357A CN116445357A CN202310482724.3A CN202310482724A CN116445357A CN 116445357 A CN116445357 A CN 116445357A CN 202310482724 A CN202310482724 A CN 202310482724A CN 116445357 A CN116445357 A CN 116445357A
- Authority
- CN
- China
- Prior art keywords
- bacillus
- colletotrichum
- strain
- bailii
- botrytis cinerea
- 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.)
- Pending
Links
- 241000193830 Bacillus <bacterium> Species 0.000 title claims abstract description 61
- 238000011282 treatment Methods 0.000 claims abstract description 28
- 239000004562 water dispersible granule Substances 0.000 claims abstract description 15
- 230000000443 biocontrol Effects 0.000 claims abstract description 8
- 230000000813 microbial effect Effects 0.000 claims abstract description 8
- 239000003337 fertilizer Substances 0.000 claims abstract description 6
- 244000000005 bacterial plant pathogen Species 0.000 claims abstract description 4
- 241000123650 Botrytis cinerea Species 0.000 claims description 24
- 201000010099 disease Diseases 0.000 claims description 17
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 17
- 238000000034 method Methods 0.000 claims description 16
- 241000193744 Bacillus amyloliquefaciens Species 0.000 claims description 15
- 241000196324 Embryophyta Species 0.000 claims description 13
- 239000003795 chemical substances by application Substances 0.000 claims description 13
- 241000222199 Colletotrichum Species 0.000 claims description 12
- 230000000853 biopesticidal effect Effects 0.000 claims description 12
- 241000223221 Fusarium oxysporum Species 0.000 claims description 8
- 241001508802 Diaporthe Species 0.000 claims description 7
- 230000004927 fusion Effects 0.000 claims description 7
- 230000005764 inhibitory process Effects 0.000 claims description 7
- 238000002360 preparation method Methods 0.000 claims description 6
- 244000068988 Glycine max Species 0.000 claims description 5
- 235000010469 Glycine max Nutrition 0.000 claims description 5
- 241000221696 Sclerotinia sclerotiorum Species 0.000 claims description 5
- 241000223602 Alternaria alternata Species 0.000 claims description 4
- 241000555706 Botryosphaeria dothidea Species 0.000 claims description 4
- 241001149955 Cladosporium cladosporioides Species 0.000 claims description 4
- 241000193403 Clostridium Species 0.000 claims description 4
- 241001123536 Colletotrichum acutatum Species 0.000 claims description 4
- 241001345409 Colletotrichum aenigma Species 0.000 claims description 4
- 241001639769 Colletotrichum fructicola Species 0.000 claims description 4
- 241001529387 Colletotrichum gloeosporioides Species 0.000 claims description 4
- 241001639768 Colletotrichum siamense Species 0.000 claims description 4
- 241000642046 Colletotrichum viniferum Species 0.000 claims description 4
- 241000802849 Dactylonectria alcacerensis Species 0.000 claims description 4
- 241000965183 Dactylonectria macrodidyma Species 0.000 claims description 4
- 241001240231 Dactylonectria torresensis Species 0.000 claims description 4
- 241001221840 Neofusicoccum parvum Species 0.000 claims description 4
- 241000579036 Neopestalotiopsis rosae Species 0.000 claims description 4
- 239000004480 active ingredient Substances 0.000 claims description 4
- 238000009629 microbiological culture Methods 0.000 claims description 4
- 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 claims description 3
- 241000223600 Alternaria Species 0.000 claims description 3
- 241000382787 Diaporthe sojae Species 0.000 claims description 3
- 241000233866 Fungi Species 0.000 claims description 3
- 241000223218 Fusarium Species 0.000 claims description 3
- 241000190144 Lasiodiplodia theobromae Species 0.000 claims description 3
- 239000002202 Polyethylene glycol Substances 0.000 claims description 3
- 239000004372 Polyvinyl alcohol Substances 0.000 claims description 3
- 244000007021 Prunus avium Species 0.000 claims description 3
- 235000010401 Prunus avium Nutrition 0.000 claims description 3
- DBMJMQXJHONAFJ-UHFFFAOYSA-M Sodium laurylsulphate Chemical compound [Na+].CCCCCCCCCCCCOS([O-])(=O)=O DBMJMQXJHONAFJ-UHFFFAOYSA-M 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
- FPAFDBFIGPHWGO-UHFFFAOYSA-N dioxosilane;oxomagnesium;hydrate Chemical compound O.[Mg]=O.[Mg]=O.[Mg]=O.O=[Si]=O.O=[Si]=O.O=[Si]=O.O=[Si]=O FPAFDBFIGPHWGO-UHFFFAOYSA-N 0.000 claims description 3
- 230000003032 phytopathogenic effect Effects 0.000 claims description 3
- 229920001223 polyethylene glycol Polymers 0.000 claims description 3
- 229920002451 polyvinyl alcohol Polymers 0.000 claims description 3
- 229940088594 vitamin Drugs 0.000 claims description 3
- 239000011782 vitamin Substances 0.000 claims description 3
- 229930003231 vitamin Natural products 0.000 claims description 3
- 235000013343 vitamin Nutrition 0.000 claims description 3
- 241000222290 Cladosporium Species 0.000 claims description 2
- 239000000203 mixture Substances 0.000 claims description 2
- 241000461774 Gloeosporium Species 0.000 claims 3
- 241001361634 Rhizoctonia Species 0.000 claims 3
- 241000223935 Cryptosporidium Species 0.000 claims 2
- 241001558929 Sclerotium <basidiomycota> Species 0.000 claims 2
- 239000002671 adjuvant Substances 0.000 claims 2
- 239000012681 biocontrol agent Substances 0.000 claims 2
- 244000000003 plant pathogen Species 0.000 claims 2
- 241000707502 Coniella Species 0.000 claims 1
- 241000138452 Coniella vitis Species 0.000 claims 1
- 241000234276 Curculigo Species 0.000 claims 1
- 241000223208 Curvularia Species 0.000 claims 1
- 241001235320 Diaporthe conorum Species 0.000 claims 1
- 241000218187 Lachnum Species 0.000 claims 1
- 241001163578 Lasiodiplodia pseudotheobromae Species 0.000 claims 1
- 244000028344 Primula vulgaris Species 0.000 claims 1
- 235000016311 Primula vulgaris Nutrition 0.000 claims 1
- 240000006474 Theobroma bicolor Species 0.000 claims 1
- NHTFLYKPEGXOAN-UHFFFAOYSA-N Trichlamide Chemical compound CCCCOC(C(Cl)(Cl)Cl)NC(=O)C1=CC=CC=C1O NHTFLYKPEGXOAN-UHFFFAOYSA-N 0.000 claims 1
- 235000009508 confectionery Nutrition 0.000 claims 1
- 239000003090 pesticide formulation Substances 0.000 claims 1
- 230000000694 effects Effects 0.000 abstract description 40
- 238000012360 testing method Methods 0.000 abstract description 26
- 235000007688 Lycopersicon esculentum Nutrition 0.000 abstract description 22
- 238000000855 fermentation Methods 0.000 abstract description 18
- 230000004151 fermentation Effects 0.000 abstract description 18
- 238000004321 preservation Methods 0.000 abstract description 10
- 244000000004 fungal plant pathogen Species 0.000 abstract description 9
- 230000000844 anti-bacterial effect Effects 0.000 abstract description 8
- 230000002265 prevention Effects 0.000 abstract description 8
- 238000011081 inoculation Methods 0.000 abstract description 7
- 241000721179 Clarias Species 0.000 abstract description 5
- 238000000338 in vitro Methods 0.000 abstract description 5
- 239000003899 bactericide agent Substances 0.000 abstract description 3
- 238000001228 spectrum Methods 0.000 abstract description 2
- 241000227653 Lycopersicon Species 0.000 abstract 3
- 238000001514 detection method Methods 0.000 description 29
- 230000001580 bacterial effect Effects 0.000 description 25
- 244000063299 Bacillus subtilis Species 0.000 description 20
- 240000003768 Solanum lycopersicum Species 0.000 description 19
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 19
- 239000002609 medium Substances 0.000 description 17
- 239000001963 growth medium Substances 0.000 description 14
- 235000014469 Bacillus subtilis Nutrition 0.000 description 11
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 10
- 239000000575 pesticide Substances 0.000 description 10
- 239000000243 solution Substances 0.000 description 10
- 241000588722 Escherichia Species 0.000 description 9
- 235000009754 Vitis X bourquina Nutrition 0.000 description 8
- 235000012333 Vitis X labruscana Nutrition 0.000 description 8
- 240000006365 Vitis vinifera Species 0.000 description 8
- 235000014787 Vitis vinifera Nutrition 0.000 description 8
- 238000012258 culturing Methods 0.000 description 8
- 241000894006 Bacteria Species 0.000 description 7
- 230000003385 bacteriostatic effect Effects 0.000 description 7
- 239000003814 drug Substances 0.000 description 7
- 239000004563 wettable powder Substances 0.000 description 7
- 108020004465 16S ribosomal RNA Proteins 0.000 description 6
- 244000052616 bacterial pathogen Species 0.000 description 6
- 239000000126 substance Substances 0.000 description 6
- 238000005406 washing Methods 0.000 description 6
- 229920001817 Agar Polymers 0.000 description 5
- 239000004382 Amylase Substances 0.000 description 5
- 102000013142 Amylases Human genes 0.000 description 5
- 108010065511 Amylases Proteins 0.000 description 5
- 241000588724 Escherichia coli Species 0.000 description 5
- 241000589902 Leptospira Species 0.000 description 5
- 108091005804 Peptidases Proteins 0.000 description 5
- 239000004365 Protease Substances 0.000 description 5
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 5
- 239000008272 agar Substances 0.000 description 5
- 235000019418 amylase Nutrition 0.000 description 5
- 238000002474 experimental method Methods 0.000 description 5
- 230000012010 growth Effects 0.000 description 5
- 230000007062 hydrolysis Effects 0.000 description 5
- 238000006460 hydrolysis reaction Methods 0.000 description 5
- 239000007788 liquid Substances 0.000 description 5
- 239000000725 suspension Substances 0.000 description 5
- 108010059892 Cellulase Proteins 0.000 description 4
- 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 description 4
- SQUHHTBVTRBESD-UHFFFAOYSA-N Hexa-Ac-myo-Inositol Natural products CC(=O)OC1C(OC(C)=O)C(OC(C)=O)C(OC(C)=O)C(OC(C)=O)C1OC(C)=O SQUHHTBVTRBESD-UHFFFAOYSA-N 0.000 description 4
- 238000012408 PCR amplification Methods 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 229920002472 Starch Polymers 0.000 description 4
- 229930006000 Sucrose Natural products 0.000 description 4
- 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 description 4
- 229940106157 cellulase Drugs 0.000 description 4
- 239000012153 distilled water Substances 0.000 description 4
- 238000004043 dyeing Methods 0.000 description 4
- 239000008103 glucose Substances 0.000 description 4
- 229960000367 inositol Drugs 0.000 description 4
- CDAISMWEOUEBRE-GPIVLXJGSA-N inositol Chemical compound O[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@H](O)[C@@H]1O CDAISMWEOUEBRE-GPIVLXJGSA-N 0.000 description 4
- 239000000463 material Substances 0.000 description 4
- 239000000843 powder Substances 0.000 description 4
- CDAISMWEOUEBRE-UHFFFAOYSA-N scyllo-inosotol Natural products OC1C(O)C(O)C(O)C(O)C1O CDAISMWEOUEBRE-UHFFFAOYSA-N 0.000 description 4
- 239000008107 starch Substances 0.000 description 4
- 235000019698 starch Nutrition 0.000 description 4
- 239000005720 sucrose Substances 0.000 description 4
- 235000004936 Bromus mango Nutrition 0.000 description 3
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 3
- 239000004367 Lipase Substances 0.000 description 3
- 102000004882 Lipase Human genes 0.000 description 3
- 108090001060 Lipase Proteins 0.000 description 3
- 240000007228 Mangifera indica Species 0.000 description 3
- 235000014826 Mangifera indica Nutrition 0.000 description 3
- 229930195725 Mannitol Natural products 0.000 description 3
- 239000001888 Peptone Substances 0.000 description 3
- 108010080698 Peptones Proteins 0.000 description 3
- 235000009184 Spondias indica Nutrition 0.000 description 3
- 239000002253 acid Substances 0.000 description 3
- 230000008485 antagonism Effects 0.000 description 3
- 238000003556 assay Methods 0.000 description 3
- 238000006243 chemical reaction Methods 0.000 description 3
- 239000011248 coating agent Substances 0.000 description 3
- 238000000576 coating method Methods 0.000 description 3
- 238000011161 development Methods 0.000 description 3
- 229940088598 enzyme Drugs 0.000 description 3
- 239000012634 fragment Substances 0.000 description 3
- 235000012055 fruits and vegetables Nutrition 0.000 description 3
- 244000053095 fungal pathogen Species 0.000 description 3
- 239000007789 gas Substances 0.000 description 3
- 239000011521 glass Substances 0.000 description 3
- 239000004519 grease Substances 0.000 description 3
- 239000008101 lactose Substances 0.000 description 3
- 235000019421 lipase Nutrition 0.000 description 3
- 239000000594 mannitol Substances 0.000 description 3
- 235000010355 mannitol Nutrition 0.000 description 3
- 235000019319 peptone Nutrition 0.000 description 3
- 108090000623 proteins and genes Proteins 0.000 description 3
- OARRHUQTFTUEOS-UHFFFAOYSA-N safranin Chemical compound [Cl-].C=12C=C(N)C(C)=CC2=NC2=CC(C)=C(N)C=C2[N+]=1C1=CC=CC=C1 OARRHUQTFTUEOS-UHFFFAOYSA-N 0.000 description 3
- 239000012192 staining solution Substances 0.000 description 3
- 239000008223 sterile water Substances 0.000 description 3
- 230000001954 sterilising effect Effects 0.000 description 3
- 208000024891 symptom Diseases 0.000 description 3
- ZCYVEMRRCGMTRW-UHFFFAOYSA-N 7553-56-2 Chemical compound [I] ZCYVEMRRCGMTRW-UHFFFAOYSA-N 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 241000193755 Bacillus cereus Species 0.000 description 2
- 241001328122 Bacillus clausii Species 0.000 description 2
- 241000194108 Bacillus licheniformis Species 0.000 description 2
- 241001290235 Ceratobasidium cereale Species 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
- 238000003794 Gram staining Methods 0.000 description 2
- DKNPRRRKHAEUMW-UHFFFAOYSA-N Iodine aqueous Chemical compound [K+].I[I-]I DKNPRRRKHAEUMW-UHFFFAOYSA-N 0.000 description 2
- 241000194105 Paenibacillus polymyxa Species 0.000 description 2
- 241000212749 Zesius chrysomallus Species 0.000 description 2
- 230000003042 antagnostic effect Effects 0.000 description 2
- 238000012512 characterization method Methods 0.000 description 2
- 238000005520 cutting process Methods 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- XJRPTMORGOIMMI-UHFFFAOYSA-N ethyl 2-amino-4-(trifluoromethyl)-1,3-thiazole-5-carboxylate Chemical compound CCOC(=O)C=1SC(N)=NC=1C(F)(F)F XJRPTMORGOIMMI-UHFFFAOYSA-N 0.000 description 2
- 238000000605 extraction Methods 0.000 description 2
- 230000006799 invasive growth in response to glucose limitation Effects 0.000 description 2
- 238000011835 investigation Methods 0.000 description 2
- 229910052740 iodine Inorganic materials 0.000 description 2
- 239000011630 iodine Substances 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 235000013336 milk Nutrition 0.000 description 2
- 239000008267 milk Substances 0.000 description 2
- 210000004080 milk Anatomy 0.000 description 2
- 239000003921 oil Substances 0.000 description 2
- 235000019198 oils Nutrition 0.000 description 2
- 230000001681 protective effect Effects 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 238000007789 sealing Methods 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 238000002791 soaking Methods 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 239000007921 spray Substances 0.000 description 2
- 238000007619 statistical method Methods 0.000 description 2
- 230000001225 therapeutic effect Effects 0.000 description 2
- 241000256852 Aculeata Species 0.000 description 1
- 108010001478 Bacitracin Proteins 0.000 description 1
- 241001465180 Botrytis Species 0.000 description 1
- 241000754798 Calophyllum brasiliense Species 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 108010084185 Cellulases Proteins 0.000 description 1
- 102000005575 Cellulases Human genes 0.000 description 1
- 244000038022 Chenopodium capitatum Species 0.000 description 1
- 235000004391 Chenopodium capitatum Nutrition 0.000 description 1
- 244000241235 Citrullus lanatus Species 0.000 description 1
- 235000012828 Citrullus lanatus var citroides Nutrition 0.000 description 1
- JJLJMEJHUUYSSY-UHFFFAOYSA-L Copper hydroxide Chemical compound [OH-].[OH-].[Cu+2] JJLJMEJHUUYSSY-UHFFFAOYSA-L 0.000 description 1
- 239000005750 Copper hydroxide Substances 0.000 description 1
- 229920000742 Cotton Polymers 0.000 description 1
- 241000192125 Firmicutes Species 0.000 description 1
- 102000004157 Hydrolases Human genes 0.000 description 1
- 108090000604 Hydrolases Proteins 0.000 description 1
- 239000012880 LB liquid culture medium Substances 0.000 description 1
- 240000008415 Lactuca sativa Species 0.000 description 1
- 235000003228 Lactuca sativa Nutrition 0.000 description 1
- 208000031888 Mycoses Diseases 0.000 description 1
- 235000019483 Peanut oil Nutrition 0.000 description 1
- 241001290151 Prunus avium subsp. avium Species 0.000 description 1
- 241000589516 Pseudomonas Species 0.000 description 1
- 241000232299 Ralstonia Species 0.000 description 1
- 241000589771 Ralstonia solanacearum Species 0.000 description 1
- 241000813090 Rhizoctonia solani Species 0.000 description 1
- 241000707465 Schizoparme Species 0.000 description 1
- 238000012300 Sequence Analysis Methods 0.000 description 1
- 239000005708 Sodium hypochlorite Substances 0.000 description 1
- 244000299461 Theobroma cacao Species 0.000 description 1
- 235000009470 Theobroma cacao Nutrition 0.000 description 1
- ABUGVBRDFWGJRD-CHOYNLESSA-N [9-[(2r,3r,4s,5r)-3,4-dihydroxy-5-methyloxolan-2-yl]-2-(2,4-dinitrophenyl)sulfanylpurin-6-yl] [hydroxy(phosphonooxy)phosphoryl] hydrogen phosphate Chemical compound O[C@@H]1[C@H](O)[C@@H](C)O[C@H]1N1C2=NC(SC=3C(=CC(=CC=3)[N+]([O-])=O)[N+]([O-])=O)=NC(OP(O)(=O)OP(O)(=O)OP(O)(O)=O)=C2N=C1 ABUGVBRDFWGJRD-CHOYNLESSA-N 0.000 description 1
- DPXJVFZANSGRMM-UHFFFAOYSA-N acetic acid;2,3,4,5,6-pentahydroxyhexanal;sodium Chemical compound [Na].CC(O)=O.OCC(O)C(O)C(O)C(O)C=O DPXJVFZANSGRMM-UHFFFAOYSA-N 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- 238000000246 agarose gel electrophoresis Methods 0.000 description 1
- 238000007605 air drying Methods 0.000 description 1
- 230000003625 amylolytic effect Effects 0.000 description 1
- 238000000540 analysis of variance Methods 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000000845 anti-microbial effect Effects 0.000 description 1
- 239000012298 atmosphere Substances 0.000 description 1
- 239000012752 auxiliary agent Substances 0.000 description 1
- 229960003071 bacitracin Drugs 0.000 description 1
- 229930184125 bacitracin Natural products 0.000 description 1
- CLKOFPXJLQSYAH-ABRJDSQDSA-N bacitracin A Chemical compound C1SC([C@@H](N)[C@@H](C)CC)=N[C@@H]1C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H]1C(=O)N[C@H](CCCN)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@H](CC=2C=CC=CC=2)C(=O)N[C@@H](CC=2N=CNC=2)C(=O)N[C@H](CC(O)=O)C(=O)N[C@@H](CC(N)=O)C(=O)NCCCC1 CLKOFPXJLQSYAH-ABRJDSQDSA-N 0.000 description 1
- 239000007640 basal medium Substances 0.000 description 1
- 235000015278 beef Nutrition 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 239000003124 biologic agent Substances 0.000 description 1
- 239000012620 biological material Substances 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 239000001768 carboxy methyl cellulose Substances 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 235000019693 cherries Nutrition 0.000 description 1
- 238000004040 coloring Methods 0.000 description 1
- IQFVPQOLBLOTPF-HKXUKFGYSA-L congo red Chemical compound [Na+].[Na+].C1=CC=CC2=C(N)C(/N=N/C3=CC=C(C=C3)C3=CC=C(C=C3)/N=N/C3=C(C4=CC=CC=C4C(=C3)S([O-])(=O)=O)N)=CC(S([O-])(=O)=O)=C21 IQFVPQOLBLOTPF-HKXUKFGYSA-L 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 229910001956 copper hydroxide Inorganic materials 0.000 description 1
- 239000013078 crystal Substances 0.000 description 1
- 238000003235 crystal violet staining Methods 0.000 description 1
- 238000012136 culture method Methods 0.000 description 1
- 230000001461 cytolytic effect Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 235000014113 dietary fatty acids Nutrition 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 238000001125 extrusion Methods 0.000 description 1
- 239000000194 fatty acid Substances 0.000 description 1
- 229930195729 fatty acid Natural products 0.000 description 1
- 150000004665 fatty acids Chemical class 0.000 description 1
- 230000000855 fungicidal effect Effects 0.000 description 1
- 239000000417 fungicide Substances 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 230000007614 genetic variation Effects 0.000 description 1
- 230000009036 growth inhibition Effects 0.000 description 1
- 101150013736 gyrB gene Proteins 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000009776 industrial production Methods 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 230000003902 lesion Effects 0.000 description 1
- 230000000670 limiting effect Effects 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000003020 moisturizing effect Effects 0.000 description 1
- 230000000877 morphologic effect Effects 0.000 description 1
- PGSADBUBUOPOJS-UHFFFAOYSA-N neutral red Chemical compound Cl.C1=C(C)C(N)=CC2=NC3=CC(N(C)C)=CC=C3N=C21 PGSADBUBUOPOJS-UHFFFAOYSA-N 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 101150012629 parE gene Proteins 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 230000007170 pathology Effects 0.000 description 1
- 239000000312 peanut oil Substances 0.000 description 1
- 239000008194 pharmaceutical composition Substances 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
- 230000001737 promoting effect Effects 0.000 description 1
- 230000017854 proteolysis Effects 0.000 description 1
- 230000002797 proteolythic effect Effects 0.000 description 1
- 238000004080 punching Methods 0.000 description 1
- 230000009257 reactivity Effects 0.000 description 1
- 230000002829 reductive effect Effects 0.000 description 1
- 230000001850 reproductive effect Effects 0.000 description 1
- 238000012552 review Methods 0.000 description 1
- 230000000630 rising effect Effects 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000008159 sesame oil Substances 0.000 description 1
- 235000011803 sesame oil Nutrition 0.000 description 1
- 235000019812 sodium carboxymethyl cellulose Nutrition 0.000 description 1
- 229920001027 sodium carboxymethylcellulose Polymers 0.000 description 1
- SUKJFIGYRHOWBL-UHFFFAOYSA-N sodium hypochlorite Chemical compound [Na+].Cl[O-] SUKJFIGYRHOWBL-UHFFFAOYSA-N 0.000 description 1
- 239000002689 soil Substances 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000005507 spraying Methods 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
- 239000012128 staining reagent Substances 0.000 description 1
- 150000005846 sugar alcohols Chemical class 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 238000011144 upstream manufacturing Methods 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
- 230000037303 wrinkles Effects 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
Classifications
-
- 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
-
- 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
- A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
- A01N63/22—Bacillus
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01P—BIOCIDAL, PEST REPELLANT, PEST ATTRACTANT OR PLANT GROWTH REGULATORY ACTIVITY OF CHEMICAL COMPOUNDS OR PREPARATIONS
- A01P1/00—Disinfectants; Antimicrobial compounds or mixtures thereof
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01P—BIOCIDAL, PEST REPELLANT, PEST ATTRACTANT OR PLANT GROWTH REGULATORY ACTIVITY OF CHEMICAL COMPOUNDS OR PREPARATIONS
- A01P3/00—Fungicides
-
- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05F—ORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C, e.g. FERTILISERS FROM WASTE OR REFUSE
- C05F11/00—Other organic fertilisers
- C05F11/08—Organic fertilisers containing added bacterial cultures, mycelia or the like
-
- 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
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/07—Bacillus
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Health & Medical Sciences (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Biotechnology (AREA)
- Organic Chemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- General Health & Medical Sciences (AREA)
- Virology (AREA)
- Plant Pathology (AREA)
- Environmental Sciences (AREA)
- Biochemistry (AREA)
- Pest Control & Pesticides (AREA)
- Tropical Medicine & Parasitology (AREA)
- General Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Medicinal Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Agronomy & Crop Science (AREA)
- Dentistry (AREA)
- Mycology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
本发明提供了一株贝莱斯芽孢杆菌及其应用。本发明的贝莱斯芽孢杆菌菌株BJ‑1的保藏编号为CGMCC No.24113。本发明的贝莱斯芽孢杆菌菌株具有对植物病原菌的抑菌活性并且具有较宽的抑菌谱。室内离体叶片接种试验结果表明,贝莱斯芽孢杆菌BJ‑1发酵液对番茄灰霉病具有较好的保护和治疗作用,1×107CFU/mLBJ‑1发酵液对番茄灰霉病的保护和治疗作用相对防效分别可达68.95%和88.28%。田间试验结果表明,贝莱斯芽孢杆菌BJ‑1水分散粒剂对番茄灰霉病具有较好的防治效果,2.0×1010CFU/克贝莱斯芽孢杆菌BJ‑1水分散粒剂处理(450克/亩)的相对防效可达80.48%。本发明的贝莱斯芽孢杆菌具有生防潜力,可用于制备针对植物病原真菌的生防菌剂和微生物菌肥。
Description
技术领域
本发明属于微生物应用领域,具体涉及一株贝莱斯芽孢杆菌(Bacillusvelezensis)及其在防治番茄灰霉病等植物病害上的应用。
背景技术
灰葡萄孢(Botrytis cinerea)寄主广泛,可侵染葡萄、草莓、樱桃、番茄、西瓜等多种果蔬,引起作物灰霉病,在世界范围内均有发生,一般情况下造成的产量损失为10%~20%,严重时高达60%及以上。化学防治是当前实际生产中防控作物灰霉病的主要措施(康立娟,张小风,王文桥,等.灰霉菌的抗药性与适合度测定.农药学学报,2003,2:39-42.)。然而由于灰葡萄孢具有繁殖快速、遗传变异大和适合度高等特点,对化学农药的抗性产生快,导致药剂的防治效果降低(Hahn M.The rising threat of fungicide resistance inplant pathogenic fungi:Botrytis as a case study.Journal of Chemical Biology,2014,7(4):133-141.)。此外,长期、反复和大量使用化学农药可引起土壤、水体和大气的污染,破坏了生态平衡。生物农药具有低毒、绿色和安全等特点,符合可持续发展理念(KaurS,Samota M K,Choudhary M,et al.How do plants defend themselves againstpathogens-Biochemical mechanisms and genetic interventions.Physiology andMolecular Biology of Plants,2022,28(2):485-504.),因此开发应用于作物灰霉病的生物农药迫在眉睫。芽孢杆菌(Bacillus spp.)在自然界分布广泛,且逆境下产生的休眠体芽孢不仅含水量极低,且具有耐酸、耐盐、耐高温、抗挤压等特点,其代谢还可产生多种酶类以及杆菌肽、环脂、类噬菌体颗粒等十几种抗菌物质,相比其他有益微生物,芽孢杆菌存活率高、繁殖能力强、环境适应能力与抗逆能力强,是目前生物农药产品中使用最广泛的一类生防菌(Ivica I,Janakiev T,/>M,et al.Plant-associated Bacillusand Pseudomonas antimicrobial activities in plant disease suppression viabiological control mechanisms-A review,Physiological and Molecular PlantPathology,2022,117:101754)。芽孢杆菌的种类多,在生物农药应用方面较多的种类包括枯草芽孢杆菌(Bacillus subtilis)、解淀粉芽孢杆菌(Bacillus amlyoliquefaciens)、多粘类芽孢杆菌(Paenibacilluspolmyxa)、蜡样芽孢杆菌(Bacillus cereus)、地衣芽孢杆菌(Bacillus licheniformis)等。贝莱斯芽孢杆菌(Bacillus velezensis)近年来由于具有较好的抑菌效果得到了研究人员的关注,如有研究表明,贝莱斯芽孢杆菌对防治立枯丝核菌引起的生菜根腐病和雷尔氏菌引起的番茄青枯病起主要作用(Cao Y,Pi HL,ChandrangsuP,et al.Antagonism of two plant-growth promoting Bacillusvelezensis isolates against Ralstonia solanacearum and Fusariumoxysporum.Scientific Reports,2018,8(1):4360.);贝莱斯芽孢杆菌BS87和RK1对草莓枯萎病的田间防治效果与氢氧化铜相当(Nam MH,Park MS,Kim HG,et al.Biologicalcontrol of strawberry Fusarium wilt caused by Fusarium oxysporumf.spfragariaeusing Bacillus velezensis BS87 and RK1formulation.Journal of Microbiology andBiotechnology,2009,19(5):520-524.);Balderas等人从芒果树叶中分离得到的贝莱斯芽孢杆菌83对芒果炭疽病的防治效果与化学处理相当(Balderas-Ruiz KA,Bustos P,Santamaria RI,et al.Bacillus velezensis 83a bacterial strain from mangophyllosphere,useful for biological Control and plant growth promotion.AMBExpress,2020,101:7-10.)。
发明内容
为了解决上述问题,本发明提供了一株贝莱斯芽孢杆菌及其应用。本发明中提供的贝莱斯芽孢杆菌对多种植物病原菌具有较好的抑制效果,对灰葡萄孢引起的番茄灰霉病在田间表现较好的防效,因此其对于防治灰葡萄孢引起的果蔬灰霉病较好的应用前景。
本发明的贝莱斯芽孢杆菌,分类命名为贝莱斯芽孢杆菌Bacillus velezensis,采集自河北省张家口怀来县,分离自葡萄枝干病害的葡萄病枝干组织,命名为贝莱斯芽孢杆菌(Bacillus velezensis)BJ-1,已于2021年12月16日保藏于中国微生物菌种保藏管理委员会普通微生物中心(地址:中国北京),保藏编号为CGMCC No.24113。
本发明的贝莱斯芽孢杆菌在LB培养基上不产色素,菌落为乳白色不透明,圆形或近圆形,边缘不光滑,表面有明显褶皱。所述贝莱斯芽孢杆菌BJ-1的生理生化特征为革兰氏染色阳性,可产纤维素酶,不产蛋白酶、淀粉酶。
本发明的贝莱斯芽孢杆菌具有对植物病原真菌的抑菌活性并且具有较宽的抑菌谱。
所述植物病原真菌为可可毛色二孢(Lasiodiplodia theobromae)、葡萄座腔菌(Botryosphaeria dothidea)、小新壳梭孢(Neofusicoccum parvum)、大豆间座壳(Diaporthe sojae)、甜樱间座壳(Diaporthe eres)、Diaporthe honkonggensis、灰葡萄孢(Botrytis cinerea)、Colletotrichum viniferum、Colletotrichum sojae、隐秘炭疽菌(Colletotrichum aenigma)、胶孢炭疽菌(Colletotrichum gloeosporioides)、果生炭疽菌(Colletotrichum fructicola)、暹罗炭疽菌(Colletotrichum siamense)、尖孢炭疽菌(Colletotrichum acutatum)、Neopestalotiopsis sp.、Neopestalotiopsis rosae、Coniella vitis、Dactylonectria alcacerensis、Dactylonectria macrodidyma、Lasiodiplodiapseudotheobromae、Dactylonectria torresensis、尖孢镰刀菌(Fusariumoxysporum)、双核丝核菌融合群A、双核丝核菌融合群G、链格孢(Alternaria alternata)、枝状枝孢菌(Cladosporium cladosporioides)、核盘菌(Sclerotinia sclerotiorum)。对上述26种植物病原真菌59个菌株的菌丝生长抑制率在53.76%-98.45%之间。该菌株在植物真菌病害生物制剂开发方面具有非常好的应用前景。
本发明还提供一种针对植物病害的生物农药,所述生物农药的活性成分为上述的贝莱斯芽孢杆菌。
所述生物农药还包括农药制剂可接受的辅料和/或助剂。
在本发明一个优选的实施方式中,所述生物农药为包含1.0×108CFU/克以上的贝莱斯芽孢杆菌BJ-1的水分散粒剂;其中,贝莱斯芽孢杆菌BJ-1为权利要求1所述的贝莱斯芽孢杆菌;
优选的,所述水分散粒剂由下述组分组成:
贝莱斯芽孢杆菌BJ-1终浓度为1.0×108-1.0×1011,优选为2.0×1010;聚乙烯醇6.0%;十二烷基硫酸钠4.0%;硫酸铵4%;聚乙二醇4%;滑石粉补足至100%,所述百分含量为质量百分含量。
同时,本发明的贝莱斯芽孢杆菌菌株对由灰葡萄孢引起的番茄灰霉病在室内和田间均表现较好的防效。室内离体叶片接种试验结果表明,贝莱斯芽孢杆菌BJ-1发酵液对番茄灰霉病具有较好的保护和治疗作用,1×107CFU/mLBJ-1发酵液对番茄灰霉病的保护和治疗作用相对防效分别可达68.95%和88.28%。田间试验结果表明,贝莱斯芽孢杆菌BJ-1水分散粒剂对番茄灰霉病具有较好的防治效果,2.0×1010CFU/克贝莱斯芽孢杆菌BJ-1水分散粒剂处理(450克/亩)的相对防效可达80.48%。本发明的贝莱斯芽孢杆菌具有生防潜力,可用于制备针对植物病原真菌的生防菌剂和微生物菌肥。
本发明的贝莱斯芽孢杆菌培养条件简单、生长迅速且容易保存,适合用于工业生产。
附图说明
图1菌株BJ-1的菌落形态和扫描电镜照片。
图2菌株BJ-1的淀粉酶检测结果;
其中,上:大肠杆菌Escherichia coliJM109,左下:枯草芽孢杆菌BacillussubtilisIPEP6,右下:菌株BJ-1。
图3菌株BJ-1的蛋白酶检测结果;
其中,上:大肠杆菌Escherichia coliJM109,左下:枯草芽孢杆菌BacillussubtilisIPEP6,右下:菌株BJ-1。
图4菌株BJ-1的纤维素酶检测结果;
其中,上:大肠杆菌Escherichia coliJM109,左下:枯草芽孢杆菌BacillussubtilisIPEP6,右下:菌株BJ-1。
图5菌株BJ-1的脂肪酶的检测结果;
其中,上:大肠杆菌Escherichia coliJM109,左下:枯草芽孢杆菌BacillussubtilisIPEP6,右下:菌株BJ-1。
图6菌株BJ-1的葡萄糖发酵的检测结果;
其中,左:大肠杆菌Escherichia coliJM109,中:菌株BJ-1,右:枯草芽孢杆菌Bacillus subtilisIPEP6。
图7菌株BJ-1的蔗糖发酵的检测结果;
其中,左:大肠杆菌Escherichia coliJM109,中:菌株BJ-1,右:枯草芽孢杆菌Bacillus subtilisIPEP6。
图8菌株BJ-1的乳糖发酵的检测结果;
其中,左:大肠杆菌Escherichia coliJM109,中:菌株BJ-1,右:枯草芽孢杆菌Bacillus subtilisIPEP6。
图9菌株BJ-1的甘露醇发酵的检测结果;
其中,左:大肠杆菌Escherichia coliJM109,中:菌株BJ-1,右:枯草芽孢杆菌Bacillus subtilisIPEP6。
图10菌株BJ-1的肌醇发酵的检测结果;
其中,左:大肠杆菌Escherichia coliJM109,中:菌株BJ-1,右:枯草芽孢杆菌Bacillus subtilisIPEP6。
图11菌株BJ-1的革兰氏染色结果。
图12基于16SrDNA系统进化树。
生物材料保藏
名称:贝莱斯芽孢杆菌(Bacillus velezensis)BJ-1;
分类命名:贝莱斯芽孢杆菌Bacillus velezensis;
保藏日:2021年12月16日;
保藏单位:中国微生物菌种保藏管理委员会普通微生物中心(地址:中国北京);
保藏编号:CGMCC No.24113。
具体实施方式
实施例1.贝莱斯芽孢杆菌(Bacillus velezensis)BJ-1的获得和鉴定
采集自北京市海淀区,分离自表现葡萄枝干病害症状的葡萄病枝干组织,采用组织分离法进行病原菌分离,方法如下:(1)观察并记录葡萄枝干病样的症状;(2)对不同发病程度的部分每间隔10cm各切取2cm的枝干,去掉葡萄枝干病样的树皮,从枝干组织的病健交界处切出5mm2的组织块,用2%的次氯酸钠消毒2分钟,70%的乙醇消毒30s,无菌水洗3次,把组织块上放灭菌的滤纸晾干,待水分吸干之后把组织块放在PDA平板上,每个PDA平板放置4~5个组织块,封口膜封口;(3)于25℃,黑暗条件下培养3d,3d后观察、统计菌落生长情况并计算相应的分离比率;(4)挑取菌落边缘少量菌丝于新的PDA或其他易诱导产孢的培养基中,待产生分生孢子后进行单孢分离。在分离病原真菌的过程中,发现一株细菌对病原真菌具有很好的拮抗作用,将其在LB培养基平板划线纯化,获得细菌的纯培养物,经鉴定,获得了一株对病原菌具有拮抗作用的菌株,将其命名为BJ-1,菌株BJ-1的菌落形态如图1所示,菌落为圆形或近圆形,乳白色,边缘不光滑,表面有不明显褶皱。
1菌株BJ-1的生理生化特征
1.菌株的生理生化鉴定
1.1淀粉酶的检测
检测培养基:淀粉酶选择培养基(LB培养基+1.0%可溶性淀粉):蛋白胨10g,酵母膏5g,NaCl 5g,可溶性淀粉10g,琼脂20g,蒸馏水1000mL,pH7.8。
检测方法:取10μL摇培的待测菌株(目标菌株和对照菌株(大肠杆菌和枯草芽孢杆菌),对照菌株大肠杆菌不具有淀粉水解酶活性)滴加到检测平板上,晾干后28℃恒温培养24-48h,然后用稀碘液覆盖染色,清水冲洗,观察透明圈大小。
检测结果:如图2所示,可以看出,在目标菌株BJ-1的周围有透明圈产生,说明该菌株能产生淀粉酶,淀粉水解阳性。对照菌株大肠杆菌的周围无透明圈,不具有淀粉水解酶活性。
1.2蛋白酶检测
检测平板:含10%的脱脂牛奶水琼脂培养基(琼脂粉,1g/100mL水),选用三元脱脂牛奶,先微波炉加热,如果有脂肪膜则去掉,再用移液枪吸取10mL加到100mL已灭菌冷凉的水琼脂培养基内。
检测方法:将待测菌株在NA培养基中摇培过夜,取10μL滴加到检测平板,晾干后28℃恒温培养,12h后观察蛋白降解圈。
检测结果:如图3所示,从图中可以看出,在目标菌株BJ-1的周围有较明显的透明圈产生,说明该菌株能产生蛋白酶,蛋白酶水解阳性。对照菌株大肠杆菌的周围无透明圈,不具有蛋白酶活性。
1.3纤维素酶检测
检测平板:半LB培养基(碳源、氮源的量减半),加0.2%的羧甲基纤维素钠。
检测方法:将待测菌株和目标菌株在NA培养基中摇培过夜,取10μL滴加到检测平板上,晾干后于28℃恒温培养24h,洗掉菌落,加入1g/L刚果红溶液染色1h,然后用1MNaCl溶液浸泡洗涤2次进行脱色,每次浸泡30min,观察菌落周围有无纤维素水解圈。
检测结果:如图4所示,可以看出,在目标菌株BJ-1的周围能产生白色透明圈,说明该菌株能产生纤维素酶,纤维素酶水解阳性。对照菌株大肠杆菌的周围无透明圈,不具有纤维素水解酶活性。
1.4脂肪酶检测
检测培养基:油脂培养基:蛋白胨10g,牛肉膏5g,花生油或香油10g,1.6%中性红水溶液1mL,琼脂15g,蒸馏水1000mL,pH7.2。
检测方法:配置油脂培养基,灭菌后备用。倒平板前,将油脂培养基充分震荡使油脂均匀分布,再倒入培养皿中。平板凝固后将待测菌株滴加到平板上,晾干后28℃恒温培养24h。观察平板底层的菌落,如果出现红色斑点,即说明脂肪酸被水解,为阳性反应。
检测结果:如图5所示,可以看出,所有待测菌株的平板底层的菌落周围无红色斑点出现,说明它不能产生脂肪酶,脂肪酶水解阴性。
1.5糖醇发酵实验
检测培养基:糖发酵基础培养基:蛋白胨10g、NaCl5g、蒸馏水1000mL,调PH7.6,加入1.6%溴甲酚紫乙醇溶液。
检测方法:糖发酵基础培养基按照分别加入葡萄糖(1%)、乳糖(0.75%)、蔗糖(0.75%)、甘露醇(0.75%)、肌醇(0.75%),分装试管后,每管放入一杜氏发酵罐,将待测菌株分别接入含糖培养基中,以不接种的培养液作为对照,30℃培养7d,观察各试管颜色的变化及杜氏发酵罐中有无气泡产生。指示剂溴甲酚紫的指示范围为pH5.2(黄色)~6.8(紫色)。
检测结果:
葡萄糖发酵实验结果如图6所示,从图中可以看出,待测菌株BJ-1的试管培养基颜色均由紫色变为黄色,杜氏管中无气泡产生,说明待测菌株能分解葡萄糖,产酸不产气。
蔗糖发酵实验结果如图7所示,从图中可以看出,待测菌株BJ-1菌株的试管培养基颜色均有紫色变为黄色,杜氏管中无气泡产生,说明待测菌株均能分解蔗糖,产酸不产气。
乳糖发酵实验结果如图8所示,从图中可以看出,待测菌株BJ-1的试管培养基颜色都没有变化,杜氏管中无气泡产生,说明待测菌株不能分解乳糖。
甘露醇发酵实验结果如图9所示,从图中可以看出,待测菌株BJ-1的试管培养基颜色均由紫色变为黄色,杜氏管中无气泡产生,说明待测菌株能分解甘露醇,产酸不产气。
肌醇发酵实验结果如图10所示,从图中可以看出,待测菌株BJ-1的试管培养基颜色没有变化,杜氏管中无气泡产生,说明待测菌株不能分解肌醇。
2菌株的革兰氏染色
2.1材料与方法
染色液和试剂:结晶紫、卢戈氏碘液、95%酒精、番红。
染色方法:待测菌株和目标菌株在LB平板上培养48h,取干净载玻片一块,在载玻片的左、右各加一滴蒸馏水,按无菌操作法取菌涂片,左边涂待测菌,右边涂目标菌株,制成浓菌悬液。再取干净载玻片一块将刚制成的待测菌菌悬液挑1-2环涂在左边制成薄的涂片,将目标菌株的浓菌液取1-2环涂在右边制成薄涂面。让涂片自然晾干,手持玻片一端,让菌膜朝上,通过火焰2-3次固定。加适量(以盖满细菌涂面)的结晶紫染色液染色1min,倾去染色液,用水小心的冲洗。滴加卢戈氏碘液,媒染1min。用水洗去碘液,将玻片倾斜,连续点滴95%乙醇通过涂面涂色20-25s至流出液无色,立即水洗。滴加番红复染3-5min。用水洗去涂片上的番红染色液。将染色的涂片放空气中晾干。
镜检:先用低倍,再用高倍,最后用油镜观察,并判断菌体的革兰氏染色反应性。
2.2实验结果
革兰氏染色的结果如图11所示,菌株BJ-1的菌体均为短杆状,两端钝圆,经染色细胞为紫色,说明是革兰氏阳性菌。
3菌株BJ-1的分子鉴定
3.1基因组DNA的提取:
将保存的菌液涂平皿后,25℃培养3d后,用接种环刮下菌体后使用LB液体培养基洗出,采用北京博迈德基因技术有限公司细菌基因组DNA快速提取试剂盒提取细菌的基因组DNA。
3.2 16S rDNA的PCR扩增:
对BJ-1进行基因组DNA的提取,并以这株细菌基因组DNA为模板,选用细菌16SrDNA通用引物27f和1492r进行PCR扩增。
PCR扩增所用引物:
27f:5’-AGAGTTTGATCCTGGCTCAG-3’
1492r:5’-TACCTTGTTACGACTT-3’
PCR扩增所用反应体系:2×Taq PCR Mix 25μL,DNA模板(100ng/μL)2μL,上下游引物(10μmol/L)各1μL,用无菌ddH2O补足50μL体系。
PCR反应程序:94℃3min;94℃30s,54℃30s,72℃90s,35个循环;72℃10min;保温:4℃。
PCR扩增产物通过电泳检测,片段长度约为1500bp(序列表中序列1)。
3.3序列分析与分子鉴定
PCR产物经1%琼脂糖凝胶电泳验证后,送至北京博迈德基因技术有限公司测序。所获得序列与NCBI网站GenBank数据库中的序列进行比对,获得与测序16S rDNA序列相似度最高的菌株种类。结果显示,BJ-1菌株的与贝莱斯芽孢杆菌Bacillus velezensis BCRC17467T的16S rDNA序列相似度为99.85%。将其初步鉴定为贝莱斯芽孢杆菌近缘种群菌株。
BJ-1菌株的16S rDNA和gyrB基因PCR扩增片段测序结果分别如序列表中序列1和序列2所示。
根据基因片段序列,利用MEGA软件,基于16S rDNA序列利用ML方法构建系统发育树,如图12所示,结果表明菌株BJ-1与贝莱斯芽孢杆菌菌株AB78(GenBank登录号:MN100588.1)聚合到一起,据此,将菌株BJ-1鉴定为贝莱斯芽孢杆菌Bacillus velezensis。将菌株BJ-1保藏于中国微生物菌种保藏管理委员会普通微生物中心(地址:中国北京),保藏日期2021年12月16日,保藏编号为CGMCC No.24113。
实施例3.贝莱斯芽孢杆菌(Bacillus velezensis)BJ-1的抑菌活性测定
采用对峙培养法,以26种植物病原真菌的59个菌株(植物病原真菌菌株信息见表1,由北京市农林科学院植病综防研究室按照形态学及分子鉴定方法和柯赫氏法则进行鉴定)为靶标菌,测定菌株BJ-1的拮抗活性。
将BJ-1在LB固体培养基上25℃培养2~3d,后挑取单菌落于1mL的LB液体培养基内,37℃200rpm/min摇培24h,备用。将59个病原菌菌株在PDA平板上培养3~5d,用打孔器在菌落边缘区域打孔制成直径为5mm的病原菌菌饼,放置到PDA平板,同时吸取4μL培养好的菌悬液滴加在直径为6mm的滤纸片上,放置距平板边缘约1cm处,1个板放4个滤纸片,并以滤纸片上滴加同样剂量的清水作为对照,重复3次,28℃培养箱中培养5d,观察抑菌带的产生,测量相对内生拮抗细菌BJ-1方向病原菌菌丝生长的半径(D)和抑菌带宽度(d),计算抑菌率。
抑制率(%)=(对照菌落半径-处理菌落半径)/对照菌落半径×100%
表1.菌株BJ-1对59株植物病原真菌的抑菌活性
/>
通过对峙培养法,测得菌株BJ-1对26种植物病原真菌59个菌株的抑菌活性,测定结果如表1所示。由实验结果可知,菌株BJ-1对供试的不同植物病原真菌都具有明显的抑制作用,对菌丝生长的抑制率在53.76%-98.45%之间,对引起葡萄蔓枯病的大豆间座壳(Diaporthesojae)OSS①菌丝生长抑制效果最佳,抑制率可达98.45%。研究结果表明,BJ-1对常见果蔬植物病原菌的抑制作用有广谱性。
实施例4.离体叶片接种法测定菌株BJ-1对番茄灰霉病的防治作用
1实验方法
1.1病原菌制备
番茄灰霉病菌(Botrytis cinerea)菌株由北京市农林科学院植保所植病研究室保存,经过常规方法鉴定。挑取保存菌种转接到PDA平板上,25℃培养箱培养。
1.2药剂配制
生防菌BJ-1原粉以无菌水为溶剂配制成浓度分别为1×105个孢子/mL、1×106个孢子/mL、5×106个孢子/mL和1×107个孢子/mL四个浓度梯度备用。
1.3试验处理
1.3.1菌株BJ-1对番茄灰霉病的保护作用的测定
药剂处理采用喷雾法,将喉头喷雾器的喷头将菌液均匀喷施到番茄叶背面,以叶面刚好溢水为准。24h后用番茄灰霉菌菌饼(直径4mm)接种,使有菌丝的一面向下和叶片接种点贴合,叶柄处用湿润脱脂棉包裹,然后放入底部铺有双层滤纸的保鲜盒保湿,25℃培养3-5d后检查结果。以上每重复6-8个叶片,每处理3次重复,以清水和对照药剂处理(100亿CFU/克枯草芽孢杆菌可湿性粉剂)为对照。
1.3.2菌株BJ-1对番茄灰霉病的治疗作用的测定
治疗作用的测定是先接种病菌,接种后24h再用不同的菌液及无菌水处理接种的番茄叶片,其所用方法及材料与保护作用测定相同。
1.4数据统计分析
采用十字交叉法测量各处理果实的病斑直径,根据公式计算相对防效。计算公式如下:
2实验结果
通过离体叶片接种方法,测得菌株BJ-1对番茄灰霉病的防治效果,结果如表2所示。结果表明,四个浓度的菌株BJ-1菌悬液对番茄灰霉病都具有一定的保护作用,随着浓度的增加,保护和治疗效果逐步提升,其中浓度为1×107CFU/mL的BJ-1发酵液保护和治疗效果最佳,分别为68.95%和88.28%。对照药剂100亿CFU/克枯草芽孢杆菌可湿性粉剂(美国拜沃股份有限公司,农药登记证号:PD20160669)对番茄灰霉病的保护和治疗效果分别为49.19%和20.08%,显著低于菌株BJ-1的保护和治疗效果。因此菌株BJ-1对番茄灰霉病的防治效果显著优于对照药剂100亿CFU/克枯草芽孢杆菌可湿性粉剂。
表2菌株BJ-1对番茄灰霉病的防治效果(室内离体叶片接种)
实施例5.2.0×1010CFU/克贝莱斯芽孢杆菌BJ-1水分散粒剂对番茄灰霉病的田间防效测定
1.材料与方法
1.1试验药剂
供试药剂为2.0×1010CFU/克贝莱斯芽孢杆菌BJ-1水分散粒剂(BJ-1母粉终浓度为2.0×1010CFU/克;聚乙烯醇6.0%;十二烷基硫酸钠4.0%;硫酸铵4%;聚乙二醇4%;滑石粉补足至100%,其中百分含量为质量百分含量);对照药剂为1000亿孢子/克枯草芽孢杆菌可湿性粉剂(江西正邦作物保护股份有限公司,农药登记证号:PD20151587)。
1.2供试植物
试验作物为番茄,品种“京彩8”,生长期为结果期。
1.3试验方法
药剂处理信息如表3,每个小区面积为20m2,每个处理重复4次。在番茄灰霉发病初期进行喷雾施药。2022年12月29日第一次施药,每7天施药1次,共施药3次,于2023年1月20日进行病害防效调查。调查标准参照中华人民共和国农业行业标准,《农药田间药效试验准则(一):杀菌剂防治蔬菜灰霉病》(GB/T 17980.28-2000)进行。
表3试验药剂试验设计
/>
病情指数和防治效果计算方法如下述公式:
1.4数据统计与分析
利用SPSS 25.0软件对试验数据进行统计分析,运用邓肯氏新复极差法(DMRT)法对试验结果数据进行差异显著性检验。
2.结果
由田间调查结果可知,供试药剂2.0×1010CFU/克贝莱斯芽孢杆菌BJ-1水分散粒剂对番茄灰霉病有很好的防治效果,平均防治效果72.38%~80.48%。供试药剂2.0×1010CFU/克贝莱斯芽孢杆菌BJ-1水分散粒剂处理1(112.5克/亩)、处理2(225克/亩)、处理3(450克/亩)与对照药剂1000亿孢子/克枯草芽孢杆菌可湿性粉剂(70克/亩)药后平均防治效果分别为72.38%、76.05%、80.48%、66.48%。
方差分析结果表明:在0.05水平上,供试药剂2.0×1010CFU/克贝莱斯芽孢杆菌BJ-1水分散粒剂处理1(112.5克/亩)、处理2(225克/亩)、处理3(450克/亩)与对照药剂1000亿孢子/克枯草芽孢杆菌可湿性粉剂(70克/亩)之间防效存在显著性差异。在0.01水平上,供试药剂2.0×1010CFU/克贝莱斯芽孢杆菌BJ-1水分散粒剂处理1(112.5克/亩)、处理2(225克/亩)、处理3(450克/亩)与对照药剂1000亿孢子/克枯草芽孢杆菌可湿性粉剂(70克/亩)防效之间存在极显著性差异。因此,2.0×1010CFU/克贝莱斯芽孢杆菌BJ-1水分散粒剂可以作为一种防治番茄灰霉病的药剂使用,于番茄灰霉病发病前或发病初期进行喷雾施药,制剂推荐用量为225~450克/亩。
以上所述实施例仅展示了本发明的几种实施方式,其描述较为具体,但对本发明而言只是说明性的,而非限制性的。对于本领域普通技术人员来说,在不脱离所附权利要求所限定的精神和范围的情况下,可做出许多修改、变化和改进,这些都属于本发明的保护范围。
Claims (10)
1.一株贝莱斯芽孢杆菌,名称为贝莱斯芽孢杆菌(Bacillus velezensis)BJ-1,其特征在于,所述贝莱斯芽孢杆菌已保存于中国微生物菌种保藏管理委员会普通微生物中心,保藏编号为为CGMCC No.24113。
2.权利要求1所述的贝莱斯芽孢杆菌在制备抑制植物病原真菌的生防菌剂或微生物菌肥或生物农药中的应用。
3.根据权利要求2所述的应用,其特征在于,所述植物病原菌为可可毛色二孢(Lasiodiplodiatheobromae)、葡萄座腔菌(Botryosphaeria dothidea)、小新壳梭孢(Neofusicoccum parvum)、大豆间座壳(Diaporthesojae)、甜樱间座壳(Diaportheeres)、Diaporthehonkonggensis、灰葡萄孢(Botrytis cinerea)、Colletotrichum viniferum、Colletotrichum sojae、隐秘炭疽菌(Colletotrichum aenigma)、胶孢炭疽菌(Colletotrichum gloeosporioides)、果生炭疽菌(Colletotrichum fructicola)、暹罗炭疽菌(Colletotrichum siamense)、尖孢炭疽菌(Colletotrichum acutatum)、Neopestalotiopsis sp.、Neopestalotiopsisrosae、Coniellavitis、Dactylonectriaalcacerensis、Dactylonectriamacrodidyma、Lasiodiplodiapseudotheobromae、Dactylonectriatorresensis、尖孢镰刀菌(Fusariumoxysporum)、双核丝核菌融合群A、双核丝核菌融合群G、链格孢(Alternaria alternata)、枝状枝孢菌(Cladosporium cladosporioides)、核盘菌(Sclerotinia sclerotiorum)中的一种或多种。
4.一种针对植物病原菌的生防菌剂,其特征在于,所述生防菌剂的活性成分为权利要求1所述的贝莱斯芽孢杆菌。
5.根据权利要求4所述的生防菌剂,其特征在于,所述植物病原菌为可可毛色二孢(Lasiodiplodia theobromae)、葡萄座腔菌(Botryosphaeria dothidea)、小新壳梭孢(Neofusicoccum parvum)、大豆间座壳(Diaporthe sojae)、甜樱间座壳(Diaportheeres)、Diaporthe honkonggensis、灰葡萄孢(Botrytis cinerea)、Colletotrichumviniferum、Colletotrichum sojae、隐秘炭疽菌(Colletotrichum aenigma)、胶孢炭疽菌(Colletotrichum gloeosporioides)、果生炭疽菌(Colletotrichum fructicola)、暹罗炭疽菌(Colletotrichum siamense)、尖孢炭疽菌(Colletotrichum acutatum)、Neopestalotiopsis sp.、Neopestalotiopsis rosae、Coniella vitis、Dactylonectriaalcacerensis、Dactylonectria macrodidyma、Lasiodiplodiapseudotheobromae、Dactylonectria torresensis、尖孢镰刀菌(Fusarium oxysporum)、双核丝核菌融合群A、双核丝核菌融合群G、链格孢(Alternaria alternata)、枝状枝孢菌(Cladosporiumcladosporioides)、核盘菌(Sclerotinia sclerotiorum)中的一种或多种。
6.一种针对植物病害的微生物菌肥,其特征在于,所述微生物菌肥的活性成分为权利要求1所述的贝莱斯芽孢杆菌。
7.根据权利要求6所述的微生物菌肥,其特征在于,导致所述植物病害的植物病原菌为可可毛色二孢(Lasiodiplodia theobromae)、葡萄座腔菌(Botryosphaeria dothidea)、小新壳梭孢(Neofusicoccum parvum)、大豆间座壳(Diaporthe sojae)、甜樱间座壳(Diaporthe eres)、Diaporthe honkonggensis、灰葡萄孢(Botrytis cinerea)、Colletotrichum viniferum、Colletotrichum sojae、隐秘炭疽菌(Colletotrichumaenigma)、胶孢炭疽菌(Colletotrichum gloeosporioides)、果生炭疽菌(Colletotrichumfructicola)、暹罗炭疽菌(Colletotrichum siamense)、尖孢炭疽菌(Colletotrichumacutatum)、Neopestalotiopsis sp.、Neopestalotiopsis rosae、Coniella vitis、Dactylonectria alcacerensis、Dactylonectria macrodidyma、Lasiodiplodiapseudotheobromae、Dactylonectria torresensis、尖孢镰刀菌(Fusariumoxysporum)、双核丝核菌融合群A、双核丝核菌融合群G、链格孢(Alternaria alternata)、枝状枝孢菌(Cladosporium cladosporioides)、核盘菌(Sclerotinia sclerotiorum)中的一种或多种。
8.一种针对植物病害的生物农药,其特征在于,所述生物农药的活性成分为权利要求1所述的贝莱斯芽孢杆菌。
9.根据权利要求8所述的生物农药,其特征在于,所述生物农药还包括农药制剂可接受的辅料和/或助剂。
10.根据权利要求8所述的生物农药,其特征在于,所述生物农药为包含1.0×108CFU/克以上的贝莱斯芽孢杆菌BJ-1的水分散粒剂;其中,贝莱斯芽孢杆菌BJ-1为权利要求1所述的贝莱斯芽孢杆菌;
优选的,所述水分散粒剂由下述组分组成:
贝莱斯芽孢杆菌BJ-1终浓度为1.0×108-1.0×1011,优选为2.0×1010;聚乙烯醇6.0%;十二烷基硫酸钠4.0%;硫酸铵4%;聚乙二醇4%;滑石粉补足至100%,所述百分含量为质量百分含量。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202310482724.3A CN116445357A (zh) | 2023-05-02 | 2023-05-02 | 一株贝莱斯芽孢杆菌及其应用 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202310482724.3A CN116445357A (zh) | 2023-05-02 | 2023-05-02 | 一株贝莱斯芽孢杆菌及其应用 |
Publications (1)
Publication Number | Publication Date |
---|---|
CN116445357A true CN116445357A (zh) | 2023-07-18 |
Family
ID=87132058
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202310482724.3A Pending CN116445357A (zh) | 2023-05-02 | 2023-05-02 | 一株贝莱斯芽孢杆菌及其应用 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN116445357A (zh) |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN117264847A (zh) * | 2023-11-01 | 2023-12-22 | 山东省烟台市农业科学研究院(山东省农业科学院烟台市分院) | 贝莱斯芽孢杆菌ytq3、复配剂及其在提高植物抗低温冻害能力中的应用 |
-
2023
- 2023-05-02 CN CN202310482724.3A patent/CN116445357A/zh active Pending
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN117264847A (zh) * | 2023-11-01 | 2023-12-22 | 山东省烟台市农业科学研究院(山东省农业科学院烟台市分院) | 贝莱斯芽孢杆菌ytq3、复配剂及其在提高植物抗低温冻害能力中的应用 |
CN117264847B (zh) * | 2023-11-01 | 2024-04-26 | 山东省烟台市农业科学研究院(山东省农业科学院烟台市分院) | 贝莱斯芽孢杆菌ytq3、复配剂及其在提高植物抗低温冻害能力中的应用 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN107099467B (zh) | 一株铜绿假单胞菌xcs007及其在防治烟草黑胫病中的应用 | |
CN112899171B (zh) | 白囊耙齿菌ll210及其应用、生防菌剂 | |
CN108998389B (zh) | 对稻黄单胞菌和稻瘟病菌具有拮抗作用的假单胞菌及应用 | |
CN112899201B (zh) | 贝莱斯芽孢杆菌及其应用以及防治香蕉枯萎病的方法 | |
CN111534460B (zh) | 一株高效拮抗禾谷镰刀菌的贝莱斯芽孢杆菌的制备方法及应用 | |
CN101993836A (zh) | 枯草芽孢杆菌菌株yb-81、菌剂及其制备方法和应用 | |
CN109456921B (zh) | 一种多粘类芽孢杆菌、应用及微生物菌剂、粉剂和颗粒剂 | |
CN101967455A (zh) | 防治小麦根部病害的解淀粉芽孢杆菌ea19及其制剂 | |
CN113717901A (zh) | 一株贝莱斯芽孢杆菌及其在防治多种蔬菜病害中的应用 | |
CN111040976B (zh) | 一株解淀粉芽孢杆菌及其应用 | |
CN101148649A (zh) | 一种坚强芽孢杆菌及其菌剂和应用 | |
CN115058358B (zh) | 一株耐盐芽孢杆菌及其应用 | |
CN110184224B (zh) | 一株表皮葡萄球菌及其应用 | |
CN108641989A (zh) | 一株甲基营养型芽孢杆菌及其应用 | |
CN116445357A (zh) | 一株贝莱斯芽孢杆菌及其应用 | |
CN113604376B (zh) | 一株甘蔗内生枯草芽孢杆菌及其应用 | |
CN109321500B (zh) | 一株解淀粉芽孢杆菌菌株及其在防治油茶炭疽病害中的应用 | |
CN114437994A (zh) | 一种生物防治菌Bacillus siamensis HT1及其在制备生防菌剂中的应用 | |
CN108165506B (zh) | 一种金黄链霉菌及其应用 | |
CN112980721B (zh) | 一株贝莱斯芽孢杆菌及其生防制剂和应用 | |
CN112760267A (zh) | 一株拮抗野油菜黄单胞菌的产酶溶杆菌cx06及其应用 | |
CN116536207A (zh) | 一种萎缩芽孢杆菌wlkysy-4、生物菌剂及应用 | |
CN106635885A (zh) | 一种昆虫病原线虫共生菌及其应用 | |
CN116496922A (zh) | 一株解淀粉芽孢杆菌及其应用 | |
CN112725193B (zh) | 一株软毛木霉菌及其应用 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination |