CN114214242B - Application of bacillus in soil-borne disease resistance and growth promotion of crops - Google Patents
Application of bacillus in soil-borne disease resistance and growth promotion of crops Download PDFInfo
- Publication number
- CN114214242B CN114214242B CN202111609289.3A CN202111609289A CN114214242B CN 114214242 B CN114214242 B CN 114214242B CN 202111609289 A CN202111609289 A CN 202111609289A CN 114214242 B CN114214242 B CN 114214242B
- Authority
- CN
- China
- Prior art keywords
- bacillus
- soil
- crops
- growth
- bacillus cereus
- 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
- 241000193830 Bacillus <bacterium> Species 0.000 title claims abstract description 18
- 230000012010 growth Effects 0.000 title claims abstract description 17
- 208000035240 Disease Resistance Diseases 0.000 title claims abstract description 8
- 240000003040 Solanum lycopersicum var. cerasiforme Species 0.000 claims abstract description 16
- 235000003953 Solanum lycopersicum var cerasiforme Nutrition 0.000 claims abstract description 13
- 201000010099 disease Diseases 0.000 claims abstract description 12
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims abstract description 12
- 235000007688 Lycopersicon esculentum Nutrition 0.000 claims description 4
- 241001660259 Cereus <cactus> Species 0.000 claims 1
- 241000227653 Lycopersicon Species 0.000 claims 1
- 241000196324 Embryophyta Species 0.000 abstract description 35
- 241000193755 Bacillus cereus Species 0.000 abstract description 29
- 244000052616 bacterial pathogen Species 0.000 abstract description 16
- 239000000589 Siderophore Substances 0.000 abstract description 13
- 230000000694 effects Effects 0.000 abstract description 13
- 238000004519 manufacturing process Methods 0.000 abstract description 7
- 230000008485 antagonism Effects 0.000 abstract description 6
- 238000004178 biological nitrogen fixation Methods 0.000 abstract description 4
- 230000007613 environmental effect Effects 0.000 abstract description 2
- 238000004321 preservation Methods 0.000 abstract description 2
- 230000000459 effect on growth Effects 0.000 abstract 1
- SEOVTRFCIGRIMH-UHFFFAOYSA-N indole-3-acetic acid Chemical compound C1=CC=C2C(CC(=O)O)=CNC2=C1 SEOVTRFCIGRIMH-UHFFFAOYSA-N 0.000 description 42
- 239000003617 indole-3-acetic acid Substances 0.000 description 21
- 239000002689 soil Substances 0.000 description 19
- 230000001580 bacterial effect Effects 0.000 description 17
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 14
- 239000007788 liquid Substances 0.000 description 14
- 230000008635 plant growth Effects 0.000 description 14
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 13
- 229910001385 heavy metal Inorganic materials 0.000 description 10
- 238000000034 method Methods 0.000 description 9
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 9
- ATNHDLDRLWWWCB-AENOIHSZSA-M chlorophyll a Chemical compound C1([C@@H](C(=O)OC)C(=O)C2=C3C)=C2N2C3=CC(C(CC)=C3C)=[N+]4C3=CC3=C(C=C)C(C)=C5N3[Mg-2]42[N+]2=C1[C@@H](CCC(=O)OC\C=C(/C)CCC[C@H](C)CCC[C@H](C)CCCC(C)C)[C@H](C)C2=C5 ATNHDLDRLWWWCB-AENOIHSZSA-M 0.000 description 7
- 239000001963 growth medium Substances 0.000 description 7
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 6
- 229930002875 chlorophyll Natural products 0.000 description 6
- 235000019804 chlorophyll Nutrition 0.000 description 6
- 229910052742 iron Inorganic materials 0.000 description 6
- 229910052761 rare earth metal Inorganic materials 0.000 description 6
- 239000006228 supernatant Substances 0.000 description 6
- 239000004570 mortar (masonry) Substances 0.000 description 5
- 230000001737 promoting effect Effects 0.000 description 5
- 238000011160 research Methods 0.000 description 5
- 238000012216 screening Methods 0.000 description 5
- 239000000243 solution Substances 0.000 description 5
- 239000000126 substance Substances 0.000 description 5
- 240000008067 Cucumis sativus Species 0.000 description 4
- 235000010799 Cucumis sativus var sativus Nutrition 0.000 description 4
- 241000209094 Oryza Species 0.000 description 4
- 235000007164 Oryza sativa Nutrition 0.000 description 4
- 241001544321 Thanatephorus cucumeris Species 0.000 description 4
- 239000011248 coating agent Substances 0.000 description 4
- 238000000576 coating method Methods 0.000 description 4
- 238000012258 culturing Methods 0.000 description 4
- 238000010790 dilution Methods 0.000 description 4
- 239000012895 dilution Substances 0.000 description 4
- 239000002609 medium Substances 0.000 description 4
- 239000002207 metabolite Substances 0.000 description 4
- 244000005700 microbiome Species 0.000 description 4
- -1 rare earth ions Chemical class 0.000 description 4
- 235000009566 rice Nutrition 0.000 description 4
- 239000007787 solid Substances 0.000 description 4
- 239000000725 suspension Substances 0.000 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 description 3
- 241000198596 Alternaria tomatophila Species 0.000 description 3
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 3
- 241000223218 Fusarium Species 0.000 description 3
- 239000012880 LB liquid culture medium Substances 0.000 description 3
- 240000003768 Solanum lycopersicum Species 0.000 description 3
- 238000002835 absorbance Methods 0.000 description 3
- 238000001816 cooling Methods 0.000 description 3
- 239000000706 filtrate Substances 0.000 description 3
- 230000002401 inhibitory effect Effects 0.000 description 3
- 230000001954 sterilising effect Effects 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 241000894006 Bacteria Species 0.000 description 2
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 2
- 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 2
- 244000061456 Solanum tuberosum Species 0.000 description 2
- 235000002595 Solanum tuberosum Nutrition 0.000 description 2
- 241001113787 Strychnos Species 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 239000011575 calcium Substances 0.000 description 2
- 235000021466 carotenoid Nutrition 0.000 description 2
- 150000001747 carotenoids Chemical class 0.000 description 2
- 230000032823 cell division Effects 0.000 description 2
- 239000012916 chromogenic reagent Substances 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- 238000000227 grinding Methods 0.000 description 2
- 150000002500 ions Chemical class 0.000 description 2
- 238000009630 liquid culture Methods 0.000 description 2
- 229910021645 metal ion Inorganic materials 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 235000015097 nutrients Nutrition 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 150000002910 rare earth metals Chemical class 0.000 description 2
- 238000005067 remediation Methods 0.000 description 2
- 230000008439 repair process Effects 0.000 description 2
- 238000002798 spectrophotometry method Methods 0.000 description 2
- 239000008223 sterile water Substances 0.000 description 2
- 231100000331 toxic Toxicity 0.000 description 2
- 230000002588 toxic effect Effects 0.000 description 2
- 238000005303 weighing Methods 0.000 description 2
- VHQALVHKNXDJRY-UHFFFAOYSA-N 1-bromo-2-chloro-4-fluoro-3-methylbenzene Chemical compound CC1=C(F)C=CC(Br)=C1Cl VHQALVHKNXDJRY-UHFFFAOYSA-N 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- 239000002028 Biomass Substances 0.000 description 1
- 230000004543 DNA replication Effects 0.000 description 1
- VTLYFUHAOXGGBS-UHFFFAOYSA-N Fe3+ Chemical compound [Fe+3] VTLYFUHAOXGGBS-UHFFFAOYSA-N 0.000 description 1
- QIVBCDIJIAJPQS-VIFPVBQESA-N L-tryptophane Chemical compound C1=CC=C2C(C[C@H](N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-VIFPVBQESA-N 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- 239000006004 Quartz sand Substances 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- QIVBCDIJIAJPQS-UHFFFAOYSA-N Tryptophan Natural products C1=CC=C2C(CC(N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-UHFFFAOYSA-N 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 238000007605 air drying Methods 0.000 description 1
- 229910021529 ammonia Inorganic materials 0.000 description 1
- 239000011324 bead Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000009835 boiling Methods 0.000 description 1
- 229910000019 calcium carbonate Inorganic materials 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 230000024245 cell differentiation Effects 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 229930002868 chlorophyll a Natural products 0.000 description 1
- 229930002869 chlorophyll b Natural products 0.000 description 1
- NSMUHPMZFPKNMZ-VBYMZDBQSA-M chlorophyll b Chemical compound C1([C@@H](C(=O)OC)C(=O)C2=C3C)=C2N2C3=CC(C(CC)=C3C=O)=[N+]4C3=CC3=C(C=C)C(C)=C5N3[Mg-2]42[N+]2=C1[C@@H](CCC(=O)OC\C=C(/C)CCC[C@H](C)CCC[C@H](C)CCCC(C)C)[C@H](C)C2=C5 NSMUHPMZFPKNMZ-VBYMZDBQSA-M 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 238000004040 coloring Methods 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000005520 cutting process Methods 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 229910001447 ferric ion Inorganic materials 0.000 description 1
- 230000035558 fertility Effects 0.000 description 1
- 239000003337 fertilizer Substances 0.000 description 1
- 238000011049 filling Methods 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 238000011065 in-situ storage Methods 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 229910052500 inorganic mineral Inorganic materials 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 239000011707 mineral Substances 0.000 description 1
- 238000005065 mining Methods 0.000 description 1
- 239000011259 mixed solution Substances 0.000 description 1
- 230000000877 morphologic effect Effects 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 239000000575 pesticide Substances 0.000 description 1
- 230000008121 plant development Effects 0.000 description 1
- 229910052573 porcelain Inorganic materials 0.000 description 1
- 235000012015 potatoes Nutrition 0.000 description 1
- 238000001243 protein synthesis Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 239000012088 reference solution Substances 0.000 description 1
- 239000007320 rich medium Substances 0.000 description 1
- 238000010008 shearing Methods 0.000 description 1
- 238000001179 sorption measurement Methods 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 230000002195 synergetic effect Effects 0.000 description 1
- 239000012085 test solution Substances 0.000 description 1
- 229910021654 trace metal Inorganic materials 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 230000014616 translation Effects 0.000 description 1
- 210000003462 vein Anatomy 0.000 description 1
- 238000004017 vitrification Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Classifications
-
- 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
Landscapes
- Life Sciences & Earth Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Health & Medical Sciences (AREA)
- Plant Pathology (AREA)
- Microbiology (AREA)
- Pest Control & Pesticides (AREA)
- Biotechnology (AREA)
- Virology (AREA)
- Agronomy & Crop Science (AREA)
- Dentistry (AREA)
- Wood Science & Technology (AREA)
- Environmental Sciences (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
The invention provides application of Bacillus to soil-borne disease resistance and growth promotion of crops, wherein the Bacillus is Bacillus cereus DW019 with a preservation number of GDMCC No:60778. the invention belongs to the technical field of biological environmental protection, and provides application of Bacillus in soil-borne disease resistance and growth promotion of crops, and the Bacillus cereus DW019 is green and pollution-free, has better antagonism to pathogenic bacteria of various diseases, has higher IAA production activity, can produce siderophores and biological nitrogen fixation, and has obvious promotion effect on growth of plants such as cherry tomatoes.
Description
Technical Field
The invention belongs to the technical field of biological environmental protection, and particularly relates to application of bacillus in soil-borne disease resistance and growth promotion of crops.
Background
The exploitation of ionic rare earth ores and the overuse of chemical fertilizers, pesticides and other human activities destroy soil properties, so that the serious pollution problem is caused. Soil is a bridge linking plants and nutrients, and damage to the soil reduces soil fertility and quality, affecting plant growth. Harmful substances such as heavy metals are enriched and toxic through a food chain, and the harmful substances form a great threat to food safety and human health. Researchers at home and abroad make various attempts to repair the heavy metal pollution of soil, such as a soil-foreign method, a vitrification method, an electric repair method and the like. The methods have the defects of high operation cost, easiness in causing soil degradation, secondary pollution, damage to soil biodiversity and the like to different degrees, so that the methods are not suitable for wide-range popularization and application, and are especially not suitable for popularization and application of agricultural lands. In contrast, the phytoremediation-microorganism synergistic remediation is a relatively simple, economical, effective and environment-friendly bioremediation method for heavy metal pollution of soil, has a great development prospect, and can be used for in-situ soil remediation.
The growth of plants and the relationship between rhizosphere and endophytic microorganisms are very close. Heavy metal stress can influence the uptake, enzyme activity, protein synthesis, DNA replication, cell division and the like of trace metal elements of soil microorganisms, so that the survival and community structure of the soil microorganisms are influenced. Research shows that even under the heavy metal environment with high concentration, stress-resistant plant growth promoting bacteria can grow and produce indoleacetic acid (IAA), and the IAA is very important for plant cell division and differentiation, and can promote plant growth, so that plants are promoted to produce larger biomass and better heavy metal absorption effect.
CN 110938565A discloses a Bacillus cereus DW019 with deposit number GDMCC No:60778 has ideal effect on removing and recovering 17 rare earth ions, and can be widely used for recovering rare earth ions and repairing rare earth ion pollution. Bacillus cereus DW019 has good adsorption effect on rare earth ions, but lacks related researches and reports on the influence of the DW on plant growth.
Disclosure of Invention
In order to solve the problems in the prior art, the invention further researches the property of Bacillus cereus DW019 and examines the influence of the Bacillus cereus DW on the growth of specific plants, and provides the application of Bacillus in soil-borne disease resistance and growth promotion of crops.
The purpose of the present invention will be further explained by the following detailed description.
The invention provides application of Bacillus to soil-borne disease resistance and growth promotion of crops, wherein the Bacillus is Bacillus cereus DW019 with a preservation number of GDMCC No:60778.
preferably, the crop comprises: rice, cucumber, tomato and cherry tomato.
Preferably, the soil-borne disease comprises: rice sheath blight, cucumber fusarium wilt and tomato early blight.
Compared with the prior art, the invention has the beneficial effects that: the invention further researches the property of Bacillus cereus DW019 and examines the influence of the Bacillus cereus DW on the growth of specific plants, and discovers that the Bacillus cereus DW019 has the effects of resisting soil-borne diseases and promoting the growth of crops; the compound has good antagonism to various pathogenic bacteria of soil-borne diseases, and can be used for preventing and inhibiting soil-borne diseases such as rice sheath blight disease (Thanatephorus cucumeris), cucumber fusarium wilt (Thanatephorus cucumeris), tomato early blight (Alternaria tomatophila) and the like; has obvious promoting effect on the growth of cherry tomato and other crops, and can produce indoleacetic acid, siderophore and NH 3 And the like to cooperatively promote the growth of crops.
Bacillus cereus DW019 not only has higher IAA production activity, but also can produce siderophores and biological nitrogen fixation. Siderophores can be chelated with ferric ions in iron-containing minerals or iron-containing organic matters to form soluble organic matters, and can also be chelated with other metal ions. Under the condition of high heavy metal ion concentration, the method can promote plants to produce more siderophores, the iron element is an element necessary for plant growth and development, and the chelated other metal ions can reduce the toxic action of heavy metal on Bacillus cereus (DW 019) to a certain extent. Heavy metal ions can inhibit IAA to a certain extent, however, the siderophores can relieve the stress of heavy metals on Bacillus cereus DW019 and promote the growth of plants. Biological nitrogen fixation systems can be reduced to ammonia-containing complexes by complex enzyme systems known as nitrogen fixation enzymes in nitrogen-fixing microorganisms and then absorbed and utilized by plants to promote plant growth.
Drawings
FIG. 1 is a graph showing the results of IAA production by 9 strains having IAA-producing ability.
Fig. 2 morphology recording of the plant just planted cherry tomato.
FIG. 3 morphology recordings of 3 groups of potted plants after 30d of cherry tomato planting with Bacillus cereus DW019.
FIG. 4 is a schematic diagram showing antagonism of Bacillus cereus DW019 against rice sheath blight disease (Thanatephorus cucumeris).
FIG. 5 is a schematic representation of antagonism of Bacillus cereus DW019 against cucumber fusarium wilt (Thanatephorus cucumeris).
FIG. 6 is a schematic representation of the antagonism of Bacillus cereus DW019 against early blight of tomato (Alternaria tomatophila).
Detailed Description
The invention will be described in further detail with reference to the drawings and examples.
The strain DW019 in the present invention refers to Bacillus cereus DW019. The media involved in the present invention are all conventional commercially available media or can be prepared according to conventional methods in the art. For example: the preparation method of the PDA liquid culture medium comprises the following steps: cutting peeled potato 200g into pieces, boiling for 30min at 100deg.C, filtering to obtain filtrate, glucose 20g, deionized water to 1000mL, and sterilizing under high pressure.
Example one screening of Bacillus and evaluation of the growth promoting Properties
The screening of bacillus includes the following steps:
1) Primary screening of bacillus: 10.0g of rhizosphere soil was weighed accurately using an analytical balance, added to a 250mL triangular flask with glass beads and 90mL of sterile water, and placed in a shaker for 10min at 150rpm and then allowed to stand for 10min. Taking 1mL of supernatant obtained after standing, placing in an EP tube, carrying out water bath at 80 ℃ for 10min, then placing in ice cubes, rapidly cooling to room temperature, sucking 100 mu L of supernatant by a pipette, coating onto LB solid medium, placing in a constant temperature incubator at 37 ℃, and allowing the strain to grow for 24h under the condition.
2) Purification of bacillus: the next day, the plate of the strain with better growth vigor and more single colonies in the step 1) is taken out from the incubator, and each time, one single colony growing on the culture medium is selected, and streaking is carried out on the LB solid culture medium. As the morphological characteristics and the distribution rules of different strains are different, it can be finally found that 31 strains are obtained from the soil of the plant root system area on the rare earth mining area through total screening, and all strains belong to bacillus. The culture was purified 3 times and numbered separately, and the culture was kept at-28℃with 30% sterile glycerol for further use.
The bacillus obtained by screening and purifying is respectively subjected to the function of producing indoleacetic acid (IAA) and the capability of producing iron carrier and NH 3 The capacity is measured by the following specific steps:
(1) Detection of indoleacetic acid (IAA): the strains to be tested were inoculated into LB liquid medium containing 0.5g/L tryptophan and shake-cultured (37 ℃ C., 150 rpm) for 36 hours, respectively, then 50. Mu.L of the suspension was added to a white porcelain plate, and 100. Mu.L of a chromogenic reagent (containing 4.5g/L FeCl) was added 3 、10.8mol/L H 2 SO 4 ) Shading treatment is carried out for 30min at normal temperature, color change is observed, and IAA is generated when the color becomes red. The culture broth was centrifuged at 8000rpm for 5min at 1:2, mixing the supernatant after centrifugation with a chromogenic reagent, coloring for 30min, and measuring the absorbance value at 550nm to see the amount of the indoleacetic acid from the quantitative angle. The results show that: among the screened bacillus strains, 9 strains with IAA production ability are provided, and the IAA production amount of the strains ranges from 3.65 mg/L to 20.27mg/L, as shown in figure 1, wherein the two strains of Y17-1 and DW019 produce relatively more indoleacetic acid, and the IAA production amounts of the strains reach 20.27mg/L and 16.06mg/L respectively.
(2) Iron producing carrier and NH producing 3 Measurement of Capacity: the strains to be tested were inoculated into a medium with no iron (KMB) and shake-cultured (37 ℃,150 rpm) for 24 hours, and centrifuged at 8000rpm for 5 minutes, and the supernatant was taken at 1:1 by volume, the removed supernatant was mixed with the CAS test solution, and if the color changed from blue to orange, this means that siderophores were produced. The quantity of siderophores produced can be determined by spectrophotometry of absorbance at 630nm using the iron-rich medium as a control group. The strains to be tested were inoculated into test tubes containing 10mL of peptone water medium, respectivelyIn the culture, concentration is 10g/L, shake culture (37 ℃,150 rpm) is carried out for 72 hours, 0.5mL Nessler's reagent is added into each tube, and the color changes from brown to yellow to indicate NH 3 And (3) generating. The results show that: 3 strains capable of producing siderophores and capable of producing NH 3 8 strains of (C), while only 3 strains capable of producing siderophores and ammonia gas, P3-1, SP1-3 and DW019, respectively. The inventor is surprised that the strain DW019 not only has higher IAA production activity, but also can produce siderophores and biological nitrogen fixation, and generally shows stronger growth promotion characteristics; while the strain Y17-1 has higher IAA-producing activity, but does not produce siderophores or NH 3 。
Example two Bacillus cereus DW019 effect determination on plant growth
From the net, 21 cherry tomato seedlings (the difference between the plants is small), 3 plants were selected for measuring the initial root length, the height of the plant, the diameter of the plant stem, the weight of the plant (not dried) and the amount of chlorophyll contained in the leaves. The specific method comprises the following steps: (1) length of root: measuring with a meter ruler with larger specification; (2) height of the plants: measuring with a meter ruler with larger specification; (3) diameter of plant stems: measuring by using a digital display vernier caliper; (4) weight of the plants: after the soil at the root of the plant is washed clean by water, the water on the surface is wiped off, and the plant is placed at a place without solar irradiation for natural air drying, and is measured by an analytical balance; (5) amount of chlorophyll contained in leaf: the determination method is a spectrophotometry method, and comprises the following specific operation steps: taking fresh cherry tomato leaves, cleaning, airing, removing veins in the leaves, shearing, placing a piece of weighing paper in a balance, peeling, placing the sheared leaves on the balance, accurately obtaining 0.2g of leaves, taking out the leaves together with the weighing paper, pouring the crushed leaves into a mortar, adding a small amount of calcium carbonate and quartz sand into the mortar by using a spoon, adding 2-3mL of 95% ethanol, fully grinding the mortar to be uniform liquid, adding 10mL of 95% ethanol, continuously grinding until tissues become white, standing for a few minutes, pouring all substances in the mortar into a funnel for filtering, using a clean volumetric flask to catch liquid below the funnel, washing chlorophyll in the mortar, filter paper and residues into the volumetric flask, finally fixing the volume to a scale mark, namely 25mL, covering a cover, and shaking repeatedly and uniformly; diluting the solution in the volumetric flask with 95% ethanol according to the ratio of 1:4, uniformly mixing, immediately pouring a proper amount of the uniformly mixed solution into a cuvette. The same volume of 95% ethanol is used as a reference solution, an ultraviolet spectrophotometer is used, zero is set on the basis of the control group, absorbance is measured at wavelengths of 6615 nm,649nm and 470nm respectively, after data are recorded, calculation can be carried out according to (2.1), (2.2), (2.3) and (2.4), and finally the chlorophyll content in plant leaves and carotenoid content in the leaves can be obtained.
Ca (chlorophyll a) =13.95A665nm-6.88A649 nm (2.1)
Cb (chlorophyll b) =24.69A649 nm-7.32A665nm (2.2)
Total chlorophyll=Ca+Cb (2.3)
Cx.c. (carotenoids) = (1000A 470nm-2.05Ca-114.8 Cb)/245 (2.4)
The rest 18 cherry tomato seedlings are planted in mine soil and randomly divided into 3 groups of 6 parallel. The morphology of the plant of the cherry tomato seedling just planted is shown in figure 2.
The first step of preparing bacterial liquid is to activate bacterial strain, adding sterile water into bacterial powder stored in vacuum freezing for dissolving, evenly stirring by means of a pipetting gun with proper specification, taking out a part of bacterial powder for dilution coating and streaking, putting the flat plate in a constant temperature incubator in an aligned manner, allowing the bacterial strain to grow for 12h in the environment, and storing the rest bacterial liquid at low temperature by means of glycerol. And inoculating, namely, randomly selecting single colonies growing on a flat plate by using an inoculating loop in an ultra-clean workbench which is sterilized in advance and is filled with sterile wind, inoculating the single colonies into an LB liquid culture medium prepared in advance, and culturing for 5 hours at 180rpm and 35 ℃. Finally, measuring the concentration of the bacterial liquid, carrying out gradient dilution on the bacterial strain solution cultured for 5 hours, and respectively adopting 10 -4 、10 -5 、10 -6 And 10 -7 The concentration of the strain solution is at the midpoint of each grid of the plate, and the concentration of the strain solution is 3 grids. And counting bacterial colonies in the next morning, and calculating the concentration of the bacterial liquid.
And (3) simultaneously adding 4mL of LB liquid culture medium, 2mL of LB liquid culture medium and bacterial liquid with the measured concentration, and 4mL of bacterial liquid with the measured concentration into each group of cherry tomato plants, and filling 1kg of collected and screened soil into each pot. The water quantity is 100mL for each pot of plant, the plant is watered twice in sunny days, the plant is watered once in rainy days or not, the plant is photographed and recorded during the planting period according to the specific situation, and the growth condition of the plant is cared for. After 30 days, the initial index was measured as described above, and the plant morphology was as shown in FIG. 3. To reduce the error, each index was measured three times and averaged.
The research result shows that the strain DW019 can greatly promote the growth of plants and has the concentration of 2.36 multiplied by 10 9 Individual/mL and 1.18X10 9 The cherry tomato plants of the DW019 strain with the volume per mL grow faster than the cherry tomato plants of the control group without the bacterial liquid and with the culture medium, have more flowers and have more branches and leaves which are visible to the naked eyes, especially when the bacterial adding amount is 4mL2.36 multiplied by 10 9 Under the condition of the bacterial liquid of each mL, the root of the plant is obviously longer than that of the plant of the control group, the plant grows higher to 33.13 cm, the plant grows to 29.72 cm, the stem is obviously thicker to 5.78mm, the chlorophyll content also reaches 10.347mg/g, the plant is extremely higher than that of the plant seedling without the bacterial liquid, the fresh weight reaches 13.050g, and the number of the results reaches 5. In total, 4mL was used at a concentration of 2.36X 10 9 The strain DW019 with the concentration of one/mL is added into the soil for planting cherry tomatoes, so that the absorption of cherry tomato seedlings to nutrient substances in the soil can be promoted, the soil environment is improved, and the growth of plants is greatly promoted. The cherry tomato plant growth condition added with the Y17-1 strain is not obviously different from the control plant growth condition without the Y17-1 strain. It can be seen that Y17-1 does not produce siderophores nor NH although it has a higher IAA-producing activity 3 Has no obvious promoting effect on the growth of plants.
Example antagonism study of Bacillus cereus DW019 with soil-borne pathogenic bacteria
The plate-facing effect of Bacillus cereus (DW 019) of the invention on various soil-borne disease pathogenic bacteria is as follows: the preparation method comprises the steps of activating various soil-borne disease pathogenic bacteria on a PDA solid culture medium (peeled potatoes are cut into pieces, boiled for 30min at 100 ℃, and filtered to obtain filtrate, 20g of glucose, 15g of agar and fixed volume to 1000 mL), and purifying and culturing twice by using the PDA liquid culture medium.
1) Bacteriostasis test of Bacillus cereus DW019 bacterial liquid on pathogenic bacteria
Respectively preparing 10 of various pathogenic bacteria of the strychnos disease -1 -10 -4 After 5d of dilution culture, operating in front of an alcohol lamp of a sterile operation table, taking 0.1ml of pathogenic bacteria suspension by using a liquid transfer device, adding the pathogenic bacteria suspension onto a PDA solid culture medium, sterilizing and cooling an inoculating rod, and uniformly coating the inoculating rod at the center of a flat plate and a position 1cm away from the flat plate; mu.L of DW019 bacteria were pipetted onto the plate (four weeks), 3 replicates were run for each concentration. Blank plates were used as controls. Covering the plate, and culturing in a culture box at 26 ℃ for 5 days; after pathogenic bacteria and Bacillus cereus DW019 grow out, the Bacillus cereus DW019 on the plate was observed to have inhibitory effect on various pathogenic bacteria. The schematic of the relevant inhibition is shown in figures 4-6.
2) Bacteriostasis test of Bacillus cereus DW019 metabolite on pathogenic bacteria
Respectively preparing 10 of various pathogenic bacteria of the strychnos disease -1 -10 -4 After dilution culture for 5d, centrifuging culture solution of Bacillus cereus DW019 at 8000rpm for 10min, collecting supernatant, collecting filtrate to obtain metabolite, operating in front of alcohol lamp, collecting 0.1ml pathogenic bacteria suspension with a pipette, adding onto PDA plate, sterilizing and cooling, and coating with inoculating rod at 1cm distance from the plate; mu.L of Bacillus cereus DW019 metabolites were pipetted onto plates (four weeks) and 3 replicates were run for each concentration. Blank plates were used as controls. Covering the plate, and culturing in a culture box at 26 ℃ for 5 days; after pathogenic bacteria and Bacillus cereus DW019 grow out, the metabolites of Bacillus cereus DW019 on the plate are observed to have inhibitory effect on various pathogenic bacteria.
The foregoing is a further detailed description of the invention in connection with the preferred embodiments, and it is not intended that the invention be limited to the specific embodiments described. It will be apparent to those skilled in the art that several simple deductions or substitutions may be made without departing from the spirit of the invention, and these should be considered to be within the scope of the invention.
Claims (1)
1. An application of bacillus in soil-borne disease resistance and growth promotion of crops, which is characterized in that: the bacillus is bacillus cereus @ andBacillus cereus) DW019, accession number GDMCC No:60778; the crops are cherry tomatoes; the soil-borne disease is tomato early blight.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111609289.3A CN114214242B (en) | 2021-12-27 | 2021-12-27 | Application of bacillus in soil-borne disease resistance and growth promotion of crops |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111609289.3A CN114214242B (en) | 2021-12-27 | 2021-12-27 | Application of bacillus in soil-borne disease resistance and growth promotion of crops |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN114214242A CN114214242A (en) | 2022-03-22 |
| CN114214242B true CN114214242B (en) | 2024-01-19 |
Family
ID=80705983
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202111609289.3A Active CN114214242B (en) | 2021-12-27 | 2021-12-27 | Application of bacillus in soil-borne disease resistance and growth promotion of crops |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN114214242B (en) |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102168051A (en) * | 2011-01-25 | 2011-08-31 | 浙江工业大学 | Bacillus cereus with higher resistance to jinggangmycin aqua and application |
| CN102321554A (en) * | 2011-08-30 | 2012-01-18 | 新疆农业科学院微生物应用研究所 | Bacillus cereus and its application of being as plant growth-promoting rhizobacteria |
| CN105296366A (en) * | 2015-11-27 | 2016-02-03 | 青岛农业大学 | Compound microbial agent capable of promoting tomato growth and development and application thereof |
| CN113373095A (en) * | 2021-07-19 | 2021-09-10 | 福建农林大学 | Application of bacillus cereus FJ2B-137 in biological control of rice sheath blight |
-
2021
- 2021-12-27 CN CN202111609289.3A patent/CN114214242B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102168051A (en) * | 2011-01-25 | 2011-08-31 | 浙江工业大学 | Bacillus cereus with higher resistance to jinggangmycin aqua and application |
| CN102321554A (en) * | 2011-08-30 | 2012-01-18 | 新疆农业科学院微生物应用研究所 | Bacillus cereus and its application of being as plant growth-promoting rhizobacteria |
| CN105296366A (en) * | 2015-11-27 | 2016-02-03 | 青岛农业大学 | Compound microbial agent capable of promoting tomato growth and development and application thereof |
| CN113373095A (en) * | 2021-07-19 | 2021-09-10 | 福建农林大学 | Application of bacillus cereus FJ2B-137 in biological control of rice sheath blight |
Also Published As
| Publication number | Publication date |
|---|---|
| CN114214242A (en) | 2022-03-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN109897806B (en) | Bacillus megaterium for promoting saline-alkali tolerant growth of crops, special microbial fertilizer for saline-alkali soil and application | |
| CN107488618B (en) | Bacillus megaterium H3 and application thereof in promoting crop growth and reducing heavy metal absorption of crops | |
| CN104962500B (en) | Phosphate-solubilizing bacteria and its separation and cultural method and application | |
| CN113755382B (en) | Bacillus aryabhattai NDFY-1 and application thereof | |
| CN111533585A (en) | Application of stenotrophomonas rhizophila in improvement of rhizosphere soil and promotion of plant growth | |
| CN110117560B (en) | Rhizosphere growth-promoting bacterium for enhancing salt tolerance of crops, microbial fertilizer and application thereof | |
| CN103658162A (en) | Method for continuously producing safe late japonica rice on farmland with soil mildly and moderately contaminated by soil heavy metals | |
| CN111484951B (en) | Bacillus for dissolving phosphorus and fixing nitrogen and application thereof in growth promotion | |
| CN104745517B (en) | One grows tobacco special molten B. mucilaginocus and its microbial inoculum | |
| CN109868242B (en) | Salt-tolerant acetoin-producing bacillus subtilis and application thereof | |
| CN113142057B (en) | Tissue culture propagation method of sphagnum fasciatum | |
| CN111484949A (en) | Heat-resistant bacillus for promoting growth, dissolving phosphorus and fixing nitrogen and application thereof | |
| CN114214242B (en) | Application of bacillus in soil-borne disease resistance and growth promotion of crops | |
| CN106399147B (en) | A kind of dedicated molten B. mucilaginocus of rape and its microbial inoculum | |
| CN102747006B (en) | Potassium decomposition bacteria, preparation method and application thereof | |
| Liu et al. | Study on the Effect of Kitchen Waste Compost Substrate on the Cultivation of Brassica chinensis L. | |
| CN109168387A (en) | A method of improvement degenerated soil is administered in combination using earthworm and organic materials | |
| CN107321774A (en) | The method of chickweed and ox chickweed to cadmium pollution soil repair ability is improved using miscegenation | |
| CN108559710B (en) | Seed coating agent for reducing arsenic absorption of crops and preparation method thereof | |
| Raphael et al. | The use of rice husk as soilless medium to grow capsicum Chinense: Effect of biochar and irrigation | |
| CN115024322B (en) | Application of monoethyl malonate in plant growth promotion | |
| CN115161217B (en) | Waste salt Tian Zhi bacillus H83 of salt-resistant degradable organic macromolecular substance and preparation and application of microbial inoculum thereof | |
| CN111567179B (en) | Method for promoting direct seeding germination of bletilla striata seeds and germination agent | |
| Toky et al. | Interaction between Rhizobium inoculation and nitrogen fertilizer application on growth and nodulation of Acacia nilotica subsp. indica | |
| CN118146993A (en) | Phosphate-dissolving selenium-dissolving bacteria and selenium-dissolving bacteria separation culture medium thereof |
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 | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |