CN111187727A - Preparation and application of bacterial liquid of bacillus methylotrophicus LW-6 for biological control of cotton fusarium wilt and verticillium wilt - Google Patents
Preparation and application of bacterial liquid of bacillus methylotrophicus LW-6 for biological control of cotton fusarium wilt and verticillium wilt Download PDFInfo
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- 239000007788 liquid Substances 0.000 title claims abstract description 58
- 241000563903 Bacillus velezensis Species 0.000 title claims abstract description 42
- 229920000742 Cotton Polymers 0.000 title claims abstract description 35
- 230000001580 bacterial effect Effects 0.000 title claims abstract description 22
- 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 title claims abstract description 18
- 241000223218 Fusarium Species 0.000 title claims abstract description 18
- 241000082085 Verticillium <Phyllachorales> Species 0.000 title claims abstract description 15
- 238000002360 preparation method Methods 0.000 title claims abstract description 14
- 238000000855 fermentation Methods 0.000 claims abstract description 46
- 230000004151 fermentation Effects 0.000 claims abstract description 46
- 238000011282 treatment Methods 0.000 claims abstract description 24
- 239000001963 growth medium Substances 0.000 claims abstract description 21
- 239000002068 microbial inoculum Substances 0.000 claims abstract description 17
- 238000003973 irrigation Methods 0.000 claims abstract description 13
- 230000002262 irrigation Effects 0.000 claims abstract description 13
- 238000002791 soaking Methods 0.000 claims abstract description 9
- 238000004321 preservation Methods 0.000 claims abstract description 7
- 244000005700 microbiome Species 0.000 claims abstract description 5
- 230000002906 microbiologic effect Effects 0.000 claims abstract 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 21
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 18
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 claims description 18
- 229920001817 Agar Polymers 0.000 claims description 10
- 229920002472 Starch Polymers 0.000 claims description 10
- 239000008272 agar Substances 0.000 claims description 10
- 238000002156 mixing Methods 0.000 claims description 10
- 239000008107 starch Substances 0.000 claims description 10
- 235000019698 starch Nutrition 0.000 claims description 10
- 239000001888 Peptone Substances 0.000 claims description 9
- 108010080698 Peptones Proteins 0.000 claims description 9
- 229940041514 candida albicans extract Drugs 0.000 claims description 9
- 229910052943 magnesium sulfate Inorganic materials 0.000 claims description 9
- 235000019341 magnesium sulphate Nutrition 0.000 claims description 9
- 235000019319 peptone Nutrition 0.000 claims description 9
- 229910000029 sodium carbonate Inorganic materials 0.000 claims description 9
- 238000000108 ultra-filtration Methods 0.000 claims description 9
- 239000012138 yeast extract Substances 0.000 claims description 9
- 241000193830 Bacillus <bacterium> Species 0.000 claims description 8
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 8
- 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 6
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- 239000000843 powder Substances 0.000 claims description 6
- 238000011218 seed culture Methods 0.000 claims description 6
- 238000009331 sowing Methods 0.000 claims description 6
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- 239000011780 sodium chloride Substances 0.000 claims description 4
- 229920002261 Corn starch Polymers 0.000 claims description 3
- 240000008042 Zea mays Species 0.000 claims description 3
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 claims description 3
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- 230000000712 assembly Effects 0.000 claims description 3
- 238000000429 assembly Methods 0.000 claims description 3
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- 239000008120 corn starch Substances 0.000 claims description 3
- 238000011081 inoculation Methods 0.000 claims description 3
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- 238000012827 research and development Methods 0.000 abstract 1
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- 241000894006 Bacteria Species 0.000 description 3
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- 206010059866 Drug resistance Diseases 0.000 description 2
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- 239000003795 chemical substances by application Substances 0.000 description 2
- 235000012343 cottonseed oil Nutrition 0.000 description 2
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- 238000010998 test method Methods 0.000 description 2
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- 239000013043 chemical agent Substances 0.000 description 1
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- 125000000118 dimethyl group Chemical group [H]C([H])([H])* 0.000 description 1
- 235000018905 epimedium Nutrition 0.000 description 1
- 239000003337 fertilizer Substances 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- 238000009629 microbiological culture Methods 0.000 description 1
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- 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
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Abstract
The invention relates to a preparation and application of a bacterial liquid of bacillus methylotrophicus LW-6 for biological control of cotton fusarium wilt, wherein the selected bacillus methylotrophicus strain source preservation mechanism is as follows: the China Committee for culture Collection of microorganisms general microbiological center Bacillus methylotrophicus, the preservation registration number is: CGMCC No.6793, the preparation of the microbial inoculum of the invention is under specific fermentation culture medium and process conditions, and mainly comprises screening stock seeds, determining starting strains, preparing liquid seed liquid, amplifying fermentation, ultrafiltering and concentrating, and preparing 200 hundred million live spore bacterial liquid/ml. The invention has the function of preventing and treating cotton wilt, and the prevention effect of preventing and treating the wilt by seed soaking treatment and drip irrigation treatment can reach 81.2 percent and the prevention effect of the wilt can reach 81.0 percent. The bacillus methylotrophicus LW-6 is used as a biological control microbial inoculum for cotton fusarium wilt and verticillium wilt, so that environmental pollution and chemical pesticide residue are isolated from the source, and a better control effect is achieved. Therefore, the invention has wide theoretical and time significance in the research and development aspects and the application aspects of new biological pesticides.
Description
Technical Field
The invention relates to preparation and application of a bacterial liquid of bacillus methylotrophicus LW-6 for biological control of cotton fusarium wilt and verticillium wilt, and belongs to the technical field of microorganisms or biological control.
Background
Cotton blight is one of the major diseases in the world, has been introduced into China from the introduction of American gossypium hirsutum in 1935, and has spread year by year. At present, the harm of blight and verticillium wilt becomes the first disease in cotton production, the disease belongs to root diseases with bacteria and spread in soil, the control is difficult, the occurrence of the disease seriously affects the yield and the quality of cotton, and the disease is one of the main obstacles for the sustainable production of cotton. At present, two mainstream methods for preventing and treating the disease exist at home and abroad, firstly, the disease-resistant variety is mainly planted, but the disease-resistant breeding progress is slow due to the lack of resistant germplasm resources, and a real effective disease-resistant variety is lacked in production; the second is chemical control, but the effect of chemical control measures for controlling the disease is very little, and the method brings serious environmental pollution and chemical pesticide residue.
Therefore, the bacillus methylotrophicus LW-6 bacterial liquid for biological control of cotton fusarium wilt is used for controlling cotton fusarium wilt in a biological control mode, beneficial microorganism bacillus methylotrophicus LW-6 is added under the condition of combining drip irrigation and water and fertilizer supplementation to control the disease, and compared with chemical agents and modes for planting disease-resistant varieties, the bacillus methylotrophicus LW-6 bacterial liquid has different advantages, and on the other hand, the biocontrol bacterial liquid disclosed by the invention is free of pollution to the environment and does not generate drug resistance.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides preparation and application of a bacterial liquid of bacillus methylotrophicus LW-6 for biological control of cotton fusarium wilt and verticillium wilt. The bacterial liquid has unique preparation process including stock seed screening, initial strain determination, liquid seed liquid preparation, amplification fermentation, ultrafiltering concentration, high live spore number of methyl nutritive bacillus LW-6 in the bacterial liquid up to 200 hundred million/ml, high biological preventing and treating effect on cotton wilt, capacity of reaching 81.2% and 81.0% through drip irrigation.
In order to achieve the purpose, the invention adopts the technical scheme that:
the preparation method comprises the following steps: preparation and application of a bacterial liquid of bacillus methylotrophicus LW-6 for biological control of cotton fusarium wilt and verticillium wilt:
(1) inoculating Bacillus methylotrophicus LW-6 stock seeds on an agar starch plate culture medium, culturing for 72 hours at 35 ℃, detecting under a microscope, and selecting large colonies with uniform formed central spores and clearly colored two ends as fermentation starting colonies;
(2) inoculating the fermentation starting bacterial colony in the step (1) to a liquid seed culture medium, wherein the liquid loading amount is as follows: shaking 500ml of the culture medium to fill 100ml of seed liquid, culturing at 35 ℃ and 200rpm for 72h to obtain fermented seed liquid;
(3) inoculating the fermentation seed liquid obtained in the step (2) into a fermentation culture medium according to the inoculation amount of 9%, and performing fermentation culture in a fermentation tank under the culture conditions that: the fermentation is terminated when the pressure in the fermentation tank is 0.07Mpa, the air volume is 1:0.8vvm, the rotating speed is 260rpm, the culture temperature is 35 ℃ and the number of live spores in the fermentation liquid in the fermentation tank reaches more than 200 hundred million/ml and the amount of spores is more than or equal to 92 percent.
(4) And (4) carrying out ultrafiltration concentration on the fermentation liquor obtained in the step (3) to remove 40% of water, wherein the fermentation liquor is characterized in that: : 3 ultrafiltration membrane assemblies with the aperture of 0.005um, 0.001um and 1nm are used in series, fermentation liquor is sequentially and simultaneously subjected to continuous ultrafiltration treatment at the temperature of 35 ℃ and the pressure of 0.3MPa for 4 hours, and the methylotrophic bacillus LW-6 liquid microbial inoculum for biological control of cotton fusarium wilt is obtained, wherein the number of live spores is more than or equal to 200 hundred million/ml.
According to the invention, the Bacillus methylotrophicus liquid in the step (1) is characterized in that the Bacillus methylotrophicus LW-6(Bacillus methylotrophicus) in the step (1) has a preservation unit as follows: CGMCC-China general microbiological culture Collection center; address: the institute of microbiology, national academy of sciences, university in the Chaoyang area, Beijing; the preservation number is: CGMCC NO. 6793; storage time: 11/07/2012, and survival was detected.
According to the invention, the bacillus methylotrophicus agent in the step (1) is characterized in that the components of the starch agar plate culture medium in the step (1) are as follows, and the components are all in percentage by weight: 15% of agar, 7% of starch, 1.8% of yeast extract powder, 01.3% of peptone, 0.1% of magnesium sulfate, 0.2% of sodium carbonate, 0.8% of sodium chloride and the balance of water, wherein the pH value is 7.2-7.3.
According to the invention, the bacillus methylotrophicus agent in the step (2) is characterized in that the liquid seed culture medium in the step (2) comprises the following components in percentage by weight: 1.0% of peptone, 2.6% of glucose, 1.3% of yeast extract, 0.8% of N aCl, 0.1% of magnesium sulfate, 0.2% of sodium carbonate and the balance of water, wherein the pH value is 7.0-7.1;
according to the invention, the fermentation medium in the step (3) preferably comprises the following components in percentage by weight: 0.7% of corn steep liquor, 0.2% of corn starch, 1.4% of bran powder, 0.8% of glucose, 0.5% of yeast extract, 0.2% of peptone, KH2PO40.005%, (NH4)2SO 40.009%, MnSO40.006%, CaCO 30.007%, MgSO4.7H20.008%, 0.1% of magnesium sulfate, 0.2% of sodium carbonate and the balance of water, wherein the pH value is 7.4-7.5.
Application of Bacillus methylotrophicus LW-6 bacterial liquid in preventing and treating cotton fusarium wilt and verticillium wilt.
The application comprises the following steps:
soaking a bacillus methylotrophicus LW-6 liquid microbial inoculum for biological control of cotton fusarium wilt for 3h before sowing, and mixing with water 1: 800 after 4 hours of mixing (after thorough mixing and filtration through a 200 mesh sieve), drip irrigation was carried out, repeated every 7 days for a total of 3 treatments.
Has the advantages that:
the bacillus methylotrophicus LW-6 for biological control of cotton fusarium wilt and verticillium wilt is prepared and applied, the preparation process, the culture medium formula and the fermentation process are unique, the number of viable spores of the prepared bacterial liquid is high, and the bacillus methylotrophicus LW-6 can be used for biological control of cotton fusarium wilt and verticillium wilt. The biological control has no pollution to the environment, is not easy to cause the drug resistance of pathogenic bacteria, is easier to obtain compared with the cultivation of disease-resistant varieties, and selects the methylotrophic bacillus LW-6 as the biocontrol bacterium because the bacterium has low nutritional requirement, strong stress resistance, fast growth and reproduction, no toxicity and harm to human and livestock, no environmental pollution, capability of generating various antibiotics and enzymes, broad-spectrum antibacterial activity, strong stress resistance and higher cotton wilt control effect.
Detailed Description
The technical solution of the present invention is further described with reference to the following examples, but the scope of the present invention is not limited thereto.
The bacillus methylotrophicus preservation registration number in the embodiment 1 is CGMCC No.6793, and the preservation organization is as follows: bacillus methylotrophicus LW-6 of common microorganism center of China Committee for culture Collection of microorganisms.
Example 1:
a bacillus methylotrophicus LW-6 liquid microbial inoculum is prepared by the following method:
(1) inoculating Bacillus methylotrophicus LW-6 stock seeds on an agar starch plate culture medium, culturing for 72 hours at 35 ℃, detecting under a microscope, and selecting large colonies with uniform formed central spores and clearly colored two ends as fermentation starting colonies;
(2) inoculating the fermentation starting bacterial colony in the step (1) to a liquid seed culture medium, wherein the liquid loading amount is as follows: shaking 500ml of the culture medium to fill 100ml of seed liquid, culturing at 35 ℃ and 200rpm for 72h to obtain fermented seed liquid;
(3) inoculating the fermentation seed liquid obtained in the step (2) into a fermentation culture medium according to the inoculation amount of 9%, and performing fermentation culture in a fermentation tank under the culture conditions that: the fermentation is terminated when the pressure of the fermentation tank is 0.07Mpa, the air volume is 1:0.8vvm, the rotating speed is 260rpm, the culture temperature is 35 ℃ and the number of live spores in the fermentation liquid in the fermentation tank reaches more than 200 hundred million/ml and the amount of spores is more than or equal to 92 percent,
(4) and (4) carrying out ultrafiltration concentration on the fermentation liquor obtained in the step (3) to remove 40% of water, wherein the fermentation liquor is characterized in that: : 3 ultrafiltration membrane assemblies with the aperture of 0.005um, 0.001um and 1nm are used in series, fermentation liquor is sequentially and simultaneously subjected to continuous ultrafiltration treatment at the temperature of 35 ℃ and the pressure of 0.3MPa for 4 hours, and the methylotrophic bacillus LW-6 liquid microbial inoculum for biological control of cotton fusarium wilt is obtained, wherein the number of live spores is more than or equal to 200 hundred million/ml.
The starch agar plate culture medium comprises the following components in percentage by mass: the components of the starch agar plate culture medium are as follows, and the weight percentages are as follows: 15% of agar, 7% of starch, 1.8% of yeast extract powder, 01.3% of peptone, 0.1% of magnesium sulfate, 0.2% of sodium carbonate, 0.8% of sodium chloride and the balance of water, wherein the pH value is 7.3.
The liquid seed culture medium comprises the following components in percentage by weight: peptone 1.0%, glucose 2.6%, yeast extract 1.3%, N aCl 0.8%, magnesium sulfate 0.1%, sodium carbonate 0.2%, and water in balance, wherein the pH value is 7.1;
the fermentation medium comprises the following components in percentage by weight: corn steep liquor 0.7%, corn starch 0.2%, bran powder 1.4%, glucose 0.8%, yeast extract 0.5%, peptone 0.2%, KH2PO40.005%, (NH4)2SO 40.009%, MnSO40.006%, CaCO 30.007%, MgSO4.7H20.008%, magnesium sulfate 0.1%, sodium carbonate 0.2%, and balance water, pH7.5
Example 2 prevention and treatment effects of Bacillus methylotrophicus LW-6 bacterial liquid on cotton verticillium wilt.
And (3) test treatment: and (3) test treatment: the bacillus methylotrophicus LW-6 microbial inoculum prepared in example (1) and water 1: 800, fully mixing, filtering by a 200-mesh filter screen, soaking the seeds for 3 hours before cotton sowing, and performing drip irrigation treatment with the microbial inoculum in a vigorous growing period (2 months after seedling emergence) after seedling emergence; blank control group: soaking seeds and drip irrigation with equal amount of clear water.
The test method comprises the following steps: selecting 2 test cells, selecting commercially available cotton seeds, performing seed soaking treatment before sowing, and after seedling emergence, entering a vigorous growth period (2 months after seedling emergence), preparing a bacillus methylotrophicus LW-6 liquid microbial inoculum prepared in the embodiment (1) and water 1: 800, filtering through a 200-mesh filter screen, and carrying out drip irrigation treatment once every 7 days, and repeating for three times. The control group was subjected to the same method. The test results are shown in the table I, and the control effect of the microbial inoculum on cotton verticillium wilt through drip irrigation can reach 81.2%.
Epimedium methylotrophic bacillus LW-6 control effect on cotton verticillium wilt
| Treatment group | Control group | |
| Number of plants sown | 1000 | 1000 |
| First survey and record the number of diseased plants | 30 | 120 |
| Second investigation and recording the number of diseased plants | 50 | 263 |
| The third investigation records the number of diseased plants | 93 | 495 |
| Ultimate incidence of disease | 9.30％ | 49.50％ |
| Final prevention and cure effect | 81.20％ |
(Note: the disease plant is not classified and recorded in unified manner)
The first table is attached with: grading standard of cotton verticillium wilt: level 0: cotton plants are healthy, have no diseased leaves and grow normally; level 1: less than one fourth of the leaves of the cotton plant develop disease, become yellow and wilting; and 2, stage: more than one fourth of the cotton plant and less than one half of the cotton plant have diseases, become yellow and wilted; and 3, level: more than one half of the cotton plant and less than three fourths of the cotton plant have diseases, become yellow and wilted; 4, level: more than three quarters of the leaves of the cotton plant develop disease, or the cotton plant dies.
Example 3 prevention and treatment effects of a Bacillus methylotrophicus LW-6 liquid microbial inoculum on cotton wilt.
1. And (3) test treatment: and (3) test treatment: mixing the liquid microbial inoculum of bacillus methylotrophicus LW-6 prepared in the example (1) with water 1: 800, fully mixing, filtering by a 200-mesh filter screen, soaking the seeds for 3 hours before cotton sowing, and performing drip irrigation treatment by adding the microbial inoculum after seedling emergence for a vigorous growing period (2 months after seedling emergence); blank control group: soaking seeds and drip irrigation with equal amount of clear water.
2. The test method comprises the following steps: selecting 2 test cells, selecting commercially available cotton seeds, performing seed soaking treatment before sowing, and after emergence, preparing bacillus methylotrophicus LW-6 bacterial liquid and water 1 by using the embodiment (1): the drip irrigation treatment with the microbial inoculum of the invention is carried out after 800-mesh mixing and filtering through a 200-mesh filter screen, the drip irrigation treatment is carried out once every 7 days, and the treatment is repeated for three times. The control group was subjected to the same method. The test results are shown in the second table, and the control effect of the bacillus methylotrophicus LW-6 liquid microbial inoculum on cotton wilt can reach 81.0 percent
Prevention and treatment effect of surface dimethyl nutritional bacillus LW-6 on cotton wilt
| Treatment group | Control group | |
| Number of plants sown | 1000 | 1000 |
| First survey and record the number of diseased plants | 38 | 149 |
| Second investigation and recording the number of diseased plants | 62 | 277 |
| The third investigation records the number of diseased plants | 102 | 524 |
| Ultimate incidence of disease | 10.20％ | 52.40％ |
| Final prevention and cure effect | 81.00％ |
And table II, supplementary notes, classification standard of cotton wilt: level 0: healthy plants have no symptom, and xylem of stems has no pathological changes; level 1: 25% of diseased plant leaves show typical symptoms, and the xylem lesion of the stem accounts for below a section 1/4; and 2, stage: 25-50% of diseased plant leaves show typical symptoms, and the xylem lesion of the stem accounts for sections 1/4-1/2; and 3, level: 50-90% of diseased plant leaves show typical symptoms, and the xylem lesion of the stem accounts for sections 1/2-3/4; 4, level: all the disease of the leaves of the diseased plants has symptoms, namely withered leaves or acute death, and the xylem lesion of the stalk accounts for more than a section 3/4.
Claims (7)
1. A preparation method and application of a bacterial liquid of bacillus methylotrophicus LW-6 for biological control of cotton fusarium wilt and verticillium wilt,
the preparation method is characterized by comprising the following steps:
(1) inoculating Bacillus methylotrophicus LW-6 stock seeds on an agar starch plate culture medium, culturing for 72 hours at 35 ℃, detecting under a microscope, and selecting large colonies with uniform formed central spores and clearly colored two ends as fermentation starting colonies;
(2) inoculating the fermentation starting bacterial colony in the step (1) to a liquid seed culture medium, wherein the liquid loading amount is as follows: shaking 500ml of the culture medium to fill 100ml of seed liquid, culturing at 35 ℃ and 200rpm for 72h to obtain fermented seed liquid;
(3) inoculating the fermentation seed liquid obtained in the step (2) into a fermentation culture medium according to the inoculation amount of 9%, and performing fermentation culture in a fermentation tank under the culture conditions that: the fermentation is terminated when the pressure of the fermentation tank is 0.07Mpa, the air volume is 1:0.8vvm, the rotating speed is 260rpm, the culture temperature is 35 ℃ and the number of live spores in the fermentation liquid in the fermentation tank reaches more than 200 hundred million/ml and the amount of spores is more than or equal to 92 percent,
(4) and (4) carrying out ultrafiltration concentration on the fermentation liquor obtained in the step (3) to remove 40% of water, wherein the fermentation liquor is characterized in that: : 3 ultrafiltration membrane assemblies with the aperture of 0.005um, 0.001um and 1nm are used in series, fermentation liquor is sequentially and simultaneously subjected to continuous ultrafiltration treatment at the temperature of 35 ℃ and the pressure of 0.3MPa for 4 hours, and the methylotrophic bacillus LW-6 liquid microbial inoculum for biological control of cotton fusarium wilt is obtained, wherein the number of live spores is more than or equal to 200 hundred million/ml.
2. The bacterial liquid of bacillus methylotrophicus LW-6 according to claim 1, wherein the bacillus methylotrophicus deposit accession number in step (1) is: CGMCC No.6793, the preservation organization is: bacillus methylotrophicus of the general microbiological center of China Committee for culture Collection of microorganisms.
3. The methylotrophic bacillus liquid as claimed in claim 1, wherein the agar starch plate culture medium in step (1) comprises the following components by weight percent: 15 percent of agar, 7 percent of starch, 1.8 percent of yeast extract powder, 01.3 percent of peptone, 0.1 percent of magnesium sulfate, 0.2 percent of sodium carbonate, 0.8 percent of sodium chloride and the balance of water, and the pH value is 7.2-7.3.
4. The bacillus methylotrophicus liquid according to claim 1, wherein the liquid seed culture medium in the step (2) comprises the following components in percentage by weight: peptone 1.0%, glucose 2.6%, yeast extract 1.3%, NaCl 0.8%, magnesium sulfate 0.1%, sodium carbonate 0.2%, and the balance of water, wherein the pH value is 7.0-7.1.
5. The bacillus methylotrophicus liquid according to claim 1, wherein the fermentation medium in the step (3) comprises the following components in percentage by weight: 0.7% of corn steep liquor, 0.2% of corn starch, 1.4% of bran powder, 0.8% of glucose, 0.5% of yeast extract, 0.2% of peptone, KH2PO40.005%, (NH4)2SO 40.009%, MnSO40.006%, CaCO 30.007%, MgSO4.7H20.008%, 0.1% of magnesium sulfate, 0.2% of sodium carbonate and the balance of water, wherein the pH value is 7.4-7.5.
6. The preparation and application of the Bacillus methylotrophicus LW-6 bacterial liquid as claimed in claim 1.
7. Use according to claim 6, characterized in that the steps are as follows:
soaking a bacillus methylotrophicus LW-6 liquid microbial inoculum for biological control of cotton fusarium wilt for 3h before sowing, and mixing with water 1: 800 after 4 hours of mixing (after thorough mixing and filtration through a 200 mesh sieve), drip irrigation was carried out, repeated every 7 days for a total of 3 treatments.
Priority Applications (1)
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