CN111205999B - Bacillus amyloliquefaciens CGMCC No.17842 and application thereof - Google Patents
Bacillus amyloliquefaciens CGMCC No.17842 and application thereof Download PDFInfo
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Abstract
The invention relates to the field of microorganisms, and particularly relates to bacillus amyloliquefaciens and application thereof. The invention provides a Bacillus amyloliquefaciens, which is preserved in the China general microbiological culture Collection center of the Committee for culture Collection of microorganisms with the preservation number of CGMCC No. 17842. The bacillus amyloliquefaciens can be prepared into a microbial agent, can also be prepared into a compound microbial agent together with other microbial agents, and can also be prepared into a fertilizer for preventing and treating root rot.
Description
Technical Field
The invention relates to the field of microorganisms, in particular to bacillus amyloliquefaciens and application thereof, and specifically relates to a bacillus amyloliquefaciens (preservation number CGMCC No.17842), a microbial agent, a compound microbial agent, a fertilizer and a method for preventing and treating root rot.
Background
Strawberry root rot is an important disease of strawberry roots, and particularly can endanger the whole strawberry garden when the strawberry is planted in continuous cropping plots for many years. The occurrence of the disease has a trend of rising year by year, and has become one of the main obstacles for the development of the strawberry industry. With the growth of strawberry industry, continuous cropping diseases of strawberries are more and more prominent, and especially, strawberry root rot, pathogenic bacteria are complex and diverse and are difficult to control. The root rot pathogens reported in major strawberry producing areas worldwide are more than twenty, and the strawberry root rot disease is a soil-borne disease which is difficult to control.
At present, chemical control is mainly used for strawberry root rot in production, but medication is relatively disordered. The frequent use of chemical agents causes the drug resistance of germs and the pesticide residue phenomenon is serious. The strawberry root rot is one of the important pathogenic bacteria of the strawberry continuous cropping disease, and the occurrence is mainly caused by the deterioration of the soil micro-ecological environment, so that the strawberry root rot is solved by specifically developing a soil micro-ecological regulator starting from the regulation of the soil micro-ecological environment, and the method is a promising control measure.
However, microbial treatment against root rot requires further improvement.
Disclosure of Invention
The present invention is directed to solving, at least to some extent, one of the technical problems in the related art. Therefore, the invention aims to provide a Bacillus amyloliquefaciens, a microbial agent, a composite microbial agent and a method for preventing and treating root rot.
The bacillus amyloliquefaciens is a bacterium with high affinity with bacillus subtilis, has the capability of inhibiting plant diseases, can produce various metabolites and mainly comprises the following components: bacteriostatic proteins, lipopeptide proteins, iturin A, surface active substances, mustards, bacillus D, macrolides, oligopeptidases, peptides, polyketides and the like. The bacillus amyloliquefaciens has good inhibition effect on pathogenic fungi of various crops and can be prepared into biological agents for preventing and treating plant diseases, so the research on the biological characteristics and the production process of the bacillus amyloliquefaciens has very important significance. In the research, the common fusarium root rot pathogenic bacteria in the near-in area are used as target fungi for screening and identifying antagonistic bacteria, the research on the biocontrol bacteria of the strawberry root rot is carried out, strains with high antagonistic activity are screened, and the production application technology is explored. The screened bacillus amyloliquefaciens (with the number of CGMCC No.17842) can be used for preparing microbial agents, complex microbial agents, fertilizers and the like and is used for preventing and treating the root rot of plants.
Specifically, the invention provides the following technical scheme:
in a first aspect of the invention, the invention provides a bacillus amyloliquefaciens which is preserved in the China general microbiological culture Collection center with the preservation number of CGMCC No. 17842.
The Bacillus amyloliquefaciens provided by the invention is obtained by separating from the surface of a strawberry root system, is identified as Bacillus amyloliquefaciens (Bacillus amyloliquefaciens) through morphological, physiological and biochemical characteristics and 16S rRNA genes, and can be used for preventing and treating root rot through laboratory dish antagonistic experiments, strawberry potting experiments and field experiments, for example, the root rot of strawberry caused by fusarium can be prevented and treated, the hypha growth of pathogenic microorganism fusarium can be inhibited, the spore production is reduced, and the disease resistance requirement in the strawberry planting process is met. The provided bacillus amyloliquefaciens can be prepared into a microbial agent and can also be compounded with other microbial agents to prepare a compound microbial agent, and the microbial agent or the compound microbial agent can be used as a fertilizer or added into the fertilizer to prevent and treat root rot, so that the yield of strawberries is remarkably increased, and the quality is remarkably improved.
In a second aspect of the invention, the invention provides a microbial agent comprising a bacillus amyloliquefaciens according to the first aspect of the invention.
According to an embodiment of the present invention, the microbial agent described above may further include the following technical features:
in some embodiments of the present invention, the microbial agent is in a dry powder form, and the effective viable count of the bacillus amyloliquefaciens is 1 × 10 per gram of the microbial agent11CFU or more, preferably 4X 1011And (4) CFU. The microbial agent is prepared into dry powder, and the effective viable count contained in each gram of microbial agent is 1 multiplied by 1011CFU or above, e.g. 2X 1011CFU or more, preferably 3X 1011CFU or more, e.g. 4X 1011CFU, thereby allowing prevention of the use of the microbial agentAnd the root rot of the crops can be treated, the root rot of the crops can be quickly and efficiently treated, and the quality of the crops can be improved.
In a third aspect of the present invention, there is provided a method of preparing a microbial agent according to the second aspect of the present invention, the method comprising: fermenting the bacillus amyloliquefaciens; obtaining the microbial agent based on a product of the fermentation process. The microbial agent is obtained by fermenting the bacillus amyloliquefaciens, can be used for preventing and treating root rot of crops, and simultaneously improves the quality and the yield of the crops.
According to an embodiment of the present invention, the method for preparing a microbial agent described above may further include the following technical features:
in some embodiments of the invention, the fermentation process comprises: carrying out liquid fermentation culture on the bacillus amyloliquefaciens so as to obtain a fermentation culture solution; inoculating the fermentation culture solution into a solid fermentation culture medium for solid fermentation culture so as to obtain a product of the fermentation treatment.
In some embodiments of the invention, the solid fermentation medium comprises: 3-10 parts of wheat bran, 2-8 parts of corn flour, 0.5-5 parts of soybean meal, 0.5-5 parts of glucose, 0.05-0.5 part of magnesium sulfate, 0.05-0.5 part of sodium chloride and water. The solid fermentation culture medium can be used for rapid fermentation to obtain the microbial agent.
In some embodiments of the invention, the method further comprises: and drying and crushing the product of the fermentation treatment to obtain the powdery microbial agent. Through drying and crushing treatment, the powdery microbial agent can be obtained, and is convenient to store and use.
In a fourth aspect of the invention, the invention provides a complex microbial inoculant, which comprises a first microbial inoculant and a second microbial inoculant different from the first microbial inoculant, wherein the first microbial inoculant is the microbial inoculant provided in any one of the embodiments of the second aspect of the invention. Different microbial agents are compounded, different microbial agents can have different or same or similar functions, and the compound microbial agent can play the aims of preventing and treating various crop diseases and improve the quality of crops.
According to an embodiment of the present invention, the complex microbial inoculum described above may further include the following technical features:
in some embodiments of the invention, the second microbial agent comprises at least one selected from the group consisting of a bacillus mucilaginosus agent, a trichoderma viride agent, and an aspergillus terreus agent. The microbial agents have great promotion effect on plant growth and better control effect on pathogenic bacteria, so the microbial agents are compounded with the first microbial agent to further improve the quality of crops.
In some embodiments of the present invention, the complex microbial inoculant comprises 1-3 parts by weight of the first microbial inoculant, and at least one selected from the following: 1-3 parts by weight of the Bacillus mucilaginosus microbial inoculum; 1-3 parts by weight of the trichoderma viride agent; 1-3 parts by weight of the aspergillus terreus microbial inoculum.
In some embodiments of the invention, the complex microbial inoculant comprises 1-1.5 parts by weight of the first microbial inoculant, and at least one selected from the following components: 1-1.5 parts by weight of the Bacillus mucilaginosus microbial inoculum; 1-1.5 parts by weight of the trichoderma viride agent; 1-1.5 parts by weight of the aspergillus terreus microbial inoculum.
In a fifth aspect of the present invention, the present invention provides a fertilizer comprising the microbial agent of the second aspect of the present invention or the complex microbial agent of the fourth aspect of the present invention.
In some embodiments of the present invention, the fertilizer further includes a base fertilizer, the base fertilizer includes at least one selected from a bio-organic fertilizer, an organic compound fertilizer, an inorganic compound fertilizer, or an organic-inorganic compound fertilizer, and the content of the microbial agent or the compound microbial agent is 1% o to 5% o of the content of the base fertilizer.
In some embodiments of the invention, the bio-organic fertilizer comprises 30-50 parts by weight of fermented cow dung, 20-40 parts by weight of carbon-based fertilizer, 10-30 parts by weight of small molecule organic material, and 5-15 parts by weight of calcium silicate.
In some embodiments of the present invention, the bio-organic fertilizer comprises 40 parts by weight of fermented cow dung, 30 parts by weight of a carbon-based fertilizer and 20 parts by weight of a small molecule organic material, and 10 parts by weight of calcium silicate.
In a sixth aspect of the invention, the invention provides an application of bacillus amyloliquefaciens in preparation of a microbial agent, a composite microbial agent or a fertilizer, wherein the bacillus amyloliquefaciens is the bacillus amyloliquefaciens in the first aspect of the invention.
In a seventh aspect of the present invention, the present invention provides a method for controlling root rot, comprising: applying an effective amount of bacillus amyloliquefaciens, a microbial agent, a compound microbial agent or a fertilizer to crops, wherein the bacillus amyloliquefaciens is the bacillus amyloliquefaciens of the first aspect of the invention, the microbial agent is the microbial agent of the second aspect of the invention, the compound microbial agent is the compound microbial agent of the fourth aspect of the invention, and the fertilizer is the fertilizer of the fifth aspect of the invention.
In some embodiments of the invention, the crop is strawberry and the root rot is that caused by fusarium.
Information on strain preservation
Bacillus amyloliquefaciens with the preservation number of CGMCC No.17842, the preservation unit is the common microorganism center of China Committee for culture Collection of microorganisms, the preservation address is the microorganism research institute of China academy of sciences No. 3 of Xilu No.1 of Beijing republic of Chaoyang, and the preservation date is 2019, 05 and 22 days.
Drawings
FIG. 1 is a picture of the antagonistic effect of Bacillus amyloliquefaciens (CGMCC No.17842) on Fusarium on a plate according to an embodiment of the invention.
FIG. 2 is a plate shape picture of Bacillus amyloliquefaciens (CGMCC No.17842) on NA medium according to an embodiment of the invention.
FIG. 3 is a phylogenetic tree of the 16srDNA gene sequence of Bacillus amyloliquefaciens CGMCC No.17842 according to an embodiment of the invention.
Detailed Description
The embodiments of the present invention will be described in detail below with reference to the accompanying drawings, which are intended to be illustrative and not to be construed as limiting the invention.
Herein, when the content of a certain substance is expressed, the mass of the substance as a percentage of the total substance content is referred to, unless otherwise specified.
The bacillus amyloliquefaciens strain capable of preventing and treating various plant diseases is obtained through a large amount of screening work, and is preserved in the common microorganism center of China Committee for culture Collection of microorganisms with the preservation number of CGMCC No. 17842. The verification shows that the bacillus amyloliquefaciens shows a very high-efficient control effect in controlling the strawberry root rot, and by taking fusarium capable of causing the strawberry root rot as an example, the inhibition rate of the bacillus amyloliquefaciens on the fusarium is over 85 percent, so that the yield of the strawberries can be remarkably increased by applying the bacillus amyloliquefaciens in the strawberry planting process. The amylolytic bacillus strain can be used for preparing microbial fertilizers for preventing and treating various plant diseases. Therefore, the bacillus amyloliquefaciens has wide application prospect.
The provided bacillus amyloliquefaciens CGMCC No.17842 can be prepared into a microbial agent which is used as a single microbial agent. To this end, in one aspect of the present invention, the present invention provides a microbial agent comprising Bacillus amyloliquefaciens CGMCC No. 17842. The provided microbial agent can be used as a fertilizer or a disease treatment drug to be applied to the prevention and treatment of root rot, can prevent and treat plant diseases and insect pests, and improves the crop yield.
The provided microbial agent can be obtained by fermentation. To this end, in another aspect of the present invention, the present invention provides a method for preparing the above microbial agent, comprising: fermenting bacillus amyloliquefaciens CGMCC No. 17842; obtaining the microbial agent based on a product of the fermentation process. In at least some embodiments of the invention, the fermentation process comprises: carrying out liquid fermentation culture on the bacillus amyloliquefaciens so as to obtain a fermentation culture solution; inoculating the fermentation culture solution into a solid fermentation culture medium for solid fermentation culture so as to obtain a product of the fermentation treatment. In at least some embodiments, the liquid fermentation culture can be performed by using an LB culture solution, and the time of the liquid fermentation culture is more than 10 hours, preferably 10 to 16 hours.
In at least some embodiments of the present invention, the fermentation culture fluid is inoculated into the solid fermentation culture medium in an inoculation amount (the fermentation culture fluid accounts for 5-10% of the solid fermentation culture medium by mass) for fermentation culture. The solid fermentation medium comprises 3-10 parts by weight of wheat bran, 2-8 parts by weight of corn flour, 0.5-5 parts by weight of soybean meal, 0.5-5 parts by weight of glucose, 0.05-0.5 part by weight of magnesium sulfate, 0.05-0.5 part by weight of sodium chloride and water, wherein the content of water in the solid fermentation medium is 50-60%.
In some preferred embodiments, the solid fermentation medium comprises 4 parts by weight of wheat bran, 3 parts by weight of corn flour, 1 part by weight of soybean meal, 1 part by weight of glucose, 0.1 part by weight of magnesium sulfate, 0.1 part by weight of sodium chloride and water, wherein the content of water in the solid fermentation medium is 55%.
Drying and pulverizing the obtained fermentation product, wherein the viable count of Bacillus amyloliquefaciens in each gram of microbial agent is at least 1 × 1011CFU, preferably 2X 1011CFU or more, for example, 3X 1011CFU，4×1011CFU。
The bacillus amyloliquefaciens CGMCC No.17842 can be prepared into a microbial agent which can be used as a single microbial agent, and can be compounded with other microbial agents to prepare a composite microbial agent. To this end, in another aspect of the present invention, the present invention provides a complex microbial inoculant comprising a first microbial inoculant which is the microbial inoculant (i.e. the microbial inoculant comprising bacillus amyloliquefaciens CGMCC No.17842) described above and a second microbial inoculant which is different from the first microbial inoculant. The second microbial agent can be other bacillus amyloliquefaciens agents, or bacillus mucilaginosus agents, trichoderma viride agents, aspergillus terreus agents and the like, as long as the purpose of preventing and treating plant diseases and insect pests or diseases can be achieved. The bacillus mucilaginosus agent refers to a microbial agent containing bacillus mucilaginosus, the trichoderma viride agent refers to a microbial agent containing trichoderma viride, and similarly, other microbial agents are also understood similarly.
In the process of preparing the complex microbial inoculum, various microorganisms can be prepared into corresponding microbial agents under appropriate conditions, and then the corresponding microbial agents are compounded. For example, Bacillus mucilaginosus can be fermented under conditions similar to those of Bacillus amyloliquefaciens to produce a Bacillus mucilaginosus preparation. In at least some embodiments, the bacillus mucilaginosus agent is obtained by fermentation of: carrying out liquid fermentation culture on the bacillus mucilaginosus, and then inoculating a fermentation culture solution onto a first solid fermentation culture medium. In at least some embodiments, the first solid fermentation medium comprises: 3-10 parts of wheat bran, 2-8 parts of corn flour, 0.5-5 parts of soybean meal, 0.5-5 parts of glucose, 0.05-0.5 part of magnesium sulfate, 0.05-0.5 part of sodium chloride and water, wherein the content of water in the solid fermentation medium is 50-60%. In some preferred embodiments, the solid fermentation medium comprises 4 parts by weight of wheat bran, 3 parts by weight of corn flour, 1 part by weight of soybean meal, 1 part by weight of glucose, 0.1 part by weight of magnesium sulfate, 0.1 part by weight of sodium chloride and water, wherein the content of water in the solid fermentation medium is 55%. In some embodiments, the fermentation broth may be inoculated at an inoculum size of 5% to 10%.
The trichoderma viride and the aspergillus terreus can be fermented to prepare trichoderma viride or aspergillus terreus by adopting similar conditions. In at least some embodiments of the present invention, Trichoderma viride and Aspergillus terreus may be fermented separately in a PDA liquid broth, and the fermentation broth may then be inoculated into a second solid fermentation cultureAnd respectively obtaining fermentation products of aspergillus terreus and trichoderma viride. Among the useful second solid fermentation media are: 2-5 parts of wheat bran, 1-5 parts of corn starch, 0.5-3 parts of broken rice residues, 0.5-3 parts of millet, 0.3-1.5 parts of straw residues and water, wherein the content of the water in the solid fermentation medium is 55% -65%. In some preferred embodiments, the second solid fermentation medium comprises: 2 parts by weight of wheat bran, 1.5 parts by weight of corn starch, 1 part by weight of rice trash, 1 part by weight of millet, 0.5 part by weight of straw residue and water, wherein the content of water in the solid fermentation medium is 60%. Respectively drying and crushing the obtained fermentation products of the aspergillus terreus and the trichoderma viride to obtain powdery aspergillus terreus microbial inoculum and powdery trichoderma viride microbial inoculum, wherein each gram of the aspergillus terreus microbial inoculum contains 3 multiplied by 10 effective viable bacteria of the aspergillus terreus10Above, preferably 5 × 1010(ii) a The effective viable count of trichoderma viride contained in each gram of trichoderma viride is 2 multiplied by 1010Above, preferably 4 × 1010。
In at least some embodiments of the present invention, the present invention provides a complex microbial inoculum, which comprises the first microbial inoculum and at least one of the bacillus mucilaginosus microbial inoculum, trichoderma viride microbial inoculum and aspergillus terreus microbial inoculum. In some embodiments, the first microbial agent can be blended with any one or more of a bacillus mucilaginosus agent, a trichoderma viride agent and an aspergillus terreus agent according to a weight ratio of 1-3: 1-3. The obtained composite microbial inoculum contains more than 1000 hundred million viable bacteria per gram, preferably more than 2000 million viable bacteria per gram, such as 3000 million viable bacteria per gram.
In at least some embodiments of the invention, the complex microbial inoculant provided by the invention comprises 1-3 parts by weight of the first microbial inoculant, and at least one selected from the following components: 1-3 parts by weight of the Bacillus mucilaginosus microbial inoculum; 1-3 parts by weight of the trichoderma viride agent; 1-3 parts of the aspergillus terreus microbial inoculum. In other embodiments of the invention, the complex microbial inoculant provided by the invention comprises 1-1.5 parts by weight of the first microbial inoculant and is selected from the following groupsAt least one of: 1-1.5 parts by weight of the Bacillus mucilaginosus microbial inoculum; 1-1.5 parts by weight of the trichoderma viride agent; 1-1.5 parts by weight of the aspergillus terreus microbial inoculum. According to the embodiment of the invention, the effective viable count of the bacillus mucilaginosus contained in each gram of the bacillus mucilaginosus agent is 1 multiplied by 1011Above CFU; the effective viable count of trichoderma viride contained in each gram of trichoderma viride is 2 multiplied by 1010Above, preferably 4 × 1010(ii) a The effective viable count of the aspergillus terreus contained in each gram of aspergillus terreus microbial inoculum is 3 multiplied by 1010Above, preferably 5 × 1010。
The microbial agent or the compound microbial agent can be used as a fertilizer or a disease treatment drug independently, and can also be added into the fertilizer for use. For example, the microbial agent or the compound microbial agent can be added into an organic compound fertilizer, an organic-inorganic compound fertilizer, an inorganic compound fertilizer and a biological organic fertilizer, the addition amount can be more than one thousandth, and the microbial fertilizer can be obtained and used for plant growth promotion or root rot prevention and treatment. To this end, in a further aspect of the present invention, the present invention provides a fertilizer comprising the above-described microbial agent or complex microbial agent. In at least some embodiments, the fertilizer is produced to contain more than 1 billion, such as 1.2 billion, of viable bacteria per gram of fertilizer.
In at least some embodiments of the invention, the provided fertilizer comprises a bio-organic fertilizer and a microbial agent or a composite microbial agent, wherein the bio-organic fertilizer comprises 30-50 parts by weight of fermented cow dung, 20-40 parts by weight of a carbon-based fertilizer, 10-30 parts by weight of a small molecular organic material and 5-15 parts by weight of calcium silicate; wherein the content of the microbial agent or the composite microbial agent is one thousandth to five thousandth of the biological organic fertilizer. In some preferred embodiments of the present invention, the provided fertilizer comprises a bio-organic fertilizer, and the microbial agent or the complex microbial agent, the bio-organic fertilizer comprises 40 parts by weight of fermented cow dung, 30 parts by weight of carbon-based fertilizer, 20 parts by weight of small molecule organic material, and 10 parts by weight of calcium silicate, wherein the content of the microbial agent or the complex microbial agent is one thousandth to five thousandth of the content of the bio-organic fertilizer.
The scheme of the invention will be explained with reference to the examples. It will be appreciated by those skilled in the art that the following examples are illustrative of the invention only and should not be taken as limiting the scope of the invention. The examples, where specific techniques or conditions are not indicated, are to be construed according to the techniques or conditions described in the literature in the art or according to the product specifications. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products commercially available.
EXAMPLE 1 isolation and screening of strains
The bacillus amyloliquefaciens (with the number of CGMCC No.17842) is obtained by separating the surface of a strawberry root system by adopting a flat plate coating method and a flat plate marking method, and comprises the following steps:
(1) separation and screening of bacillus: selecting healthy strawberry plants from a field block with root rot disease in a Shandong Junan strawberry greenhouse, gently shaking off root soil, washing the root soil with clear water, placing the healthy strawberry plants on weighing paper, lightly scraping strawberry root systems with a blade, collecting and uniformly mixing all the plants, weighing 1g of the plants, placing the plants into 100ml of sterile water, oscillating the plants at 150rpm and 30 ℃ for 30min, performing gradient dilution, selecting 3 gradients for coating, enabling 3 gradients to be parallel to each other, culturing the plants in an incubator at 30 ℃ for 2d, selecting strains with different colony forms, scribing the strains on an NA culture medium, and regularly observing the growth condition of colonies. Then, the strains are purified by adopting a plate marking method, and are respectively numbered and stored.
(2) Screening of antagonistic strains of strawberry root rot pathogenic bacteria
Primary screening: preparing a PDA culture medium by adopting a plate confronting method, punching a fungus cake with the diameter of 5mm at the edge of strawberry root rot pathogenic bacteria (fusarium) by using a puncher, transplanting the fungus cake into the center of a plate, inoculating bacterial strains around the plate by using toothpicks, culturing at a constant temperature of 25 ℃, observing the inhibition effect of the bacterial strains on the pathogenic bacteria day by day, and screening the bacterial strains with high-efficiency antagonistic effect on the fusarium.
Re-screening: a microbial strain with antagonistic effect on fusarium and the best antagonistic effect is obtained by comparing and screening again, and is shown in figure 1.
Example 2 identification of Bacillus amyloliquefaciens
Example 2 the microbial strain obtained in example 1 was further identified as bacillus amyloliquefaciens. The microbiological and molecular biological characteristics are mainly identified, including:
(1) microbiological characteristics: the colony on the NA culture medium is round, milky white and yellowish, and has a rough and wrinkled surface, and the bacterial cells are rod-shaped and can move after being cultured on the NA culture medium for 2 days at 28 ℃. Gram-positive staining (E.coli control) resulted in the results shown in FIG. 2.
(2) Molecular biological Properties
The 16S rRNA sequencing result is shown in SEQ ID NO: 1:
ACGCTGGCGGCGTGCCTAATACATGCAAGTCGAGCGGACAGATGGGAGCT
TGCTCCCTGATGTTAGCGGCGGACGGGTGAGTAACACGTGGGTAACCTGC
CTGTAAGACTGGGATAACTCCGGGAAACCGGGGCTAATACCGGATGGTTG
TTTGAACCGCATGGTTCAGACATAAAAGGTGGCTTCGGCTACCACTTACA
GATGGACCCGCGGCGCATTAGCTAGTTGGTGAGGTAACGGCTCACCAAGG
CGACGATGCGTAGCCGACCTGAGAGGGTGATCGGCCACACTGGGACTGAG
ACACGGCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCGCAATG
GACGAAAGTCTGACGGAGCAACGCCGCGTGAGTGATGAAGGTTTTCGGAT
CGTAAAGCTCTGTTGTTAGGGAAGAACAAGTGCCGTTCAAATAGGGCGGC
ACCTTGACGGTACCTAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGC
CGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGAATTATTGGGCGTAAAG
GGCTCGCAGGCGGTTTCTTAAGTCTGATGTGAAAGCCCCCGGCTCAACC(SEQ ID NO:1)
the phylogenetic tree of the 16srDNA gene sequence of the strain is shown in FIG. 3.
The strain is identified as bacillus amyloliquefaciens, and is preserved with the preservation number of CGMCC No.17842, the preservation unit is the common microorganism center of China Committee for culture Collection of microorganisms, and the preservation address is the institute of microbiology of China academy of sciences No. 3, North West Lu No.1 Hopkin.
Example 3 anti-disease Effect of Bacillus amyloliquefaciens
1. Disease-resistant effect of plate
Preparing a PDA culture medium by adopting a plate confronting method, punching a fungus cake with the diameter of 5mm at the edge of strawberry root rot pathogenic bacteria (fusarium) by using a puncher, transplanting the fungus cake into the center of a plate, inoculating bacillus amyloliquefaciens strains around the plate by using toothpicks, comparing antagonistic effects of the bacillus amyloliquefaciens strains with exogenously purchased bacillus amyloliquefaciens (W1, W2 and W3 respectively), bacillus mucilaginosus (W4) and bacillus subtilis (W5 and W6 respectively), culturing the culture dish at the constant temperature of 25 ℃ after inoculation, and measuring the antagonistic rate.
The antagonism rate is calculated by the following formula, wherein the control colony diameter in the formula refers to the colony diameter of corresponding pathogenic bacteria treated by the PDA culture medium.
Antagonism (%) (control colony diameter-treated colony diameter)/control colony diameter x 100 (formula)
The measurement results are shown in table 1 below:
TABLE 1 antagonistic effect on Fusarium on plates
| Numbering | Treatment of | Antagonism rate |
| CK (blank control) | PDA culture medium | / |
| T1 | Bacillus amyloliquefaciens (CGMCC No.17842) | 86％ |
| T2 | Bacillus amyloliquefaciens purchased outside (W1) | 60％ |
| T3 | Bacillus amyloliquefaciens purchased outside (W2) | / |
| T4 | Bacillus amyloliquefaciens purchased outside (W3) | / |
| T5 | Bacillus mucilaginosus (W4) | / |
| T6 | Bacillus subtilis (W5) | 50％ |
| T7 | Bacillus subtilis (W6) | 40％ |
As can be seen from Table 1, aiming at the plate antagonistic effect of fusarium which is a pathogenic bacterium of strawberry root rot, the antagonistic rate of the bacillus amyloliquefaciens provided by the invention on the fusarium which is a pathogenic bacterium of strawberry root rot is improved by 26% compared with that of the bacillus amyloliquefaciens purchased from outsourcing. Compared with the antagonism rate of the bacillus subtilis, the antagonism rate of the bacillus subtilis also has 36 to 44 percent improvement.
2. Disease-resistant effect of pot culture test
The disease treatment effect of different microbial strains is verified by the following method, which comprises the following steps:
(1) preparation of a suspension of pathogenic spores. Respectively inoculating fusarium on a PDA (personal digital assistant) plate, culturing for 5-7d in a constant-temperature incubator at 25 ℃, washing off spores on the plate by using sterile water after the strain grows over the whole plate, centrifuging, collecting pathogenic bacteria spores, counting under a microscope by using a blood counting plate, and diluting spore suspension to 120 spores/ml for later use.
(2) Preparing an antagonistic bacteria suspension. Inoculating antagonistic bacteria (including Bacillus amyloliquefaciens, Bacillus mucilaginosus and Bacillus subtilis) from a plate into a 100ml LB liquid shake flask, fermenting at 30 deg.C, pH6.8, and rotating at 140rpm, and culturing for 24 hr. Adjusting the concentration of the bacterial suspension to 10 after 24 hours8cfu/ml, used for treatment of experimental groups.
(3) And (5) disease resistance test. The test is carried out in a test greenhouse of the research and development center limited company of the Linyi city of Shandong province, the tests are divided into a control group 1, a control group 2 and an experimental group 1-7, each group comprises 20 pots, each pot is planted with a healthy strawberry plant with consistent growth and no leaf spots, the pot is 30cm multiplied by 20cm multiplied by 30cm (the diameter of the pot mouth is multiplied by the diameter of the pot bottom is multiplied by the height of the pot), and 3.5kg of sterilized soil is filled in the pot.
Wherein different bacterial suspensions of the experiment groups 1 to 7 are respectively inoculated into root soil of each pot plant 15 days after seedling revival, pathogenic bacteria spore suspension (the concentration is 100cfu/ml) is inoculated after 48 hours, the test period is 30 days (calculated from 15 days after seedling revival), the watering amount of each treatment is the same every day, and the management is unified. Wherein the experimental group 1 is accessed with bacillus amyloliquefaciens (with the number of CGMCC No.17842) bacterial suspension, the experimental groups 2 to 4 are respectively accessed with bacillus amyloliquefaciens exogenously (W1) bacterial suspension, bacillus amyloliquefaciens exogenously (W2) bacterial suspension and bacillus amyloliquefaciens exogenously (W3) bacterial suspension, the experimental group 5 is accessed with bacillus mucilaginosus exogenously (W4) bacterial suspension, and the experimental group 6 and the experimental group 7 are respectively accessed with bacillus subtilis exogenously (W5) bacterial suspension and bacillus subtilis exogenously (W6) bacterial suspension.
In addition, control 1 was not treated at all; control group 2 received only diseaseA suspension of original bacteria spores with a concentration of 106cfu/ml. The incidence was calculated with reference to the following methods, respectively.
Dividing the root rot disease into 6 grades, wherein 0 grade is that the root system is not attacked; grade 1 is that the incidence of root system is less than or equal to 30 percent and the leaves are normal; grade 2 is 30%, the incidence rate of roots is less than or equal to 60%, and the leaves are normal; grade 3 is 60%, the incidence rate of roots is less than or equal to 80%, and leaves turn yellow; grade 4 is that the incidence rate of root systems is more than 80 percent, and leaves wither; grade 5 indicates death of the whole plant and dry leaves.
Disease incidence (%) (∑ (disease grade × number of plants at that grade)/(total number of plants × highest grade value of disease) × 100
The specific experimental results are shown in table 2 below:
TABLE 2 antagonistic Effect of the potting
| Numbering | Treatment of | Incidence of disease |
| Control group 1 | Blank space | 5％ |
| Control group 2 | Pathogenic bacteria (fusarium) spore suspension | 88％ |
| Experimental group 1 | Bacillus amyloliquefaciens (number CGMCC No.17842) | 10％ |
| Experimental group 2 | Bacillus amyloliquefaciens purchased outside (W1) | 25％ |
| Experimental group 3 | Bacillus amyloliquefaciens purchased outside (W2) | 35％ |
| Experimental group 4 | Bacillus amyloliquefaciens purchased outside (W3) | 43％ |
| Experimental group 5 | Bacillus mucilaginosus (W4) | 33％ |
| Experimental group 6 | Bacillus subtilis (W5) | 55％ |
| Experimental group 7 | Bacillus subtilis (W6) | 57％ |
The result shows that in the sterilized soil, the bacillus amyloliquefaciens provided by the invention has obvious disease prevention effect on the strawberry root rot of pathogenic fusarium, the morbidity is only 10%, and the disease index is obviously reduced compared with the morbidity of 25-43% of the purchased bacillus amyloliquefaciens.
Example 4 preparation of Bacillus amyloliquefaciens inoculum
The method for preparing the microbial inoculum by utilizing the bacillus amyloliquefaciens (with the number of CGMCC No.17842) provided by the invention comprises the following steps:
(1) inoculating Bacillus amyloliquefaciens (with the number of CGMCC No.17842) into a 100ml LB liquid shake flask for independent fermentation at the temperature of 30 ℃, the pH value of 6.8 and the rotation speed of 140rpm, and culturing for 12 hours to obtain fermentation liquor.
And then, transferring the fermentation liquor onto a solid fermentation culture medium A, wherein the inoculation amount is 5% (namely the mass of the inoculated fermentation liquor accounts for 5% of the mass of the solid fermentation culture medium), and fermenting for 5 days at 30 ℃ to obtain a fermentation product. Wherein the formula of the solid fermentation medium A comprises the following components in parts by weight: corn flour: bean pulp: glucose: magnesium sulfate: sodium chloride was 4:3:1:1:0.1:0.1, water content was 55%.
(2) And drying and crushing the fermentation product to obtain the microbial agent for preventing and treating strawberry root rot pathogenic bacteria. The effective viable count of the contained bacillus amyloliquefaciens GGMCCNo.17842 is 3 multiplied by 1011CFU/g。
The method for verifying the field disease-resistant effect of the prepared microbial inoculum comprises the following steps:
the test is carried out in the Shandong province Linyi city, 1 strawberry greenhouse is selected, 2 mu is selected, and a control group 1, a control group 2, a control group 3 and a test group are respectively arranged, wherein the control group 1 does not use a microbial inoculum (namely a blank group), the control group 2 uses a market-purchased microbial inoculum (S1), the control group 3 uses a market-purchased microbial inoculum (S2), and the test group uses the prepared microbial inoculum. Other field management is the same. The results are shown in Table 3 below. Wherein fresh weight (g/plant) represents the weight of strawberry plants; the number of results (number/plant) represents the number of strawberries knotted per plant; yield (g/plant) represents the weight of strawberry fruit; sweetness (%) was obtained by detection with a brix meter. In the following examples, the fresh weight, the number of results, the yield, the sweetness and the like have the same meanings and the same calculation methods as those of the examples, and the sweetness is measured by a brix meter.
TABLE 3 strawberry greenhouse test results
Compared with a control group, the microbial inoculum provided by the invention has the advantages that the physiological indexes of strawberry plants are improved, fruits are full, malformed fruits are few, the taste is sweet, the mouthfeel is good, the morbidity is only 8%, the morbidity is obviously reduced compared with an outsourcing market microbial inoculum, and the sweetness is also obviously increased compared with the outsourcing market microbial inoculum.
Example 5 Complex microbial Agents
The embodiment provides a compound microbial inoculum, which is obtained by mixing different microbial inocula according to a certain proportion. The bacillus amyloliquefaciens CGMCC No.17842 microbial inoculum and at least one of bacillus mucilaginosus microbial inoculum, trichoderma viride microbial inoculum and aspergillus terreus microbial inoculum are mixed according to the weight ratio of 1-3: 1-3: 1-3: 1-3, preferably, the bacillus amyloliquefaciens CGMCC No.17842 microbial inoculum and at least one of bacillus mucilaginosus microbial inoculum, trichoderma viride microbial inoculum and aspergillus terreus microbial inoculum are mixed according to the mass ratio of 1-1.5: 1-1.5: 1-1.5: 1 to 1.5 by mass. Is prepared by the following steps:
1. respectively preparing bacillus amyloliquefaciens (with the number of CGMCC No.17842) and bacillus mucilaginosus microbial inoculum
Respectively inoculating bacillus amyloliquefaciens and bacillus mucilaginosus (W4) into a 100ml LB liquid shake flask for independent fermentation, wherein the temperature is 30 ℃, the pH value is 6.8, the rotating speed is 140rpm, and the culture is carried out for 18 hours. Then transferring the mixture to a solid fermentation medium A, wherein the inoculation amount is 5%, fermenting for 7 days at 30 ℃, and the formula of the solid fermentation medium A comprises the following components in parts by weight: corn flour: bean pulp: glucose: magnesium sulfate: sodium chloride 4:3:1:1:0.1:0.1, water content 55%. Drying and crushing the fermentation product to obtain powdery bacillus amyloliquefaciens microbial inoculum and powdery bacillus mucilaginosus microbial inoculum. Wherein each gram of bacillus amyloliquefaciens microbial inoculum contains bacillus amyloliquefaciens with the effective viable count of 3 multiplied by 1011CFU, the effective viable count of the bacillus mucilaginosus contained in each gram of bacillus mucilaginosus agent is 1 multiplied by 1011CFU。
2. Respectively preparing trichoderma viride and aspergillus terreus
Separately fermenting Aspergillus terreus and Trichoderma viride in 100ml PDA liquid shake flask at 27 deg.C and rotation speed of 120rpm, and culturing for 28 hr to obtain mycelium pellet. Then inoculating the mycelium pellets on a solid fermentation medium B respectively, wherein the inoculation amount is 8 percent, fermenting at 30 ℃, and illuminating for 2 percentCulturing in dark for 24 hours after 4 hours, and continuously fermenting for 9 days to respectively obtain fermentation products of aspergillus terreus and trichoderma viride. Wherein the formula of the solid fermentation medium B comprises the following components by weight: corn starch: crushing rice dregs: millet: the straw residue is 2:1.5:1:1:0.5, and the water content is 60%. Drying and crushing the fermentation product to obtain powdery aspergillus terreus and trichoderma viride. Wherein the effective viable count of the aspergillus terreus contained in each gram of aspergillus terreus microbial inoculum is 5 multiplied by 1010The effective viable count of trichoderma viride in each gram of trichoderma viride is 4 multiplied by 1010。
3. And mixing the prepared bacillus amyloliquefaciens microbial inoculum, the prepared bacillus mucilaginosus microbial inoculum, the prepared trichoderma viride microbial inoculum and the prepared aspergillus terreus microbial inoculum according to the compounding proportion shown in the table 4 to obtain the composite microbial inoculum.
TABLE 4 examples of complex microbial agents
The prepared composite microbial inoculum is subjected to field disease resistance verification, and the test is carried out in Linyi City in Shandong province, and comprises the following steps:
selecting 1 strawberry greenhouse for 2 mu, and respectively setting a control group 1, a control group 2 and an experimental group, wherein the control group 1 does not use a microbial inoculum and only carries out conventional fertilization, and the control group 2 also uses a composite microbial inoculum purchased in the market besides the conventional fertilization. In addition to conventional fertilization, each experimental group also applied the complex microbial inoculum prepared above. Other field management is the same. The results are shown in table 5 below:
TABLE 5 strawberry greenhouse test results
From the results given in table 5, it can be seen that, compared with the control, the compound microbial inoculum of the invention has the advantages of improved physiological indexes of strawberry plants, full fruits, less malformed fruits, sweet taste, good mouthfeel, the morbidity rate of less than 10%, obviously reduced morbidity rate compared with the outsourcing market microbial inoculum, and generally increased sweetness compared with the market microbial inoculum.
Example 6 application of Bacillus amyloliquefaciens in preparation of fertilizers
Example 6 provides a fertilizer prepared by the steps of:
1. the complex microbial inoculum was prepared according to the method of example 5.
2. Weighing different materials according to the fermented cow dung content of 40%, the carbon-based fertilizer content of 30%, the small molecular organic material content of 20% and the calcium silicate content of 10%, mixing to obtain a biological organic fertilizer, and uniformly adding the composite microbial inoculum obtained in the step 1, wherein the composite microbial inoculum accounts for 1-1.5 per mill of the mass of the biological organic fertilizer, so as to obtain the corresponding fertilizer. Examples of fertilizers prepared using different bioorganic fertilizers are listed in table 6 below, and the fertilizers prepared in this example are designated by bioorganic fertilizer numbers for the purpose of distinguishing them from the fertilizers prepared in example 7 below.
TABLE 6 Fertilizer examples
The prepared fertilizer is subjected to field growth promotion and disease resistance verification, and the test is carried out in the Taoist and town of Junan county in Linyi city, Shandong province, and the method comprises the following steps:
selecting 1 strawberry greenhouse for 2 mu, and respectively arranging a blank control group and a control group, wherein the fertilizer used in the blank control group does not contain microbial inoculum compared with each experimental group, and the control group uses the market purchased microbial inoculum to replace the compound microbial inoculum for preparing the fertilizer. Each experimental group used each fertilizer prepared above, and other field management was the same. The results are shown in Table 7 below.
TABLE 7 results of field tests
Example 7 application of Bacillus amyloliquefaciens in preparing fertilizers
Example 7 provides a fertilizer prepared as follows:
the compound microbial inoculum prepared in the example 5 is uniformly mixed with anti-caking agent in the coating working section of organic compound fertilizer, organic-inorganic compound fertilizer and inorganic compound fertilizer, and then added according to 1-5 per mill (accounting for the proportion of the corresponding organic compound fertilizer, organic-inorganic compound fertilizer and inorganic compound fertilizer), and the corresponding fertilizer is obtained after stable production. As shown in tables 8 and 9. For ease of distinction, the fertilizers prepared in tables 8-11 are named as the corresponding base fertilizers.
In table 8, the organic-inorganic compound fertilizer refers to 15-40(16-12-12/S) of organic-inorganic compound fertilizer (wherein 15-40 refers to the ratio of organic fertilizer to inorganic fertilizer, and 16-12-12/S refers to the ratio of nitrogen, phosphorus and potassium).
In Table 9, the inorganic compound fertilizer means 15-15-15 (the contents of NPK are 15%, 15%, and 15%, respectively).
TABLE 8 examples of organic-inorganic Compound fertilizers
| Examples of organic-inorganic compound fertilizers | Complex microbial inoculum | Proportion of complex microbial inoculum (‰) |
| Example 1 of organic-inorganic Compound fertilizers | Complex microbial inoculum 1 | 2 |
| Example 2 of organic-inorganic Compound fertilizers | Complex microbial inoculum 2 | 3 |
| Example 3 of organic-inorganic Compound fertilizers | Complex microbial inoculum 3 | 4 |
| Example 4 of organic-inorganic Compound fertilizers | Complex microbial inoculum 4 | 5 |
| Example 5 of organic-inorganic Compound fertilizers | Complex microbial inoculum 5 | 1 |
| Example 6 of organic-inorganic Compound fertilizers | Complex microbial inoculum 6 | 2 |
| Example 7 of organic-inorganic Compound fertilizers | Complex microbial inoculum 7 | 3 |
| Example 8 of organic-inorganic Compound fertilizers | Complex microbial inoculum 8 | 5 |
| Example 9 of organic-inorganic Compound fertilizers | CompoundingBacterial agent 9 | 4 |
| Example 10 of organic-inorganic Compound fertilizers | Complex microbial inoculum 10 | 2 |
| Example of organic-inorganic Compound fertilizers 11 | Complex microbial inoculum 11 | 2 |
| Example 12 of organic-inorganic Compound fertilizers | Complex microbial inoculum 12 | 4 |
| Example 13 of organic-inorganic Compound fertilizers | Complex microbial inoculum 13 | 3.5 |
| Example 14 of organic-inorganic Compound fertilizers | Complex microbial inoculum 14 | 2.5 |
| Example 15 of organic-inorganic Compound fertilizers | Complex microbial inoculum 15 | 4 |
| Example 16 of organic-inorganic Compound fertilizers | Compound bacterial agent 16 | 5 |
| Example 17 of organic-inorganic Compound fertilizers | Complex microbial inoculum 17 | 4 |
TABLE 9 inorganic Compound fertilizer mix examples
The prepared fertilizer is subjected to field disease resistance verification, and the experiment is carried out in a test area of a strawberry shed at the town of the Shandong Junan county, and comprises the following steps:
3 treatments are arranged in the shed, each treatment is carried out in 3 small areas, each small area is 66.6 square meters, and the fertilizer application amount is 120 Kg/mu. Wherein the control group 1 is applied by conventional fertilization, the control group 2 is applied by microbial fertilizer purchased in the market, and the other experimental groups are applied by the prepared fertilizer. The results are shown in tables 10 and 11 below.
TABLE 10 field test results of organic-inorganic compound fertilizer strawberry in greenhouse
TABLE 11 field test results of strawberry greenhouse with inorganic compound fertilizer
As can be seen from the results given in tables 10 and 11, the physiological indexes of strawberry plants are significantly improved and the field incidence of strawberry root rot is significantly reduced by using the fertilizer of the present invention compared to the control.
In the description herein, references to the description of the term "one embodiment," "some embodiments," "an example," "a specific example," or "some examples," etc., mean that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the invention. In this specification, the schematic representations of the terms used above are not necessarily intended to refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples. Furthermore, various embodiments or examples and features of different embodiments or examples described in this specification can be combined and combined by one skilled in the art without contradiction.
Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention, and that variations, modifications, substitutions and alterations can be made to the above embodiments by those of ordinary skill in the art within the scope of the present invention.
SEQUENCE LISTING
<110> research and development center of Zhonghua agriculture (Linyi)
<120> Bacillus amyloliquefaciens CGMCC number 17842 and application thereof
<130> PIDC3194244
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 599
<212> DNA
<213> Artificial Sequence
<220>
<223> Bacillus amyloliquefaciens
<400> 1
acgctggcgg cgtgcctaat acatgcaagt cgagcggaca gatgggagct tgctccctga 60
tgttagcggc ggacgggtga gtaacacgtg ggtaacctgc ctgtaagact gggataactc 120
cgggaaaccg gggctaatac cggatggttg tttgaaccgc atggttcaga cataaaaggt 180
ggcttcggct accacttaca gatggacccg cggcgcatta gctagttggt gaggtaacgg 240
ctcaccaagg cgacgatgcg tagccgacct gagagggtga tcggccacac tgggactgag 300
acacggccca gactcctacg ggaggcagca gtagggaatc ttccgcaatg gacgaaagtc 360
tgacggagca acgccgcgtg agtgatgaag gttttcggat cgtaaagctc tgttgttagg 420
gaagaacaag tgccgttcaa atagggcggc accttgacgg tacctaacca gaaagccacg 480
gctaactacg tgccagcagc cgcggtaata cgtaggtggc aagcgttgtc cggaattatt 540
gggcgtaaag ggctcgcagg cggtttctta agtctgatgt gaaagccccc ggctcaacc 599
Claims (13)
1. A Bacillus amyloliquefaciens (Bacillus amyloliquefaciens) is preserved in the China general microbiological culture Collection center with the preservation number of CGMCC No. 17842.
2. A microbial agent, comprising the bacillus amyloliquefaciens of claim 1.
3. The microbial agent according to claim 2, wherein the microbial agent is in a dry powder form, and the effective viable count of the bacillus amyloliquefaciens is at least 1 x 10 per gram of the microbial agent11CFU。
4. The microbial agent according to claim 2, wherein the microbial agent is in a dry powder form, and the effective viable count of the bacillus amyloliquefaciens is 4 x 10 per gram of the microbial agent11CFU。
5. A complex microbial inoculant comprising a first microbial inoculant and a second microbial inoculant different from the first microbial inoculant, wherein the first microbial inoculant is the microbial inoculant of any one of claims 2 to 4;
the second microbial agent comprises at least one of bacillus mucilaginosus agent, trichoderma viride agent and aspergillus terreus agent.
6. The complex microbial inoculant according to claim 5, wherein the complex microbial inoculant comprises 1-3 parts by weight of the first microbial inoculant and at least one of the following components:
1-3 parts by weight of the Bacillus mucilaginosus microbial inoculum;
1-3 parts by weight of the trichoderma viride agent;
1-3 parts of the aspergillus terreus microbial inoculum.
7. The complex microbial inoculant according to claim 5, wherein the complex microbial inoculant comprises 1-1.5 parts by weight of the first microbial inoculant, and at least one of the following microbial inoculant:
1-1.5 parts by weight of the Bacillus mucilaginosus microbial inoculum;
1-1.5 parts by weight of the trichoderma viride agent;
1-1.5 parts by weight of the aspergillus terreus microbial inoculum.
8. A fertilizer, characterized in that the fertilizer comprises the microbial agent of any one of claims 2 to 4 or the complex microbial agent of any one of claims 5 to 7.
9. The fertilizer of claim 8, wherein said fertilizer further comprises a base fertilizer,
the basic fertilizer comprises at least one of a biological organic fertilizer, an inorganic compound fertilizer, an organic compound fertilizer or an organic-inorganic compound fertilizer, and the content of the microbial agent or the compound microbial agent is 1-5 per mill of the content of the basic fertilizer.
10. The fertilizer according to claim 9, wherein the bio-organic fertilizer comprises 30-50 parts by weight of fermented cow dung, 20-40 parts by weight of carbon-based fertilizer, 10-30 parts by weight of small-molecule organic material and 5-15 parts by weight of calcium silicate.
11. The use of the bacillus amyloliquefaciens of claim 1 in the preparation of a microbial agent, a complex microbial agent or a fertilizer.
12. A method of controlling root rot, comprising:
applying an effective amount of bacillus amyloliquefaciens, a microbial agent, a complex microbial agent or a fertilizer to a crop, wherein the bacillus amyloliquefaciens is the bacillus amyloliquefaciens of claim 1, the microbial agent is the microbial agent of any one of claims 2 to 4, the complex microbial agent is the complex microbial agent of any one of claims 5 to 7, and the fertilizer is the fertilizer of any one of claims 8 to 10; the root rot is caused by fusarium.
13. The method of claim 12, wherein the crop is strawberry.
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| CN105132336A (en) * | 2015-10-10 | 2015-12-09 | 中国热带农业科学院环境与植物保护研究所 | Bacillus amyloliquefaciens and microbial inoculum and application thereof |
| CN106350470A (en) * | 2016-11-18 | 2017-01-25 | 甘肃农业大学 | Bacillus amyloliquefaciens and microbial inoculum, and production method and application of microbial inoculum |
| CN110205273A (en) * | 2019-06-11 | 2019-09-06 | 山东碧蓝生物科技有限公司 | A kind of bacillus amyloliquefaciens and its application with growth promotion and resistant effect |
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| CN105132336A (en) * | 2015-10-10 | 2015-12-09 | 中国热带农业科学院环境与植物保护研究所 | Bacillus amyloliquefaciens and microbial inoculum and application thereof |
| CN106350470A (en) * | 2016-11-18 | 2017-01-25 | 甘肃农业大学 | Bacillus amyloliquefaciens and microbial inoculum, and production method and application of microbial inoculum |
| CN110205273A (en) * | 2019-06-11 | 2019-09-06 | 山东碧蓝生物科技有限公司 | A kind of bacillus amyloliquefaciens and its application with growth promotion and resistant effect |
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