CN117511785A - Streptomyces GS2 and application thereof - Google Patents
Streptomyces GS2 and application thereof Download PDFInfo
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- CN117511785A CN117511785A CN202311447674.1A CN202311447674A CN117511785A CN 117511785 A CN117511785 A CN 117511785A CN 202311447674 A CN202311447674 A CN 202311447674A CN 117511785 A CN117511785 A CN 117511785A
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- streptomyces
- dimei
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- phytophthora
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
- A01N63/28—Streptomyces
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01P—BIOCIDAL, PEST REPELLANT, PEST ATTRACTANT OR PLANT GROWTH REGULATORY ACTIVITY OF CHEMICAL COMPOUNDS OR PREPARATIONS
- A01P21/00—Plant growth regulators
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01P—BIOCIDAL, PEST REPELLANT, PEST ATTRACTANT OR PLANT GROWTH REGULATORY ACTIVITY OF CHEMICAL COMPOUNDS OR PREPARATIONS
- A01P3/00—Fungicides
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
- C12P17/16—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms containing two or more hetero rings
- C12P17/165—Heterorings having nitrogen atoms as the only ring heteroatoms
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/465—Streptomyces
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/10—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in agriculture
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Biotechnology (AREA)
- General Health & Medical Sciences (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- Environmental Sciences (AREA)
- Virology (AREA)
- General Engineering & Computer Science (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Plant Pathology (AREA)
- Pest Control & Pesticides (AREA)
- Biomedical Technology (AREA)
- Medicinal Chemistry (AREA)
- Tropical Medicine & Parasitology (AREA)
- Agronomy & Crop Science (AREA)
- Dentistry (AREA)
- Botany (AREA)
- Mycology (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
Streptomyces GS2 and application thereof belong to the technical field of agricultural microorganisms. In order to screen Streptomyces capable of being used as a biological bactericide and producing beneficial compounds, the invention provides Streptomyces (Streptomyces sp.) GS2, and experiments on plate opposition, potting experiments and germination experiments prove that the Streptomyces has strong inhibition effect on various plant pathogenic microorganisms, has good control effect on plant diseases caused by various pathogenic microorganisms, and can promote plant growth. In addition, the invention extracts the fermentation product of streptomyces GS2, and discovers that the strain can produce Li Dimei element compounds and can be used as a novel Li Dimei element compound production strain for the production and preparation of Li Dimei element compounds.
Description
Technical Field
The invention belongs to the technical field of agricultural microorganisms, relates to streptomyces GS2 and application thereof, and in particular relates to streptomyces GS2 and application thereof in promoting plant growth, preventing and treating plant diseases and preparing Li Dimei element compounds.
Background
Plant diseases caused by plant pathogenic bacteria have serious influence on agricultural production. The traditional plant disease control is mainly based on chemical pesticides, but the environmental pollution is caused by the massive use and abuse of the chemical pesticides, and the influence of the residues of the pesticides in crop products on human health is also attracting more and more attention. The biological control by adopting the microorganism has the characteristics of low cost and environmental friendliness, is an important component of agricultural comprehensive control, and has become a research focus at home and abroad. Screening and constructing microbial strains with excellent performance, and developing biocontrol microbial agents has become an important development direction for controlling plant diseases.
Streptomyces is a kind of microorganism resource with important value, can produce abundant and various secondary metabolites with antifungal activity, can secrete some hydrolytic enzymes to damage fungal cell walls, and can produce active substances such as indoleacetic acid for promoting plant growth, thereby playing an important role in preventing and controlling plant diseases. Some actinomycete resources, such as Streptomyces griseoviridis K61, streptomyces lydicus WYEC, and Streptomyces saraceticus KH400, have been successfully developed as biocontrol agents for the control of plant diseases. Therefore, streptomyces has important research significance in the development of biocontrol agents.
Disclosure of Invention
In order to screen Streptomyces capable of being used as a biological bactericide and producing beneficial compounds, the invention provides a Streptomyces (Streptomyces sp.) GS2 strain which can promote plant growth, prevent and treat plant diseases caused by plant pathogenic bacteria, can ferment to produce Li Dimei element compounds, and has wide application prospect.
In order to solve the technical problems and realize the corresponding technical effects, the invention provides the following technical scheme:
the first object of the present invention is to provide a Streptomyces sp GS2 strain which is preserved in China general microbiological culture Collection center with the preservation number of CGMCC No.28479, the preservation date of 2023, 9 months and 19 days, and the preservation unit address of Beijing Kogyo area, beichen Xiyu No. 1, 3, national academy of sciences microbiological institute.
A second object of the present invention is to provide the use of the above Streptomyces GS2 for promoting plant growth.
In one embodiment of the invention, the Streptomyces GS2 concentration is 10 5 cfu/mL～10 8 The cfu/mL range can promote plant growth.
In one embodiment of the present invention, the plant is any one of soybean, rape, cucumber, rice, pepper, pumpkin, potato, wheat, and corn.
A third object of the present invention is to provide the use of the above-mentioned Streptomyces GS2 for controlling plant diseases caused by any one of plant pathogens of the genus Sclerotinia (Sclerotinia sp.), the genus Rhizoctonia (Rhizoctonia sp.), the genus colletotrichum (colletotrichum sp.), the genus Botrytis sp.), the genus Alternaria sp, the genus Phytophthora (Phytophthora sp.), and the genus Pythium.
In one embodiment of the present invention, the plant disease is a plant disease caused by any one of phytophthora infestans (Phytophthora infestans), phytophthora capsici (Phytophthora capsici), phytophthora sojae (Phytophthora sojae), pythum ultimum, pythium aphanidermatum (Pythium aphanidermatum), sclerotinia rot pathogen (Sclerotinia sclerotiorum), alternaria alternata (Alternaria alternata), rhizoctonia solani pathogen (Rhizoctonia solani), cucumber anthracnose pathogen (Colletotrichum orbiculare), tomato gray mold pathogen (Botrytis cinerea).
The fourth object of the present invention is to provide a microbial inoculum comprising the aforementioned Streptomyces GS2.
A fifth object of the present invention is to provide the use of the above-mentioned microbial agents for promoting plant growth.
In one embodiment of the present invention, the plant is any one of soybean, rape, cucumber, rice, pepper, pumpkin, potato, wheat, and corn.
A sixth object of the present invention is to provide the use of the above microbial inoculum for controlling plant diseases caused by any one of plant pathogenic bacteria of the genus Sclerotinia (Sclerotinia sp.), the genus Rhizoctonia (Rhizoctonia sp.), the genus Colletotrichum (Colletotrichum sp.), the genus Botrytis sp.), the genus Alternaria (Alternaria sp.), the genus Phytophthora (Phytophthora sp.), and the genus Pythium (Pythium sp.).
In one embodiment of the present invention, the plant disease is a plant disease caused by any one of phytophthora infestans (Phytophthora infestans), phytophthora capsici (Phytophthora capsici), phytophthora sojae (Phytophthora sojae), pythum ultimum, pythium aphanidermatum (Pythium aphanidermatum), sclerotinia rot pathogen (Sclerotinia sclerotiorum), alternaria alternata (Alternaria alternata), rhizoctonia solani pathogen (Rhizoctonia solani), cucumber anthracnose pathogen (Colletotrichum orbiculare), tomato gray mold pathogen (Botrytis cinerea).
In one embodiment of the invention, the microbial inoculum is applied by any one or more of soil mixing, root irrigation, spraying, seed soaking and coating.
In one embodiment of the invention, the microbial inoculum further comprises agriculturally acceptable adjuvants.
The seventh object of the present invention is to provide an application of the streptomyces GS2 in preparing Li Dimei element compounds, wherein the structural formula of the Li Dimei element compounds is shown in formula I:
in the formula I, R is CH 3 Or H, when R=CH 3 The Li Dimei element compound is Li Dimei element, and the Li Dimei element compound is demethyl Li Dimei element when r=h.
In one embodiment of the present invention, the specific method for preparing Li Dimei-based compounds is as follows:
1) Inoculating streptomycete GS2 into a seed culture medium, culturing in a shaking flask to obtain seed liquid, inoculating the seed liquid into a fermentation culture medium, and culturing in the shaking flask to obtain fermentation liquor;
2) Centrifuging the fermentation broth obtained in the step 1), collecting thalli, adding an extracting agent for ultrasonic extraction, and collecting the extracting agent to obtain a crude extract;
3) Separating and purifying the crude extract to obtain Li Dimei element and demethyl Li Dimei element.
In one embodiment of the invention, the seed medium and the fermentation medium of step 1) are both ISP3 liquid medium.
In one embodiment of the invention, the specific method of the step 1) is that the activated streptomycete GS2 is inoculated into ISP3 liquid culture medium, and the seed liquid is obtained by shaking culture for 2-4d at 28 ℃; inoculating the seed solution into ISP3 liquid culture medium according to the volume percentage of 2-10%, and shake culturing at 28 ℃ for 5-10d to obtain fermentation liquor.
In one embodiment of the present invention, the extractant in step 2) is methanol and the sonication time is 30-50min.
In one embodiment of the invention, the specific method of the separation and purification in the step 3) is that the crude extract obtained in the step 2) is mixed with silica gel powder, and then the mixture is subjected to dry column chromatography, dichloromethane/methanol gradient elution, gel chromatography and dichloromethane/methanol elution in sequence, and then the mixture is separated and purified by semi-preparative HPLC to obtain Li Dimei element and demethyl Li Dimei element.
The invention has the beneficial effects that:
the streptomyces GS2 provided by the invention has a strong inhibition effect on various plant pathogenic microorganisms, particularly has an inhibition rate of nearly 100% on phytophthora sojae, phytophthora capsici and sclerotium disease pathogenic bacteria of rape, and has a good control effect on plant diseases caused by the pathogenic microorganisms. And Streptomyces GS2 can also promote plant growth. Compared with the traditional chemical prevention and treatment means, the streptomyces GS2 can safely, efficiently and pollution-free prevent and treat plant diseases, reduce pollution to ecological environment, improve the quality safety of agricultural products, ensure the health of human beings, and has higher application value and better application prospect when being used as a biological microbial inoculum for development.
In addition, the streptomyces GS2 can be fermented to generate the lidamycin compound, the Li Dimei element and the demethyl Li Dimei element are enveloped, and the streptomyces GS2 can be used as a new strain for producing the Li Dimei element compound for producing and preparing the Li Dimei element compound.
Drawings
FIG. 1 is a graph showing the results of measurement of antagonistic activity of Streptomyces GS2 against plant pathogenic bacteria;
FIG. 2 is a graph showing the results of measuring the growth promoting effect of Streptomyces GS2 on wheat and corn; wherein, a in fig. 2 is a graph of the measurement result of the growth promoting effect of streptomyces GS2 on wheat, and b in fig. 2 is a graph of the measurement result of the growth promoting effect of streptomyces GS2 on corn;
FIG. 3 is a chart showing the HR-ESI-MS spectrum of Li Dimei element;
FIG. 4 is Li Dimei element 1 H-NMR spectrum;
FIG. 5 is Li Dimei element 13 C-NMR spectrum;
FIG. 6 is an HR-ESI-MS spectrum of normidecamycin;
FIG. 7 is a diagram of the desmethyllidamycin 1 A spectrum of H-NMR;
FIG. 8 shows the effect of the normidecamycin 13 C-NMR spectrum.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention will be described in further detail with reference to specific embodiments. The experimental methods used in the examples below were conventional, and the materials, reagents, methods and apparatus used, unless otherwise indicated, were all conventional in the art and commercially available to those skilled in the art.
The plant pathogenic bacteria used in the invention: phytophthora infestans (Phytophthora infestans), phytophthora capsici (Phytophthora capsici), phytophthora sojae (Phytophthora sojae), pythum ultimum (Pythum) Pythium ultimum, pythium aphanidermatum (Pythium aphanidermatum), sclerotinia sclerotiorum (Sclerotinia sclerotiorum), alternaria alternata (Alternaria alternata), rhizoctonia solani (Rhizoctonia solani), rhizoctonia cerealis (Colletotrichum orbiculare) and Botrytis cinerea (Botrytis cinerea) are stored in the university of northeast agriculture.
The preparation method of the V8 liquid culture medium used in the invention comprises the following steps:
340mL of V8 mixed fruit and vegetable juice is added with 3.4g of calcium carbonate powder, the mixture is centrifuged for 5min at 4 ℃ and 4000rpm, 300mL of fruit and vegetable juice without solid suspension is measured, 2700mL of deionized water is added to prepare 3L of V8 liquid medium, and the mixture is sterilized for 30min at 121 ℃. (the V8 mixed fruit and vegetable juice is purchased from the United states and is "Jinbao V8 vegetable juice 100%"340mL tin standard).
The PDA culture medium used in the invention is prepared by the following steps:
peeling 200g of potato, manually cutting into small pieces, boiling with boiling water for 30 minutes, filtering with gauze, and collecting juice; adding 20g of agar and 20g of glucose into the juice obtained by filtration, and fixing the volume to 1000mL; sterilizing at 121deg.C for 30 min.
The preparation method of the carrot culture medium used in the invention comprises the following steps:
200g of carrot, manually cutting into carrot blocks with the length of about 2 multiplied by 2cm, boiling for 30 minutes in boiling water, filtering with gauze, and collecting juice; adding 20g of agar into the juice obtained by filtration, and fixing the volume to 1000mL; sterilizing at 121deg.C for 30 min.
Example 1: inhibition of plant pathogenic bacteria by Streptomyces GS2
The strain GS2 was isolated from forest soil of Mianyang City, sichuan province, determined by 16S rRNA sequence, and the DNA sequence generated by sequencing was BLAST (https;// BLAST. NCBI. Lm. Gov/BLAST. Cgi) aligned by bacterial 16S rRNA of NCBI of public database, and the strain was determined to be Streptomyces by homology detection and alignment. The strain GS2 is preserved to China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC No.28479, the preservation date of 2023, 9 and 19 days, and the preservation unit address of Beijing Chaoyang area North Chenxi Lu No. 1, 3 and China academy of sciences microbiological study.
The antagonistic activity of Streptomyces GS2 against plant pathogenic bacteria was measured by plate-facing culture. Plant pathogenic bacteria include Phytophthora infestans (Phytophthora infestans), phytophthora capsici (Phytophthora capsici), phytophthora sojae (Phytophthora sojae), pythum ultimum (Pythum) Pythium ultimum, pythium aphanidermatum (Pythium aphanidermatum), sclerotinia sclerotiorum (Sclerotinia sclerotiorum), alternaria alternata (Alternaria alternata), rhizoctonia solani (Rhizoctonia solani), rhizoctonia cucumerica (Colletotrichum orbiculare), botrytis cinerea (Botrytis cinerea). Wherein phytophthora sojae is cultured by carrot solid culture medium, and other pathogenic bacteria are cultured by PDA solid culture medium. The Streptomyces GS2 is inoculated on the left side of a corresponding solid culture medium plate, cultured at 28 ℃ for 4 days until single colony of the Streptomyces GS2 grows to a proper size, a bacterial cake is punched out of pathogenic bacteria by a puncher (d=5mm), a toothpick is stuck on the right side of the solid culture medium plate, the Streptomyces GS2 is opposite to the pathogenic bacteria, and a plate which is not inoculated with the Streptomyces GS2 is used as a control. And then the whole plate is placed at 28 ℃ for culture, pathogenic microorganisms in the control plate grow on the whole plate, and the bacteria inhibition rate of the streptomycete GS2 is calculated statistically. The calculation formula of the bacteriostasis rate is as follows:
as shown in FIG. 1, the experimental results show that the Streptomyces GS2 has obvious inhibition effect on the tested plant pathogenic bacteria, and the inhibition rates of the phytophthora infestans (Phytophthora infestans), phytophthora capsici (Phytophthora capsici), phytophthora sojae (Phytophthora sojae), pythum ultimum (Pythum ulisum), pythum aphanidermatum (Pythium aphanidermatum), sclerotinia sclerotiorum (Sclerotinia sclerotiorum), alternaria alternata (Alternaria alternata), rhizoctonia solani (Rhizoctonia solani), cucumber anthracnose pathogen (Colletotrichum orbiculare) and tomato gray mold pathogen (Botrytis cinerea) are respectively 100%, 91%, 93.1%, 51.1%, 51.4%, 91.7%, 79.4%, 71%, 74.8% and 65.9%.
Example 2: application of streptomyces GS2 in controlling phytophthora sojae
(1) Preparation of Streptomyces GS2 spore suspension and Phytophthora sojae spore suspension
Preparation of Streptomyces GS2 spore suspension: scribing the Streptomyces GS2 plate, filling the ISP3 plate, scraping off all spores after the spores grow well, adding the spores into a 250mL triangular flask containing 10mL of sterile water and glass beads, and oscillating at 250rpm for 30min, so as to scatter and mix the spores uniformly. Sequentially diluting the mixed spore suspension to 10 -7 Select 10 -5 、10 -6 And 10 -7 And (3) coating 100 mu L of plates for each gradient, culturing until all single colonies grow out, counting the number of colonies, and obtaining the number of spores in the plates, wherein each gradient is spore suspension with corresponding concentration, and properly diluting the spore suspension according to experimental requirements.
Preparation of phytophthora sojae spore suspension: transferring Phytophthora sojae preserved at 4deg.C onto carrot solid culture medium, culturing at 28deg.C for 3d, transferring again, culturing under the same conditions for 5-6d, perforating plate full of Phytophthora sojae with a puncher, placing the punched bacterial pieces into carrot liquid culture medium, standing for 24h, pouring out liquid culture medium, adding sterile water, cleaning for three times, soaking with sterile water, standing for culturing for 12h, releasing zoospore, and filtering to obtain filtrate. Regulating concentration of released zoospores to 1×10 5 CFU/mL。
(2) Surface disinfection of soybean seeds
The soybean seed surface disinfection steps are as follows: selecting seeds with uniform size and full grains, putting the seeds into alcohol with the volume fraction of 70%, soaking for 1min, and pouring the alcohol; washing 3 times with sterile water; soaking in 5% sterile NaClO solution for 3min; finally, the surface is washed by sterile water for a plurality of times, so as to ensure that no disinfectant remains on the surface.
(3) Soybean seedling stage potting biocontrol test
The disease prevention experiment of soybean in seedling stage is provided with 4 treatment groups, which are respectively pathogen treatment, common treatment of streptomyces GS2 and pathogen, streptomyces GS2 treatment and aseptic water treatment. The disease prevention experiment of the phytophthora sojae in the seedling stage comprises the following specific experimental methods: preparation of Streptomyces GS2 spore suspension (spore concentration 10) 7 CFU/mL), uniformly mixing 1mL of spore suspension with 100g of soil in a soil mixing mode, planting soybean seeds into prepared soil, growing four plants in each pot until seedlings grow to a V3 stage, and preparing the soybean seeds with the concentration of 10 5 CFU/mL phytophthora sojae spores infest soybeans by root irrigation. After 10d, the disease occurrence is investigated, and the disease occurrence rate and the control effect are calculated according to the following formula.
The result shows that: after the phytophthora sojae pathogen spore suspension is inoculated, the phytophthora sojae treatment group is inoculated for one week, the disease symptoms begin to appear, the obvious unrecoverable wilting of the leaves appears, the falling phenomenon of part of the leaves appears, the leaf spots are light brown in color and irregular in shape, the disease incidence rate of plants is 100%, and the disease index is 41.9+/-3.5; the phytophthora sojae disease-receiving treatment group mixed with the streptomyces sojae GS2 spore suspension in the soil has good growth vigor and basically has no disease-sensing phenomenon, and the plant is obviously stronger than the plant treatment without adding the active fungus spore suspension, the morbidity is 13.9%, the disease index is 14.2+/-1.5, and the control effect reaches 76.5%. Compared with the CK control group, the experimental treatment group which is only inoculated with the streptomyces GS2 and not inoculated with pathogenic bacteria has the advantage that the plant height of soybean plants is improved by 14.6%.
The results show that the streptomyces GS2 not only can effectively prevent and control the infection of phytophthora sojae, but also can promote the growth of soybean.
Example 3: application of streptomyces GS2 in controlling phytophthora capsici
(1) Preparation of Streptomyces GS2 spore suspension and Phytophthora capsici spore suspension
Preparation of Streptomyces GS2 spore suspension: as in example 2.
Preparation of phytophthora capsici spore liquid: the activation step of phytophthora capsici is the same as that of phytophthora sojae, after transfer culture, punching phytophthora capsici, placing the punched fungus cake in 10% V8 liquid culture medium, culturing at 28deg.C for 3d, pouring into the liquid culture medium, soaking with sterile water, replacing once every 12 hr, continuously replacing for one week, respectively placing in 4 deg.C and 28deg.C for 30min, filtering to obtain filtrate, and regulating spore concentration to 1×10 5 CFU/mL。
(2) Surface disinfection of pepper seeds:
the surface disinfection steps of the capsicum seed are as follows: selecting seeds with uniform size and full grains, putting the seeds into alcohol with the volume fraction of 70%, soaking for 1min, and pouring the alcohol; washing 3 times with sterile water; soaking in 5% sterile NaClO solution for 3min; finally, the surface is washed by sterile water for a plurality of times, so as to ensure that no disinfectant remains on the surface.
(3) Pepper seedling stage potting biocontrol test
The disease prevention experiment in the seedling stage of the capsicum is provided with 4 treatment groups which are respectively pathogen treatment, common treatment of streptomyces GS2 and pathogen, streptomyces GS2 treatment and aseptic water treatment. The disease prevention experiment of the phytophthora capsici at the seedling stage comprises the following specific experimental methods: the pepper is planted in advance, cultivated at room temperature, and transplanted into soil, 4 plants are cultivated in each pot until the period of 7-8 leaves, and the root inoculation concentration is 10 5 CFU/mL phytophthora capsici spores are cultured for 24 hours, and then the root is inoculated with 10 percent of concentration 8 CFU/mL Streptomyces GS2 spore suspension. After 7d, the disease condition is investigated according to the classification standard of the phytophthora capsici, and the disease index and the control effect are calculated according to the following formula.
Classification standard of phytophthora capsici: level 0: no disease; stage 1: the root and stem are slightly blackened, and leaves are not wilted or restorative wilting; 2 stages: the root stem is blackened by 1-2cm, the leaves are irrecoverable and wilt, and the leaves at the lower part are occasionally fallen off; 3 stages: the root stem is blackened by more than 2cm, and the leaves are obviously wilted or fallen leaves are obvious; 4 stages: the roots are blackened and overflowed, and all leaves are fallen or the whole plant is wilted except the growing points; 5 stages: plants die.
The result shows that: after the phytophthora capsici is inoculated for 24 hours, active bacterial spore suspension is inoculated, the disease occurrence phenomenon occurs after bagging treatment for about 3d and 5-7d, the disease occurrence phenomenon occurs when only a phytophthora capsici pathogenic bacteria treatment group is inoculated, the leaves are browned in a large area and are seriously fallen, the whole plant is wilted, the whole plant grows slowly, the disease occurrence rate is up to 100%, and the disease index is 46.1+/-1.9; the leaves of the disease-grafting group inoculated with the active bacterial spore suspension are slightly yellow, the leaves at the lower part are slightly fallen off, the whole state of the plant is healthy, the morbidity is 20.2 percent, the disease index is 23.4+/-2.1, and the control effect reaches 61.3 percent. Compared with the CK control group, the experimental treatment group which is only inoculated with the streptomyces GS2 and not inoculated with pathogenic bacteria has the advantage that the plant height of the pepper plants is improved by 10.9 percent.
The results show that the streptomyces GS2 not only can effectively prevent and treat the infection of phytophthora capsici, but also can promote the growth of the capsicum.
Example 4: application of streptomyces GS2 in preventing and controlling cucumber diseases caused by Pythium aphanidermatum
Activating and growing the preserved Pythium aphanidermatum strain on a 10% V8 culture medium, and culturing in the dark in a constant temperature incubator at 25 ℃ for 1-2 days. A mycelia block with a size of 10X 15mm was cut with a scalpel and placed in 15mL of sterilized tap water with the mycelia side of the mycelia block facing upward, and water was changed every 30 minutes. Repeating for 3 times, adding sterilized water, making the sterilized water just submerge mycelium surface, culturing in a constant temperature incubator at 25deg.C for 20-24 hr, inducing zoospore, and regulating concentration to 10 with sterile water 5 Individual spores/mL. Filling sterilized sand and nutrient soil into plastic pot with aperture of 10cm, adding water to saturation, flattening surface, and uniformly stamping 1 sowing pit per pot for standby. The cucumber seeds are surface-sterilized with 10% sodium hypochlorite for 5min, washed with sterile water for 4-5 times and then dried. 1 seed is put in each pit, a layer of sterilizing substrate soil is covered on the seeds, and the seeds are placed in a greenhouse for cultivation. Root-filling treatment of each seedling with Streptomyces GS2, wherein the spore inoculation amount of Streptomyces GS2 is 10 6 The zoospores of Pythium aphanidermatum are irrigated into the root of the cucumber after 3 days, and the inoculation amount of the zoospores in each pit is about 10 5 And (3) spores. After 11 days, calculating the disease index and the prevention and treatment effect of the plants, wherein the calculation formula is as follows:
representative values for each stage are as follows: 0 = plant remains green and healthy; 1 = leaf sheath discoloration and lower leaf yellowing; 2 = plant survival, but leaf complete yellowing or death; 3 = whole plant death.
The result shows that: the disease index of the cucumber treated by the streptomyces GS2 is lower than that of a disease control inoculated with the Pythium aphanidermatum only, the average disease index of the treated group is 26.7, and the relative prevention effect is 59.3%.
Example 5: application of streptomyces GS2 in preventing and treating sclerotinia sclerotiorum caused by S.sclerotiorum
Selecting rape with 2-3 true leaves with good growth vigor, selecting rape with uniform plant size and leaf size, and suspending spore of Streptomyces GS2 (10) 7 Individual spores/mL) were evenly sprayed on rape leaves and left to stand overnight in a natural air-dry room. Then, a piece of sclerotium pathogenic bacteria cake with the same bacterial age is placed in the middle position of each leaf, the diameter of the bacterial cake is 5mm, and the mycelium is placed downwards. After inoculation, the rape is placed in an environment with the temperature of 24 ℃ and the relative humidity of more than 80 percentCulturing for 5 days. Grading investigation is carried out according to the morbidity of the blank control, and grading standards are as follows:
level 0: no symptoms appear; stage 1: the area of the disease spots on the blades is less than or equal to 5 percent; 3 stages: 6% < the area of lesions on leaves is 10% or less; 5 stages: 10% < the area of lesions on leaves is less than or equal to 20%;7 stages: 20% < the area of lesions on leaves is less than or equal to 40%; stage 9: the area of lesions on the leaves was >40%.
And calculating the disease index according to the disease area of the leaf blade, and further counting the prevention and treatment effect.
The calculation formulas of the disease index and the prevention and treatment effect in the in-vitro leaf are as follows:
the result shows that: the control effect of the streptomyces GS2 on sclerotinia sclerotiorum caused by sclerotinia sclerotiorum is 68.2 percent.
Example 6: application of streptomyces GS2 in preventing and controlling botrytis cinerea diseases caused by B.cinerea
Selecting tomato fruits with similar sizes and maturity, and determining biocontrol effect of streptomyces GS2 on tomato gray mold by adopting a wound inoculation method, wherein the biocontrol effect is specifically as follows: soaking tomato fruit in 75% ethanol for 5min, soaking in 3% sodium hypochlorite solution for 5min, washing with sterile water for 3 times, and wiping with sterile filter paper. Holes with the diameter of 5mm are lightly punched on the surface of the tomato by adopting a puncher with the diameter of 5mm, and the surface skin is prevented from cracking. Streptomyces GS2 spore suspension (10) 6 And (3) inoculating the liquid to the wound of the fruit, picking pathogenic bacteria with the diameter of 5mm by using an inoculating needle after the liquid is completely absorbed by the fruit, and inoculating the pathogenic bacteria to the wound of the fruit. And finally, placing the tomato fruits in a sterile porcelain plate, placing 2 absorbent cotton stained with water in a blank place for moisturizing, sealing by using a preservative film, placing in a constant temperature incubator at 25 ℃ for culturing for 7 days, measuring the area of the disease spots, and calculating the control effect.
Control% = (control plaque area-treatment plaque area) ×100/control plaque area
The result shows that: streptomyces GS2 has good in-vitro control effect on tomato gray mold caused by Botrytis cinerea, and the control effect can reach 76.2%.
Example 7: application of streptomyces GS2 in preventing and controlling rice sheath blight caused by R.solani
Soaking rice seeds for 24 hours, placing the rice seeds in an incubator at 25 ℃ for germination acceleration, sowing the rice seeds in a plastic basin filled with sterilized soil after exposure to white, watering regularly, transplanting the rice seeds in plastic barrels after 20 days, and planting 3 holes in each barrel and 3 plants in each hole. Selecting rice plants with consistent growth vigor at the end of tillering, and inoculating rice sheath blight pathogenic bacteria cake (diameter 5 mm) at the equal position by using a toothpick stem embedding method. Spore suspension of Streptomyces GS2 was then sprayed (10 6 and/mL) until the leaves drip down. Spraying is carried out 1 time every 7 days, spraying is carried out 2 times continuously, and the length of the lesion is investigated and recorded after 21 days. Disease severity was recorded according to a scale of 0 to 9. Level 0: healthy and disease-free; stage 1: the area of the lesion is smaller than 1/4 of the area of the leaf sheath; 3 stages: the area of the lesion accounts for 1/4 to 1/2 (excluding 1/2) of the area of the leaf sheath; 5 stages: the area of the lesion accounts for 1/2 to 3/4 (excluding 3/4) of the area of the leaf sheath; 7 stages: the area of the lesion is more than or equal to 3/4 of the area of the leaf sheath; stage 9: the lesions reached the top of the plants and all leaves were severely infested.
As a result, it was found that Streptomyces GS2 had no pathogenic effect on rice and the control effect on Rhizoctonia solani was 58.7%.
Example 8: application of streptomyces GS2 in preventing and controlling leaf spot disease of rape caused by alternaria alternata
Rape leaves with proper size, regular shape and easy cultivation and can be infected by Alternaria alternata, so rape leaves with similar size, shape and growth state are selected as test leaves, the rape leaves are picked, soaked in sterile water for 30min and naturally dried for standby, spore suspension of streptomycete GS2 is sprayed on the front and back sides of the rape leaves, so that tiny water drops are formed without flowing down, the rape leaves are naturally dried, spread in a culture dish containing filter paper, and a proper amount of sterile water is added to keep moist. Taking Alternaria alternata fungus cake with the diameter of 9mm by a puncher, inoculating the Alternaria alternata fungus cake in the center of a leaf blade, enabling a mycelium surface to contact with the leaf blade, culturing in a plant illumination incubator under the control of 28 ℃ and illumination darkness for 12 hours respectively, alternately culturing, observing the disease condition of the leaf blade after 5d, measuring the diameter of a disease spot on the leaf blade by a crisscross method, and photographing and recording.
As a result, the Streptomyces GS2 has a good control effect on rape leaf spot, and the control effect is 57.9%.
Example 9: application of streptomyces GS2 in promoting plant growth
Streptomyces GS2 is covered on the surface of the whole ISP3 solid medium by a streaking mode, after the spores grow well, all spores are scraped off and added into a 250mL triangular flask containing 10mL of sterile water and glass beads, and the shaking is carried out at 250rpm for 30min, so that the spores are scattered and uniformly mixed. Sequentially diluting the mixed spore suspension to 10 -7 Select 10 -5 、10 -6 And 10 -7 The three concentrations are obtained by coating 100 mu L of plates for each gradient, culturing until single colonies grow out completely, counting the number of colonies, and obtaining the number of spores in the plates, wherein each gradient is spore suspension with corresponding concentration. Diluting spore suspension to obtain spore with concentration of 10 5 、10 6 、10 7 、10 8 And 10 9 CFU/mL spore suspension.
Selecting normal color wheat and corn seeds, soaking in sterile water for 6 hr, washing with 70% ethanol solution for 1min, washing with 3% NaClO solution for 5min, and washing with 2% Na 2 S 2 O 3 The solution was washed 2 times,finally, the seed surface was washed 3 times with sterile water. Soaking the sterilized seeds in a solution of different concentrations (10 5 -10 9 CFU/mL) was used as a control with sterile water for 30min in GS2 spore suspension. Placing the soaked seeds evenly in flat plates paved with double-layer filter paper, adding 5mL of sterilized water into each flat plate as water required by seed germination, carrying out dark culture on the wheat germinated flat plates at 18 ℃ for 7 days, measuring root length and bud length of the wheat, and carrying out dark culture on the corn germinated flat plates at 30 ℃ for 7 days, and measuring root length and bud length of the corn.
As shown in FIG. 2 and Table 1, it can be seen from the experimental results that at low concentrations, streptomyces GS2 significantly increased the sprout length and root length of wheat and corn, and the stronger the growth promoting effect with increasing spore concentration, but when the spore concentration reached 10 9 CFU/mL can inhibit the growth of wheat and corn. Therefore, the concentration of Streptomyces GS2 is 10 5 CFU/mL～10 8 Plant growth can be promoted in the CFU/mL range.
TABLE 1 Streptomyces GS2 has effect on promoting the growth of wheat and corn
Example 5: application of streptomycete GS2 in preparation of Li Dimei element compounds
(1) Fermenting and culturing streptomyces GS2 to prepare a fermentation liquid:
fermenting strains: the fermentation strain is Streptomyces GS2.
Slant culture: ISP2 medium (yeast extract 4.0g, malt extract 10.0g, glucose 4.0g, agar 20.0g, distilled water 1000mL, pH 7.0-7.2) was used to sterilize at 121deg.C for 20min, and after inoculation, the culture was carried out at 28deg.C for 6-8 days.
Seed culture: seed medium composition: glucose 4.0g, malt extract powder 10.0g, yeast powder 4.0g, caCO 3 0.2 g, 1000mL of distilled water, pH value of 7.0-7.2, subpackaging 250mL for each bottle by using a 1000mL triangular bottle, and sterilizing at 121 ℃ for 20min. Then 10mL of sterile water was used to remove the residueThe spores of the slant-cultured Streptomyces GS2 were washed off and made into spore suspension at a concentration of 10 7 -10 8 Each bottle was filled with 2mL of spore suspension, and the mixture was placed on a shaking table at 28℃and 250rpm for 48 hours.
Fermentation culture: fermentation medium ISP 3: oat flour 20g/L FeSO 4 ·7H 2 O 0.001g/L，MnCl 2 ·4H 2 O 0.001g/L，ZnSO 4 ·7H 2 O0.001 g/L, distilled water, pH7.0-7.2, and sterilizing at 121deg.C for 20min. Inoculating the seed solution into 5L fermenter according to 10% inoculum size, culturing at 28deg.C with stirring rate of 100rpm and ventilation rate of 120m 3 And/h, culturing for 5-10 days.
(2) Crude extracts were obtained:
centrifuging the fermentation liquor obtained in the step (1) to obtain thalli and supernatant, extracting active ingredients in the thalli by methanol ultrasound for 30-50min, recovering methanol solvent, and concentrating to obtain crude extract.
(3) Preparation of the compound:
mixing the crude extract obtained in the step (2) with silica gel powder (100-200 mesh), loading on a column by a dry method, and performing gradient elution by using dichloromethane/methanol, wherein the volume ratio of the dichloromethane to the methanol is 100:0, 90:10, 80:20, 70:30, 60:40 and 50:50 in sequence; thereby respectively obtaining 6 elution components of Fraction A-Fraction F, wherein the component Fraction E is subjected to gel Sephadex LH20 chromatography and dichloromethane is used for preparing the composition: methanol (v/v) was 1:1 is eluent, and the eluent eluted in turn is divided into three parts (one part for every 1/3 and three parts) according to the proportion to obtain 3 components of Fraction E1-Fraction E3; wherein Fraction E1 was purified by semi-preparative HPLC (column Agilent SB-C18column, 250X 9.4mm i.d.,5 μm) using methanol: water (0.1% triethylamine) was used as a mobile phase at a flow rate of 1.5ml/min, and Li Dimei element (retention time t) was obtained by separation and purification R =16.3 min) and desmethyllidamycin (retention time t R ＝18.2min)。
(4) Structural identification of compound Li Dimei and desmethyl Li Dimei:
the HRESIMS spectrum of Li Dimei is shown in FIG. 3, the NMR spectrum is shown in FIGS. 4-5, the HRESIMS spectrum of normidecamycin is shown in FIG. 6, and the NMR spectrum is shown in FIGS. 7-8. The structures of Li Dimei and demethyl Li Dimei are determined by 1D and 2D NMR, MS, etc. spectroscopy as follows:
li Dimei: r=ch 3
Desmethyllidamycin: r=h
Li Dimei the extract is white powder, and has formula C 47 H 74 N 4 O 10 High resolution mass spectrometry (hresis): m/z [ M+H ]] + 855.5483(calcd for C 47 H 74 N 4 O 10 ，855.5478)。
The demethyl lidamycin is white powder with a molecular formula of C 46 H 71 N 4 O 10 High resolution mass spectrometry (hresis): m/z [ M+H ]] + 841.5322(calcd for C 46 H 73 N 4 O 10 ，841.5321)。
Li Dimei and demethyl Li Dimei elements 1 H and 13 c NMR (MeOD) data were as follows.
Li Dimei: 1 H NMR(400MHz,MeOD)δH 5.58(1H,m,H-23),5.51(1H,m,H-27),5.45(1H,m,H-22),5.50(1H,m,H-26),5.42(1H,m,H-19),4.85(1H,m,H-11),3.64(1H,m,H-21),4.03(1H,m,H-29),4.02(1H,m,H-35),3.80(1H,t,J＝6.6Hz,H-25),3.65(1H,m,H-33),3.80(1H,m,H-9),3.58(1H,m,H-17),3.51(2H,m,H2-40),3.48(1H,m,Ha-38),3.43(1H,m,H-8),3.35(1H,m,Hb-38),3.07(1H,m,H-5),2.62(1H,m,H-13),2.49(1H,m,H-10),2.32(1H,m,Ha-20),1.66(1H,m,H-28),1.83(1H,m,Ha-32),1.13(1H,m,Hb-32),2.21(1H,m,H-24),2.20(1H,m,Hb-20),2.07(1H,m,Ha-36),2.07(2H,m,H2-37),1.90(1H,m,Hb-36),1.84(2H,m,H2-14),1.54(2H,m,H2-15),1.72(3H,s,H3-42),1.70(1H,m,Hb-7),1.37(1H,m,Ha-7),2.30(2H,m,H2-34),1.62(3H,s,H3-46),1.58(3H,s,H3-44),2.20(1H,m,H-16),1.39(3H,s,H3-41),1.37(1H,m,Ha-6),1.72(1H,m,Hb-6),1.54(2H,m,H2-15),1.84(2H,m,H2-14),0.93(3H,d,J＝6.8Hz,H3-45),0.76(3H,d,J＝6.7Hz,H3-43)； 13 C NMR(100MHz,MeOD)δC 203.86(C-3),192.62(C-39),180.97(C-1),155.79(C-47),141.28(C-18),140.44(C-12),139.68(C-30),134.95(C-23),134.29(C-22),134.34(C-26),129.80(C-27),123.97(C-31),123.01(C-19),120.19(C-11),102.99(C-2),83.72(C-17),77.62(C-9),78.32(C-25),75.59(C-21),73.69(C-29),71.15(C-8),70.21(C-33),57.61(C-35),54.43(C-4),50.69(C-40),48.24(C-38),44.99(C-13),43.99(C-24),43.71(C-10),42.3(C-28),41.65(C-34),37.83(C-32),37.96(C-15),37.38(C-16),36.74(C-20),33.55(C-5),32.22(C-36),29.60(C-7),29.97(C-14),29.60(C-7),24.14(C-37),23.48(C-6),23.37(C-42),17.87(C-41),17.13(C-43),16.50(C-45),11.99(C-46),11.96(C-44)。
desmethyllidamycin: 1 H NMR(400MHz,MeOD)δH 5.63(1H,m,H-31),5.59(1H,m,H-23),5.51(1H,m,H-27),5.57(1H,m,H-30),5.50(1H,m,H-22),5.50(1H,m,H-26),5.33(1H,t,J＝6.8Hz,H-19),4.85(1H,m,H-11),4.06(1H,m,H-29),4.03(1H,m,H-21),4.03(1H,m,H-35),3.84(1H,t,J＝6.7Hz,H-25),3.65(1H,m,H-33),3.64(1H,m,H-9),3.57(1H,m,H-17),3.50(2H,m,H2-40),3.48(1H,m,Ha-38),3.43(1H,m,H-8),3.35(1H,m,Hb-38),3.06(1H,m,H-5),2.63(1H,m,H-13),2.50(1H,m,H-10),2.34(1H,m,Ha-20),2.19(1H,m,Hb-20),1.65(1H,m,H-32),2.23(1H,m,H-24),2.21(1H,m,H-28),2.07(1H,m,Ha-36),2.06(2H,m,H2-37),1.90(1H,m,Hb-36),1.54(2H,m,H2-15),1.85(2H,m,H2-14),1.73(3H,s,H3-42),1.55(2H,m,H2-7),2.28(1H,m,Ha-34),2.21(1H,m,Hb-34),1.58(3H,s,H3-44),1.84(1H,m,H-16),1.40(3H,s,H3-41),1.73(2H,m,H2-6),0.93(3H,d,J＝6.8Hz,H3-45),0.76(3H,d,J＝6.7Hz,H3-43)； 13 C NMR(100MHz,MeOD)δC 203.88(C-3),192.44(C-39),181.02(C-1),155.75(C-47),141.22(C-12),139.65(C-18),136.91(C-30),134.90(C-23),134.71(C-26),134.27(C-22),129.50(C-27),128.02(C-31),123.99(C-19),120.19(C-11),102.99(C-2),83.71(C-17),77.53(C-25),75.59(C-9),73.66(C-21),73.14(C-29),71.13(C-8),69.86(C-33),57.54(C-35),54.44(C-4),50.69(C-40),48.21(C-38),44.98(C-13),77.53(C-25),43.76(C-10),42.15(C-28),41.66(C-34),41.46(C-32),37.90(C-15),37.90(C-16),36.71(C-20),33.54(C-5),32.14(C-36),29.92(C-14),29.57(C-7),24.11(C-37),23.38(C-42),23.38(C-6),17.86(C-41),17.11(C-43),16.35(C-45),11.96(C-44)。
while the invention has been described with reference to the preferred embodiments, it is not limited thereto, and various changes and modifications can be made therein by those skilled in the art without departing from the spirit and scope of the invention as defined in the appended claims.
Claims (10)
1. Streptomyces sp GS2 is characterized by having a preservation number of CGMCC No.28479.
2. Use of streptomyces GS2 according to claim 1 for promoting plant growth.
3. The use of the Streptomyces GS2 according to claim 1 for controlling plant diseases, wherein the plant diseases are plant diseases caused by any one of plant pathogenic bacteria of the genus Sclerotinia (Sclerotinia sp.), the genus Rhizoctonia (Rhizoctonia sp.), the genus Celerotinia (Colletotrichum sp.), the genus Botrytis sp.), the genus Alternaria (Alternaria sp.), the genus Phytophthora (Phytophthora sp.), and the genus Pythium.
4. A microbial agent comprising the Streptomyces GS2 according to claim 1.
5. The use of the microbial inoculum of claim 4 for promoting plant growth.
6. The use of the microbial inoculum according to claim 4 for controlling plant diseases, wherein the plant diseases are plant diseases caused by any one of plant pathogenic bacteria of the genus Sclerotinia (Sclerotinia sp.), the genus Rhizoctonia (Rhizoctonia sp.), the genus Colletotrichum (Colletotrichum sp.), the genus Botrytis sp.), the genus Alternaria (Alternaria sp.), the genus Phytophthora (Phytophthora sp.), and the genus Pythium (Pythium sp.).
7. The use according to any one of claims 5 or 6, wherein the microbial inoculum is applied by any one or a combination of two or more of soil mixing, root irrigation, spraying, seed soaking and coating.
8. The application of the streptomyces GS2 in preparing Li Dimei element compounds as claimed in claim 1, wherein the structural formula of the Li Dimei element compounds is shown in the formula I:
in the formula I, R is CH 3 Or H, when R=CH 3 The Li Dimei element compound is Li Dimei element, and the Li Dimei element compound is demethyl Li Dimei element when r=h.
9. The use according to claim 8, wherein the specific method for preparing Li Dimei-based compounds is as follows:
1) Inoculating streptomycete GS2 into a seed culture medium, culturing in a shaking flask to obtain seed liquid, inoculating the seed liquid into a fermentation culture medium, and culturing in the shaking flask to obtain fermentation liquor;
2) Centrifuging the fermentation broth obtained in the step 1), collecting thalli, adding an extracting agent for ultrasonic extraction, and collecting the extracting agent to obtain a crude extract;
3) Separating and purifying the crude extract to obtain Li Dimei element and demethyl Li Dimei element.
10. The use according to claim 9, wherein the extractant of step 2) is methanol; the specific method of the separation and purification in the step 3) is that the crude extract obtained in the step 2) is mixed with silica gel powder, and then the mixture is put on a column by a dry method, dichloromethane/methanol gradient elution, gel chromatography and dichloromethane/methanol elution are sequentially carried out, and Li Dimei element and demethyl Li Dimei element are obtained through semi-preparative HPLC separation and purification.
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