A small molecule volatile compound for enhancing plant disease resistance and its application

Abstract

The present invention relates to a small molecule volatile compound that enhances plant disease resistance and its application. Specifically, the present invention provides the use of a compound of Formula I or a pharmaceutically acceptable salt thereof for enhancing plant resistance to pathogens; or for preparing a composition or formulation for enhancing plant resistance to pathogens. The present invention also discovers for the first time that the compound of Formula I enhances plant resistance to pathogens by activating the plant's own immune response, without affecting the normal growth of the plant or the pathogen. #imgabs0#

Description

Technical Field

[0001] The present invention relates to the field of agronomy, and in particular to a small molecule volatile compound for enhancing plant disease resistance and application thereof. Background Art

[0002] Plant pathogens have a significant negative impact on agricultural production and applications. To promote plant growth or enhance plant disease resistance, agricultural compounds are often used to assist in solving the problem. However, these agricultural compounds often cause a certain degree of environmental pollution.

[0003] Agricultural pathogenic bacteria stress, such as rice blast, significantly impacts agricultural production. Using certain growth-promoting rhizobacteria (PGPRs) has been found to be an effective approach to address plant growth efficiency under biotic and abiotic stresses.

[0004] Small volatile compounds released by growth-promoting rhizobacteria act as important signaling molecules to induce a variety of plant responses, such as 2,3-butanedione, which induces plant immune responses under low-phosphorus conditions. However, the characteristics of many other growth-promoting rhizobacteria and their stress response mechanisms in plants remain unclear, or their ability to improve stress resistance is limited.

[0005] Therefore, there is an urgent need in the art to develop strains or small molecule volatiles released by them or metabolites or agricultural preparations secreted by them that can significantly enhance plant resistance to disease stress, resistance to abiotic stress and / or significantly improve plant growth performance. Summary of the Invention

[0006] The purpose of the present invention is to provide a strain or the small molecule volatiles released by it or the metabolites or agricultural preparations secreted by it that can significantly enhance the plant's resistance to disease stress, resistance to abiotic stress and / or significantly improve the plant's growth performance.

[0007] The first aspect of the present invention provides a use of a compound of formula I or a pharmaceutically acceptable salt thereof for enhancing plant resistance to pathogens; or for preparing a composition or formulation for enhancing plant resistance to pathogens. The structure of the compound of formula I is shown below:

[0008]

[0009] Where,

[0010] R1 is H, amino, halogen, -OH, C1-C3 alkyl;

[0011] R2 is H, amino, halogen, -OH, C1-C3 alkyl;

[0012] R3 is H, -N(Ra )2, halogen, -OH, substituted or unsubstituted C1-C3 alkyl, wherein the "substituted" means that the H in the group is substituted by one or more substituents selected from the group consisting of amino, halogen, -OH, R a Selected from the group consisting of H, C1-C3 alkyl, or a combination thereof.

[0013] In another preferred embodiment, the halogen includes F, Cl, Br or I.

[0014] In another preferred embodiment, the compound has a structure of Formula Ia, Ib or Ic:

[0015]

[0016] In the formula, R1, R2, and R3 are as defined above.

[0017] In another preferred embodiment, R1 is H, amino, or -OH.

[0018] In another preferred embodiment, R2 is H, amino, or -OH.

[0019] In another preferred embodiment, R1 is amino.

[0020] In another preferred embodiment, R2 is amino.

[0021] In another preferred embodiment, R3 is -N(R a )2 or substituted C1-C3 alkyl, wherein the "substituted" means that the H in the group is substituted by a substituent selected from the group consisting of amino, R a Selected from the group consisting of H, C1-C3 alkyl, or a combination thereof.

[0022] In another preferred embodiment, R3 is a substituted methyl group, wherein the "substituted" means that the H in the group is substituted by a substituent selected from the following group: amino group.

[0023] In another preferred embodiment, the compound is selected from the following group:

[0024]

[0025] In another preferred embodiment, the plant's resistance to pathogens is enhanced by activating the plant's own immune response.

[0026] In another preferred embodiment, the pathogens include agricultural pathogens, such as plant pathogens.

[0027] In another preferred embodiment, the pathogen includes Pseudomonas syringaepv.Tomato.

[0028] In another preferred embodiment, the pathogen is selected from the group consisting of Pseudomonas syringae pv. Tomato DC3000 (Pto DC3000), Pseudomonas syringae pv. Actinidiae ICMP9617 (Pac ICMP9617), Pseudomonas aeruginosa (CGMCC No.: 1.15148), or a combination thereof.

[0029] In another preferred embodiment, the compound of formula I is derived from rhizosphere growth-promoting bacteria, such as Psudomonas aeruginosasp., Paenibacillus alvei or Streptomyces, or from grapes and bees.

[0030] In another preferred embodiment, the composition is an agricultural composition.

[0031] In another preferred embodiment, the composition comprises (a) a compound of formula I; and (b) an agriculturally acceptable carrier.

[0032] In another preferred embodiment, the composition further comprises other substances for enhancing the resistance of plants to pathogens.

[0033] In another preferred embodiment, the other substances for enhancing the resistance of plants to pathogens include: flg22, elf18, Chitin, and avirulent proteins such as Avrpto.

[0034] In another preferred embodiment, the dosage form of the composition or preparation is selected from the following group: solution, emulsion, suspension, powder, foam, paste, granule, aerosol, or a combination thereof.

[0035] In another preferred embodiment, the plants include agricultural plants, horticultural plants, and forestry plants.

[0036] In another preferred embodiment, the plants include woody plants and herbaceous plants.

[0037] In another preferred embodiment, the plants include Solanaceae, Cruciferae, Poaceae, Leguminosae, Musaceae, Bananaceae, Chenopodiaceae, and Liliaceae.

[0038] In another preferred embodiment, the plants include Solanum, Oryza, Nicotiana, Arabidopsis, Capsicum, Rapeseed, and Medicago.

[0039] In another preferred embodiment, the plants include Arabidopsis thaliana, rice, rapeseed, tomato, tobacco, pepper, rapeseed, and alfalfa.

[0040] A second aspect of the present invention provides a composition comprising:

[0041] (i) a compound of formula I;

[0042] (ii) other substances used to enhance plant resistance to pathogens;

[0043] (iii) an agriculturally acceptable carrier;

[0044] The structure of the compound of formula I is shown below:

[0045]

[0046] Where,

[0047] R1 is H, amino, halogen, -OH, C1-C3 alkyl;

[0048] R2 is H, amino, halogen, -OH, C1-C3 alkyl;

[0049] R3 is H, -N(R a )2, halogen, -OH, substituted or unsubstituted C1-C3 alkyl, wherein the "substituted" means that the H in the group is substituted by one or more substituents selected from the group consisting of amino, halogen, -OH, R a Selected from the group consisting of H, C1-C3 alkyl, or a combination thereof.

[0050] In another preferred embodiment, the composition comprises an agricultural composition.

[0051] In another preferred embodiment, the composition includes an antibacterial agent.

[0052] In another preferred embodiment, the dosage form of the composition is selected from the following group: solution, emulsion, suspension, powder, foam, paste, granule, aerosol, or a combination thereof.

[0053] In another preferred embodiment, the composition contains 0.0001-99 wt%, preferably 0.1-90 wt% of component (a), based on the total weight of the composition.

[0054] In another preferred embodiment, the composition contains 0.0001-99 wt%, preferably 0.1-90 wt% of component (b), based on the total weight of the composition.

[0055] In another preferred embodiment, the content of component (a) in the agricultural composition is 0.0001-99 wt%, preferably 0.001-90 wt%, more preferably 0.01-50%.

[0056] In another preferred embodiment, the concentration of component (a) in the agricultural composition is 5-800 μM, preferably 20-400 μM, more preferably 50-200 μM.

[0057] In another preferred embodiment, the weight ratio of component (a) to component (b) is 100:1-0.01:1, preferably, 10:1-0.1:1, and more preferably, 2:1-0.5:1.

[0058] In another preferred embodiment, in the composition, the component (a) and the component (b) account for 0.01-99.99 wt %, preferably 0.1-90 wt %, and more preferably 1-80 wt % of the total weight of the composition.

[0059] In another preferred embodiment, the other substances for enhancing the resistance of plants to pathogens include: flg22, elf18, Chitin, and avirulent proteins such as Avrpto.

[0060] The third aspect of the present invention provides a use of the composition according to the second aspect of the present invention for enhancing the resistance of plants to pathogens.

[0061] A fourth aspect of the present invention provides a method for enhancing plant resistance to pathogens, comprising the steps of:

[0062] The plant is administered with a compound of formula I, or the composition according to the second aspect of the present invention, wherein the structure of the compound of formula I is shown below:

[0063]

[0064] Where,

[0065] R1 is H, amino, halogen, -OH, C1-C3 alkyl;

[0066] R2 is H, amino, halogen, -OH, C1-C3 alkyl;

[0067] R3 is H, -N(R a )2, halogen, -OH, substituted or unsubstituted C1-C3 alkyl, wherein the "substituted" means that the H in the group is substituted by one or more substituents selected from the group consisting of amino, halogen, -OH, R a Selected from the group consisting of H, C1-C3 alkyl, or a combination thereof.

[0068] In another preferred embodiment, the application is selected from the group consisting of spraying, irrigation, drip irrigation, atomization, coating, injection or other methods known to those skilled in the art.

[0069] In another preferred embodiment, the administration can be a one-time administration, repeated administration or continuous administration.

[0070] In another preferred embodiment, the application method is application to plants or soil surrounding plants.

[0071] In another preferred embodiment, the administered dosage is 0.01-5 μmol / strain, preferably, 0.1-3 μmol / strain, and more preferably, 0.4-2 μmol / strain.

[0072] In another preferred embodiment, the administered dosage is 1-800 μM / plant, preferably, 10-400 μM / plant, and more preferably, 50-200 μM / plant.

[0073] It should be understood that within the scope of the present invention, the above-mentioned technical features of the present invention and the technical features described in detail below (such as in the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be listed here one by one. BRIEF DESCRIPTION OF THE DRAWINGS

[0074] The following drawings are used to illustrate specific embodiments of the present invention and are not used to limit the scope of the present invention defined by the claims.

[0075] Figure 1 This figure shows that 2'-aminoacetophenone helps Arabidopsis thaliana resist invasion by the pathogen Pseudomonas syringae pv. tomato DC3000. [A] Phenotypic evidence of 2'-aminoacetophenone-induced resistance to the pathogen DC3000 in Arabidopsis thaliana. n = 12, white scale represents 1 cm. [B] Quantitative results of pathogen colonization. n > 12, Student's t-test was used to test significance between samples, with p < 0.05 indicating significant differences. Three independent experiments were performed to confirm the same results.

[0076] Figure 2This study shows that 2'-aminoacetophenone does not affect the normal growth of plants or pathogens. [A] Number of lateral roots in 5-day-old Arabidopsis seedlings treated with 2'-aminoacetophenone 7 days after treatment. n > 7, Student t-test was used to test for significance between samples. p < 0.01 indicates an extremely significant difference, and *** indicates p < 0.001. [B] Leaf surface area in 5-day-old Arabidopsis seedlings treated with 2'-aminoacetophenone 7 days after treatment. n > 7, Student t-test was used to test for significance between samples. p < 0.05 indicates a significant difference, and ns indicates no significant difference. [C] Main root length in 5-day-old Arabidopsis seedlings treated with 2'-aminoacetophenone 7 days after treatment. n = 7, Student t-test was used to test for significance between samples. p < 0.05 indicates a significant difference, and ns indicates no significant difference. [D] DC3000 cells were treated with 2'-aminoacetophenone. OD values ​​were measured at different time points to examine the effect of 2'-aminoacetophenone on DC3000 growth. Error bars represent the standard deviation of four biological replicates, and the same results were confirmed by three independent experiments.

[0077] Figure 3 This figure shows that 2'-aminoacetophenone helps rice resist invasion by the pathogen Pseudomonas syringae pv. tomato DC3000. [A] Phenotype of 2'-aminoacetophenone-resistant rice against the pathogen DC3000. n > 12. [B] Quantitative results of pathogen colonization. n > 12. Student's t-test was used to test significance between samples. p < 0.01 indicates extremely significant differences, and *** indicates p < 0.001. Results were confirmed by two independent experiments.

[0078] Figure 4 This figure shows that 2'-aminoacetophenone protects rapeseed against the pathogen Pseudomonas syringae pv. tomato DC3000. [A] Phenotypic evidence of 2'-aminoacetophenone protecting rapeseed against the pathogen DC3000. n = 9. [B] Quantitative results of pathogen colonization. n > 12. Student's t-test was used to test significance between samples. p < 0.01 indicates extremely significant differences, and *** indicates p < 0.001. Results were verified by two independent experiments.

[0079] Figure 52'-Aminoacetophenone helps Arabidopsis thaliana resist infection with Pseudomonas aeruginosa sp. (CGMCC 1.1514). 2'-Aminoacetophenone helps Arabidopsis thaliana resist the phenotype of the pathogen Pseudomonas aeruginosa sp. 2'-Aminoacetophenone pretreatment and pathogen infection treatments were the same as those used for DC3000. n = 15, white scale represents 2 cm. [B] Quantitative results of pathogen colonization. n = 14, Student's t-test was used to test significance between samples, * indicates a significant difference at p < 0.05. Three independent experiments confirmed the same results.

[0080] Figure 6 2-aminoacetophenone and 4'-aminoacetophenone, isomers of 2'-aminoacetophenone, protect Arabidopsis thaliana against infection by the pathogen Pseudomonas syringae pv. tomato DC3000. [A] Arabidopsis plants were pretreated with 100 μL of a 1 mM compound for two days and then treated with DC3000 at an OD of 0.01. Two days later, samples were collected for pathogen colonization analysis. Statistical analysis showed that 2-aminoacetophenone significantly reduced DC3000 colonization in Arabidopsis plants. n > 24, one-way anova was used to test for significance between samples; ** indicates p < 0.01. Results were confirmed by two independent experiments. [B] Results of pathogen colonization in Arabidopsis plants pretreated with 100 μL of a 1 mM compound for two days after infection. n > 24, one-way anova was used to test for significance between samples; ** indicates p < 0.01. Results were confirmed by two independent experiments. DETAILED DESCRIPTION

[0081] After extensive and in-depth research, the inventors unexpectedly discovered that compounds of Formula I can effectively enhance plant resistance to pathogens. Furthermore, the inventors unexpectedly discovered that compounds of Formula I enhance plant resistance to pathogens by activating the plant's own immune response, without affecting the normal growth of the plant or the pathogen. Based on this, the inventors completed the present invention.

[0082] As used herein, the term "flg22" refers to a flagellin protein, which is a peptide segment containing 22 amino acids.

[0083] As used herein, the term "elf18" refers to a bacterial translation elongation factor, a polypeptide containing 18 amino acids.

[0084] As used herein, the term "chitin" refers to a component of fungal cell walls, which is chitin, also known as chitin.

[0085] As used herein, the term "Avrpto" refers to a protein secreted by pathogens that can induce plant immunity.

[0086] Group Definition

[0087] As used herein, the term "C1-C3 alkyl" refers to a straight or branched chain alkyl group having 1 to 3 carbon atoms, such as methyl, ethyl, n-propyl, isopropyl, or the like.

[0088] As used herein, the term "halogen" refers to fluorine, chlorine, bromine, or iodine.

[0089] The compounds of the present invention may contain one or more asymmetric centers and thus occur as racemates, racemic mixtures, single enantiomers, diastereomeric compounds, and single diastereomers. The presence of asymmetric centers depends on the nature of the various substituents on the molecule. Each such asymmetric center will independently produce two optical isomers, and all possible optical isomers and diastereomeric mixtures and pure or partially purified compounds are included within the scope of the present invention. The present invention includes all isomeric forms of the compounds.

[0090] Active ingredient

[0091] As used herein, the terms "active ingredient of the present invention" and "compound of formula I of the present invention" are used interchangeably and refer to active ingredients derived from rhizospheric growth-promoting bacteria, such as Psudomonas aeruginosa sp. or from grapes and bees, which have the function of enhancing the resistance of plants to pathogens.

[0092] In the present invention, the active ingredient of the present invention has the general formula shown in Formula I:

[0093]

[0094] Where,

[0095] R1 is H, amino, halogen, -OH, C1-C3 alkyl;

[0096] R2 is H, amino, halogen, -OH, C1-C3 alkyl;

[0097] R3 is H, -N(R a )2, halogen, -OH, substituted or unsubstituted C1-C3 alkyl, wherein the "substituted" means that the H in the group is substituted by one or more substituents selected from the group consisting of amino, halogen, -OH, R a Selected from the group consisting of H, C1-C3 alkyl, or a combination thereof.

[0098] In another preferred embodiment, the compound has a structure of Formula Ia, Ib or Ic:

[0099]

[0100] In the formula, R1, R2, and R3 are as defined above.

[0101] In another preferred embodiment, the compound is selected from the following group:

[0102]

[0103] In another preferred embodiment, the compound of formula I or a pharmaceutically acceptable salt thereof is chemically synthesized or isolated from rhizosphere growth-promoting bacteria.

[0104] 2'-aminoacetophenone, a key bacterial quorum sensing (QS) agent, plays an irreplaceable role in guiding bacterial aggregation and biofilm formation. Studies have shown that 2'-aminoacetophenone can be used as a diagnostic marker for the human lung infection caused by the pathogen Pseudomonas aeruginosa, and its concentration can be used as an important indicator for diagnosing disease severity.

[0105] In addition to being reported to be efficiently synthesized in bacteria, 2'-aminoacetophenone has also been detected in many other species, including grapes, grains, and ants. 2'-aminoacetophenone also has a very low molecular weight of 135.16 Daltons. Commercially available 2'-aminoacetophenone is an oily liquid with a strong aroma, high volatility, and low price (average price of approximately 150 RMB per milliliter). 2'-aminoacetophenone can be used as a food additive to enhance the flavor of foods, such as wine, or as a fragrance in perfumes.

[0106] 4'-Aminoacetophenone is a nontoxic derivative of aminoacetophenone that can be used to prepare a cyanide antidote. Its molecular weight is 135.16 Daltons. Commercially available 4'-aminoacetophenone is a pale yellow, aromatic, solid substance with a low price (averaging approximately 7 yuan per gram).

[0107] 2-Aminoacetophenone is a naturally occurring, small-molecule volatile product and an important intermediate in the production of chemical products, with a molecular weight of 135.16 Daltons. Commercialized 2-aminoacetophenone (2-aminoacetophenone hydrochloride) is a white crystal with a faint aroma and a yellowish color upon dissolution in water. Its price is slightly higher than that of 2'-aminoacetophenone and 4'-aminoacetophenone (averaging approximately 600 yuan per gram).

[0108] However, current research on the bacterial small molecules 2'-aminoacetophenone, 4'-aminoacetophenone, and 2-aminoacetophenone in plants is still unclear. In a preferred embodiment of the present invention, using the model plant Arabidopsis thaliana as an example, the present invention discovered that 2'-aminoacetophenone, 4'-aminoacetophenone, and 2-aminoacetophenone can effectively help Arabidopsis thaliana resist invasion by the pathogen Pseudomonas syringae pv. TomatoDC3000, achieving disease resistance. This finding was also verified by treating rice and rapeseed with 2'-aminoacetophenone.

[0109] In the present invention, 2'-aminoacetophenone, 4'-aminoacetophenone, and 2-aminoacetophenone can all be found in rhizospheric growth-promoting bacteria, such as 2'-aminoacetophenone can be detected in Psudomonas aeruginosa sp.; 4'-aminoacetophenone can be detected in Paenibacillus alvei (T19); and 2-aminoacetophenone can be detected in Streptomyces.

[0110] Agricultural compositions or preparations

[0111] The active substances of the present invention (such as compounds of formula I or pharmaceutically acceptable salts thereof) can be prepared into agricultural preparations by conventional methods, such as solutions, emulsions, suspensions, powders, foams, pastes, granules, aerosols, natural and synthetic materials impregnated with active substances, microcapsules in polymers, and coatings for seeds.

[0112] These preparations can be produced by known methods, for example, by mixing the active substance with an extender, i.e., a liquid, liquefied gas, or solid diluent or carrier, and optionally a surfactant, i.e., an emulsifier and / or dispersant and / or foam former. When, for example, water is used as the extender, an organic solvent can also be used as an auxiliary agent.

[0113] When a liquid solvent is used as a diluent or carrier, it is basically suitable, such as: aromatic hydrocarbons, for example xylene, toluene or alkylnaphthalene; chlorinated aromatic or chlorinated aliphatic hydrocarbons, for example chlorobenzene, vinyl chloride or dichloromethane; aliphatic hydrocarbons, for example cyclohexane or paraffin, for example mineral oil fractions; alcohols, for example ethanol or ethylene glycol and their ethers and lipids; ketones, for example acetone, methyl ethyl ketone, methyl isobutyl ketone or cyclohexanone; or less commonly used polar solvents, for example dimethylformamide and dimethyl sulfoxide, and water.

[0114] As for the diluent or carrier of liquefied gas, it refers to the liquid that will become gas at normal temperature and pressure, such as aerosol propellants, such as halogenated hydrocarbons and butane, propane, nitrogen and carbon dioxide.

[0115] Solid carriers can be ground natural minerals such as kaolin, clay, talc, quartz, atavistic clay, montmorillonite, or diatomaceous earth, and ground synthetic minerals such as highly dispersed silicic acid, alumina, and silicates. Solid carriers for particles are crushed and graded natural zircons such as calcite, marble, pumice, sepiolite, and dolomite, as well as particles synthesized from inorganic and organic coarse powders, and particles of organic materials such as sawdust, coconut shells, corn cobs, and tobacco stems.

[0116] Nonionic and anionic emulsifiers can be used as emulsifiers and / or foam formers. Examples include polyoxyethylene fatty acid esters, polyoxyethylene fatty alcohol ethers, such as alkylaryl polyethylene glycol ethers, alkyl sulfonates, alkyl sulfates, aryl sulfonates, and albumin hydrolysates. Dispersants include, for example, lignin sulfite waste liquor and methylcellulose.

[0117] Binders such as carboxymethylcellulose and natural and synthetic polymers in the form of powders, granules or emulsions, such as gum arabic, polyvinyl alcohol and polyvinyl acetate, may be used in the formulations.

[0118] Colorants such as inorganic dyes, such as iron oxide, cobalt oxide and Prussian blue; organic dyes, such as azo dyes or metal phthalocyanine dyes; and trace nutrients such as salts of iron, manganese, boron, copper, cobalt, aluminum and zinc can be used.

[0119] In the present invention, the "agricultural formulation" is generally an agricultural plant growth regulator, which contains a compound of formula I or a pharmaceutically acceptable salt thereof as an active ingredient for enhancing plant resistance to pathogens; and an agriculturally acceptable carrier.

[0120] As used herein, the "agriculturally acceptable carrier" is an agriculturally acceptable solvent, suspending agent, or excipient for delivering the compound of Formula I of the present invention or a pharmaceutically acceptable salt thereof to a plant. The carrier can be a liquid or a solid. Suitable agriculturally acceptable carriers for use in the present invention are selected from the group consisting of water, a buffer, DMSO, a surfactant such as Tween-20, or a combination thereof. Any agriculturally acceptable carrier known to those skilled in the art can be used in the present invention.

[0121] The agricultural preparation of the present invention can be prepared into a mixture with other substances that enhance plant resistance to pathogens and exist in their commercial preparations or in dosage forms prepared from these preparations. These other substances that enhance plant resistance to pathogens include (but are not limited to): flg22 (a flagellin protein, a peptide segment containing 22 amino acids), elf18 (a bacterial translation elongation factor, a polypeptide containing 18 amino acids), Chitin (a component of fungal cell walls, also known as chitin), and non-toxic proteins such as Avrpto (a protein secreted by pathogens that can induce plant immunity).

[0122] In addition, the agricultural preparations of the present invention can also be mixed with synergists in their commercial preparations or in dosage forms prepared from these preparations. These synergists are compounds that enhance the effects of the active substances. Since the active substances themselves are active, it is not necessary to add synergists.

[0123] The agricultural formulation of the present invention can be in a variety of forms, as long as the active ingredients can effectively reach the plant body. From the standpoint of ease of preparation and application, the preferred agricultural formulation is a spray or solution formulation.

[0124] The agricultural formulations of the present invention typically contain 0.0001-99 wt %, preferably 0.1-90 wt %, of the active substance of the present invention, based on the total weight of the agricultural formulation. The concentration of the active substance of the present invention in commercial formulations or dosage forms can vary over a wide range. The concentration of the compound of the present invention in commercial formulations or dosage forms can range from 0.0000001-100% (g / v), preferably between 0.0001 and 1% (g / v).

[0125] Methods for enhancing plant resistance to pathogens

[0126] The present invention provides a method for enhancing the resistance of plants to pathogens, comprising the steps of applying a compound of formula I or a pharmaceutically acceptable salt thereof, or applying a corresponding agricultural composition or preparation to the plants.

[0127] Application can be carried out by various known methods, for example, by spraying, misting, dusting or spreading the compound of formula I or its pharmaceutically acceptable salt or an agricultural composition or preparation containing the compound of formula I or its pharmaceutically acceptable salt on plant leaves or propagation materials, or by otherwise contacting the plant with the compound of formula I or its pharmaceutically acceptable salt or an agricultural preparation containing the compound of formula I or its pharmaceutically acceptable salt.

[0128] In a preferred embodiment, the compound of formula I or its pharmaceutically acceptable salt or the agricultural composition or formulation containing the compound of formula I or its pharmaceutically acceptable salt can also be delivered to the plant by spraying (such as spraying from an airplane) or irrigation.

[0129] The main advantages of the present invention include:

[0130] (1) The present invention first discovered that the compound of formula I or a pharmaceutically acceptable salt thereof can effectively enhance the resistance of plants to pathogens (especially tomato bacterial spot fungus).

[0131] (2) The present inventors have discovered for the first time that the compound of formula I or a pharmaceutically acceptable salt thereof enhances the resistance of plants to pathogens by activating the immune response of the plants themselves, without affecting the normal growth of the plants and the pathogens.

[0132] (3) The method of the present invention is highly safe and extremely low cost.

[0133] (4) The present invention has a wide range of applications and is easy to operate.

[0134] (5) The product of the present invention is easy to obtain.

[0135] The present invention will be further described below with reference to specific examples. It should be understood that these examples are intended to illustrate the present invention only and are not intended to limit the scope of the present invention. The experimental methods in the following examples, for which specific conditions are not specified, were generally performed under conventional conditions such as those described in Sambrook et al., Molecular Cloning: A Laboratory Manual (New York: Cold Spring Harbor Laboratory Press, 1989), or according to the conditions recommended by the manufacturer. Unless otherwise specified, the materials and reagents used in the examples are all commercially available products.

[0136] The experimental methods described in this example, unless otherwise specified, are all conventional means and methods in the art. The experimental kit consumables and reagents used, unless otherwise specified, are also conventional reagents and consumables in the art and can be purchased from commercial products.

[0137] The following examples involve Arabidopsis thaliana model Col-0 obtained from ABRC (Arabidopsis Biological Resource Center), rice model Oryza sativa L. japonica obtained from AmkhaSeed, rapeseed model Brassica napus L. obtained from Longping High-Tech (purchased from Taobao), 2'-aminoacetophenone purchased from Sigma with a purity of >99%, and 4'-aminoacetophenone and 2-aminoacetophenone purchased from Sigma with a purity of >99%.

[0138] Pseudomonas aeruginosa (CGMCC No.: 1.15148) was purchased from the CGMCC collection center, Pseudomonas syringae pv. Tomato DC3000 (Pto DC3000) can be obtained from the reference: doi: 10.15252 / embj.2019102602, and Pseudomonas syringae pv. Actinidiae ICMP9617 (Pac ICMP9617) can be obtained from the reference: 10.5423 / PPJ.NT.12.2017.0281.

[0139] General Methods

[0140] 1. Plant Growth

[0141] Arabidopsis thaliana was grown under the following conditions: temperature 22°C, humidity 65%, light intensity 100umol / m 2 / s 1 The photoperiod was 10 hours of light and 14 hours of darkness. When the Arabidopsis plants were about 4-5 weeks old, they were treated with 2'-aminoacetophenone and then injected or sprayed with DC3000 bacterial solution.

[0142] 2,2'-aminoacetophenone treatment

[0143] Prepare a 100mM stock solution of 2'-aminoacetophenone in water. Aliquot the solution into separate centrifuge tubes, seal with slit film, and store at 4°C. Dilute the stock solution to a final concentration of 200μM. Apply an appropriate volume of 2'-aminoacetophenone in water depending on the experimental purpose and system. For soil-based systems, seal the treated plants with 2'-aminoacetophenone with plastic wrap and keep them sealed for 2 days before treating for pathogens.

[0144] 3. Processing of DC3000

[0145] DC3000 was stored at -80°C in 50% glycerol. Activate DC3000 by streaking onto fresh LB plates containing 25 μg / mL of medium and incubate at 28°C. Suspend DC3000 in 10 mM magnesium chloride solution and dilute to an OD of 0.001-0.02. Add 0.02% surfactant, mix thoroughly, and spray evenly onto the surface of plant leaves. Seal the DC3000-treated plants with plastic wrap for 6 hours. Samples were taken and the DC3000 colonization efficiency was recorded at 0 and 3 days.

[0146] 4. Treatment of 4'-aminoacetophenone

[0147] The treatment method for 4'-aminoacetophenone is the same as that for 2'-aminoacetophenone. Prepare a 100mM stock solution in water and divide it into different PCR tubes. Add 100uL of the stock solution to each tube and store it at 4 degrees for future use. Petri dish system: 5 to 6 5-6 day old seedlings are grown on one side of a two-divided Petri dish. 100uL of 1mM 4'-aminoacetophenone is added to the other side of the culture medium. The Petri dish is re-wrapped with airtight sealing film and returned to the original growth chamber. After 48 hours of compound treatment, activate the DC3000 pathogen and adjust the concentration to a final OD of 0.01. Add 0.02% of the surfactant Silwet L77, mix well and spray evenly on the surface of the plant leaves. Return the plants after pathogen treatment to the incubator and take samples for bacterial colonization detection at the specified time.

[0148] 5. Treatment of 2-aminoacetophenone

[0149] The treatment method for 2-aminoacetophenone is the same as that for 2'-aminoacetophenone. Prepare a 100mM stock solution of water-soluble and divide it into different PCR tubes. Add 100uL of stock solution to each tube and store at 4 degrees for future use. Petri dish system: 5 to 6 5-6 day old seedlings are grown on one side of a two-divided Petri dish. 100uL of 1mM 2-aminoacetophenone is added to the other side of the culture medium. The Petri dish is re-wrapped with airtight sealing film and returned to the original growth chamber. After 48 hours of compound treatment, activate the DC3000 pathogen and adjust the concentration to a final OD of 0.01. Add 0.02% of the surfactant Silwet L77, mix well and spray evenly on the surface of the plant leaves. Return the plants after pathogen treatment to the incubator and take samples for bacterial colonization detection at the specified time.

[0150] Example 1 2'-aminoacetophenone enhances the resistance of Arabidopsis thaliana to the pathogen DC3000

[0151] Arabidopsis thaliana was grown under short-day conditions. Healthy seedlings, approximately 4-5 weeks old, were treated with 2'-aminoacetophenone and DC3000. A 2'-aminoacetophenone aqueous solution was prepared to a final concentration of 200 μM. 50 mL of this solution was poured into individual pots (4-5 seedlings per pot). Six pots were used in each experiment, with approximately 24 seedlings used. For the control group, an equal volume of water was added. The treated seedlings were sealed with plastic wrap and left for two days before the DC3000 treatment. To activate the DC3000 pathogen, 0.02% of the surfactant Silwet L77 was added to a final concentration of 0.2. The mixture was evenly mixed and sprayed onto the plant leaves. The leaves were then wrapped in plastic wrap for 6 hours before being removed. On the third day after DC3000 treatment, samples were collected for DC3000 colonization experiments. On average, four biological replicates were collected for each sample group, and each biological replicate collected samples from four different plant leaf holes. On the fifth day, photos were taken to record disease symptoms. The colonization experiment showed that the number of DC3000 colonization in Arabidopsis treated with 2'-aminoacetophenone was significantly less than that in the control group, indicating that 2'-aminoacetophenone can help Arabidopsis effectively resist the invasion of DC3000. Figure 1 The symptoms of infection further confirmed the results of the colonization experiment, indicating that plants treated with 2'-aminoacetophenone could significantly reduce the infection efficiency of DC3000. The plants in the control group showed a more yellow and wilted state, while the plants treated with 2'-aminoacetophenone showed a healthier phenotype. Figure 1 , A, B.

[0152] Example 2 2'-aminoacetophenone enhances rice resistance to pathogen DC3000

[0153] Rice seeds were germinated in culture soil and allowed to grow normally until the seedlings reached a height of approximately 10 cm. Twelve seedlings were planted in each pot, and healthy seedlings were treated with 2'-aminoacetophenone and DC3000. A 2'-aminoacetophenone aqueous solution was prepared to a final concentration of 200 μM. 50 mL of the 2'-aminoacetophenone solution was poured into a single pot. Six pots were used in each experiment, and approximately 70 seedlings were used. An equal volume of water was added to the control group. The treated seedlings were sealed with plastic wrap and treated for two days before the DC3000 treatment was initiated. The DC3000 pathogen was activated, adjusted to a final OD of 0.2, and 0.02% of the surfactant Silwet L77 was added. The mixture was evenly sprayed onto the plant leaves, which were then wrapped in plastic wrap for 6 hours before being removed. Disease symptoms were photographed and recorded on days 3-5. The disease symptoms of rice showed that 2'-aminoacetophenone-treated plants could significantly reduce the infection efficiency of DC3000. The plants in the control group showed a larger area of ​​yellowing and wilting, while the plants treated with 2'-aminoacetophenone showed a healthier phenotype. Figure 3 , A, B.

[0154] Example 3 2'-aminoacetophenone enhances rapeseed's resistance to the pathogen DC3000

[0155] Rapeseed seeds were germinated in culture soil and allowed to grow normally until they had three leaves. Four seedlings were planted in each pot, and healthy seedlings were treated with 2'-aminoacetophenone and DC3000. A 2'-aminoacetophenone aqueous solution was prepared to a final concentration of 200 μM. 50 mL of the 2'-aminoacetophenone aqueous solution was poured into each pot. Six pots were used in each experiment, and approximately 12 seedlings were used. An equal volume of water was added to the control group. The treated seedlings were sealed with plastic wrap and treated for two days before the DC3000 treatment was initiated. The DC3000 pathogen was activated, adjusted to a final concentration of 0.2, and 0.02% of the surfactant Silwet was added. The mixture was evenly sprayed onto the plant leaves, which were then wrapped in plastic wrap for 6 hours before being removed. Disease symptoms were photographed and recorded on days 3-5. The disease symptoms of rapeseed showed that plants treated with 2'-aminoacetophenone could significantly reduce the infection efficiency of DC3000. The plants in the control group showed a larger area of ​​yellowing and wilting, while the plants treated with 2'-aminoacetophenone showed a healthier phenotype. Figure 4 , A, B.

[0156] Example 4 2'-aminoacetophenone does not change the normal growth of plants and pathogens

[0157] Five-day-old Arabidopsis seedlings were transplanted to one side of a 1% agar-containing Petri dish, separating two separate plates. Five to six seedlings were placed in each dish. 100 μL of a 1 mM 2'-aminoacetophenone aqueous solution was added to the other dish. The dishes were sealed with airtight sealing film and incubated in a light-controlled incubator for 7 days. The seedlings were characterized using a camera, and the images were processed and analyzed using ImageJ software. The results showed that 2'-aminoacetophenone significantly increased the number of lateral roots in the plants. Figure 2 A, but it has no significant effect on the leaf surface area and taproot length of the plant, see Figure 2 , B, C. DC3000 was treated with 2'-aminoacetophenone at a concentration of 200 μM. An equal volume of sterile water was added to the control group. DC3000 was cultured in a 28-degree incubator and the OD value of DC3000 was measured at the specified time points. The results showed that compared with the control group, 2'-aminoacetophenone had no significant effect on the normal production of DC3000. Figure 2D. This indicates that 2'-aminoacetophenone enhances plant resistance to pathogens not by inhibiting pathogen growth or affecting plant growth, but by activating the plant's own immune response to enhance plant resistance to pathogen infection.

[0158] Example 5 2'-aminoacetophenone enhances the resistance of tomato and tobacco to DC3000

[0159] Tomato experiments: 7-day-old tomato seedlings were transplanted onto a medium containing 15 mL of 200 μM 2'-aminoacetophenone for two days. Both the treated and untreated tomatoes were sprayed with DC3000 at an OD of 0.04. After 2-3 days, an infected phenotype was observed in the tomatoes. Compound-treated tomatoes exhibited resistance to DC3000. Tobacco experiments: Healthy 4-5-week-old tobacco leaves were pretreated with flg22 (positive control), 2'-aminoacetophenone, and water (control). Nine hours later, the pretreated leaves were injected with DC3000 at an OD of 0.001. One to two days later, 2'-aminoacetophenone helped the tobacco resist the cell death phenotype caused by DC3000.

[0160] Example 6 2'-aminoacetophenone enhances the resistance of Arabidopsis thaliana to Pseudomonas aeruginosa (CGMCC No.: 1.15148

[0161] Healthy Arabidopsis thaliana seedlings, approximately 4-5 weeks old, were treated with 2'-aminoacetophenone and Pseudomonas aeruginosa. A 2'-aminoacetophenone aqueous solution was prepared to a final concentration of 200 μM. 50 mL of the solution was poured into individual pots (4-5 seedlings per pot). Six pots were used in each experiment, with approximately 12 seedlings used. For the control group, an equal volume of water was added. The treated seedlings were sealed with plastic wrap and left for two days before the Pseudomonas aeruginosa treatment began. To activate the Pseudomonas aeruginosa pathogen, 0.02% of the surfactant Silwet L77 was added to a final concentration of 0.2. The mixture was evenly mixed and sprayed onto the plant leaves. The leaves were then wrapped with plastic wrap for 6 hours before being removed. On the third day after treatment with Pseudomonas aeruginosa, samples were collected for colonization experiments. On average, four biological replicates were collected for each sample group, and each biological replicate collected samples from four different plant leaf holes. On the fifth day, photos were taken to record the disease symptoms. The colonization experiment showed that the colonization of Pseudomonas aeruginosa in Arabidopsis treated with 2'-aminoacetophenone was significantly less than that in the control group, indicating that 2'-aminoacetophenone can help Arabidopsis effectively resist the invasion of Pseudomonas aeruginosa. Figure 1 The symptoms of the disease further confirmed the results of the colonization experiment, indicating that plants treated with 2'-aminoacetophenone could significantly reduce the infection efficiency of Pseudomonas aeruginosa. The plants in the control group showed more yellowing of leaves, while the plants treated with 2'-aminoacetophenone showed a healthier phenotype. Figure 5 , A, B.

[0162] Example 7 Response of 2'-aminoacetophenone isomers to the pathogen DC3000

[0163] Arabidopsis thaliana plants, approximately 5-6 days old, were pretreated with 2'-aminoacetophenone (2-aminoacetophenone) and 4'-aminoacetophenone (4'-aminoacetophenone), the isomers of 2'-aminoacetophenone, at the same concentration as 2'-aminoacetophenone: 100 μL of a 1 mM aqueous solution. A control group was treated with an equal volume of water. Two days later, the plants were treated with the pathogen DC3000 (OD = 0.01) to observe their response to the pathogen.

[0164] The results are as follows Figure 6As shown in (AB), the results show that plants pretreated with 2-aminoacetophenone and 4'-aminoacetophenone showed enhanced resistance to the pathogen DC3000. Plants treated with the compounds showed significantly fewer DC3000 colonies than the control.

[0165] All documents mentioned in this application are incorporated herein by reference, just as if each document were incorporated herein by reference individually. It should also be understood that after reading the above teachings of the present invention, those skilled in the art may make various changes or modifications to the present invention, and that such equivalents also fall within the scope of the claims appended hereto.

Claims

1. A use of a compound or a pharmaceutically acceptable salt thereof, characterized in that: For enhancing the resistance of plants to pathogens; or preparing a composition or preparation for enhancing the resistance of plants to pathogens, wherein the compound is selected from the following group:

2. The use according to claim 1, characterized in that The pathogenic bacteria include Pseudomonas syringae pv. Tomato.

3. The use according to claim 1, characterized in that The pathogen is selected from the group consisting of Pseudomonassyringae pv. Tomato DC3000, Pseudomonas aeruginosa, or a combination thereof.

4. The use according to claim 1, wherein The plants include Solanaceae and Cruciferae.

5. The use according to claim 1, characterized in that The plants include Arabidopsis thaliana, rapeseed, and tomato.

6. A composition, characterized in that include: (i) a compound; (ii) other substances used to enhance plant resistance to pathogens; (iii) an agriculturally acceptable carrier; The compound is selected from the group consisting of:

7. Use of the composition according to claim 6, characterized in that: Used to enhance plant resistance to pathogens.

8. A method for enhancing plant resistance to pathogens, characterized in that: Including steps: Applying a compound to the plant, or applying the composition of claim 6, wherein the compound is selected from the group consisting of:

9. The method according to claim 8, wherein The dosage of the administration is 0.01-5 μmol / plant.

10. The method according to claim 8, wherein The administration may be a one-time administration, repeated administration, or continuous administration.

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