Application of compound MSP-10 in preparation of medicine for preventing and treating magnaporthe oryzae

Abstract

The invention discloses application of a compound MSP-10 in preparation of a medicine for preventing and treating Magnaporthe oryzae, and belongs to the field of prevention and treatment of plant fungal diseases, and the compound MSP-10 can be used for preventing and treating Magnaporthe oryzae or / and other pathogenic fungi. The compound MSP-10 disclosed by the invention has a certain prevention and treatment effect after being mixed with a magnaporthe oryzae spore solution and sprayed for 0 hour, which indicates that the compound MSP-10 disclosed by the invention can be used as a protection medicament to have a better effect in the early stage of magnaporthe oryzae infection; moreover, the rice sprayed with the compound is good in growth situation and free of obvious adverse effects, which indicates that the compound is safe to the rice and has practical significance in preventing rice blast.

Description

TECHNICAL FIELD

[0001] The application belongs to the field of plant fungal disease prevention and treatment, and particularly relates to application of a compound MSP-10 in preparation of a medicine for preventing and treating Magnaporthe oryzae. BACKGROUND

[0002] Rice is one of the most important food crops in the world. However, plant diseases pose a significant threat to food crop production, among which rice blast caused by Magnaporthe oryzae is one of the most devastating crop diseases in the world and has a great destructive power on global rice production. Rice blast has a broad host range, and different strains can infect at least 50 species of grasses, including core food crops such as rice, wheat, barley and oats. Magnaporthe oryzae

[0003] The main form of Magnaporthe oryzae infection in the field is conidia, which are spread and transmitted by media such as air flow, water flow, raindrops and insects. The conidia that fall on rice leaves germinate under suitable conditions to produce a specific dome-shaped infection structure, the appressorium. The appressorium accumulates about 3.2 M high-concentration glycerol during development, which can generate a turgor pressure as high as 8.0 MPa, promoting the appressorium to protrude a narrow infection peg at the base to infect the host leaf. It is worth noting that Magnaporthe oryzae has characteristics such as wide geographical distribution, strong environmental adaptation and fast genetic variation, which not only increase the difficulty of disease control, but also make rice blast a hot and difficult problem in plant pathology research and disease management practice. Therefore, the development and implementation of comprehensive control strategies for rice blast are of great significance to ensure the safety of food crop production and maintain the stability of global food supply.

[0004] Traditional disease control strategies include breeding disease-resistant and high-yield crop varieties, eliminating sources of pathogens, implementing chemical control, strengthening field management, optimizing water and fertilizer management, and properly handling diseased grains and rice straw. Among them, chemical control, as a crucial link, is of great concern and favor to agricultural scientists and agricultural practitioners because it still shows significant control effect during the period when crop diseases are serious. However, in recent decades, the overuse and frequent use of chemical pesticides have led to rapid degradation of farmland soil properties and continuous decline of the health status of farmland ecosystems. At the same time, long-term pesticide selection pressure has prompted Magnaporthe oryzae to evolve into many physiological races with resistance, which undoubtedly increases the complexity and challenge of rice blast control.

[0005] ​In view of the above serious problems, it is currently urgently needed to accelerate the research and development process of environmentally friendly and green ecological pesticides by in-depth exploration of potential pathogenic related targets of Magnaporthe oryzae, with the aim of effectively resisting the invasion of Magnaporthe oryzae, protecting rice crops from serious damage, promoting sustainable agricultural development, ensuring the yield and quality of rice, and achieving the ideal effect of pesticide control.

[0006] Peroxisome is a key organelle in fungal cells, which undertakes multiple core metabolic functions including fatty acid beta-oxidation, reactive oxygen species (ROS) removal and glyoxylate cycle, and is known as the "metabolic center" of the cell. Its biogenesis depends on a class of proteins encoded by PEX genes, called Peroxin. Currently, 37 Peroxins have been identified in fungi, plants and animals. MoPex19 is a soluble chaperone that can specifically recognize and bind newly formed Peroxisomal Membrane Proteins (PMPs), guiding their directional transport to the peroxisomal membrane, thereby promoting membrane assembly. If MoPex19 is absent, most PMPs cannot be correctly positioned and integrated into the membrane, resulting in failure of peroxisome assembly and complete loss of function, indicating that the protein is upstream in the biogenesis pathway and has a core regulatory function.

[0007] MoPex19 is a key protein for Peroxisomal Membrane Proteins (PMPs) transport and peroxisome maintenance in Magnaporthe oryzae, Magnaporthe oryzae ) and is involved in the biogenesis of peroxisomes. MoPex19 The deletion mutant exhibits various abnormalities in fungal development and pathogenicity-related morphogenesis, and completely loses pathogenicity to the host. However, there is currently no pesticide targeting MoPex19. Therefore, there is a need to develop innovative pesticides targeting MoPex19 to effectively resist the invasion of Magnaporthe oryzae and ensure the safe production of core food crops such as rice.

[0008] It should be noted that the information disclosed in the above background section is only used to strengthen the understanding of the background of the present disclosure, and therefore can include information that does not constitute prior art known to those of ordinary skill in the art. SUMMARY

[0009] The purpose of the present application is to provide the use of compound MSP-10 in the preparation of a medicine for preventing and treating Magnaporthe oryzae, which solves the problem of drug resistance to pathogenic related targets of existing medicines. The compound has the advantages of remarkable effect, longer action time, safety, simple application method, etc. in preventing and treating Magnaporthe oryzae.

[0010] In order to achieve the above purpose, the present application provides the use of compound MSP-10 in the preparation of a medicine for preventing and treating Magnaporthe oryzae, Magnaporthe oryzae) and other pathogenic fungi, the chemical structure of the compound MSP-10 is shown as follows: ; The other pathogenic fungi include any one or more than two of Botrytis cinerea ( Botrytis cinerea ), Fusarium fujikuroi ( Fusarium fujikuroi ), Mycosphaerella melonis ( Colletotrichum orbiculare ), Alternaria alternata ( Alternaria alternata ) and Phoma exigua ( Colletotrichum acutatum ).

[0011] The second object of the present application is to provide the use of the compound MSP-10 in the preparation of a medicine for inhibiting the mycelium growth or / and spore germination or / and appressorium development of Magnaporthe grisea or / and other pathogenic fungi, the other pathogenic fungi including any one or more than two of Botrytis cinerea, Fusarium fujikuroi, Mycosphaerella melonis, Alternaria alternata and Phoma exigua.

[0012] Preferably, the compound MSP-10 can reduce the turgor pressure of appressorium of Magnaporthe grisea, or / and slow down the transportation of glycogen from conidium to appressorium, or / and slow down the rate of glycogen degradation in appressorium.

[0013] Preferably, the concentration of the compound MSP-10 is 10 ~ 50 ppm. The compound MSP-10 in the present application can effectively inhibit the pathogenicity of Magnaporthe grisea spores in the concentration range of 10 ~ 50 ppm.

[0014] More preferably, the concentration of the compound MSP-10 is 30 ~ 50 ppm. The compound MSP-10 in the present application has a significant inhibitory effect on Magnaporthe grisea in vitro when the concentration is 30 ~ 50 ppm.

[0015] More preferably, the concentration of the compound MSP-10 is 50 ppm, and the compound MSP-10 can completely inhibit Magnaporthe grisea in vitro at this concentration.

[0016] The third object of the present application is to provide a medicine for preventing and treating Magnaporthe grisea or / and other pathogenic fungi, the other pathogenic fungi including any one or more than two of Botrytis cinerea, Fusarium fujikuroi, Mycosphaerella melonis, Alternaria alternata and Phoma exigua, comprising the compound MSP-10.

[0017] Preferably, it further comprises an agriculturally acceptable carrier.

[0018] A fourth object of the present application is to provide a method for preventing Magnaporthe grisea or / and other pathogenic fungi, wherein the medicine for preventing Magnaporthe grisea or / and other pathogenic fungi is sprayed on the leaves of plants; the other pathogenic fungi include any one or two or more of Botrytis cinerea, Fusarium fujikuroi, Guazatine, Alternaria alternata and Phakopsora pachyrhizi.

[0019] Preferably, the concentration of the compound MSP-10 is 10 ~ 50 ppm.

[0020] More preferably, the concentration of the compound MSP-10 is 30 ~ 50 ppm.

[0021] The application of the compound MSP-10 in the present application in the preparation of a medicine for preventing Magnaporthe grisea solves the problem of drug resistance of existing medicines to pathogenic related targets, and has the following advantages: (1) The present application first discovers that the compound MSP-10 has a significant effect in inhibiting Magnaporthe grisea and preventing rice blast, and can effectively inhibit the pathogenicity of Magnaporthe grisea spores in the concentration range of 10 ~ 50 ppm, especially the compound MSP-10 at 50 ppm can completely prevent rice blast, and the application method is simple; (2) The compound MSP-10 used in the present application has a significantly better prevention effect on Magnaporthe grisea of isolated rice leaves than that of isolated barley leaves at the same treatment concentration; (3) The compound MSP-10 of the present application has a certain prevention effect when mixed with Magnaporthe grisea spore solution and sprayed at 0 h, indicating that the compound MSP-10 of the present application can be used as a protective agent at the early stage of Magnaporthe grisea infection and has better effect; moreover, the sprayed compound has good growth trend of rice without obvious adverse effects, indicating that the compound is safe to rice and has practical significance for preventing rice blast; (4) The present application discovers that the compound MSP-10 can inhibit the hyphal growth of Magnaporthe grisea, increase the rate of appressorium collapse, significantly reduce the turgor pressure of appressorium, and slow down the transport of glycogen from conidium to appressorium and the rate of glycogen degradation in appressorium; (5) The compound MSP-10 of the present application not only can inhibit Magnaporthe grisea, but also has a significant inhibitory effect on other various pathogenic fungi, showing its broad-spectrum antifungal activity, which means that the use of the compound MSP-10 can simultaneously deal with various diseases caused by different pathogenic fungi in agricultural production practice, reducing the types and frequency of pesticide use, reducing the cost of agricultural production, and being conducive to environmental protection and ecological balance, and not easy to cause environmental pollution and ecological destruction, which meets the demand of modern agriculture for green and environmentally friendly pesticides. BRIEF DESCRIPTION OF DRAWINGS

[0022] Figure 1 Predicted map of compound MSP-10 and MoPex19 binding sites.

[0023] Figure 2 Inhibition of rice blast mycelial growth by compound MSP-10 at different concentrations.

[0024] Figure 3 Comparison of rice blast mycelial growth diameter (A) and growth inhibition rate (B) under treatment of compound MSP-10 at different concentrations; in the figure, different concentrations of compound MSP-10 inhibition rates with significant differences are represented by different letters a, b, c, d, e or f; p < 0.05; data in the figure is obtained by One-way ANOVA.

[0025] Figure 4 Disease conditions of detached barley leaves (A) and detached rice leaves (B) when spore solution of rice blast is added with different concentrations of compound MSP-10.

[0026] Figure 5 Disease conditions of living rice leaves when 50 ppm compound MSP-10 is sprayed at different time periods after inoculation of spore solution of rice blast.

[0027] Figure 6 Comparison of relative lesion area of living rice leaves when 50 ppm compound MSP-10 is sprayed at different time periods after inoculation of spore solution of rice blast.

[0028] Figure 7 Formation conditions of appressorium at different time points when 50 ppm compound MSP-10 is added exogenously to spore solution of rice blast.

[0029] Figure 8 Comparison of appressorium formation rate at different time points when 50 ppm compound MSP-10 is added exogenously to spore solution of rice blast.

[0030] Figure 9 Collapse conditions of appressorium under 2 different concentrations of glycerol when 50 ppm compound MSP-10 is added exogenously to spore solution of rice blast.

[0031] Figure 10 Comparison of appressorium collapse rate under 2 different concentrations of glycerol when 50 ppm compound MSP-10 is added exogenously to spore solution of rice blast.

[0032] Figure 11The glycan transport and degradation in different time points of the appressorium induced by adding 50 ppm compound MSP-10 to the spore liquid of Magnaporthe oryzae (A); the proportion of glycan contained in conidia in different time points induced (B) and the proportion of glycan contained in appressorium (C).

[0033] Figure 12 The lipid droplet transport and degradation in different time points of the appressorium induced by adding 50 ppm compound MSP-10 to the spore liquid of Magnaporthe oryzae (A); the proportion of lipid droplet contained in conidia in different time points induced (B) and the proportion of lipid droplet contained in appressorium (C).

[0034] Figure 13 The inhibition of the growth of different plant pathogenic fungi by 50 ppm compound MSP-10.

[0035] Figure 14 The colony growth diameter of different plant pathogenic fungi after treatment by 50 ppm compound MSP-10; the black column chart is the colony diameter of the pathogenic fungi without adding MSP-10 and under normal growth (control), and the white column chart is the colony diameter of the pathogenic fungi after treatment by MSP-10, n = 3.

[0036] Figure 15 The influence of different concentrations of compound MSP-10 on the growth of rice; the scale = 2 cm.

[0037] Note: In the figure, * represents p < 0.05, ** represents p < 0.01, and *** represents p < 0.001. DETAILED DESCRIPTION

[0038] The technical solutions in the embodiments of the present application will be described below in a clear and complete manner. Obviously, the described embodiments are only some of the embodiments of the present application, but not all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative labor fall within the scope of protection of the present application.

[0039] It should be noted that: the specific conditions not indicated in the embodiments are carried out according to the conventional conditions or the conditions suggested by the manufacturer. The instruments not indicated by the manufacturer are conventional products that can be obtained by market purchase. The raw materials and reagents not indicated by the manufacturer are market goods or can be prepared by known methods.

[0040] In this invention, all features defined in the form of numerical ranges or percentage ranges, such as numerical values, quantities, contents, and concentrations, are used only for simplicity and convenience. Accordingly, the description of numerical ranges or percentage ranges should be considered as covering and specifically disclosing all possible sub-ranges and individual numerical values ​​(including integers and fractions) within those ranges.

[0041] The features mentioned in this invention can be combined arbitrarily, and all possible combinations should be considered within the scope of this specification, provided that there is no contradiction in the combination of these features. Each feature disclosed in the specification can be replaced by any alternative feature that provides the same, equivalent, or similar purpose. Therefore, unless otherwise specified, the disclosed features are merely general examples of equivalent or similar features.

[0042] MoPex19 is the rice blast fungus ( Magnaporthe oryzae Key proteins for the transport and maintenance of peroxisome membrane proteins (PMPs) in the middle peroxisome. MoPEX19 The deletion mutant exhibited various abnormalities in fungal development and pathogenicity-related morphogenesis, completely losing its pathogenicity to the host. These results indicate that MoPex19 plays a crucial role in maintaining peroxisomal and peroxisomal-derived structures, and is essential for the metabolism, development, and pathogenicity of rice blast fungus (MoPex19, which Is Essential for Maintenance of Peroxisomal Structure and Woronin Bodies, Is Required for Metabolism and Development in the Rice BlastFungus, PLoS One. 2014 Jan 14;9(1):e85252). However, there are currently no reports on drugs targeting MoPex19 for the control of rice blast.

[0043] The inventors of this invention used MoPex19 (MoPex19 gene information available in NCBI, MGG_00971) as a target and obtained the lead compound MSP-10 through high-throughput screening of a large number of compounds. To investigate the specific binding of MSP-10 to this target, AlphaFold3 was used to perform structural simulations of the interaction sites between MSP-10 and the MoPex19 protein. The predicted results are as follows: Figure 1 As shown. Analysis indicates that the binding sites of the MoPex19 protein to the compound MSP-10 are TYR-269, ARG-268, ASP-265, THR-261, and SER-314.

[0044] Further application of compound MSP-10 in the study of rice blast fungus prevention and treatment is proved by the experiments involved in the following examples.

[0045] The experimental materials used in the following examples of the present application are as follows: 1. The rice blast fungus used in the examples of the present application is a wild type strain Guy11 (American ATCC strain center) preserved in the laboratory; 2. The rice blast fungus culture medium used in the laboratory is complete culture medium (abbreviated as CM), and the CM culture medium formula is: 10 g of glucose, 2 g of proteose peptone-140, 1 g of casein amino acid, 1 g of yeast extract, 6 g of NaNO3, 1.52 g of KH2PO4, 0.52 g of KCl, 0.52 g of MgSO4·7H2O, 0.1 mg of biotin, 0.1 mg of vitamin B, 0.1 mg of thiamine, 0.1 mg of riboflavin, 0.1 mg of nicotinic acid, 0.1 mg of p-aminobenzoic acid, 1.5 mg of Na2MoO4·5H2O, 1.6 mg of CuSO4·5H2O, 1.7 mg of CoCl2·6H2O, 5 mg of MnCl2·4H2O, 5 mg of FeSO4·7H2O, 11 mg of H3BO3, 22 mg of ZnSO4·7H2O, 50 mg of Na4EDTA·2H2O and 15 g of agar powder, and deionized water is added to 1 L, NaOH is used to adjust the pH to 6.5, and 121℃ high-temperature high-pressure sterilization is performed for 15 min; 3. The compound MSP-10 is 4-[(5Z)-5-[(4-methoxyphenyl) methylene]-4-oxo-2-thioxo-1,3-thiazolidin-3-yl]-N-phenyl butyramide, which is purchased from Shanghai Tao Technology Biotechnology Co., Ltd., with the catalog number HIT ID: HIT 103730647, and the molecular structural formula of the compound MSP-10 is as follows: ; 4. The preparation method and preservation of the compound MSP-10 mother liquor are as follows: 10 mg of the compound MSP-10 is added to 4 mL of DMSO to make the final concentration of the medicament 2500 ppm. The medicament is short-term stored at-20℃, and long-term stored at-80℃ in a divided manner.

[0046] Example 1 Toxicity determination and results of compound MSP-10 on rice blast fungus The toxicity of the compound MSP-10 on rice blast fungus is determined by the mycelial growth rate method, and the specific process is as follows: Rice blast fungus Guy11 was inoculated on 7 cm CM flat, and cultured in an incubator at 25°C, 16 h light, 8 h dark for 7 days; different doses (0, 10, 20, 30, 40 and 50 ppm) of compound MSP-10 were added to the solid CM medium cooled to 40-50°C to make drug-containing plates; 3 mL of sterile water was added to the plate inoculated with rice blast fungus Guy11 for 7 days under the sterile environment of a clean bench, and the conidia were washed off with a sterilized and cooled coating rod, filtered into a 1.5 mL centrifuge tube with 3 layers of filter paper, and the number of conidia was counted under a light microscope with a hemocytometer, and the conidial liquid was diluted to 1×10 5 spores / mL, and 5 μL of the diluted conidial liquid was added to the center of the drug-containing plate with a 10 μL pipette; each concentration was repeated 3 times; the CM medium containing the same concentration of DMSO was used as a control; the culture dishes were inverted in an incubator at 25°C, 16 h light, 8 h dark for 3 days, and the growth of rice blast fungus mycelium in the experimental group was observed.

[0047] The mycelial diameter was measured by the cross method, and the mycelial growth rate method was used to calculate the inhibition rate of rice blast mycelium under different concentrations of compound MSP-10, and the calculation formula was as follows: Mycelial growth inhibition rate (%) = (average diameter of control group colony - average diameter of treatment group colony) / (average diameter of control group colony - 0.5) × 100.

[0048] The DPS statistical software was used for analysis, and a virulence regression equation was established to calculate the EC 50 of compound MSP-10 on rice blast fungus, and the correlation coefficient r was evaluated to evaluate the virulence level of compound MSP-10 on rice blast fungus.

[0049] Table 1 In vitro virulence of different concentrations of compound MSP-10 on rice blast fungus

[0050] Results analysis: the virulence of different concentrations of compound MSP-10 on rice blast fungus was tested. As shown in Figure 2 , the five concentration treatments of compound MSP-10 set in the experiment had a certain inhibitory effect on the growth of rice blast fungus mycelium, and the higher the concentration of the test, the smaller the average diameter of the colony, and the higher the mycelial growth inhibition rate Figure 3 . Within this concentration gradient range, the inhibition rate of compound MSP-10 on rice blast fungus was between 10% and 80%, indicating that the experimental data was accurate and reliable; the EC 50The value is 34.7035 ppm (Table 1). When the concentration of compound MSP-10 is 50 ppm, the most significant inhibition effect on Magnaporthe oryzae is obtained, and the relative inhibition rate is 76.40% (Table 1).

[0051] Example 2 Determination of pathogenicity of compound MSP-10 on Magnaporthe oryzae and results The pathogenicity of compound MSP-10 on Magnaporthe oryzae was determined by in vitro inoculation method and in vivo rice spraying method, as follows: 1. In vitro inoculation method (1) In vitro barley inoculation The Guy11 strain was inoculated on a 7 cm quantitative CM plate, and after being placed in a culture box (temperature 28°C, light 16 h, dark 8 h) for 7 days, 3 mL ddH2O was added to the CM plate, and the conidia were washed off with a sterilized and cooled coating rod, and filtered into a 1.5 mL centrifuge tube with 3 layers of filter paper. Compound MSP-10 was mixed with the spore solution to prepare spore solutions with different drug concentrations (0, 10, 20, 30, 40 and 50 ppm), and the final concentration of the spore solution was 5×10 4 spores / mL, and the difference in the dose of compound MSP-10 in different treatment groups was supplemented with DMSO to make the final concentration of DMSO consistent; a 10 cm square culture dish was prepared, and the water-absorbing paper was cut into the size of the square dish and placed in the square dish, and then a piece of filter paper of the same size was covered after being wetted with an appropriate amount of water. The barley leaves (5-7 cm) planted for 8-10 days were cut and placed in order in the square dish, 3 drops (20 μL) of spore solution were added to each leaf, and each treatment was repeated 3 times. The spore solution of the control group contained the same concentration of DMSO. The culture dish with leaves was carefully placed in a culture box (temperature 28°C, light 16 h, dark 8 h), and the disease incidence was observed and photographed after 4 days of culture.

[0052] Result analysis: The inhibitory effect of gradient concentrations of compound MSP-10 on Magnaporthe oryzae on in vitro barley leaves was tested. As shown in (A) of Figure 4 , 10-50 ppm of compound MSP-10 had a certain inhibitory effect on Magnaporthe oryzae infection on in vitro barley leaves; and within this range, the higher the concentration, the stronger the inhibitory effect; among them, 40-50 ppm of the compound MSP-10 had a significant inhibitory effect on Magnaporthe oryzae spores on in vitro barley leaves; and 50 ppm of the compound MSP-10 could completely inhibit the occurrence of Magnaporthe oryzae on in vitro barley leaves.

[0053] (2) In vitro rice inoculation The healthy second leaf (about 15 days of culture) of three-leaf stage rice (cultivated by China Rice Research Institute, variety: Oryza sativa CO39) was cut and inoculated in vitro according to the method of barley inoculation in vitro.

[0054] Result analysis: The inhibitory effect of compound MSP-10 at gradient concentrations on rice blast in vitro was tested. As shown in (B) of Figure 4 , among the five concentration treatments of compound MSP-10 set in the experiment, 10 ~ 50 ppm of compound MSP-10 had a certain inhibitory effect on rice blast in vitro; and within this range, the higher the concentration, the stronger the inhibitory effect; among them, 20 ~ 50 ppm of the compound MSP-10 had an obvious inhibitory effect on rice blast in vitro; 40 ppm of the compound MSP-10 could completely inhibit the occurrence of rice blast in vitro.

[0055] As shown in (A) and (B) of Figure 4 , under the same concentration of compound, the control effect of compound MSP-10 on rice blast in vitro was stronger than that on barley leaves in vitro.

[0056] 2. In vivo rice spray method First, the Oryza sativa CO39 variety of rice was sown (30 seeds per pot), and cultured to two weeks old under sufficient light outdoors; the Magnaporthe oryzae was cultured on CM medium for 7 ~ 9 d to obtain spore solution; the compound MSP-10, gelatin solution and spore solution were mixed to prepare spore solution containing drug and gelatin (the final concentration of spores was 5 × 10 4 spores / mL, and the final concentration of gelatin was 0.2%), so that the final concentration of compound MSP-10 was 50 ppm, and the dose difference of compound MSP-10 was supplemented with DMSO to make the final concentration of DMSO consistent. 2 mL of mixed spore solution containing compound and gelatin was evenly sprayed on the rice leaves, and 3 pots of rice were inoculated in each treatment group; the spore solution containing the same concentration of DMSO and gelatin, and the gelatin solution containing 50 ppm of compound MSP-10 but no spores were used as toxicity test controls; the inoculated in vivo rice was placed in a 22°C incubator for dark culture for 48 h, and then placed in a 25°C incubator for 16 h / 8 h light-dark cycle for 3-4 d; the disease incidence of rice leaves was observed and photographed, and the lesion area was calculated by Image J software, and the lesion rate was calculated.

[0057] Lesion rate (%) = lesion area / leaf area × 100 The specific optimization experiment steps are as follows: the concentration of compound MSP-10 for inhibiting rice blast is 50 ppm, 50 ppm of compound MSP-10 is applied at different time points after inoculation of rice blast spore solution, and three groups of experiments are set up: (1) -24 h: compound MSP-10 is sprayed 24 hours before inoculation of rice blast spore solution; (2) 0 h: compound MSP-10 is applied at the same time of inoculation of rice blast spore solution, that is, compound MSP-10 is mixed with rice blast spore solution and sprayed for inoculation; (3) +24 h: compound MSP-10 is sprayed 24 hours after inoculation of rice blast spore solution.

[0058] Each treatment uses 3 pots of rice as repeats, and blank control and DMSO solvent control are set.

[0059] Result analysis: the effect of applying compound MSP-10 at different time points on the prevention and treatment of rice blast in vivo is tested. At the same time, compared with the control group without applying compound MSP-10, the -24 h, 0 h and +24 h treatment groups can significantly inhibit rice blast in vivo (P<0.05) Figure 5 and Figure 6 ), which has practical significance for preventing rice blast. At the same time, it is observed that the rice in the control groups applying only the same concentration of DMSO or the same concentration of compound MSP-10 grows well without obvious adverse effects, indicating that the compound may be safe for rice (P>0.05) Figure 5 ), which has practical significance for preventing rice diseases.

[0060] Example 3 Determination and results of the effect of compound MSP-10 on the development of appressorium of rice blast fungus 1. Determination method of appressorium formation rate After Guy11 is inoculated on a 7 cm CM flat plate and grown in an incubator at 28°C with light for 16 h and darkness for 8 h for 7 days, 3 mL of ddH2O is added to the flat plate, and the conidiophores growing on the aerial hyphae are gently scraped and washed with a coating rod. After filtration through 3 layers of filter paper, the spore solution mother liquor is obtained, and ddH2O is added to dilute it to a final concentration of 5×10 4Spores / mL spore solution, compound MSP-10 and spore solution were mixed to prepare a spore solution containing the drug, so that the final concentration of compound MSP-10 was 50 ppm, and the dose difference of compound MSP-10 was supplemented with DMSO to make the final concentration of DMSO consistent; the same concentration of DMSO was used as a control; 70% ethanol solution was used to clean the artificial hydrophobic membrane, which was placed on a glass slide, 3 artificial hydrophobic membranes were placed on each glass slide, 20 μL of diluted spore solution was added to each hydrophobic membrane, and the prepared glass slide was transferred to a humidification box (add appropriate amount of sterile water), and placed in a 22°C dark incubator for induction. The adhesion cell formation rate was counted at 4 h, 8 h, 16 h and 24 h, respectively, and photographed, and each group of tests was repeated 3 times.

[0061] Results analysis: The effect of 50 ppm compound MSP-10 on the adhesion cell formation rate of Magnaporthe oryzae at different time points was tested. As shown in Figure 7 and Figure 8 , 50 ppm compound MSP-10 had a certain inhibitory effect on the adhesion cell formation at the 4 different time points set in the test. At different stages of Magnaporthe oryzae adhesion cell development (4 h, 8 h, 16 h and 24 h), the exogenous addition of 50 ppm compound MSP-10 significantly reduced the adhesion cell formation rate.

[0062] 2. Method for measuring adhesion cell turgor pressure The process of inducing adhesion cells is basically the same as the adhesion cell formation determination. The glycerol solution with a concentration of 2 M and 3 M was prepared, and after 24 h of induction, the hydrophobic membrane was taken out, and the surface water was gently absorbed with a pipette or water-absorbing paper. An equal volume of glycerol solution was quickly added, and the glass slide was placed at room temperature for 5 min. Then the collapse rate of adhesion cells was quickly counted, i.e. the proportion of collapsed adhesion cells to the total adhesion cells. Each strain was repeated 3 times at each glycerol concentration.

[0063] Results analysis: The effect of 50 ppm compound MSP-10 on the adhesion cell turgor pressure under two different glycerol concentrations was tested. As shown in Figure 9 and Figure 10 , 50 ppm compound MSP-10 had a certain effect on the adhesion cell turgor pressure under the 2 different glycerol concentrations set in the test. Under the glycerol concentrations of 2 M and 3 M, the exogenous addition of 50 ppm compound MSP-10 significantly increased the adhesion cell collapse rate, which indicated that the exogenous addition of 50 ppm compound MSP-10 significantly reduced the turgor pressure of Magnaporthe oryzae adhesion cells.

[0064] 3. Method for determining adhesion cell KI / I2 sugar origin staining KI / I2 solution (60 mg / mL KI, 10 mg / mL I2) was prepared and the process of inducing appressorium was consistent with the above. After 0 h, 8 h, 16 h and 24 h of induction, the hydrophobic membrane with spore solution was taken out from the humidifying box, the surface moisture was gently absorbed by pipette or water-absorbing paper, and the same volume of KI / I2 solution was added by pipette. The cover glass was covered, and the conidia and appressorium glycogen staining was counted and photographed under the optical microscope. Each test was repeated 3 times.

[0065] Result analysis: The effects of 50 ppm compound MSP-10 on glycogen transport and degradation of Magnaporthe grisea at different time points were tested. As shown in Figure 11 , exogenous addition of 50 ppm compound MSP-10 had no effect on glycogen synthesis, but significantly slowed down the transport of glycogen in conidia to appressorium; after 16 h of hydrophobic membrane inoculation, exogenous addition of 50 ppm compound MSP-10 significantly slowed down the rate of glycogen degradation in appressorium, and after 24 h of hydrophobic membrane inoculation, the glycogen degradation in appressorium returned to normal.

[0066] 4. Appressorium Bodipy lipid droplet staining determination method Before inducing appressorium, tricyclazole was added at a ratio of 1 μL of 10 μg / μL to 1 mL of spore solution, in order to inhibit the formation of appressorium melanin and facilitate fluorescence observation. The lipid droplet dye Bodipy (Boron dipyrromethene) was diluted at a ratio of 1:1000. After 0 h, 4 h, 8 h and 24 h of induction, the hydrophobic membrane with spore solution was taken out from the humidifying box, the surface moisture was gently absorbed by pipette or water-absorbing paper, and the same volume of diluted Bodipy dye was added by pipette. The cover glass was covered, and the conidia and appressorium glycogen staining was counted and photographed under the fluorescence microscope. Each test was repeated 3 times.

[0067] Result analysis: The effects of 50 ppm compound MSP-10 on lipid droplet transport and degradation of Magnaporthe grisea at different time points were tested. As shown in Figure 12 , exogenous addition of 50 ppm compound MSP-10 had no significant effect on lipid droplet synthesis, lipid droplet transport and lipid droplet degradation.

[0068] Example 4 Inhibition experiment of compound MSP-10 on other pathogenic fungi This example also detected the toxic effects of compound MSP-10 on other plant pathogenic fungi. The concentration of compound MSP-10 was 50 ppm, and the experimental steps were the same as in Example 1. Other plant pathogenic fungi included Botrytis cinerea B. cinerea , Botrytis cinerea and Fusarium fujikuroi F. fujikuroi .Fusarium fujikuroi Anthracnose of melons ( ) C. orbiculare : Colletotrichum orbiculare Alternaria ( A. alternata : Alternaria alternata ) and Anthrax bacillus fusiforme ( C. acutatum : Colletotrichum acutatum (See Table 2).

[0069] Table 2 Plant pathogens

[0070] The results are as follows Figure 13 and Figure 14 As shown, compound MSP-10 exhibits inhibitory effects on the plate growth of various plant pathogenic fungi, inhibiting the mycelial growth of the five different genera of plant pathogens to varying degrees, especially *Botrytis cinerea* and *Anthracnose cucurbita*, with significant inhibitory effects. These results demonstrate the broad-spectrum antibacterial activity of MSP-10, which may be due to its specific targeting of the Pex19 protein. Given the high homology of Pex19 proteins in different pathogens, MSP-10 can bind to Pex19 homologous proteins in various pathogens, thereby achieving a significant antibacterial effect.

[0071] Example 5 Oryza sativa CO39 rice seeds were placed in a humidified container at 37°C for 48 hours to germinate, and then sown in culture pots (30 seeds / pot) and cultured outdoors in a netted room for 14 days. Compound MSP-10 and gelatin solution were mixed to prepare a drug-containing gelatin solution (final gelatin concentration 0.2%), resulting in final MSP-10 concentrations of 25, 50, and 100 ppm. The dosage difference of MSP-10 was compensated with DMSO to ensure a consistent final DMSO concentration. 2 mL of the mixed spore solution containing the compound and gelatin was evenly sprayed onto rice leaves, with 3 pots of rice inoculated for each treatment group. A gelatin solution containing the same volume of ddH2O and a gelatin solution containing 100 ppm DMSO were used as negative controls; a gelatin solution containing 100 ppm tricyclazole was used as a positive control. After spraying, the live rice plants were placed at 25°C and cultured under a light-dark cycle of 16 h / 8 h for 6-7 days; the growth of the rice was observed and photographed.

[0072] like Figure 15 As shown, compound MSP-10 showed no significant difference in rice growth compared to the control, and did not exhibit any obvious negative effects on rice. This suggests that MSP-10 may be safe for rice while effectively controlling rice blast.

[0073] In summary, the compound MSP-10 has a broad-spectrum bacteriostatic property, and can play an important role in agricultural production as an effective plant pathogen inhibitor. It not only has a significant control effect on rice blast fungus, but also shows good inhibition effect on other various plant pathogens, and does not have obvious negative effect on rice, showing good biological safety. Therefore, the compound MSP-10 is expected to become a new choice of efficient and safe plant pathogen control, and provides strong support for the sustainable development of agricultural production.

[0074] Although the content of the present application has been described in detail by the above preferred embodiments, it should be recognized that the above description should not be considered as a limitation of the present application. After reading the above content, various modifications and alternatives of the present application will be apparent to those skilled in the art. Therefore, the protection scope of the present application should be defined by the appended claims.

Claims

1. Compound MSP-10 was used in the preparation of a fungicide for the control of rice blast fungus. Magnaporthe oryzae Its use in drugs containing ) or / and other pathogenic fungi is characterized by, The chemical structural formula of the compound MSP-10 is shown below: ； The other pathogenic fungi include: *Botrytis cinerea* (… Botrytis cinerea ), Fusarium oxysporum ( Fusarium fujikuroi Anthracnose of melons ( ) Colletotrichum orbiculare Alternaria ( Alternaria alternata ) and Anthrax bacillus fusiforme ( Colletotrichum acutatum Any one or more of the following.

2. The use of compound MSP-10 in the preparation of a drug for inhibiting the mycelial growth and / or spore germination and / or appressorium development of *Oryza sativa* or / and other pathogenic fungi, characterized in that... The other pathogenic fungi include any one or more of the following: Botrytis cinerea, Fusarium oxysporum, Anthracnoseus cucurbita, Alternaria alternata, and Anthracnoseus oxysporum.

3. The application as described in claim 1 or 2, characterized in that, The compound MSP-10 can reduce the turgor pressure of the appressorium of *Bacillus oryzae*, and / or slow down the transport of glycogen from *Bacillus oryzae* conidia to the appressorium, and / or slow down the rate of glycogen degradation in the appressorium.

4. The application as described in claim 1 or 2, characterized in that, The concentration of the compound MSP-10 is 10 to 50 ppm.

5. The application according to claim 4, characterized in that, The concentration of the compound MSP-10 is 30~50 ppm.

6. A drug for controlling rice blast fungus and / or other pathogenic fungi, characterized in that, Including compound MSP-10; The other pathogenic fungi include any one or more of the following: Botrytis cinerea, Fusarium oxysporum, Anthracnoseus cucurbita, Alternaria alternata, and Anthracnoseus oxysporum.

7. The drug according to claim 6, characterized in that, It also includes agriculturally acceptable carriers.

8. A method for controlling rice blast fungus and / or other pathogenic fungi, characterized in that, Spray the pesticide as described in claim 6 or 7 for controlling rice blast fungus and / or other pathogenic fungi onto the leaves of the plant. The other pathogenic fungi include any one or more of the following: Botrytis cinerea, Fusarium oxysporum, Anthracnoseus cucurbita, Alternaria alternata, and Anthracnoseus oxysporum.

9. The method according to claim 8, characterized in that, The concentration of the compound MSP-10 is 10 to 50 ppm.

10. The method according to claim 9, characterized in that, The concentration of the compound MSP-10 is 30 to 50 ppm.

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