Use of an immunosuppressive active substance

Abstract

The application belongs to the technical field of agricultural microbiology and plant protection, and discloses application of an immunosuppressive active substance, and aims to solve the problem that there is no immunosuppressive substance for effectively suppressing plant immunity, improving the colonization efficiency of microorganisms on crops, and improving the biological prevention and control effect in the prior art. The immunosuppressive substance comprises the following applications: application in inhibiting plant immunity, application in promoting plant microbial colonization, application in plant disease biological prevention and control, and application in preparation of agricultural microbial agents, biological prevention and control agents and / or growth promoting agents. The application is suitable for crop health and sustainable agricultural production, and can achieve the effect of plant disease biological prevention and control.

Description

Technical Field

[0001] This invention relates to the fields of agricultural microbiology and plant protection technology, and specifically to the application of an immunosuppressive active substance, particularly the application of an immunosuppressive active substance produced by Pseudomonas. Background Technology

[0002] Pseudomonas spp. are important symbiotic microorganisms widely distributed in the rhizosphere and phyllosphere of plants, possessing the ability to promote plant growth, induce resistance, and inhibit plant pathogens. However, in the natural environment, the crop immune system often recognizes and rejects foreign microorganisms, limiting the colonization efficiency of beneficial Pseudomonas on crops, thus affecting their biocontrol and growth-promoting effects.

[0003] Recent studies have revealed that some microorganisms can regulate plant immune responses by secreting metabolites or effector molecules to establish stable symbiotic relationships. However, there is currently a lack of clear identification methods, preparation methods, and application strategies for immunosuppressive substances that can effectively enhance the colonization efficiency of Pseudomonas aeruginosa on crops. Therefore, providing a substance that can inhibit crop immunity, enhance Pseudomonas aeruginosa colonization, and ultimately improve the effectiveness of biocontrol is of great significance. Summary of the Invention

[0004] The purpose of this invention is to provide an application of an immunosuppressive active substance, which solves the problem of the lack of immunosuppressive substances that can effectively suppress crop immunity, improve the colonization efficiency of microorganisms on crops, and improve the effectiveness of biocontrol.

[0005] To achieve the above objectives, the present invention is implemented using the following technical solution:

[0006] In a first aspect, the present invention provides the application of an immunosuppressive active substance in the preparation of an inhibitor to suppress plant immunity, wherein the immunosuppressive active substance is Orfamide A, and the structure of Orfamide A is as follows:

[0007] .

[0008] Furthermore, the inhibition of plant immunity includes inhibiting the expression of FRK1, PR1, and WRKY22 genes and / or inhibiting reactive oxygen species bursts.

[0009] Furthermore, the method for preparing the immunosuppressive active substance includes the following steps:

[0010] The cultured Pseudomonas was activated and centrifuged to collect the precipitate, thus obtaining the cultured Pseudomonas.

[0011] After adding solvent to Pseudomonas, centrifuge, collect the culture supernatant, and dry to obtain crude extract;

[0012] The crude extract was dissolved in a solvent and then purified by liquid chromatography to obtain an immunosuppressive active substance.

[0013] Optionally, the activation and centrifugation of the Pseudomonas culturatus and the collection of the precipitate includes the following steps:

[0014] The Pf-5 strain was activated on KB solid medium containing 100 µg / mL ampicillin (Amp100);

[0015] The activated Pf-5 strain was cultured on KB liquid medium in Amp100 under light shaking conditions, then transferred to new KB liquid medium with added resin and cultured under heavy shaking conditions. The precipitate was then collected by centrifugation.

[0016] Optionally, the shaking conditions are 200 rpm / min, shaking at 30°C for 16 hours.

[0017] Optionally, the volume ratio of the resin to KB liquid culture medium is 1:40.

[0018] Optionally, the shaking conditions are 200 rpm / min, 30°C, and shaking for 24 hours.

[0019] Optionally, all solvents are methanol, and the methanol added to the crude extract is chromatographic grade methanol.

[0020] Optionally, the drying process is rotary drying.

[0021] Secondly, the present invention provides the application of an immunosuppressive active substance in the preparation of a reagent to promote plant microbial colonization.

[0022] Furthermore, the plant microbial colonization includes the colonization of aboveground microorganisms.

[0023] Furthermore, the microorganisms include Pseudomonas.

[0024] Optionally, the Pseudomonas is the defensive Pseudomonas Pf-5.

[0025] Thirdly, the present invention provides the application of an immunosuppressive active substance in the preparation of biological control reagents for wheat and tomato diseases and reagents for promoting soybean growth.

[0026] Furthermore, the application of immunosuppressive active substances in the preparation of biological control reagents for wheat and tomato diseases and reagents for promoting soybean growth includes the application of reagents that promote the biological control of wheat and tomato diseases by microorganisms and promote soybean growth.

[0027] Furthermore, the wheat diseases include wheat scab, the tomato diseases include tomato bacterial wilt, and the soybean growth promotion is to promote the growth of soybean leaves and roots.

[0028] Fourthly, the present invention provides the application of an immunosuppressive active substance in the preparation of agricultural microbial preparations, biocontrol agents, and / or growth promoters.

[0029] Compared with the prior art, the beneficial effects achieved by the present invention are as follows:

[0030] The immunosuppressive active substance provided by this invention can effectively inhibit plant immunity, promote the colonization efficiency of microorganisms on plants, enhance the control effect of plant diseases, and has the potential to prepare agricultural microbial preparations, biological control agents and growth promoters.

[0031] Experiments on the inhibitory effects of ophamid A on the expression of immune-related genes revealed that ophamid A produced by Pseudomonas thaliana Pf-5 inhibited the upregulation of Arabidopsis thaliana defense response genes FRK1, PR1, and WRKY22 induced by a 22-amino acid flagellin fragment (flavoprotein 22). The inhibitory effect of ophamid A on the reactive oxygen species (ROS) burst induced by flagellin 22 in Arabidopsis thaliana (Col-0) and rice (Zhonghua 11, ZH11) indicates that ophamid A produced by Pseudomonas thaliana Pf-5 can significantly inhibit flagellin 22-induced ROS bursts in plants. Given that flagellin 22's receptor, flagellin-sensing protein 2 (FLS2), co-receptor BAK1, the simultaneous inhibition of downstream responses in this pathway by ophamid A strongly suggests that its target is BAK1. Furthermore, in vitro protein binding assays (MST) showed that olfamed A directly binds to the Arabidopsis BAK1 protein; sequence alignment indicated that BAK1 is highly conserved in crops such as Arabidopsis, rice, maize, and soybean. Therefore, olfamed A can broadly inhibit the immune response triggered by flagellin 22 in different plants by targeting the evolutionarily conserved BAK1 receptor.

[0032] Experiments on the effects of Ophamid A on the colonization of Pseudomonas aeruginosa Pf-5 in crop roots showed that exogenous addition of Ophamid A significantly promoted the colonization level of Pf-5 strain on plants.

[0033] The results of the efficacy of Aofamide A in promoting the defense against Pseudomonas Pf-5 against wheat scab and tomato bacterial wilt showed that the application of the Pf-5 + Aofamide A compound inoculant significantly reduced the incidence of wheat scab and tomato bacterial wilt compared with the application of Pf-5 alone. The results of Aofamide A in promoting the defense against Pf-5 on soybean growth showed that the application of the Pf-5 + Aofamide A compound inoculant resulted in more significant growth improvement in soybeans compared with the application of Pf-5 alone, indicating that the aforementioned Pseudomonas and its related active substances have good potential for biological control under actual agricultural production conditions. Attached Figure Description

[0034] To more clearly illustrate the technical solutions in the embodiments of this disclosure or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of this disclosure. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0035] Figure 1 This is a diagram showing the identification results of the immunosuppressive active substance produced by Pseudomonas Pf-5 in Example 1 of the present invention, wherein... Figure 1 Image A is the liquid phase diagram for the preparation and purification of Pf-5 lipopeptide compounds. Figure 1 In section B, the detection liquid phase diagram is shown after purification of A from Orphamed. Figure 1 C represents the standard curve prepared by the Ophamid A standard.

[0036] Figure 2 This is a diagram illustrating the inhibitory effect of Orphamid A on flagellin 22-induced expression of plant immune-related genes in Example 2 of the present invention. Figure 2 In the figure, A represents the relative expression level of the plant innate immune defense gene FRK1 under different treatment conditions. Figure 2 In the figure, B represents the relative expression level of the systemically acquired resistance marker gene PR1 under different treatment conditions; Figure 2 C represents the relative expression level of WRKY22, a transcription factor gene involved in plant innate immunity, under different treatment conditions.

[0037] Figure 3 This is a diagram illustrating the inhibitory effect of Orphamid A on flagellin 22-induced reactive oxygen species burst in plants, as shown in Example 2 of this invention. Figure 3 Figure A shows the inhibitory effect of Orphamid A on flagellin 22-induced reactive oxygen species burst in Arabidopsis thaliana. Figure 3 Figure B shows the inhibitory effect of Ophamid A on flagellin 22-induced reactive oxygen species burst in rice. Figure 3 In the middle (C), we see the inhibitory effect of Orphamid A on flagellin 22-induced reactive oxygen species burst in tomato. Figure 3 Figure D shows the inhibitory effect of Ophamid A on flagellin 22-induced reactive oxygen species burst in soybean.

[0038] Figure 4 This is a diagram illustrating the interaction between Orphamed A and the plant immune receptor BAK1 in Example 2 of the present invention. Figure 4 The diagram in Figure A is a microthermophoretic (MST) interaction diagram of Orphamid A and the plant immune receptor BAK1. Figure 4In section B, the conservation analysis of the plant immune protein BAK1 in different crops is shown. "*" (asterisk) indicates that the amino acids in this column are completely conserved in all sequences. ":" (colon) indicates that the amino acids in this column are highly conserved, i.e., chemically similar. "." (dot) indicates that the amino acids in this column are somewhat similar, but their conservation is weaker than that of the amino acids in the colon column.

[0039] Figure 5 This is a graph showing the experimental results (CFU count) of Ophamid A promoting the colonization of Pseudomonas aeruginosa Pf-5 in crop roots in Example 3 of the present invention.

[0040] Figure 6 This is a graph showing the efficacy of Orphamid A in promoting the defense against *Pseudomonas* Pf-5 against wheat scab, as described in Example 4 of the present invention. Figure 6 Photo A in the image is from the Oda experiment. Figure 6 In the figure, B represents the control efficacy in field experiments under different treatments.

[0041] Figure 7 This is a graph showing the effect of Orphamid A in promoting the prevention of bacterial wilt of tomato by Pseudomonas Pf-5, as described in Example 4 of the present invention. Figure 7 Photo A in the middle is a picture of a potted plant experiment. Figure 7 In the figure, B represents the control efficacy of pot experiments under different treatments.

[0042] Figure 8 This is a diagram showing the results of the Ophamid A of the present invention promoting the growth of soybeans protected against Pseudomonas Pf-5, wherein... Figure 8 Image A shows a photograph of leaf growth in a soybean pot experiment. Figure 8 Photograph B shows root growth in soybean pot experiments under different treatments. Detailed Implementation

[0043] The above content is further illustrated below with specific embodiments, but it should not be construed as limiting the scope of the invention to the following embodiments. All technologies implemented based on the above content of this invention fall within the scope of this invention.

[0044] It should be understood that all experimental procedures not detailed in the experiment are routine experimental procedures well known to those skilled in the art.

[0045] Example 1

[0046] This embodiment provides the extraction and identification of immunosuppressive substances that defend against Pseudomonas Pf-5.

[0047] Extraction method of immunosuppressive active substance against Pseudomonas Pf-5: Amp100-resistant Pf-5 strain was activated on 10 mL of King's B (KB) solid medium containing Amp100 (working concentration of 100 µg / mL ampicillin). After overnight incubation on 20 mL of KB liquid medium containing Amp100 and gentle shaking, the strain was transferred to 2 L of KB liquid medium and shaken at 200 rpm / min and 30 °C for 24 h. Resin was added at a ratio of 5 mL per 200 mL of medium, and the mixture was shaken for another 24 h. The precipitate was collected by centrifugation, resuspended in methanol, and shaken at 30 °C for 4 h. The supernatant was collected by centrifugation, evaporated to dryness, and then redissolved in chromatographic grade methanol. After filtration, the supernatant was purified by liquid chromatography to obtain the immunosuppressive active substance. The resin used in this experiment is Amberlite XAD16 polystyrene synthetic adsorbent resin, catalog number 03.01.XAD 16. The shaking conditions are 200 rpm / min and shaking at 30℃ for 16 h.

[0048] High-performance liquid chromatography (HPLC) was used to determine immunosuppressive active substances. The results of the determination of immunosuppressive active substances are as follows: Figure 1 As shown, where, Figure 1 Image A is the liquid phase diagram for the preparation and purification of Pf-5 lipopeptide compounds. Figure 1 In section B, the detection liquid phase diagram is shown after purification of A from Orphamed. Figure 1 C represents the standard curve prepared from the alphamid A standard, used for subsequent determination of alphamid A concentration. The results show that the immunosuppressive active substance against Pseudomonas Pf-5 was identified as alphamid A. The structural formula of alphamid A is as follows:

[0049] .

[0050] Example 2

[0051] This embodiment provides a study on the inhibition of plant immunity by Orphamid A produced by Pseudomonas Pf-5.

[0052] This study investigated the effect of ophomede A on the inhibition of immune-related gene expression. Specifically, it examined the effect of ophomede A on flagellin 22-induced plant immune responses. To assess the inhibitory effect of ophomede A on flagellin 22-induced plant immune responses, Arabidopsis thaliana (Col-0) was planted in soil for 28 days. Plants were then treated with sterile water, ophomede A aqueous solution, flagellin 22, and ophomede A + flagellin 22. Treatment involved injecting 1 mL of the prepared aqueous solution into the underside of leaves using a sterile syringe. After treatment, plant samples were collected, total RNA was extracted, and cDNA was synthesized via reverse transcription. Real-time quantitative PCR was used to detect the expression levels of defense-related genes FRK1 and PR1, with ACTIN2 (α-actin-2) used as an internal control gene. The relative gene expression level was determined using a 2-1... ⁻ΔΔCt The method calculates the inhibitory effect of ophomede A on the upregulation of flagellin 22-induced defense genes by comparing the changes in gene expression induced by ophomede A in the ophomede A treatment group and the control group.

[0053] The inhibitory effect of Ophamid A on the expression of immune-related genes is as follows: Figure 2 As shown, where, Figure 2 In the figure, A represents the relative expression level of the plant innate immune defense gene FRK1 under different treatment conditions. Figure 2 In the middle B, the relative expression level of the systemically acquired resistance marker gene PR1 under different treatment conditions is represented. Figure 2 In the figure, C represents the relative expression level of WRKY22, a transcription factor gene involved in plant innate immunity, under different treatment conditions. The results showed that Ophamid A, produced by the defense against Pseudomonas Pf-5, inhibited the upregulation of flagellin 22-induced FRK1, PR1, and WRKY22 genes.

[0054] Method for detecting the inhibitory effect of alphamid A on plant immune response: To detect the inhibitory effect of alphamid A on reactive oxygen species burst in plant immune response, healthy leaves of Arabidopsis thaliana (Col-0), rice (Zhonghua 11, ZH11), soybean (Tianlong No. 8), and tomato (RG-PtoS) were selected. Leaf discs were prepared using a perforator and placed in sterile deionized water for overnight equilibration in the dark. Equal volumes of sterile water and alphamid A solution were added to each well. The treatment group consisted of a solution group, an aqueous solution group containing flagellin 22, and an aqueous solution group containing both olfamed A and flagellin 22. Luminol and horseradish peroxidase were then added to each treatment group. The changes in luminescence signal were continuously monitored using a chemiluminescence detection system. The relative luminescence intensity (RLU) per unit time was used to reflect the level of reactive oxygen species (ROS) burst. The inhibitory effect of olfamed A on ROS burst in plants was evaluated by comparing the differences in ROS levels between the flagellin 22 and flagellin 22 + olfamed A treatment groups.

[0055] The inhibitory effect of Ophamid A on reactive oxygen species (ROS) bursts in Arabidopsis thaliana, rice, soybean, and tomato plants is as follows: Figure 3 As shown, this demonstrates that defense against Ophamid A produced by Pseudomonas Pf-5 can significantly inhibit flagellin 22-induced plant reactive oxygen species bursts.

[0056] Method for detecting the interaction between ophamid A and the plant immune receptor BAK1: To detect the direct binding ability of ophamid A to BAK1 protein, micro-thermophoresis (MST) was used. Recombinantly expressed and purified plant immune receptor BAK1 protein was labeled with a fluorescent dye as the target molecule, and ophamid A was serially diluted with buffer (from high to low concentration, a total of 16 concentration points) as the ligand molecule. Equal volumes of fluorescently labeled BAK1 protein were mixed with each concentration of ophamid A and incubated at room temperature for 10 minutes. The mixture was then aspirated into a capillary tube and placed in an MST instrument to measure the thermophoretic signal. A binding curve was fitted with ophamid A concentration on the x-axis and normalized fluorescence signal on the y-axis, and the equilibrium dissociation constant KD was calculated. A blank control without ophamid A was also included. The interaction analysis diagram of ophamid A and the plant immune receptor BAK1 is shown below. Figure 4 As shown, Figure 4 A in the figure represents the interaction diagram of ophomede A and the plant immune receptor BAK1 by microthermophoresis (MST). The results show that ophomede A has direct binding activity with BAK1 protein (KD=0.52μM). Figure 4 B in the figure represents the conservation analysis of the plant immune protein BAK1 in different crops (Arabidopsis thaliana, rice, wheat, tomato, etc.). Multiple sequence alignment is used to show the degree of amino acid conservation of BAK1 homologs in key structural domains.

[0057] Example 3

[0058] This embodiment provides a study on the use of Orphamid A produced by Pseudomonas Pf-5 to promote the colonization of Pseudomonas on plants.

[0059] Experiment on the colonization of Pseudomonas aeruginosa by Ophamid A produced by Pf-5: To analyze the effect of Ophamid A on the colonization ability of Pf-5 in plant roots, Pf-5 was used as the experimental material. The strain was cultured in liquid medium to the logarithmic growth phase, and the cells were collected by centrifugation and washed. The cultured plants were aseptic Arabidopsis seedlings with similar growth period of 7 days and inoculated with Pf-5 and Pf-5 + Ophamid A. Treatment groups: (1) 1 / 2 MS liquid medium + Pf-5 with resistance to Amp100; (2) 1 / 2 MS liquid medium + Pf-5 with resistance to Amp100 + Ophamid A. Experimental treatment: Arabidopsis thaliana seedlings and each treatment group were co-incubated at 25℃ and 100 rpm / min on a shaker for 30 minutes. The seedlings were then removed from the mixed culture using forceps; the bacterial suspension on the surface of each plant was absorbed with sterile absorbent paper; and the seedlings were transplanted onto 1 / 2 MS solid medium and incubated again for 48 hours. Seven days after inoculation, plant tissue samples were collected, and the samples were serially diluted and spread onto the medium. The number of colonies formed (in CFU) was counted, and the colony level was expressed as colony count per unit.

[0060] The experimental results (CFU count) of the effect of Ophamid A on promoting the colonization of Pseudomonas Pf-5 in crop roots are as follows: Figure 5 As shown, the results showed that exogenous addition of Ophamid A significantly promoted the colonization level of Pf-5 strain on plants.

[0061] Example 4

[0062] This embodiment provides a study on the efficacy of Ophamid A produced by Pseudomonas Pf-5 in promoting the prevention of wheat scab and tomato bacterial wilt, as well as promoting soybean plant growth.

[0063] To verify the biocontrol efficacy of Pseudomonas aeruginosa preparations under crop production conditions, a field trial was conducted in a typical wheat-growing area. Wheat fields with uniform growth were selected as the test subjects. Pseudomonas aeruginosa Pf-5 and Pf-5 + ophamid A were prepared as inoculants. During the wheat heading and flowering stage, the inoculants were applied to the crop plants via foliar spraying, with a water treatment serving as a control. The experiment was divided into three groups: a control group sprayed with water, a Pf-5 group sprayed with Pseudomonas aeruginosa Pf-5, and a Pf-5 + ophamid A group sprayed with Pf-5 + ophamid A. These treatments could be applied once or multiple times, preferably with a second application 3-7 days after the first application. In this example, the second application was performed five days after the first application.

[0064] The severity of disease on a single plant was evaluated using a disease grading system of 0-4, where: 0, no visible lesions; 1, diseased spikelet proportion <25%; 2, diseased spikelet proportion 25-50%; 3, diseased spikelet proportion 50-75%; and 4, diseased spikelet proportion >75%.

[0065] The disease index is calculated using the following formula: Disease index = Σ (disease level × corresponding number of plants) / (highest level × total number of plants surveyed) × 100.

[0066] Further calculations of field control efficacy were performed using the following formula: control efficacy (%) = (control disease index − treatment disease index) / control disease index × 100%.

[0067] Orphamid A enhances the protective efficacy against wheat scab by Pseudomonas Pf-5. Figure 6 As shown, where, Figure 6 Photo A in the image is from the Oda experiment. Figure 6 Figure B represents the control efficacy in field experiments under different treatments. The results show that the application of Pseudomonas Pf-5 + Ofamid A compound inoculant significantly reduced the incidence of wheat scab compared to the application of Pf-5 Pseudomonas inoculant alone.

[0068] To verify the control effect of Ofamid A on bacterial wilt of tomato under pot conditions, uniformly grown tomato seedlings (4-6 true leaves) were transplanted into flowerpots, one seedling per pot, and managed routinely. Pseudomonas Pf-5 and Pf-5 + Ofamid A were prepared as inoculants. Seven days after transplanting, the tomato seedlings were inoculated with a Ralstonia solanacearum suspension (1×10⁻⁶) via root drenching. 8 (CFU / mL), 10 mL per plant. The first treatment was administered within 24 hours of inoculation by drenching the roots of the plants with the above-mentioned inoculum, 10 mL per plant. A water treatment served as a control. The experiment was divided into three groups: the control group sprayed with water, the Pf-5 group sprayed with Pseudomonas Pf-5, and the Pf-5 + Ophamid A group sprayed with Pf-5 + Ophamid A. In this example, a second spray was performed seven days after the first spray.

[0069] The severity of disease on individual plants was evaluated using a disease grading system ranging from 0 to 4, where: 0, no wilting symptoms; 1, 1%-25% leaf wilting; 2, 26%-50% leaf wilting; 3, 51%-75% leaf wilting; and 4, 76%-100% leaf wilting or plant death. The disease index was calculated using the following formula: Disease Index = Σ(Disease Grade × Corresponding Number of Plants) / (Highest Grade × Total Number of Plants Surveyed) × 100. The control effect was further calculated using the following formula: Control Efficacy (%) = (Control Disease Index − Treatment Disease Index) / Control Disease Index × 100%.

[0070] Orphamid A enhances the protective efficacy against Tomato bacterial wilt caused by Pseudomonas Pf-5. Figure 7 As shown, where, Figure 7 Photo A in the middle is a picture of a potted plant experiment. Figure 7 Figure B represents the control efficacy of pot experiments under different treatments. The experimental results show that the application of Pseudomonas Pf-5 + Aofamide A compound inoculant significantly reduced the incidence of bacterial wilt in tomatoes compared with the application of Pf-5 Pseudomonas inoculant alone.

[0071] To verify the effect of Aofamide A on promoting the growth of soybeans by *Pseudomonas aeruginosa* Pf-5, soybean seeds of uniform growth (such as Zhonghuang 13) were sown in pots with 3 seedlings per pot and managed routinely. *Pseudomonas aeruginosa* Pf-5, alone or in combination with Aofamide A, was prepared as a microbial agent. When the soybean seedlings reached the stage where the first trifoliate leaf was fully unfolded, the microbial agent was applied to the plants via foliar spraying, with a water treatment serving as a control. The experiment consisted of three groups: a control group sprayed with water, a Pf-5 group sprayed with the Pf-5 microbial agent, and a Pf-5 + Aofamide A group sprayed with a mixed microbial agent. Each treatment was followed by a second spraying every two days after the first application, and multiple sprayings were performed. Fourteen days after treatment, the growth of the above-ground leaves and underground roots of the soybean plants was observed and recorded, and representative plant photos of leaves and roots were taken using a digital camera. Figure 8 The study demonstrated the effect of Aofamide A on the overall growth of soybean plants protected against Pseudomonas Pf-5. Compared with the control group, soybean plants treated with Pf-5 alone had slightly larger leaves and slightly longer roots. In contrast, soybean plants treated with the combined treatment of Pf-5 and Aofamide A had significantly larger and darker green leaves and a significant increase in overall plant biomass, indicating that Aofamide A can synergistically promote soybean growth with Pf-5. Figure 8 In the image, A represents a photograph of the leaf growth in a soybean pot experiment. It can be seen that the Pf-5+Ofamid A treatment group has the largest leaf area and the darkest leaf color. Figure 8 B in the figure shows root growth photos of soybean pot experiments under different treatments. The root system of the combined treatment group is more developed, with elongated taproots and significantly better number and length of lateral roots than the control group and the single Pf-5 application group.

[0072] This indicates that the aforementioned Pseudomonas and its related active substances have good potential for biological control under actual agricultural production conditions.

[0073] Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, and not all embodiments. Those skilled in the art can obtain other embodiments based on these embodiments without creative effort, and these embodiments all fall within the protection scope of the present invention.

Claims

1. The application of an immunosuppressive active substance in the preparation of an inhibitor to suppress plant immunity, characterized in that, The immunosuppressive active substance is Ophamid A; the inhibition of plant immunity includes inhibition of... FRK1, PR1 and WRKY22 Gene expression and / or inhibition of reactive oxygen species bursts.

2. The application of the immunosuppressive active substance according to claim 1 in the preparation of inhibitors for suppressing plant immunity, characterized in that, The method for preparing the immunosuppressive active substance includes the following steps: The cultured Pseudomonas was activated and centrifuged to collect the precipitate, thus obtaining the cultured Pseudomonas. After adding solvent to Pseudomonas, centrifuge, collect the culture supernatant, and dry to obtain crude extract; The crude extract was dissolved in a solvent and then purified by liquid chromatography to obtain an immunosuppressive active substance.

3. The application of an immunosuppressive active substance in the preparation of a reagent to promote plant microbial colonization, characterized in that, The immunosuppressive active substance is Ophamid A; the plant microbial colonization includes microbial colonization of the aboveground parts of the plant; the microorganisms include Pseudomonas.

4. The application of an immunosuppressive active substance in the preparation of biological control reagents for wheat and tomato diseases and reagents for promoting soybean growth, characterized in that, The immunosuppressive active substance is Ophamid A; the wheat diseases include wheat scab, the tomato diseases include tomato bacterial wilt, and the promotion of soybean growth refers to promoting the growth of soybean leaves and roots.

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