Method for disease control and rejuvenation after transplantation of ancient and famous trees

Abstract

The invention relates to the field of prevention and treatment of ancient plants, and particularly discloses a method for disease prevention and rejuvenation after transplantation of ancient and famous trees. The method comprises the following steps: taking tissues and rhizosphere soil of diseased plants of ancient and famous trees, sequencing amplification products after extracting DNA (Deoxyribose Nucleic Acid), counting the types and quantity of OTUs in the diseased plants and the rhizosphere soil, analyzing the composition and diversity of microbial flora, and determining growth critical values of harmful pathogenic bacteria and induced pathogenic bacteria; and according to the growth condition of the microbial flora, sequentially performing macro-environment physical measure prevention and control to improve the growth environment of diseased plants, chemical prevention and control to inhibit harmful bacteria, and biological prevention and control to form a beneficial flora effect. By adopting the method, diseases and pests of the ancient peonies can be effectively prevented and treated, a benign microbial ecological system is established around the transplanted plants, and the autoimmunity of the plants is enhanced, so that the purposes of comprehensively preventing and treating diseases and promoting rejuvenation of the ancient peonies are achieved.

Description

Technical Field

[0001] The present application relates to the field of garden cultivation, and more specifically, to a method for preventing and controlling diseases and rejuvenating ancient and famous trees after transplantation. Background Art

[0002] As dual natural and cultural heritage, ancient and famous trees hold irreplaceable value in ecological conservation and cultural heritage. Ancient peonies, in particular, have attracted widespread attention for their rich history and unique ornamental value. However, in the context of modern urbanization, these trees face severe challenges to their survival due to environmental changes, natural disasters, and other factors. In particular, during transplantation, drastic changes in the rhizosphere soil environment can lead to poor growth and frequent pests and diseases, severely impacting their physiological stability and lifespan. Therefore, the scientific and effective conservation and management of ancient peonies has become a crucial issue in the field of landscaping.

[0003] Currently, the main measures for managing the health of ancient tree peonies include traditional pesticide control, fertilization and irrigation, and physical isolation. For example, regular spraying of chemical pesticides is used to control pests and diseases; organic fertilizers are added to improve soil fertility; or protective netting is installed to prevent external damage. While these methods can alleviate some problems in the short term, they have many limitations in practical application. Specifically, chemical pesticides may cause environmental pollution and the risk of pesticide resistance; biological control is environmentally friendly but slow in effectiveness; physical and mechanical control is simple to operate but has limited applicability; and relying solely on external nutrient inputs makes it difficult to fundamentally optimize the rhizosphere microecological environment.

[0004] These conventional methods generally suffer from the inability to precisely regulate the structure of the rhizosphere microbial community, resulting in the ancient tree peony remaining susceptible to adverse stresses after transplantation and lacking long-term, stable physiological regulatory mechanisms to support its healthy recovery and sustainable development. This necessitates the exploration of new, more environmentally friendly and efficient approaches to address the current challenges faced by ancient and famous trees during transplantation and rejuvenation. Summary of the Invention

[0005] In order to solve the above technical problems, the present application provides a method for preventing and controlling diseases and rejuvenating ancient and famous trees after transplantation.

[0006] This application provides the following technical solutions: In one aspect, the present application provides a method for disease prevention and rejuvenation of ancient and famous trees after transplantation, comprising: 1. Take tissues and rhizosphere soil from diseased plants of ancient and famous trees, extract DNA, and sequence the amplified products. Count the types and numbers of OTUs in the diseased plants and rhizosphere soil, and analyze the composition and diversity of the microbial flora. 2. Based on the growth of microbial flora, determine the critical point at which microorganisms exert harmful effects. Combined with the current growth environment and pH test results, implement physical environmental prevention and control measures to improve the environment and make it suitable for the growth of ancient and famous trees; 3. Based on the detected pathogenic microorganisms and the characteristics of the diseases of ancient and famous trees, chemical control is carried out during the growth of the plants through multiple root irrigation treatments and application of biological fertilizers to inhibit harmful bacteria; 4. Biological control is carried out by applying bacterial agents and fertilizers containing beneficial bacteria to the plants to form a beneficial bacterial community effect.

[0007] By adopting the above technical solution, it is possible to comprehensively analyze the composition and diversity of the microbial flora in the diseased plants of ancient and famous trees and their rhizosphere soil, accurately determine the critical point of action of harmful microorganisms, and formulate targeted macro-environmental physical prevention and control measures in combination with environmental factors, effectively improving soil conditions to promote plant growth. At the same time, chemical control is achieved through multiple root irrigation treatments and the application of biological fertilizers, which significantly inhibits the reproduction and spread of harmful bacteria. In addition, after chemical control, the abundance of microbial flora in the rhizosphere soil decreases, and harmful bacteria are effectively suppressed, but many beneficial bacteria are also affected. Therefore, it is necessary to use bacterial agents and fertilizers containing beneficial bacteria for biological control, increase the synthesis of beneficial flora in the rhizosphere soil, and play its beneficial role, establish a benign microbial ecosystem around the plant, and enhance the plant's own resistance, so as to achieve the purpose of comprehensive disease prevention and control and promote the rejuvenation of ancient and famous trees.

[0008] Furthermore, the above-mentioned ancient trees and famous trees include ancient peonies, ancient cypresses, ancient pines, and ancient ginkgo trees.

[0009] By adopting this technical solution, it can be specifically applied to specific types of ancient and valuable trees, such as ancient peonies, ancient cypresses, ancient pines, and ancient ginkgo trees. This limitation further clarifies the scope of application of the method, facilitating refined management and prevention based on the characteristics and needs of different trees, thereby improving the overall effectiveness of disease prevention and rejuvenation.

[0010] Furthermore, in the above step (1), the method for obtaining the composition and diversity of the microbial flora includes: Tissues and rhizosphere soil of diseased plants of ancient and famous trees were taken as samples, and genomic DNA was extracted. The extracted DNA was amplified by PCR using universal primers 16S and ITS. The amplified products were purified and sequenced. The sequence data obtained by sequencing were clustered according to a similarity of 97% and divided into different OTUs. The OTUs were identified and the classification information was determined. The composition and diversity of the microbial flora were analyzed based on the types and numbers of the OTUs.

[0011] By employing this technical approach, we can accurately capture the composition and diversity of the microbial communities in diseased ancient and renowned trees and their rhizosphere soil. Specifically, genomic DNA extracted from the roots, stems, leaves, and rhizosphere soil of diseased trees was amplified and sequenced using universal primers 16S and ITS. OTUs were then clustered and identified based on 97% similarity, accurately reflecting the structural characteristics of the microbial community. This method improves the reliability and resolution of the analysis results, laying the foundation for the subsequent development of targeted disease prevention and control strategies.

[0012] Furthermore, in the above step (2), the macro-environmental physical measures prevention and control method includes: Bury the roots of the plants with a composite material containing vermiculite, perlite, peat soil and sand, and add humic acid conditioners to adjust the soil pH to a weak alkaline level and increase soil permeability; During the growth of the plants, spray leaf fertilizer, increase soil permeability by watering and adding drainage pipes to ensure that the soil is not short of water or has water accumulation; Prune branches in time to retain healthy branches, and carry out oxygen-free carbonization incineration on the pruned branches.

[0013] By adopting the above technical solution, the growth environment of ancient and famous trees can be effectively improved. The specific effects are as follows: Soil improvement: Use a composite material containing vermiculite, perlite, peat soil and sandy soil to bury the roots of the plants, and add humic acid conditioners to adjust the soil pH to a weak alkaline and increase soil permeability, which helps to optimize the soil structure, improve soil aeration and water retention capacity, and thus promote the healthy development of the root system of ancient and famous trees. Preferably, the mass ratio of vermiculite, perlite, peat soil and sandy soil is 1:1:(1.5-2.5):(2.5-3.5).

[0014] Microenvironment improvement: Spray foliar fertilizer during the growth of the plant, and increase soil permeability by watering and adding drainage pipes to ensure that the soil is not short of water or has water accumulation. This can meet the water needs of the plant while preventing root rot and further ensure the normal metabolic activities of the plant.

[0015] Pruning: Timely pruning to retain healthy branches, and anaerobic carbonization and incineration of the pruned branches can not only reduce the risk of disease and pest transmission and prevent the spread of diseases to healthy plants; it can also concentrate nutrients to the dominant parts, avoid useless branches consuming nutrients, and improve the overall vitality and stress resistance of the plant.

[0016] Furthermore, in the above step (3), the method of chemical control during the growth of the plant includes 4-5 root irrigation treatments, each with an interval of 5-10 days, and fertilizing with biological fertilizer once after each root irrigation treatment.

[0017] The applicant's detection of pathogenic microorganisms revealed that under conditions of high temperature and high humidity, the pathogenic fungus Alternaria is the main pathogen causing plant rot, Aspergillus niger causes root mold rot, Aspergillus tubingensis is the pathogen of leaf spot, and Mucor racemosus is the pathogen of brown spot. The simultaneous harmful effects of these fungi cause the symptoms to be severe. The bacterial diseases are mostly caused by high temperature, high humidity, and uncomposted organic fertilizers, with the main pathogen being Ralstonia solanacearum. Bacterial wilt is one of the world's top ten plant pathogens and a typical vascular disease that can affect roots, stems, and leaves. Chemical methods are used to control these harmful pathogens. By adopting the above technical solution, 4-5 root irrigation treatments are carried out at intervals of 5-10 days, and applying biological fertilizer after each root irrigation, the growth of harmful bacteria can be effectively suppressed while promoting the establishment of beneficial bacterial communities. This method, through multiple treatments during plant growth, gradually improves the rhizosphere microecological environment, enhances the disease resistance of ancient and famous trees, and thus achieves effective disease control and plant health rejuvenation.

[0018] Furthermore, the root irrigation treatment includes: First root irrigation treatment: Use diluted carbendazim suspension or chlorothalonil wettable powder combined with thiamethoxam to evenly irrigate the roots for disinfection; Second root irrigation: For plants with black spot, ring rot or anthracnose, use carbendazim wettable powder combined with mancozeb wettable powder, or thiophanate-methyl wettable powder combined with mancozeb wettable powder for root irrigation. Third root irrigation: Use thiophanate-methyl wettable powder or mancozeb wettable powder combined with Jiamei Bonus or benomyl for root irrigation to prevent and control root and soil diseases. The fourth root irrigation treatment: Zhongshengmycin and copper rosinate were mixed for root irrigation, and a mixed solution of carbendazim, azoxystrobin and pyraclostrobin was sprayed on branches and leaves; The fifth root irrigation treatment: use a mixture of zhongshengmycin and pyraclostrobin for root irrigation, and use a mixture of methylthiocarb and azoxystrobin to spray on branches and leaves.

[0019] By adopting the above technical solution, we can achieve effective prevention and control of diseases and rejuvenation of ancient and famous trees after transplantation. The specific effects are as follows: The first root irrigation treatment uses a diluted suspension of carbendazim or chlorothalonil wettable powder combined with thiamethoxam for uniform root disinfection. This effectively kills various fungi and insect larvae in the soil, reducing the risk of initial infection and creating favorable conditions for subsequent growth. The second root irrigation treatment targets plants affected by scab, ring rot, or anthracnose. This combination of specifically formulated agents can precisely target common fungal diseases, significantly improving cure rates and reducing recurrence rates.

[0020] The third root drench treatment utilized a combination of thiophanate-methyl wettable powder or mancozeb wettable powder with either Jiamei Hongli or benomyl. This not only enhanced the fungicidal efficacy but also promoted root development, boosting absorption capacity and protecting against potential threats. The fourth root drench treatment combined a combination of zhongshengmycin and copper rosinate to deeply control pathogens in the root zone. Foliar sprays of a mixture of carbendazim, azoxystrobin, and pyraclostrobin were also applied to fully cover the aboveground areas, forming a three-dimensional protective system and enhancing overall resilience. Zhongshengmycin exhibits high activity against bacterial and some fungal plant diseases, while also increasing yield. It also protects against soft rot, bacterial wilt, leaf blight, ring rot, anthracnose, leaf spot, and rotten core. Finally, the fifth root drench treatment utilized a combination of zhongshengmycin and pyraclostrobin to further consolidate the previous gains. A combination of thiophanate-methyl and azoxystrobin was simultaneously applied to strengthen the surface protective barrier, ensuring long-term, stable restoration of health. Azoxystrobin has excellent activity against nearly all fungal diseases, including powdery mildew, rust, glumebode, web blotch, downy mildew, and rice blast. Pyraclostrobin has protective, therapeutic, and leaf-penetrating properties, and is highly effective against root rot, black rot, brown spot, leaf spot, fusarium head blight, and wilt.

[0021] Furthermore, in the fourth root irrigation treatment, the agents used are: The root irrigation agent is a mixture of 600-800 times of 3% bromomycin and 800-1200 times of 20% copper rosinate; the agent applied to branches and leaves is a mixture of 700-900 times of 80% carbendazim, 1400-1600 times of 25% myclobutanil and 800-1000 times of 15% pyraclostrobin.

[0022] By adopting this technical solution, the root drench agent is a mixture of 600-800 times the concentration of 3% zhongshengmycin and 800-1200 times the concentration of 20% copper rosinate. It can effectively inhibit the growth and reproduction of various pathogenic fungi and bacteria, reduce the risk of root infection, and enhance plant immunity, thereby improving the plant's resistance to stress. The agent applied to branches and leaves contains a mixture of three ingredients: 700-900 times the concentration of 80% carbendazim, 1400-1600 times the concentration of 25% azoxystrobin, and 800-1000 times the concentration of 15% pyraclostrobin. This not only comprehensively covers the prevention and control needs of different types of foliar diseases, but also reduces the possibility of drug resistance from single drug use, ensuring stable and long-lasting treatment effects.

[0023] Furthermore, in the fifth root irrigation treatment, the agents used are: The root irrigation agent is a mixture of 600-800 times of 3% Zhongshengmycin and 1400-1600 times of 15% pyraclostrobin; The agent applied to branches and leaves is a mixed solution formed by mixing 700-900 times of 50% methotrexate and 1400-1600 times of 25% myclobutanil.

[0024] By adopting the above technical solution, it is possible to achieve effective prevention and control of diseases and rejuvenation of ancient and famous trees after transplantation. Specifically, using a mixed solution formed by compounding 600-800 times of 3% zhongshengmycin with 1400-1600 times of 15% pyraclostrobin for root irrigation can specifically inhibit the growth and reproduction of various pathogenic fungi and bacteria, effectively reduce the incidence of diseases in the roots and soil, and promote healthy growth of plants. At the same time, applying a mixed solution formed by compounding 700-900 times of 50% methyl thiophanate with 1400-1600 times of 25% azoxystrobin to branches and leaves further enhances the prevention ability of leaf and stem diseases, improves the overall disease resistance of the plant, reduces drug residues while ensuring good prevention and control effects.

[0025] Furthermore, the biological control in the above step (4) includes the combined application of EM composite microbial agent, Trichoderma agent, Bacillus subtilis agent and AM microbial fertilizer.

[0026] By adopting the above technical solution, bacterial fertilizer can promote the production of stimulants, regulating and promoting plant growth and development; microbial agents can promote the production of gibberellins, auxins, and other active substances. The bacterial species in the bacterial fertilizer and microbial agent have the ability to secrete antibiotics and various active enzymes, inhibiting or killing pathogenic fungi and bacteria, and enhancing crop resistance to disease and stress. The combination of EM compound microbial agent + Trichoderma agent + Bacillus subtilis agent + AM microbial fertilizer, combining different beneficial bacteria, not only provides beneficial bacteria and forms a beneficial bacterial community, but also, combined with physical environmental regulation, further facilitates the beneficial effects of the bacterial community. Specifically, the application of EM compound microbial agent promotes the proliferation of beneficial microorganisms in the soil, enhances the ability of plant roots to absorb nutrients, and improves plant immunity. The application of Trichoderma agent can inhibit the growth of various soil-borne pathogenic fungi, reducing the incidence of diseases. The use of Bacillus subtilis agent further strengthens the antagonistic effect against harmful bacteria and fungi while promoting plant growth and development. Applying AM bacterial fertilizer directly to the roots of plants helps establish a symbiotic relationship, improves the balance of rhizosphere microecology, and increases nutrient utilization.

[0027] Under the combined effect, this method can build a virtuous cycle system that is conducive to the growth of ancient and famous trees from multiple levels, achieving the dual goals of preventing and controlling diseases and rejuvenation.

[0028] Furthermore, the biological control in step (4) above includes: Use AM fertilizer to apply directly to the roots of plants; Mix 200-500 times diluted EM fungal agent with sugar substance in a mass ratio of 1:0.8-1.2 and apply; Use 400-600 times diluted EM fungus agent, 800-1200 times diluted Trichoderma harzianum fungus agent and sugar substances to mix and apply; EM fungal agent diluted 400-600 times, Trichoderma harzianum fungal agent 800-1200 times, and Bacillus subtilis 700-900 times were mixed with sugar substances and applied.

[0029] By adopting the above technical solutions, the health and survival rate of ancient and famous trees after transplantation can be effectively improved. The details are as follows: Directly inoculating and applying AM fertilizer to the roots of century-old peonies can promote root development, enhance nutrient absorption, and improve stress resistance, thereby improving the overall growth of these ancient and valuable trees. AM arbuscular mycorrhizae, obtained from the Chinese Arbuscular Mycorrhizal Fungi Resource Bank, are rapidly propagated in the laboratory to create a finely granulated mixture of a substrate rich in fungal spores and mycelium and plant root segments. After air-drying, an equal volume of humus is added and mixed thoroughly to create the AM fertilizer specifically for ancient peonies.

[0030] Mixing a 200-500-fold diluted EM inoculant with sugars in a specific ratio before application helps quickly establish beneficial microbial communities, inhibiting the growth of harmful bacteria while providing plants with the necessary carbon source and further strengthening their immune function. EM is a composite inoculant containing over 80 microorganisms from 10 genera across five families, including photosynthetic bacteria, yeasts, lactic acid bacteria, actinomycetes, and mycorrhizal bacteria, all in a single state.

[0031] The combined application of a 400-600-fold diluted EM agent with an 800-1200-fold diluted Trichoderma harzianum agent and sugars not only enhances antagonism against a variety of pathogenic fungi but also promotes the formation of soil aggregate structure, improving soil aeration and water retention, and fostering a microecological environment more suitable for the growth of ancient and famous trees. Trichoderma harzianum colonizes plant roots and produces compounds that stimulate plant growth and induce plant defense responses, improving the root microenvironment and enhancing plant growth and disease resistance. Adding a 700-900-fold diluted Bacillus subtilis synergizes with the aforementioned ingredients. Bacillus subtilis has strong protease, lipase, and amylase activities, promoting the degradation of nutrients in the soil and enabling plants to more fully absorb and utilize nutrients, achieving the goal of comprehensive prevention and control.

[0032] In summary, this application has the following beneficial effects: This application conducts high-precision sequencing analysis of the microbial flora in diseased plants of ancient and famous trees and their rhizosphere soil, which can accurately grasp their composition and diversity, and provide a scientific basis for subsequent targeted prevention and control. Combining physical and chemical measures to improve the growth environment not only optimizes the soil structure and pH value, but also effectively prevents water stagnation and disease spread, significantly improving the overall adaptability of the plants. The comprehensive use of multiple root irrigation treatments and the application of biological fertilizers has achieved effective inhibition of harmful bacteria, and promoted the healthy recovery of plants by introducing beneficial bacteria, enhancing long-term disease prevention and stress resistance.

[0033] This application is based on ancient peony as the research object, and it is found that after the above-mentioned prevention and control measures in the previous year, the diseases of the plants in the second year are significantly alleviated, and the vast majority of diseased plants have been effectively cured, and their growth and development performance is normal. By carrying out soil microbial detection on the rhizosphere soil before and after prevention and control, the results show that after prevention and control, the fungal species in the rhizosphere soil of the plants are reduced, and the test results of the proportion of harmful bacteria are significantly reduced, which shows that the early prevention and control method is effective. Moreover, the abundance and evenness of beneficial bacteria in the rhizosphere of the ancient peony soil after prevention and control are better than those of healthy plants, thus illustrating that the present application method can effectively prevent and control the diseases and insect pests of ancient peony, establish a benign microbial ecosystem around the plant after transplantation, enhance the plant's own resistance, thereby achieving the purpose of comprehensive disease prevention and control and promoting the rejuvenation of ancient peony. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 These are the growth states of the diseased ancient peony, which is the experimental object in the performance test of this application, including: A: moldy branches; B: dead branches; C: yellow leaves; D: blighted leaves; and E: leaf spots.

[0035] Figure 2 This is the growth status of the diseased ancient peony after improved prevention and control in the performance test of this application, among which A: new buds appear on the branches; B: the branches resume growth; C: the leaves grow normally; D: the stems and leaves grow normally; E: the leaves have normal color. DETAILED DESCRIPTION

[0036] The embodiments of the present invention will be described in detail below with reference to the examples. However, those skilled in the art will understand that the following examples are only used to illustrate the present invention and should not be construed as limiting the scope of the present invention. Specific conditions not specified in the examples are carried out according to conventional conditions or conditions recommended by the manufacturer. Reagents or instruments used without indicating the manufacturer are all conventional products that can be purchased commercially.

[0037] The following is a detailed description of the specific embodiments of the present invention. It should be understood that the specific embodiments described herein are only used to illustrate and explain the present invention and are not intended to limit the present invention. Example

[0038] Example 1 This embodiment provides a method for disease prevention and rejuvenation of ancient tree peonies after transplantation, comprising the following steps: 1. The roots, stems, leaves and rhizosphere soil of different diseased plants of transplanted ancient peony were collected and identified using universal primers 16S and ITS. Genomic DNA was extracted using the TIANGEN extraction kit. The PCR products were purified and sequenced. The obtained OTUs were identified using the microbial classification database. The microbial flora of different diseased plants and rhizosphere soil were different, and statistical analysis of the data was performed.

[0039] 2. Based on the statistical growth of detected microorganisms, determine the zero point where microorganisms exert harmful effects. The disease often occurs in summer, because high temperature and high humidity are more conducive to the harmful effects of microbial flora. Combined with summer temperature, humidity, light and pH testing, physical measures for prevention and control of the environment are carried out, as follows: a. Soil improvement: The original soil is weakly alkaline. Use vermiculite + perlite + peat soil + sandy soil in a ratio of 1:1:2:3 to bury the roots of the ancient peony. Add humic acid conditioners to adjust the soil pH to weakly alkaline and increase soil permeability.

[0040] b. Improvement of microenvironment: Provide semi-shade treatment above the ancient peony in summer, spray foliar fertilizer to promote growth in advance, water thoroughly 2-3 times in June depending on the specific situation, and generally do not water during the rainy season from July to August. Add drainage ditches along the edge of the rhizosphere, and add 4-6 80cm drainage pipes to the root gaps in the rhizosphere soil to increase water permeability, accelerate the evaporation of water in the soil, and prevent water accumulation.

[0041] c. Pruning: Prune off dead branches, rotten leaves, diseased branches, weak branches, and horizontal branches in a timely manner, and carry out anaerobic carbonization and incineration to prevent the spread of diseases to healthy plants and avoid useless branches consuming nutrients. Flower buds should not be damaged during pruning, and stronger and healthier branches should be retained as much as possible.

[0042] (3) Based on the detected types of microorganisms causing the disease and the characteristics of the ancient peony disease, during the growth of the plant, chemical control is carried out by repeatedly irrigating the roots and applying biological fertilizers to inhibit harmful bacteria. This is done once every week, and biological fertilizers are applied once after each root irrigation treatment: a. First treatment: Use 800 times diluted 40% carbendazim suspension to evenly irrigate the roots for disinfection. b. Second treatment: For plants infected with scab, ring rot, or anthracnose, use 50% carbendazim wettable powder + 70% mancozeb wettable powder for root irrigation. c. The third treatment is root irrigation: use a 500-fold dilution of 50% thiophanate-methyl wettable powder, mixed with Jiamei Bonus and benomyl to prevent and control root and soil diseases; d. The fourth treatment was root irrigation: 700 times 3% Zhongshengmycin + 1000 times 20% copper rosinate was used for root irrigation, and 800 times 80% carbendazim + 1500 times 25% azoxystrobin + 1000 times 15% pyraclostrobin was used for spraying branches and leaves; e. The fifth treatment was root irrigation: 700 times 3% Zhongshengmycin + 1500 times 15% pyraclostrobin was used for root irrigation, and 800 times 50% methyltroponin + 1500 times 25% azoxystrobin was used for spraying branches and leaves.

[0043] (4) Then, biological control is carried out by applying microbial agents and fertilizers containing beneficial bacteria to the plants to form a beneficial bacterial community effect, specifically including: a. AM fertilizer is directly inoculated and applied to the roots of the century-old peony; b. After diluting the EM solution 350 times, add brown sugar at a mass ratio of 1:1 and apply it three times in a row, with a one-week interval between each application. c. Mix 500 times the EM solution with brown sugar (mass ratio of 1:1) and 1000 times the Trichoderma harzianum solution with brown sugar (mass ratio of 1:1) and apply it twice in a row, with a one-week interval between each application. d. 500 times EM agent mixed with brown sugar (mass ratio 1:1) 1 + 1000 times Trichoderma hardis agent mixed with brown sugar (mass ratio 1:1) + 800 times Bacillus subtilis and brown sugar (mass ratio 1:1), apply continuously for 3 times, each time with an interval of 1 week.

[0044] Example 2 The difference between this embodiment and embodiment 1 is that in step (3), when performing root irrigation treatment, the first root irrigation treatment uses 600 times solution of 75% chlorothalonil wettable powder combined with 1000 times solution of 25% thiamethoxam to perform uniform root irrigation disinfection treatment.

[0045] Example 3 The difference between this embodiment and embodiment 1 is that in step (3), when the root irrigation treatment is performed, the second root irrigation treatment is performed using 70% thiophanate-methyl wettable powder + 65% mancozeb wettable powder.

[0046] Example 4 The difference between this embodiment and embodiment 1 is that in step (3), when the root irrigation treatment is performed, the third root irrigation treatment is performed using a 500-fold solution of 70% mancozeb wettable powder mixed with Jiamei Bonus.

[0047] Example 5 The difference between this embodiment and embodiment 1 is that in step (4), the method of applying the bacterial agent for the last time is: using a mixture of 500 times EM bacterial agent and Bacillus subtilis, and using it continuously for 3 times, with an interval of 1 week between each application.

[0048] Example 6 This embodiment provides a method for disease prevention and rejuvenation of ancient ginkgo after transplantation, comprising the following steps: 1. The roots, stems, leaves and rhizosphere soil of different diseased plants of transplanted ancient ginkgo were collected and identified using universal primers 16S and ITS. Genomic DNA was extracted using the TIANGEN extraction kit, and the PCR products were purified and sequenced. The obtained OTUs were identified using the microbial classification database. The microbial flora of different diseased plants and rhizosphere soil were different, and statistical analysis of the data was performed.

[0049] Based on the statistical growth of microorganisms detected, the zero point at which microorganisms exert harmful effects is determined. The disease mostly occurs in summer because high temperature and high humidity are more conducive to the harmful effects of microbial flora. Combined with summer temperature, humidity, light and pH detection, physical measures of the environment are taken to prevent and control the disease.

[0050] (3) According to the detected types of microorganisms causing the disease and the characteristics of the ancient peony disease, during the growth of the plant, chemical control is carried out to inhibit harmful bacteria by multiple root irrigation treatments and application of biological fertilizers. This is done once every week, and biological fertilizers are applied once after each root irrigation treatment.

[0051] (4) Biological control is carried out by applying bacterial agents and fertilizers containing beneficial bacteria to the plants. EM compound bacterial agent + Trichoderma agent + Bacillus subtilis agent + AM bacterial fertilizer are selected to form a beneficial bacterial community effect.

[0052] Performance testing Detection method / test method The method provided in Example 1 is used to prevent and control diseases and rejuvenate diseased plants of ancient peony. 1. Experimental subjects: Diseased ancient peony plants with mildew, dead branches, yellow leaves, blight and leaf spots, such as Figure 1 shown.

[0053] 2. For the samples of the above-mentioned diseased ancient tree peonies, DNA was extracted, amplified, and sequenced to obtain OTUS. The data was identified using the microbial classification database, and the microbial flora of different diseased plants and rhizosphere soils was detected. The data were statistically analyzed. The results are shown in Table 1 (fungal results) and Table 2 (bacterial results).

[0054] Table 1 Analysis of fungal detection results of diseased ancient peony samples Table 2. Analysis of bacterial detection results of diseased ancient peony samples 3. Based on the statistical growth of the detected microorganisms, the zero point at which the microorganisms exert harmful effects was determined. The disease mostly occurs in summer, because high temperature and high humidity are more conducive to the harmful effects of microbial flora. Therefore, the growth environment of the diseased ancient peony in summer was tested, and the results are shown in Table 3: Table 3. Environmental testing data of diseased ancient tree peonies in summer (15:00) 4. The method of step (2) in Example 1 was adopted to carry out macro-environmental physical measures for prevention and control. The improved environmental detection data are shown in Table 4, which is more conducive to the growth of ancient tree peony.

[0055] Table 4. Environmental monitoring data of diseased ancient tree peonies after physical treatment in summer (15:00) 5. The diseased ancient peony was chemically controlled according to the method of step (3) in Example 1.

[0056] 6. The diseased ancient peony was biologically controlled according to the method of step (4) in Example 1.

[0057] (7) After the above method was used to prevent and control the diseased ancient peony, the growth in the second year was as follows Figure 2 At the same time, the rhizosphere soil of the diseased ancient peony was sampled again for soil microbial testing and compared with that of the healthy ancient peony. The fungi and bacteria contained in the soil are shown in Tables 5 and 6: Table 5. Analysis of fungal detection results in rhizosphere soil of improved ancient tree peony Table 6. Analysis of bacterial detection results in rhizosphere soil of improved ancient tree peony Depend on Figure 1 and Figure 2 As shown, after a series of prevention and control measures, the disease of the diseased ancient peony was significantly alleviated in the second year, most of them have been effectively cured, and their growth and development performance is normal. The rhizosphere soil of the diseased ancient peony after improvement and prevention was sampled again for soil microbial detection (with healthy ancient peony as a control). The results showed (see Table 5 and Table 6) that compared with the diseased ancient peony before improvement and prevention, the fungal species of the improved ancient peony were significantly reduced, and the proportion of harmful bacteria in the test results was significantly reduced. It can be seen that the early prevention and control method of this application is effective. Compared with the healthy ancient peony in the control group, the 11 bacteria detected in the diseased ancient peony after improvement and prevention each accounted for 8%, indicating that most of them are artificially added biological agents, and the abundance and evenness of the beneficial bacteria community of the improved diseased ancient peony are better than those of the healthy ancient peony plant soil bacteria community, which further confirms that the application is based on the microbial flora of the rhizosphere soil of the diseased plant and the growth environment. The systematic method of accurately applying physical environment regulation + chemical control + biological control plays a positive role in the rejuvenation of the ancient peony.

[0058] This specific embodiment is merely an explanation of the present application and is not a limitation of the present application. After reading this specification, those skilled in the art may make non-creative modifications to the present embodiment as needed, but as long as they are within the scope of the claims of the present application, they are protected by the patent law.

Claims

1. A method for disease prevention and rejuvenation of ancient and famous trees after transplantation, characterized in that: It includes: The tissues and rhizosphere soil of diseased ancient and famous trees were collected, DNA was extracted and the amplified products were sequenced. The types and numbers of OTUs in the diseased plants and rhizosphere soil were counted, and the composition and diversity of the microbial flora were analyzed. Based on the growth of microbial flora, the critical point at which microorganisms exert harmful effects is determined. Combined with the current growth environment and pH test results, physical environmental prevention and control measures are implemented to improve the environment and make it suitable for the growth of ancient and famous trees. Based on the detected types of disease-causing microorganisms and the characteristics of the diseases of ancient and famous trees, chemical control is carried out during the growth of the plants through multiple root irrigation treatments and application of biological fertilizers to inhibit harmful bacteria; Biological control is carried out by applying bacterial agents and fertilizers containing beneficial bacteria to plants to form a beneficial bacterial community effect.

2. The method for disease prevention and rejuvenation of ancient and famous trees after transplantation according to claim 1, characterized in that: The ancient trees and famous trees include ancient peonies, ancient cypresses, ancient pines, and ancient ginkgo trees.

3. The method for disease prevention and rejuvenation of ancient and famous trees after transplantation according to claim 1, characterized in that: In step (1), the method for obtaining the composition and diversity of the microbial flora includes: Tissues and rhizosphere soil of diseased plants of ancient and famous trees were taken as samples, and genomic DNA was extracted. The extracted DNA was amplified by PCR using universal primers 16S and ITS. The amplified products were purified and sequenced. The sequence data obtained by sequencing were clustered according to similarity and divided into different OTUs. The OTUs were identified and the classification information was determined. The composition and diversity of the microbial flora were analyzed based on the types and numbers of the OTUs.

4. The method for disease prevention and rejuvenation of ancient and famous trees after transplantation according to claim 1, characterized in that: In step (2), the macro-environmental physical measures prevention and control method includes: Bury the roots of the plants with a composite material containing vermiculite, perlite, peat soil and sand, and add humic acid conditioners to adjust the soil pH to a weak alkaline level and increase soil permeability; During the growth of the plants, spray leaf fertilizer, increase soil permeability by watering and adding drainage pipes to ensure that the soil is not short of water or has water accumulation; Prune branches in time to retain healthy branches, and carry out oxygen-free carbonization incineration on the pruned branches.

5. The method for disease prevention and rejuvenation of ancient and famous trees after transplantation according to claim 1, characterized in that: In step (3), the method of chemical control during plant growth includes 4-5 root irrigation treatments, each with an interval of 5-10 days, and applying biological fertilizer once after each root irrigation treatment.

6. The method for disease prevention and rejuvenation of ancient and famous trees after transplantation according to claim 1, characterized in that: The root irrigation treatment comprises: First root irrigation treatment: Use diluted carbendazim suspension or chlorothalonil wettable powder combined with thiamethoxam to evenly irrigate the roots for disinfection; Second root irrigation treatment: For plants with black spot disease, ring rot or anthracnose, use carbendazim wettable powder combined with mancozeb wettable powder, or thiophanate-methyl wettable powder combined with mancozeb wettable powder for root irrigation treatment; The third root irrigation treatment: use thiophanate-methyl wettable powder or mancozeb wettable powder, compounded with Jiamei Bonus or benomyl for root irrigation treatment to prevent and control root and soil diseases; The fourth root irrigation treatment: Zhongshengmycin and copper rosinate were mixed for root irrigation, and a mixed solution of carbendazim, azoxystrobin and pyraclostrobin was sprayed on branches and leaves; The fifth root irrigation treatment: use a mixture of zhongshengmycin and pyraclostrobin for root irrigation, and use a mixture of methylthiocarb and azoxystrobin to spray on branches and leaves.

7. The method for disease prevention and rejuvenation of ancient and famous trees after transplantation according to claim 6, characterized in that: In the fourth root irrigation treatment, the medicament used is: The root irrigation agent is a mixture of 600-800 times of 3% Zhongshengmycin and 800-1200 times of 20% copper rosinate; The agent applied to branches and leaves is a mixture of 700-900 times of 80% carbendazim, 1400-1600 times of 25% myclobutanil and 800-1000 times of 15% pyraclostrobin.

8. The method for disease prevention and rejuvenation of ancient and famous trees after transplantation according to claim 6, characterized in that: In the fifth root irrigation treatment, the medicament used is: The root irrigation agent is a mixture of 600-800 times of 3% Zhongshengmycin and 1400-1600 times of 15% pyraclostrobin; The agent applied to branches and leaves is a mixture of 700-900 times of 50% metoclopramide and 1400-1600 times of 25% myclobutanil.

9. The method for disease prevention and rejuvenation of ancient and famous trees after transplantation according to claim 1, characterized in that: The biological control in step (4) includes the combined application of EM composite microbial agent, Trichoderma agent, Bacillus subtilis agent and AM microbial fertilizer.

10. The method for disease prevention and rejuvenation of ancient and famous trees after transplantation according to claim 9, characterized in that: The biological control in step (4) includes: Use AM fertilizer to apply directly to the roots of plants; Mix 200-500 times diluted EM fungal agent with sugar substance in a mass ratio of 1:0.8-1.2 and apply; Use 400-600 times diluted EM fungus agent, 800-1200 times diluted Trichoderma harzianum fungus agent and sugar substances to mix and apply; EM fungal agent diluted 400-600 times, Trichoderma harzianum fungal agent 800-1200 times, and Bacillus subtilis 700-900 times were mixed with sugar substances and applied.

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