Dse synthetic flora for increasing yield and quality and resisting disease of fruit trees and application and application method thereof

Abstract

This invention relates to the field of microbial technology, and particularly to DSE synthetic microbial communities for increasing fruit tree yield, improving quality, and enhancing disease resistance, along with their application and application methods. The synthetic microbial communities of this application have effects on improving the quality and efficiency of various fruit trees and preventing diseases, including promoting the dry and fresh weight of grapevines, providing good protection against grape downy mildew, increasing mango yield, providing good protection against mango anthracnose, increasing the single fruit weight, diameter, and sugar content of passion fruit, providing good protection against passion fruit anthracnose, increasing the chlorophyll content, plant height, fresh weight, and dry weight of bananas, and providing good protection against banana wilt. This demonstrates that the synthetic microbial communities have a good control effect on fruit tree diseases, and field tests have proven that the control effect of the synthetic microbial communities on fruit tree diseases is stable. Furthermore, when applying the synthetic microbial communities, only one application is needed during the seedling or growth stage of the fruit tree to achieve disease prevention, making it a simple application method.

Description

Technical Field

[0001] This invention relates to the field of microbial technology, and in particular to DSE synthetic microbial communities for increasing fruit tree yield and quality and improving disease resistance, their application, and application methods. Background Technology

[0002] Guangxi Zhuang Autonomous Region is an important subtropical fruit-producing area in southern my country, with its planting scale and output of cash crops such as grapes, mangoes, and bananas ranking among the top in the country. However, while the hot and humid climate promotes industrial development, it also leads to a high incidence of diseases, making disease control a key bottleneck restricting the improvement of industry quality and efficiency. Currently, the over-reliance on chemical pesticides in production not only exacerbates the evolution of pesticide resistance in pathogens but also poses a serious threat to agricultural ecosystems and the quality and safety of agricultural products due to pesticide residues and environmental pollution. Against this backdrop, biological control strategies based on microbial agents have become a key research direction for replacing or reducing the use of chemical pesticides due to their advantages such as good environmental compatibility, diverse targets, and low likelihood of inducing resistance.

[0003] Biocontrol fungi, as one of the core resources of microbial agents, can effectively inhibit the infection and spread of pathogens through mechanisms such as competition, parasitism, antagonism, and induction of systemic resistance in plants. Targeting the disease outbreak patterns and actual control needs of characteristic fruit diseases in Guangxi, conducting research on the germplasm resources, functional evaluation, and formulation of local high-efficiency biocontrol fungi, and constructing a green control technology system based on fungal control, is an important scientific and technological support for promoting pesticide reduction.

[0004] Dark-colored septate endophytic fungi (DSE) are a newly emerging microbial resource that can colonize plant tissues, promoting the host's absorption of various nutrients, conferring excellent growth traits, and enhancing the host's resistance to diseases and pests, as well as its resilience under stress. They possess advantages such as strong colonization ability, long-lasting activity, and broad-spectrum resistance, making their development and utilization promising. Previous studies have found that DSE fungi... Acidomelania saccharicola LZ3 Ochroconis guangxiensis X22 Cladosporium chlorocephalum Strains such as LS1 can significantly promote banana growth and reduce the incidence of banana wilt disease. They can also effectively induce increased activity of related antioxidant enzymes in banana plants, significantly enrich differentially expressed genes in the biosynthesis of phenylpropanoids, plant-pathogen interactions, and the biosynthetic pathways of phenylalanine, tyrosine, and tryptophan. Furthermore, they significantly increase the content of lignin, phenylalanine ammonia-lyase, chitinase, and malondialdehyde in banana plants, thereby forming a physical barrier against pathogen invasion. Studies have shown that synthetic bacterial communities are significantly superior to single strains in terms of efficacy and stability, especially in the later stages of disease control. However, because different strains have different disease control mechanisms, mixed use may result in synergistic or detrimental effects. Therefore, it is necessary to combine different strains to identify synthetic bacterial communities with complementary advantages, synergistic effects, and stable efficacy. Summary of the Invention

[0005] In view of the above, it is necessary to identify synthetic microbial communities that complement each other's strengths, synergistically enhance each other's effects, and have stable effects, so as to achieve biological control that has good preventive effect against a variety of common diseases of fruit trees, reduce the incidence of fruit tree diseases, reduce the use of chemical pesticides, improve fruit tree yield and quality, reduce costs and increase efficiency, and effectively achieve a dual improvement in economic and social benefits.

[0006] DSE-synthesizing microbial community for increasing fruit tree yield, improving quality, and enhancing disease resistance, wherein the DSE-synthesizing microbial community is composed of Cladosporium ( Cladophialophora immunda LC3, *Cladosporium guilloché* (Guangxi) Cladophialophora guangxiense ) HX2, Acidospora saccharum ( Acidomelania saccharicola LZ3 and Guangxi ochre mold ( Ochroconis guangxiensis It consists of 22 x 22.

[0007] The Cladosporium ( Cladophialophora immunda LC3, its classification name is: Cladophialophora immunda The Chinese classification name is: Cladosporium, the accession number is CGMCC NO. 41540; the depositary institution is: China General Microbiological Culture Collection Center; the deposit address is: No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing; the deposit date is: October 12, 2024.

[0008] The Guangxi Cladosporium ( Cladophialophora guangxiense HX2, its classification name is: Cladophialophora guangxiense The Chinese classification name is: *Cyclopyrum guangxiense*, the accession number is CGMCC NO.41498, the depositary institution is: China General Microbiological Culture Collection Center; the deposit address is: No.3, No.1 Beichen West Road, Chaoyang District, Beijing; the deposit date is: September 5, 2024.

[0009] The sucrose dark mold ( Acidomelania saccharicola LZ3, its category name is: Acidomelania saccharicola The accession number is CGMCC NO.18805, the depositary institution is the China General Microbiological Culture Collection Center, the depositary address is No.3, No.1 Beichen West Road, Chaoyang District, Beijing, and the deposit date is October 25, 2019.

[0010] The Guangxi ochre mold ( Ochroconis guangxiensis X22, its classification name is: Ochroconis guangxiensisThe Chinese classification name is: Guangxi Ochre, the accession number is CGMCC NO. 19656, the depositary institution is: China General Microbiological Culture Collection Center, the depositary address is: No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, and the deposit date is: June 8, 2020.

[0011] Furthermore, the DSE-synthesizing microbial community includes Cladosporium ( Cladophialophora immunda LC3, *Cladosporium guilloché* (Guangxi) Cladophialophora guangxiense ) HX2, Acidospora saccharum ( Acidomelania saccharicola LZ3 and Guangxi ochre mold ( Ochroconis guangxiensis The mycelial mass ratio of X22 is 1:1:1:1.

[0012] The present invention also includes the application of the DSE synthetic microbial community in the preparation of fruit tree biopesticides.

[0013] Furthermore, the fruit tree diseases controlled by the aforementioned biological pesticides are: grape downy mildew, mango anthracnose, passion fruit anthracnose, and / or banana wilt.

[0014] This invention also includes the application of the DSE synthetic microbial community in improving grape dry weight, grape fresh weight, mango yield, passion fruit single fruit weight, passion fruit diameter, passion fruit sugar content, banana chlorophyll content, banana plant height, banana fresh weight and / or banana dry weight.

[0015] This invention also includes a method for applying the DSE-synthetic microbial community, wherein the planting method is as follows: the strains of the DSE-synthetic microbial community are cultured separately on PDA plates and then transferred to PDB medium for shaking culture. The mycelium is collected, and the mycelium is mixed and prepared into a 5×10⁻⁶ solution. 5 A synthetic bacterial culture solution is obtained by dissolving bacteria at CFU / mL; then the synthetic bacterial culture solution is applied to the fruit trees.

[0016] Furthermore, the fruit tree is a grape or a mango, and the application method is to spray the grape or mango plants with the synthetic bacterial solution.

[0017] Furthermore, the fruit tree is passion fruit, and the application method is as follows: one week before transplanting the passion fruit, the passion fruit plants are soaked in a synthetic microbial solution.

[0018] Furthermore, the fruit tree is a banana, and the application method is as follows: the synthetic microbial solution is irrigated at the roots of the banana plant.

[0019] The present invention has the following beneficial effects.

[0020] 1. The DSE synthetic microbial community of this invention has good effects on increasing yield and improving quality, as well as disease resistance in fruit trees. Through repeated experiments, we found that the synthetic microbial community of this application is the most stable in fruit tree cultivation. Experiments on different petri dishes and pots revealed that the synthetic microbial community of this application has effects on improving quality and efficiency, and preventing diseases in various fruit trees: it promotes both the dry and fresh weight of grapevines and has good control effects against downy mildew in grapes, with a control effect of 65.8% in petri dishes and 56.2% in the field; it increases the yield of mangoes and has good control effects against anthracnose in mango fruits, with a control effect as high as 60.2% 10 days after harvest, 47.5% 12 days after harvest, and 4% 16 days after harvest. The synthetic microbial community exhibits a 6.8% effect; it increases the single fruit weight, diameter, and sugar content of passion fruit, and provides excellent control over passion fruit anthracnose, with a control efficacy as high as 86.9%. It also increases the chlorophyll content, plant height, fresh weight, and dry weight of bananas, and provides good control over banana wilt disease, with a leaf control efficacy as high as 57.1% and a bulb control efficacy of 50.6%. This demonstrates that the synthetic microbial community reduces fruit tree diseases, decreases the use of chemical pesticides, and improves fruit yield and quality, resulting in significant cost savings and increased efficiency, effectively achieving both economic and social benefits. Furthermore, the synthetic microbial community only requires one application during the seedling or growth stage of the fruit tree to achieve disease prevention, making it a simple application method. Attached Figure Description

[0021] Figure 1 This is a schematic diagram of the synthetic bacterial community after mixing.

[0022] Figure 2 The diagram shows the resistance effect of synthetic microbial communities on mango anthracnose. Among them, A represents the control group (CK) treated mangoes on day 10 post-harvest, B represents the experimental group treated mangoes with synthetic microbial communities on day 10 post-harvest, C represents the control group (CK) treated mangoes on day 16 post-harvest, and D represents the experimental group treated mangoes with synthetic microbial communities on day 16 post-harvest.

[0023] Figure 3 The figure shows the effect of synthetic microbial communities on the quality of passion fruit; where A represents passion fruit treated with the control group (CK) on day 7 post-harvest, and B represents passion fruit treated with synthetic microbial communities in the experimental group on day 7 post-harvest.

[0024] Figure 4 The image shows the disease resistance effect of the synthesized microbial community on passion fruit plants with anthracnose. In the image, A represents passion fruit leaves treated with the control group (CK) on the day of harvest, and B represents passion fruit leaves treated with the synthesized microbial community on the day of harvest.

[0025] Figure 5 The figure shows the experimental results of the synthetic microbial community on banana potted plants; where A is the blank group (CK) and B is the experimental group of banana potted plants treated with the synthetic microbial community.

[0026] Figure 6 The figure shows the experimental results of the synthetic microbial community on the root longitudinal section of banana bonsai; where A is the control group (CK) treatment and B is the experimental group synthetic microbial community treatment.

[0027] Information on the preservation of biological materials.

[0028] The strain preservation information for LC3 of this application is: Cladosporium ( Cladophialophora immunda LC3, its classification name is: Cladophialophora immunda The Chinese classification name is: Cladosporium, the accession number is CGMCC NO.41540, the depositary institution is: China General Microbiological Culture Collection Center, the deposit address is: No.3, No.1 Beichen West Road, Chaoyang District, Beijing, and the deposit date is: October 12, 2024.

[0029] The strain preservation information for HX2 in this application is: *Cladosporium guangxiense* (… Cladophialophora guangxiense HX2, its classification name is: Cladophialophora guangxiense The Chinese classification name is: *Cyclopyrum guangxiense*, the accession number is CGMCC NO. 41498, the depositary institution is: China General Microbiological Culture Collection Center; the deposit address is: No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing; the deposit date is: September 5, 2024.

[0030] The strain preservation information for LZ3 in this application is: *Dark mold sucrose* (… Acidomelania saccharicola LZ3, its classification naming Acidomelania saccharicola The Chinese classification name is: *Dictyophora sucralose*, the accession number is CGMCC NO. 18805, the depositary institution is: China General Microbiological Culture Collection Center; the deposit address is: No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing; the deposit date is: October 25, 2019.

[0031] The strain preservation information for X22 in this application is: Guangxi Ochratus ( Ochroconis guangxiensis X22, its classification name is: Ochroconis guangxiensis The Chinese classification name is: Guangxi Ochre, the accession number is CGMCC NO.19656, the depositary institution is: China General Microbiological Culture Collection Center, the deposit address is: No.3, No.1 Beichen West Road, Chaoyang District, Beijing, and the deposit date is: June 8, 2020. Detailed Implementation

[0032] All features disclosed in this specification, or all steps in all disclosed methods or processes, may be combined in any way, except for mutually exclusive features and / or steps.

[0033] Unless otherwise stated, each feature disclosed in this specification is merely one example of a series of equivalent or similar features.

[0034] Example 1.

[0035] This example studies the disease resistance effect of the DSE strain on grapes, as detailed below.

[0036] 1. Plate test of grapes.

[0037] (1) Strains treatment: Single strain X22, single strain HX2, and experimental group were combined to form a microbial community (the mycelia of strain LZ3, strain HX2, strain LC3 and strain X22 were mixed in a mass ratio of 1:1:1:1, and the effective viable count of the mixed microbial community was 5×10⁻⁶). 5 CFU / mL, the mixed mycelial solution as follows Figure 1 As shown), the control group synthesized bacterial community (composed of mycelia of strains LS1, TK731, DH16 and LC3 in a mass ratio of 1:1:1:1, with an effective viable count of 5 × 10⁻⁶). 5 CFU / mL) and blank control; among them, the strain LS1 in the synthetic flora of the control group is classified as ( Cladosporium chlorocephalum The Chinese classification name is *Cladosporium chloritidis*, with accession number CGMCC NO. 16498, and it has been published in patent application CN109652320A; the strain TK731 in the control group synthetic flora is classified as... Acremonium The genus *Sp.*, Chinese taxonomic name: *Cladosporium*; strain DH16 in the control group synthetic flora is classified as... Exophiala spinifera The genus is classified in Chinese as *Exophyllum*. Both strains DH16 and TK731 were published in the article "A Study on the Diversity and Ecological Function of Dark-colored Septate Endophytic Fungi in Two Habitats in Guangxi" (by Xie Ling).

[0038] (2) Experiment location: Institute of Plant Protection, Guangxi Academy of Agricultural Sciences.

[0039] (3) Experimental method: Strains LZ3, HX2, LC3 and X22 were selected for activation and then inoculated onto oat medium (wherein, the oat medium consisted of 1.0 g of MgSO4·7H2O, 1 0.5g KH2PO4, 1.0g NaNO3, 10g oat flour, and 11g agar powder were diluted to 1L with distilled water. Single-strain treatments were inoculated with HX2 and X22, with 4 bacterial blocks per plate. For experimental groups, 4 strains (LZ3+HX2+LC3+X22) were simultaneously inoculated onto the culture medium, with 4 bacterial blocks per plate. For the control group, 4 strains (LS1+TK731+DH16+LC3) were simultaneously inoculated onto the culture medium, with 4 bacterial blocks per plate. After 10 days of culture, grape tissue culture seedlings (variety: Thomson seedless) with uniform growth were selected and inoculated onto the colonies (1 seedling per plate). Grape tissue culture seedlings cultured on uninoculated oat medium served as blank controls. Each treatment had 5 plates, and the results were repeated 3 times. Plant growth was continuously observed during the treatment period, and growth indicators were investigated after 30 days. Later, the grape seedlings were transplanted into culture cups for further cultivation and inoculated against grape downy mildew. The preparation and inoculation treatment of the downy mildew pathogen are as follows: Collect the 3rd-5th tender leaves from new grape shoots, wash them with distilled water and blot dry with absorbent paper. Place the leaves, back side up, in a culture dish lined with moist filter paper. Inoculate the underside of the leaves with a suspension containing sporangia of the grape downy mildew pathogen. Incubate for 24 hours in an incubator under 22°C for 12 hours of light during the day and 20°C for 12 hours in darkness at night. Afterward, blot off excess bacterial solution with filter paper and continue culturing for 3-4 days until the new sporangia mature. Place the entire leaf in a centrifuge tube, centrifuge the moldy material into the tube, add pure water and shake to distribute and mix the mycelium evenly, resulting in a final concentration of 10%. 4 A sporangium suspension of 1 sporangium / mL was prepared. The suspension was sprayed evenly onto the underside of the leaves of the potted plants using a sprayer. The plants were then covered with plastic film to maintain moisture for 24 hours. Infection and disease incidence were observed 4-8 days after inoculation, and the control efficacy was calculated.

[0040] (4) Survey methods.

[0041] ① Investigate disease susceptibility: Investigate all leaves of all grapevines in the petri dish from top to bottom, recording the number of diseased leaves at each level and the total number of leaves according to the leaf grading standards. The leaf grading standards are as follows: Grade 0: No lesions; Grade 1: Lesions cover less than 5% of the total leaf area; Grade 3: Lesions cover 6%–25% of the total leaf area; Grade 5: Lesions cover 26%–50% of the total leaf area; Grade 7: Lesions cover 51%–75% of the total leaf area; Grade 9: Lesions cover more than 76% of the total leaf area. Based on the grading, calculate the grape downy mildew disease index and control efficacy. The formulas for calculating the disease index and control efficacy are as follows.

[0042] Disease index = [∑(representative value of each disease level × number of diseased leaves at that level) / (total number of leaves surveyed × highest level value)] × 100.

[0043] Prevention and control effect (%) = [(disease index of blank group - disease index of treated group) / disease index of blank group] × 100%.

[0044] ② Investigate grape growth: Remove the culture medium from the grapevine roots, clean them, weigh each replicate of grape seedlings, record the fresh weight, then place the plants in an oven at 60℃ to dry, and then investigate the dry weight. Perform variance analysis on the above data.

[0045] 2. Grape field plot experiment.

[0046] (1) Strains treatment: Single strain X22, single strain HX2, and experimental group were combined to form a microbial community (the mycelia of strain LZ3, strain HX2, strain LC3 and strain X22 were mixed in a mass ratio of 1:1:1:1, and the effective viable count of the mixed microbial community was 5×10⁻⁶). 5 CFU / mL, the mixed mycelial solution as follows Figure 1 As shown), the control group synthesized bacterial community (composed of mycelia of strains LS1, TK731, DH16 and LC3 in a mass ratio of 1:1:1:1, with an effective viable count of 5 × 10⁻⁶). 5 (CFU / mL) and blank control.

[0047] (2) Test location: Anyang Town, Du'an Yao Autonomous County, Hechi City, Guangxi Zhuang Autonomous Region.

[0048] (3) Experimental design: Strains LZ3, HX2, LC3, X22, LS1, TK731 and DH16 were selected and cultured on PDA plates for about 10 days, then transferred to PDB medium and cultured with shaking (28℃, 120 r / min) for 7~14 days. The mycelium was collected by filtration with sterile gauze and rinsed several times with sterile water. HX2 and X22 were homogenized and crushed to prepare single bacterial suspensions.

[0049] The experimental group treatment involved mixing four types of mycelia in a mass ratio of 1:1:1:1, homogenizing them into a mixed mycelial suspension, and preparing 5×10⁻⁶ solutions. 5 Bacterial solution with CFU / mL.

[0050] The control group synthetic microbial treatment involved mixing four types of mycelia in a mass ratio of 1:1:1:1, homogenizing them into a mixed microbial suspension, and preparing 5×10⁻⁶ solutions. 5 Bacterial solution with CFU / mL.

[0051] Treatment with strain HX2: HX2 mycelium was prepared into 5×105 Bacterial solution with CFU / mL.

[0052] Treatment with strain X22: X22 mycelium was prepared into 5×10 5 Bacterial solution with CFU / mL.

[0053] The control group (CK) was a treatment that involved conventional fertilization but no fungicide application.

[0054] The application method was as follows: Foliar spraying was carried out on grape seedlings in the field during the grape berry growth period (May 22, 2025). Each treatment consisted of 3 plots, with 5 plants per plot. A second spraying was carried out on June 12, 2025. The incidence of downy mildew in the field was observed, and the disease situation was investigated promptly to calculate the control efficacy.

[0055] (4) Investigation method: The disease index was investigated and the prevention efficacy was calculated by referring to the grape plate test in step 1. The results are shown in Table 1.

[0056]

[0057] Note: In the table, different lowercase letters indicate significant differences in the same column of data (p<0.05), and the same lowercase letters indicate no significant differences in the same column of data (p>0.05). Different uppercase letters indicate extremely significant differences in the same column of data (p<0.01), and the same uppercase letters indicate no significant differences in the same column of data (p>0.01). The same applies to the following tables.

[0058] Table 1 shows that both single bacterial strains and synthetic microbial communities had significant growth-promoting effects on grapes. Compared with the control group, the increases in grape plant dry weight were as follows: strain HX2 treatment increased by 69.7%, strain X22 treatment increased by 87.9%, the synthetic microbial community treatment increased by 60.6%, and the experimental group synthetic microbial community treatment increased by 39.4%. Compared with the control group, all treatments showed significant control effects against downy mildew. The treatments with the highest control efficacy in both the plate test and the field test were the experimental group synthetic microbial community, at 65.8% and 56.1%, respectively. Other treatments did not show as significant control effects against downy mildew as the experimental group synthetic microbial community. Among them, the plate control efficacy of strain HX2 treatment was 42.6%. However, the control efficacy in the field was ineffective. The plate control efficacy of strain X22 was as high as 61.4%, while the field control efficacy was only 25.6%, which was significantly lower than that of the synthetic microorganism in the experimental group. This indicates that the single strain has high resistance to downy mildew in the plate test, but when applied in the field, the disease resistance effect is unstable due to changes in the cultivation environment, and there may be no control effect. In addition, the comparison of the plate control efficacy and field control efficacy of the synthetic microorganism in the control group and the experimental group shows that the control efficacy stability of the synthetic microorganism against downy mildew varies among different strains. The control effect in the plate test and the field control effect are significantly different. The test results show that the synthetic microorganism of this application has a good resistance to downy mildew in grapes.

[0059] Example 2.

[0060] This embodiment studies the effects of the DSE strain on mango quality improvement, yield increase, and disease resistance, as detailed below.

[0061] 1. Strains Treatment: The experimental group was composed of mycelia of strains LZ3, HX2, LC3, and X22 mixed in a mass ratio of 1:1:1:1, with an effective viable count of 5 × 10⁻⁶. 5 CFU / mL, the mixed mycelial solution as follows Figure 1 As shown), the control group synthesized bacterial community (composed of mycelia of strains LS1, TK731, DH16 and LC3 in a mass ratio of 1:1:1:1, with an effective viable count of 5 × 10⁻⁶). 5 (CFU / mL) and blank control.

[0062] 2. Test location: Baiyu Town, Tianyang District, Baise City, Guangxi Zhuang Autonomous Region.

[0063] 3. Experimental methods: Strains LZ3, HX2, LC3, X22, and XX were cultured on PDA plates for about 10 days, then transferred to PDB medium and cultured with shaking (28℃, 120r / min) for 7-14 days. The mycelium was collected by filtration through sterile gauze and rinsed several times with sterile water.

[0064] The experimental group treatment involved mixing four types of mycelia in a mass ratio of 1:1:1:1, homogenizing them into a mixed mycelial suspension, and preparing 5×10⁻⁶ solutions. 5 CFU / mL bacterial culture.

[0065] The control group synthetic microbial treatment involved mixing four types of mycelia in a mass ratio of 1:1:1:1, homogenizing them into a mixed microbial suspension, and preparing 5×10⁻⁶ solutions. 5 CFU / mL bacterial culture.

[0066] The control group (CK) was a treatment that involved conventional fertilization but no fungicide application.

[0067] The application method is as follows: During the fruit enlargement period (June 9, 2025), foliar spraying is carried out on mango plants in the field. Each treatment area covers 1 acre, and only one spraying treatment is performed. Fruits are harvested on July 5, 2025 when they mature, and disease incidence is investigated in a timely manner to calculate the control effect.

[0068] 4. Survey methods.

[0069] (1) Investigation of anthracnose susceptibility: Mangoes were stored at room temperature. The investigation of anthracnose susceptibility during mango storage was conducted at 10, 12, and 16 days after storage. Investigation method: The number of diseased fruits at each grade was investigated based on the total number of fruits in each replicate treatment. The incidence of anthracnose was investigated in accordance with GB / T17980.99-2004 "Guidelines for Field Efficacy Trials of Pesticides (II) Part 99: Control of Mango Anthracnose During Storage with Fungicides"; the grading standards for anthracnose are as follows: Grade 0: No symptoms; Grade 1: Lesion area accounts for less than 5% of the fruit area; Grade 3: Lesion area accounts for 6-15% of the fruit area; Grade 5: Lesion area accounts for 16-25% of the fruit area; Grade 7: Lesion area accounts for 26-50% of the fruit area; Grade 9: Lesion area accounts for more than 51% of the fruit area. The anthracnose disease severity index and control efficacy were calculated based on the grading. The calculation formulas for the severity index and control efficacy are as follows.

[0070] Disease index = [∑(representative value of each disease level × number of fruits with disease at that level) / (total number of fruits surveyed × highest level value)] × 100.

[0071] Prevention and control effect (%) = [(disease index of blank group - disease index of treated group) / disease index of blank group] × 100%.

[0072] (2) Mango quality survey: After harvesting, the yield of mangoes was investigated by plot (weighed using a balance); the Brix value was investigated according to NY / T 2637-2014 "Determination of Soluble Solids Content in Fruits and Vegetables - Refractometer Method" issued by the Ministry of Agriculture and Rural Affairs: The mangoes were peeled and pitted, and the pulp was squeezed out with gauze or homogenized and filtered to obtain pure juice. A handheld digital saccharimeter (Brix meter) was calibrated with distilled water. 2-3 drops of juice were dropped onto the prism surface, the cover was closed, and the value was allowed to stand for a few seconds until it stabilized before reading the °Brix value (percentage). The results are shown in Table 2 and Figure 2 As shown.

[0073]

[0074] As shown in Table 2, in terms of yield increase rate, the synthetic microbial community in the control group had a significantly higher yield increase rate than the synthetic microbial community in the experimental group. In terms of post-harvest tribulation (12 and 17 days), there were no significant differences between the synthetic microbial community treatment in the control group, the synthetic microbial community treatment in the experimental group, and the control group (CK). At 10 days post-harvest, the control efficacy of the synthetic microbial community treatment in the experimental group reached 60.2%, while that in the control group was only 23.2%. At 12 days post-harvest, the control efficacy of the synthetic microbial community treatment in the experimental group reached 47.5%, while the synthetic microbial community in the control group had no control effect on mangoes. At 16 days post-harvest, the control efficacy of the synthetic microbial community treatment in the experimental group still reached 46.8%. This indicates that the synthetic microbial community treatment in the experimental group had a good resistance to mango anthracnose and maintained a high level of control efficacy even at 16 days post-harvest.

[0075] The effect of synthetic microbial flora on the resistance of mango anthracnose is as follows: Figure 2 As shown, Figure 2 In the diagram, A represents the control group (CK) treated mangoes on day 10 post-harvest, B represents the experimental group treated mangoes with microbial communities on day 10 post-harvest, C represents the control group (CK) treated mangoes on day 16 post-harvest, and D represents the experimental group treated mangoes with microbial communities on day 16 post-harvest. Figure 2 As shown in Figure A, on day 10 post-harvest, the control group (CK) mangoes generally showed obvious black spots, lesions, and browning of the peel; from Figure 2 As shown in Figure B, the mangoes treated with the bacterial flora in the experimental group had brighter, more uniformly colored peels with almost no obvious lesions, and their overall health was far superior to that of the control group; from Figure 2 As shown in Figure C, most of the mangoes in the control group (CK) were severely rotten, with large areas of mold and softness, and the lesions were black and gray, almost all of them losing their commercial value. Figure 2 As shown in Figure D, the mangoes treated with the combined microbial community in the experiment still maintained a good appearance, with no serious rot or mold. Only a few fruits had slight marks, and the preservation effect was significant. The experimental results show that the synthetic microbial community of this application has a good effect on improving the quality, increasing the yield, and resisting anthracnose in mangoes.

[0076] Example 3.

[0077] This embodiment studies the effects of the DSE strain on improving the quality, increasing the yield, and enhancing the disease resistance of passion fruit, as detailed below.

[0078] 1. Strains Treatment: The experimental group was composed of mycelia of strains LZ3, HX2, LC3, and X22 mixed in a mass ratio of 1:1:1:1, with an effective viable count of 5 × 10⁻⁶. 5 CFU / mL, the mixed mycelial solution as follows Figure 1 (as shown) and blank control.

[0079] 2. Test location: Wuping Town, Jingxi City, Baise City, Guangxi Zhuang Autonomous Region.

[0080] 3. Experimental methods: Strains LZ3, HX2, LC3, X22, LS1, TK731, and DH16 were cultured on PDA plates for about 10 days, then transferred to PDB medium and cultured with shaking (28℃, 120r / min) for 7-14 days. The mycelium was collected by filtration through sterile gauze and rinsed several times with sterile water.

[0081] The experimental group treatment involved mixing four types of mycelia (strain LZ3, strain HX2, strain LC3, and strain X22) in a mass ratio of 1:1:1:1, homogenizing them into a mixed bacterial suspension, and preparing 5×10⁻⁶ solutions. 5 CFU / mL bacterial culture.

[0082] The control group synthetic microbial treatment involved mixing four types of mycelia (strain LC3, strain LS1, strain TK731, and strain DH16) in a mass ratio of 1:1:1:1, homogenizing them into a mixed microbial suspension, and preparing 5×10⁻⁶ solutions. 5 CFU / mL bacterial culture.

[0083] The control group (CK) was a treatment that involved conventional fertilization but no fungicide application.

[0084] The application method was as follows: One week before transplanting passion fruit, the roots were soaked in water for 30 minutes as a control. Each treatment consisted of 3 rows, with 50 plants per row, and single plants per row. The plant spacing was 1.0 meter, and the row spacing was 2.0 meters. The root soaking treatment was only conducted once on March 13, 2025. Fruit samples were harvested on August 12, 2025, at maturity. 15 fruits were harvested per row, for a total of 45 fruits per treatment. Anthracnose incidence was investigated on the same day, control efficacy was calculated, and the weight of individual fruits and fruit stems were measured. Sugar content was measured seven days after harvesting and allowing the fruit to stand at room temperature.

[0085] 4. Survey methods.

[0086] (1) Investigation of anthracnose susceptibility in passion fruit: The investigation was conducted in accordance with the "Guidelines for Field Efficacy Experiments of Pesticides (1)" (compiled by the Institute for the Control of Pesticides, Ministry of Agriculture, 2000). The grading standards are as follows: Grade 0: no symptoms; Grade 1: lesion area less than 2% of leaf / fruit area; Grade 3: lesion area 3%-8% of leaf / fruit area; Grade 5: lesion area 9%-15% of leaf / fruit area; Grade 7: lesion area 16%-25% of leaf / fruit area; Grade 9: lesion area more than 25% of leaf / fruit area. The anthracnose disease index and control efficacy were calculated based on the grading. The calculation formulas for the disease index and control efficacy are as follows.

[0087] Disease index = [∑(representative value of each disease level × number of fruits with disease at that level) / (total number of fruits surveyed × highest level value)] × 100.

[0088] Prevention and control effect (%) = [(disease index of blank group - disease index of treated group) / disease index of blank group] × 100%.

[0089] (2) Investigation of passion fruit quality: The weight of a single fruit and the weight of the fruit stem were measured. The sugar content was measured after the fruit was harvested and left at room temperature for seven days. The results are shown in Table 3. Figure 3 and Figure 4 As shown.

[0090]

[0091] As shown in Table 3, after passion fruit seedlings were treated with DSE microbial community before transplanting, the fruit diameter of the experimental group treated with the microbial community was significantly higher than that of the control group (CK) (P<0.05), and the single fruit weight and sugar content were also higher than those of the control group (CK), but the differences were not significant (P>0.05). The control efficacy against passion fruit anthracnose was as high as 86.9%, which was significantly higher than that of the control group with synthetic microbial community (P<0.05). The single fruit weight of the control group with synthetic microbial community was lower than that of the control group (CK), and the fruit diameter was slightly higher than that of the control group (CK), but the differences were not significant (P>0.05). The sugar content was higher than that of the control group (CK), and the differences were significant (P<0.05). The control efficacy against passion fruit anthracnose was only 73.6%, which was significantly lower than that of the experimental group treated with the microbial community (P<0.05).

[0092] The effects of synthetic microbial communities on passion fruit fruit are as follows: Figure 3 As shown in the figure, A represents passion fruit treated with the control group (CK) on day 7 post-harvest, and B represents passion fruit treated with the experimental group's microbial community on day 7 post-harvest; from Figure 2 As shown in A, on day 7 post-harvest, the passion fruit in the control group (CK) was greener in color and smaller in size; from Figure 2 As shown in Figure B, on the 7th day post-harvest, the passion fruit treated with the experimental group of microbial communities had a yellowish color and the fruit size was more uniform and larger than that of the control group.

[0093] The effect of synthetic microbial flora on the resistance of passion fruit plants to anthracnose is as follows: Figure 4 As shown; where A represents passion fruit treated with the control group (CK) on the day of harvest, and B represents passion fruit leaves treated with the experimental group's microbial community on the day of harvest; from Figure 4 As shown in Figure A, on the day of harvest, the leaves of the control group (CK) passion fruit were yellowish and covered with anthracnose lesions; from Figure 4 As shown in Figure B, on the day of harvest, the passion fruit leaves treated with the combined microbial community were dark green with some anthracnose lesions scattered on them. The experimental results show that the synthetic microbial community of this application has a good effect on improving the quality, increasing the yield and resisting anthracnose in passion fruit.

[0094] Example 4.

[0095] This embodiment studies the growth-promoting and disease-resistant effects of the DSE strain on bananas, as detailed below.

[0096] 1. Strains Treatment: Single strains X22, HX2, and LZ3 were combined to form a bacterial colony (the mycelia of strains LZ3, HX2, LC3, and X22 were mixed in a mass ratio of 1:1:1:1, and the effective viable count of the mixed colony was 5 × 10⁻⁶). 5 CFU / mL, the mixed mycelial solution as follows Figure 1 As shown), the control group synthesized bacterial community (composed of mycelia of strains LS1, TK731, DH16 and LC3 in a mass ratio of 1:1:1:1, with an effective viable count of 5 × 10⁻⁶). 5 (CFU / mL) and blank control.

[0097] 2. Experiment location: Institute of Plant Protection, Guangxi Academy of Agricultural Sciences.

[0098] 3. Experimental design: Strains LZ3, HX2, LC3, X22, LS1, TK731 and DH16 were cultured on PDA plates for about 10 days, then transferred to PDB medium and cultured with shaking (28℃, 120r / min) for 7-14 days. The mycelium was collected by filtration with sterile gauze and rinsed several times with sterile water. HX2 and X22 were homogenized and broken up to prepare single-strain suspensions.

[0099] The experimental group treatment involved mixing four types of mycelia in a mass ratio of 1:1:1:1, homogenizing them into a mixed mycelial suspension, and preparing 5×10⁻⁶ solutions. 5 CFU / mL bacterial culture.

[0100] The control group synthetic microbial treatment involved mixing four types of mycelia in a mass ratio of 1:1:1:1, homogenizing them into a mixed microbial suspension, and preparing 5×10⁻⁶ solutions. 5CFU / mL bacterial culture.

[0101] Treatment with strain HX2: HX2 mycelium was prepared into 5×10 5 CFU / mL bacterial culture.

[0102] Treatment with strain X22: X22 mycelium was prepared into 5×10 5 CFU / mL bacterial culture.

[0103] Treatment with strain LZ3: LZ3 mycelium was prepared into 5×10 5 CFU / mL bacterial culture.

[0104] The control group (CK) was a treatment that involved conventional fertilization but no fungicide application.

[0105] The application method was as follows: Sterilized substrate was mixed with field soil at a 1:1 ratio and placed in 8cm×8cm seedling cups. One banana seedling with 4-5 leaves and uniform growth was planted in each cup, with 5 seedlings per treatment, replicated 3 times. The cups were placed in an artificial climate chamber for cultivation. Ten days after transplanting, the seedlings were irrigated with the bacterial solution (February 11, 2026), with 50mL of solution applied to each seedling's roots. Before irrigation, several small holes were made around the roots with a bamboo stick to facilitate bacterial solution penetration. A control group was treated with 50mL of water. On March 6, 2026, 1×10⁶ seedlings were inoculated. 6 CFU / mL banana wilt pathogen spore suspension, 50mL per seedling.

[0106] 4. Survey methods.

[0107] (1) Observe the incidence of banana wilt disease: The disease incidence of banana wilt disease is observed. The disease grading standards for the above-ground parts (leaves) are as follows: Grade 0: No symptoms on leaves; Grade 1: 1-2 lower leaves are yellowed, or the yellowed area is ≤25%; Grade 3: 3-4 leaves are yellowed, or 25%≤yellowing area≤50%; Grade 5: The outer leaves are yellowed, the yellowed area is >50%, and the leaves are wilted; Grade 7: Most leaves are yellowed and wilted, only the leaf heart is normal; Grade 9: The plant wilts, dies. The disease grading standards for the bulbs are as follows: Grade 0: The bulbs are healthy and have no discoloration; Grade 1: The discolored area of ​​the bulb is <20% of the bulb area; Grade 2: 20% < the discolored area of ​​the bulb is <40% of the bulb area; Grade 3: 40% < the discolored area of ​​the bulb is <60% of the bulb area; Grade 4: 60% < the discolored area of ​​the bulb is <80% of the bulb area; Grade 5: The discolored area of ​​the bulb is >80% of the bulb area. The prevention and control efficacy is calculated based on the grading standards. The formulas for calculating the disease index and the prevention and control effect are as follows.

[0108] Disease index = [∑(number of diseased plants × representative value) / (total number of plants × highest disease level representative value)] × 100.

[0109] Prevention and control effect (%) = [(disease index of blank group - disease index of treated group) / disease index of blank group] × 100%.

[0110] (2) Investigation of banana quality: Observe the growth and disease incidence of banana seedlings in each treatment group. After 30 days, measure plant height, stem diameter and fresh weight. The results are shown in Table 4 and Figure 5 , Figure 6 .

[0111]

[0112] Table 4 shows that the chlorophyll content of single strain X22, single strain HX2, experimental group combined microbial community, and control group combined microbial community was higher than that of the blank group (CK) (P<0.05), while the chlorophyll content of single strain X22 was lower than that of the blank group (CK). The chlorophyll content of single strain HX2 and experimental group combined microbial community was significantly higher than that of the blank group (CK) (P<0.05). The plant height of other treatment groups was not significantly different from that of the blank group (CK) (P>0.05). The plant height of single strain X22, single strain HX2, single strain LZ3, experimental group combined microbial community, and control group combined microbial community was higher than that of the blank group (CK). Except for the plant height of the experimental group combined microbial community treatment, which was significantly higher than that of the blank group (CK) (P<0.05), the plant height of other treatments was not significantly different from that of the blank group (CK) (P>0.05). The fresh weight of the combined microbial groups was higher than that of the control group (CK). Specifically, the fresh weight of the HX2 treatment group, the combined microbial group treatment, and the synthetic microbial group treatment in the control group were significantly higher than that of the control group (CK) (P<0.05). The fresh weight of other treatments did not differ significantly from that of the control group (CK) (P>0.05). Regarding the leaf control efficacy against banana wilt disease: the combined microbial group in the experimental group > single strain LZ3 > single strain X22 > synthetic microbial group in the control group > single strain HX2. The bulb control efficacy of the combined microbial group in the experimental group was significantly higher than that of other treatment groups (P<0.05). Regarding the bulb control efficacy against banana wilt disease: the combined microbial group in the experimental group > synthetic microbial group in the control group > single strain LZ3 > single strain X22 > single strain HX2. The bulb control efficacy of the combined microbial group in the experimental group was significantly higher than that of other treatment groups (P<0.05). Therefore, our combined microbial group in the experimental group has a good control effect against banana wilt disease.

[0113] The experimental results of synthetic microbial communities on potted banana plants are as follows: Figure 5 As shown, Figure 5 In the figure, A represents the banana potted plants treated with the control group (CK), and B represents the banana potted plants treated with the experimental group's microbial community. As can be seen from the figure, the banana potted seedlings inoculated with the experimental group's microbial community showed significantly higher growth than the control group (CK).

[0114] The experimental results of synthetic microbial communities on the longitudinal section of the roots of potted banana plants are as follows: Figure 6 As shown, Figure 6 In the figure, A represents the control group (CK) and B represents the experimental group of synthesized microbial communities. As can be seen from the figure, the banana roots inoculated with the experimental group of synthesized microbial communities were thicker and more vigorous than those in the uninoculated banana roots. The experimental results show that the synthetic microbial communities of this application have good effects on improving the quality, increasing the yield and resisting Fusarium wilt in bananas.

[0115] In summary, the synthetic microbial community of this application (composed of strains LZ3, HX2, LC3, and X22) has the effects of improving quality and yield and preventing diseases in various fruit trees: it promotes both the dry and fresh weight of grapevines and has good control efficacy against downy mildew, with a plate control efficacy of 65.8% and a field control efficacy of 56.1%; it increases the yield of mangoes and has good control efficacy against anthracnose in mango fruits, with a control efficacy of up to 60.2% 10 days after harvest, 47.5% 12 days after harvest, and [missing data] 16 days after harvest. The efficacy reached 46.8%; it improved the single fruit weight, diameter, and sugar content of passion fruit, and had a good control effect on passion fruit anthracnose, with a control effect as high as 86.9%; it improved the chlorophyll content, plant height, fresh weight, and dry weight of bananas, and had a good control effect on banana wilt disease, with a leaf control effect as high as 57.2% and a bulb control effect as high as 54.3%; thus, it is shown that the synthetic microbial community of this application has a good control effect on fruit tree diseases, and the plate, pot, and field tests have proved that the synthetic microbial community has a stable effect on the control of fruit tree diseases.

[0116] The embodiments described above are merely illustrative of several implementations of the present invention, and while the descriptions are specific and detailed, they should not be construed as limiting the scope of the invention. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these modifications and improvements all fall within the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be determined by the appended claims.

Claims

1. The application of DSE synthetic microbial communities in the control of grape downy mildew, mango anthracnose, and passion fruit anthracnose, characterized in that, The DSE-synthetic microbial community is composed of Cladosporium (… Cladophialophora immunda LC3, *Cladosporium guilloché* ( Cladophialophora guangxiense ) HX2, Acidospora saccharum ( Acidomelania saccharicola LZ3 and Guangxi ochre mold ( Ochroconis guangxiensis X22 is composed of mycelium in a mass ratio of 1:1:1:1; The Cladosporium ( Cladophialophora immunda The accession number of LC3 is CGMCC NO. 41540; the depositary institution is the China General Microbiological Culture Collection Center; the depositary address is No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing; the deposit date is October 12, 2024. The Guangxi Cladosporium ( Cladophialophora guangxiense The accession number of HX2 is CGMCC NO.41498, the depositary institution is the China General Microbiological Culture Collection Center, the depositary address is No.3, No.1 Beichen West Road, Chaoyang District, Beijing, and the deposit date is September 5, 2024. The sucrose dark mold ( Acidomelania saccharicola LZ3 has the accession number CGMCC NO.18805, the depositary institution is the China General Microbiological Culture Collection Center, the depositary address is No.3, No.1 Beichen West Road, Chaoyang District, Beijing, and the deposit date is October 25, 2019. The Guangxi ochre mold ( Ochroconis guangxiensis X22 has the accession number CGMCC NO. 19656, the depositary institution is the China General Microbiological Culture Collection Center, the depositary address is No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, and the deposit date is June 8, 2020.

2. The application as described in claim 1, characterized in that, The DSE-synthesizing microbial community described above can increase the dry weight of grapes, the fresh weight of grapes, the yield of mangoes, the single fruit weight of passion fruit, the fruit diameter of passion fruit, and / or the sugar content of passion fruit.

3. The application as described in claim 1, characterized in that, The application method of the DSE synthetic microbial flora is as follows: the strains in the DSE synthetic microbial flora as claimed in claim 1 are respectively cultured on PDA plates, then transferred to PDB medium for shaking culture, mycelia are collected, and the mycelia are mixed to prepare a microbial liquid with a concentration of 5x10 5 CFU / mL of the synthetic microbial flora; and then the synthetic microbial flora liquid is applied to fruit trees.

4. The application as described in claim 3, characterized in that, The fruit tree is a grape or a mango, and the application method is to spray the grape or mango plants with the synthetic bacterial solution.

5. The application as described in claim 3, characterized in that, The fruit tree is passion fruit, and the application method is as follows: one week before transplanting the passion fruit, the roots of the passion fruit plant are soaked in a synthetic microbial solution.

6. The application as described in claim 3, characterized in that, The fruit tree is a banana, and the application method is to irrigate the roots of the banana plant with the synthetic microbial solution.

Images